Podcasts about G protein

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G protein

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Best podcasts about G protein

Latest podcast episodes about G protein

Vegan Podcast
Vegan High Protein - Full Day of Eating mit 150 g Protein #1069

Vegan Podcast

Play Episode Listen Later Nov 26, 2023 25:01


Video anschauen: https://youtu.be/CuEeR9EeqHsErnährung für Vegane Sportler & Full Day of Eating mit 150g vegan Eiweiß Ganzes Interview - KlickKlicke, um zu Veganen Muskelaufbau Rezepten zu kommen (High Protein vegan): https://www.vegan-athletes.com/vegan-protein-shake-test/Klicke, um alle Tipps für einen unkomplizierten Einstieg in die vegane Ernährung zu kommen: https://www.vegan-athletes.com/vegan-fuer-anfaenger/Du treibst regelmäßig Sport und fragst Dich, wie Du Deinen Proteinbedarf vegan decken kannst. Dann kommt diese Episode gerade richtig.Christian verrät Dir, auf was es bei einer veganen Ernährung ankommt und wie Du ganz leicht - ohne Proteinshakes - Deinen Proteinbedarf decken kannst. Er teilt mit Dir seine Top Lebensmittel und warum dieser Proteinhype kritisch gegenübersteht.Lies auch unsere Artikel über die die besten Proteinshakes und die Bedeutsamkeit von Aminosäuren durch.Melde Dich für den Grüne Lebenskraft Kongress an! Er findet vom 20. - 26.11.2023 statt und ist komplett kostenfrei. (klick)Du willst mehr erfahren? Schreibe eine E-Mail an: christian@christian-wenzel.comMehr mr.broccoli: Podcast auf Spotify Apple Podcast Mehr Podcast Abonniere meinen YouTube Kanal*Affiliate LinkAchtung betreffend Nahrung, Geräten und Supplements:Vorliegend habe ich meine eigene Erfahrung und die von Interviewpartnern genannt. Das sind die Effekte, die ich bei mir gespürt habe. Diese können bei jedem unterschiedlich ausfallen.Natürlich kann kein Lebensmittel, keine Nahrungsergänzung oder Superfoods sowie Inspirationen aus diesem Podcast alleine für sich eine Heilwirkung erzielen oder versprechen.Die beschriebenen Erfahrungen sind keine wissenschaftlichen Erkenntnisse und keine Tatsachenbehauptungen. Sämtliche Inhalte dieser Podcast Episoden sind keine Heilaussagen und ausschließlich informativ, sie dienen keinesfalls als Ersatz für eine ärztliche Behandlung.Die Aussagen der Interview Gäst:innen stehen für sich. Diese spiegeln nicht zwingend die Meinung des Herausgebers.

Vegan Podcast
Vegan High Protein - 50 g dieses Lebensmittel bringen 30 g Protein

Vegan Podcast

Play Episode Listen Later Nov 23, 2023 22:07


Video anschauen: https://youtu.be/CuEeR9EeqHsErnährung für Vegane Sportler & Full Day of Eating mit 150g vegan Eiweiß Ganzes Interview - KlickKlick zu Vegane Muskelaufbau Rezepte High Protein vegan: https://www.vegan-athletes.com/vegan-protein-shake-test/Du treibst regelmäßig Sport und fragst Dich, wie Du Deinen Proteinbedarf vegan decken kannst. Dann kommt diese Episode gerade richtig. Christian verrät Dir, auf was es bei einer veganen Ernährung ankommt und wie Du ganz leicht - ohne Proteinshakes - Deinen Proteinbedarf decken kannst. Er teilt mit Dir seine Top Lebensmittel und warum dieser Proteinhype kritisch gegenübersteht. Lies auch unsere Artikel über die die besten Proteinshakes und die Bedeutsamkeit von Aminosäuren durch. Melde Dich für den Grüne Lebenskraft Kongress an! Er findet vom 20. - 26.11.2023 statt und ist komplett kostenfrei. (klick)Du willst mehr erfahren? Schreibe eine E-Mail an: christian@christian-wenzel.comMehr mr.broccoli: Podcast auf Spotify Apple Podcast Mehr Podcast Abonniere meinen YouTube Kanal*Affiliate LinkAchtung betreffend Nahrung, Geräten und Supplements:Vorliegend habe ich meine eigene Erfahrung und die von Interviewpartnern genannt. Das sind die Effekte, die ich bei mir gespürt habe. Diese können bei jedem unterschiedlich ausfallen.Natürlich kann kein Lebensmittel, keine Nahrungsergänzung oder Superfoods sowie Inspirationen aus diesem Podcast alleine für sich eine Heilwirkung erzielen oder versprechen.Die beschriebenen Erfahrungen sind keine wissenschaftlichen Erkenntnisse und keine Tatsachenbehauptungen. Sämtliche Inhalte dieser Podcast Episoden sind keine Heilaussagen und ausschließlich informativ, sie dienen keinesfalls als Ersatz für eine ärztliche Behandlung.Die Aussagen der Interview Gäst:innen stehen für sich. Diese spiegeln nicht zwingend die Meinung des Herausgebers.

ChemTalk
Episode 37: Dr. Robert Lefkowitz on his Nobel Prize Winning Work on G-Protein Coupled Receptors

ChemTalk

Play Episode Listen Later Sep 27, 2023 25:44


With curiosity and creativity, there are no limits. We can be scientists. We can be doctors. We can be authors. Our renowned guest today has done all three. On this riveting episode of Let's Talk Chemistry edited by Presley Vu, hosts Yeongseo Son and Grace Go talk about their interview with Nobel Prize Laureate Dr. Robert Lefkowitz, a James B. Duke Professor of Medicine and Professor of Biochemistry and Chemistry at Duke University. He elaborates on his Nobel Prize winning work on G-protein coupled receptors. We hope you enjoy! If you like to know more about Dr. Robert Lefkowitz, you can check out his research here or reach him by email at lefko001@receptor-biol.duke.edu. Check out his book “A Funny Thing Happened on the Way to Stockholm: The Adrenaline-Fueled Adventures of an Accidental Scientist.”

PaperPlayer biorxiv neuroscience
Monomeric amyloid-β inhibits microglial inflammatory activity in the brain via an APP/heterotrimeric G protein-mediated pathway

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Jul 26, 2023


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.07.24.550398v1?rss=1 Authors: Kwon, H., Santhosh, D., Huang, Z. Abstract: Microglia, the resident immune cell of the brain, play critical roles in brain development, function, and disease. However, how microglial activity is regulated in this process remains to be elucidated. Here we report an amyloid precursor protein (APP) and heterotrimeric G protein-mediated pathway that negatively regulates microglial inflammatory activation during cerebral cortex development. Disruption of this pathway results in dysregulated microglial activity, excessive extracellular matrix proteinase production, cortical basement membrane breach, and laminar assembly disruption. We further show that this pathway is activated by amyloid {beta} (A{beta}), the cleavage product of APP that accumulates in large quantities as plaques in the Alzheimer's disease brain. Specifically, we find A{beta} monomers potently suppress inflammatory cytokine transcription and secretion by brain microglia, in an APP and heterotrimeric G protein-dependent manner. These results discover a previously unknown activity of A{beta} as a negative regulator of brain microglia as well as a new pathway that mediates the signal transduction. They shed new light on the cell-cell communication mechanisms that regulate brain immune homeostasis and may facilitate further insight into Alzheimer's disease pathogenesis. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

CRNA School Prep Academy Podcast
G Protein Pathway Part #2 With PKA: ICU Drugs & Mechanism Of Action

CRNA School Prep Academy Podcast

Play Episode Listen Later Jul 5, 2023 23:18


Mastering the complexities of the G Protein Pathway and deciphering the molecular mechanisms of PKA not only equips you to tackle challenging CRNA interview questions with confidence, but it also unveils a simplified understanding of ICU drugs. Welcome back to the second part of our series on G Protein-coupled Receptor Pathways! Today, your host Jenny Finnell takes you on an exciting and simplified journey into the intricate world of protein kinase A (PKA) and its fascinating involvement in critical care drugs. Buckle up because Jenny will be diving deep into the molecular level to uncover the mechanisms of action behind commonly used ICU drugs like epinephrine and norepinephrine. Get ready for a content-heavy episode as Jenny masterfully breaks down the complexities of PKA and its role in the G Protein Pathway. Join us in understanding how critical care drugs work and ace that CRNA interview! Get access to planning tools, mock interviews, valuable CRNA Faculty guidance, and mapped-out courses that have been proven to accelerate your CRNA success! Become a member of CRNA School Prep Academy here! https://www.crnaschoolprepacademy.com/join Book a mock interview, personal statement, resume and more at http://www.NursesTeachNurses.com Join the CSPA email list here! https://www.cspaedu.com/podcast-email Grab the Study Notes for this episode here: https://www.cspaedu.com/ldmbjald Send Jenny an email or make a podcast request!Hello@CRNASchoolPrepAcademy.com

PaperPlayer biorxiv cell biology
CaaX-motif adjacent residues control G protein prenylation under suboptimal conditions

PaperPlayer biorxiv cell biology

Play Episode Listen Later Jul 4, 2023


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.07.04.547731v1?rss=1 Authors: Tennakoon, M., Thotamune, W., Payton, J. L., Karunarathne, A. Abstract: Prenylation is a universal and irreversible post-translational modification that supports membrane interactions of proteins involved in various cellular processes, including migration, proliferation, and survival. Thus, dysregulation of prenylation contributes to multiple disorders, including cancers, vascular diseases, and neurodegenerative diseases. During prenylation, prenyltransferase enzymes tether metabolically produced isoprenoid lipids to proteins via a thioether linkage. Pharmacological inhibition of the lipid synthesis pathway by statins has long been a therapeutic approach to control hyperlipidemia. Building on our previous finding that statins inhibit membrane association of G protein {gamma} (G{gamma}) in a subtype-dependent manner, we investigated the molecular reasoning for this differential. We examined the prenylation efficacy of carboxy terminus (Ct) mutated G{gamma} in cells exposed to Fluvastatin and prenyl transferase inhibitors and monitored the subcellular localization of fluorescently tagged G{gamma} subunits and their mutants using live-cell confocal imaging. Reversible optogenetic unmasking-masking of Ct residues was used to probe their contribution to the prenylation process and membrane interactions of the prenylated proteins. Our findings suggest that specific Ct residues regulate membrane interactions of the G{gamma} polypeptide statin sensitivity and prenylation efficacy. Our results also show that a few hydrophobic and charged residues at the Ct are crucial determinants of a protein's prenylation ability, especially under suboptimal conditions. Given the cell and tissue-specific expression of different G{gamma} subtypes, our findings explain how and why statins differentially perturb heterotrimeric G protein signaling in specific cells and tissues. Our results may provide molecular reasoning for repurposing statins as Ras oncogene inhibitors and the failure of using prenyltransferase inhibitors in cancer treatment. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

New Matter: Inside the Minds of SLAS Scientists
SLAS Europe 2023 | Interviews with Tony B. Award and Student Poster Award Winners

New Matter: Inside the Minds of SLAS Scientists

Play Episode Listen Later Jun 12, 2023 25:30


Recorded straight from the SLAS Europe 2023 exhibit hall floor – New Matter host, Hannah Rosen, Ph.D., and SLAS Scientific Director Lesley Mathews, Ph.D., speaks with winners of the Tony B. Award and the Student Poster Award!Congratulations to our winners!(Listed in interview order)Tony B. Travel AwardÁdám Wolf, Ph.D. Student (Óbuda University)Towards robotic laboratory automation Plug & Play: The “LAPP” frameworkMarisa Parra López, Predoctoral Researcher (University of Granada)A multidisciplinary approach for the identification of Tsg101 UEV ligands with potential as novel broad spectrum antiviralsNatalia Karczewska, Research Biologist (Institute of Bioorganic Chemistry, Polish Academy of Sciences)Evaluation of the bioactivity of compounds using Cell Painting assay and alternative High-Content assaysStudent Poster AwardMorgan Dennis, Ph.D. Student (University of Nottingham )How Can Membrane-based Assays Revolutionise Drug Discovery for G Protein-coupled Receptors?Qian Wei, Ph.D. (Oslo University Hospital)High-Throughput Screening of Small Molecules Targeting FoxP3 in Regulatory T Cells for Cancer TreatmentWhat is the Tony B. Travel Award?This annual travel grant program provides funding for the SLAS International Conference and Exhibition and the SLAS Europe Conference and Exhibition for students, graduate students, post-doctoral associates, and junior faculty with less than four years in their first academic appointments.  What is the Student Poster Award?Student posters presented during the SLAS International Conference and Exhibition and SLAS Europe Conference and Exhibition are judged on quality and relevance. Student poster presenters are all eligible for the Student Poster Competition, in which the top three posters are recognized. Winners, up to three, receive a cash prize of $500.Visit our Student Resources to learn more about these awards.Stay connected with SLASOnline at www.slas.orgFacebookTwitter @SLAS_OrgLinkedInInstagram @slas_orgYouTubeAbout SLASSLAS (Society for Laboratory Automation and Screening) is an international professional society of academic, industry and government life sciences researchers and the developers and providers of laboratory automation technology. The SLAS mission is to bring together researchers in academia, industry and government to advance life sciences discovery and technolUpcoming SLAS Events: SLAS 2023 Microscales Innovation in Life Sciences Symposium September 14-15, 2023 La Jolla, CA, USA SLAS 2023 Sample Management Symposium October 12-13, 2023 Washington, D.C., USA SLAS 2023 Data Sciences and AI Symposium November 14-15, 2023 Basel, Switzerland SLAS2024 International Conference and Exhibition February 3-7, 2024 Boston, MA, USA

PaperPlayer biorxiv cell biology
Biomolecular Condensates defined by Receptor Independent Activator of G protein Signaling: Properties and Regulation

PaperPlayer biorxiv cell biology

Play Episode Listen Later May 3, 2023


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.05.02.539089v1?rss=1 Authors: Vural, A., Lanier, S. M. Abstract: Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

PaperPlayer biorxiv cell biology
β-arrestin-dependent and -independent endosomal G protein activation by the vasopressin type 2 receptor

PaperPlayer biorxiv cell biology

Play Episode Listen Later Apr 2, 2023


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.01.535208v1?rss=1 Authors: Daly, C., Guseinov, A. A., Hahn, H., Tikhonova, I., Thomsen, A. R. B., Plouffe, B. Abstract: The vasopressin type 2 receptor (V2R) is an essential GPCR in renal regulation of water homeostasis. Upon stimulation, the V2R activates Gs and Gq/11, which is followed by robust recruitment of {beta}-arrestins and receptor internalization into endosomes. Unlike canonical GPCR signaling, the {beta}-arrestin association with the V2R does not terminate Gs activation, and thus, Gs-mediated signaling is sustained while the receptor is internalized. Here, we demonstrate that this V2R ability to co-interact with G protein/{beta}-arrestin and promote endosomal G protein signaling is not restricted to Gs, but also involves Gq/11. Furthermore, our data implies that {beta}-arrestins potentiate Gs/Gq/11activation at endosomes rather than terminating their signaling. Surprisingly, we found that the V2R internalizes and promote endosomal G protein activation independent of {beta}-arrestins to a minor degree. These new observations challenge the current model of endosomal GPCR signaling and suggest that this event can occur in both {beta}-arrestin-dependent and -independent manners. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Superhelden Ohne Cape
Larissas Erfolgsgeschichte

Superhelden Ohne Cape

Play Episode Listen Later Feb 19, 2023


In dieser Woche spreche ich mit Dr. Larissa Neumann über ihre Genesungsgeschichte. Bei ihr wurden Autoantikörper gegen G-Protein gekoppelte Rezeptoren (GPCR) & gegen α1-/ß1-/ß2-adrenerge, M3-/M4-muskarinerge und AT1-Rezeptoren gefunden. Diese Antikörper werden oft auch bei ME/CFS nachgewiesen. Links mentioned in this episode: Larissa Neumann: IG: dr.larissaneumann www.amarevitalita.com Superhelden Ohne Cape: IG: superheldenohnecape superheldenohnecape@yahoo.com This podcast is hosted by ZenCast.fm

PaperPlayer biorxiv cell biology
Functional Optimization of Light-Activatable Opto-GPCRs: Illuminating the Importance of the Proximal C-terminus in G-protein Specificity

PaperPlayer biorxiv cell biology

Play Episode Listen Later Jan 28, 2023


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.01.27.525823v1?rss=1 Authors: Leemann, S., Kleinlogel, S. Abstract: G-protein coupled receptors (GPCRs) are the largest family of human receptors that transmit signals from natural ligands and pharmaceutical drugs into essentially every physiological process. One main characteristic of GPCRs is their ability to specifically couple with different families of G-proteins, thereby triggering specific downstream signaling pathways. While an abundance of structural information is available on GPCR interactions with G-proteins, little is known about the GPCR domains functionally mediating G-protein specificity, in particular the proximal C-terminus, the structure which cannot be predicted with high confidentiality due to its flexibility. In this study, we exploited OptoGPCR chimeras between light-gated GPCRs (opsins) and ligand-gated GCPRs to systematically investigate the involvement of the C-terminus steering G-protein specificity. We employed rhodopsin-beta2-adrenoceptor and melanopsin-mGluR6 chimeras. We discovered a dominant role of the proximal C-terminus, dictating G-protein selectivity in the melanopsin-mGluR6 chimera, whereas it is the intracellular loop 3, which steers G-protein tropism in the rhodopsin-beta2-adrenoceptor. From the functional results and structural predictions, melanopsin and mGluR6 use a different mechanism to bRhod and b2AR to couple to a selective G-protein. Collectively, this work adds knowledge to the GPCR domains mediating G-protein selectivity, ultimately paving the way to optogenetically elicited specific G-protein signaling on demand. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

PaperPlayer biorxiv cell biology
Endometrial adhesion G protein-coupled receptors are dynamically expressed across the menstrual cycle and expression is altered by ovarian stimulation

PaperPlayer biorxiv cell biology

Play Episode Listen Later Dec 5, 2022


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.12.04.519044v1?rss=1 Authors: Kalakota, N. R., Lemenze, A., George, L., Zhao, Q., Wu, T., Morelli, S. S., Douglas, N. C., Babwah, A. V. Abstract: Ovarian stimulation (OS), utilized for the development of multiple ovarian follicles for IVF, induces supraphysiologic levels of E2 and an early rise in P4 that disrupt endometrial differentiation and decreases implantation rates or result in placental insufficiency and pregnancy complications. To improve pregnancy rates and reduce the risk of pregnancy complications associated with IVF, it is crucial to advance our molecular understanding of the molecular regulation of endometrial differentiation. Previous studies from our laboratory suggest G protein-coupled receptors (GPCRs) are important regulators of endometrial differentiation. To investigate this further, using a retrospective dataset, we identified all GPCRs expressed across the proliferative and secretory phase of the menstrual cycle and found that many members of the adhesion G protein-coupled receptor (ADGR) family are dynamically expressed. For each ADGR subfamily exhibiting differentially-expressed genes across the cycle, their expression was investigated by RT-PCR in the non-pregnant mouse uterus and decidua on E7.5 of pregnancy. For those genes expressed in the E7.5 decidua, their expression was further quantified by qPCR across early mouse pregnancy. The RT-PCR screen revealed expression of 13 ADGRs (4 of the 9 subfamilies) in E7.5 decidua and among these genes, many were differentially expressed between E0.5 and E5.5 or 6.5 and between E5.5 and E6.5. The dynamic expression of the ADGRs across the menstrual cycle and in early mouse pregnancy, suggests these ADGRs are E2- and/or P4-regulated genes. We therefore hypothesized that for these ADGR genes, mRNA expression would be disrupted in an OS cycle. This hypothesis was tested on endometrial biopsies collected in the secretory phase from prospective cohorts of women in natural and OS cycles. Consistent with the retrospective dataset, our data revealed that members of the ADGR gene family are expressed in the secretory phase of the natural menstrual cycle and for the first time, we show that their expression is altered by ovarian stimulation. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

PaperPlayer biorxiv neuroscience
G protein-biased LPAR1 agonism promotes prototypic antidepressant effects

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Nov 3, 2022


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.11.02.514841v1?rss=1 Authors: Kajitani, N., Okada-Tsucioka, M., Inoue, A., Miyano, K., Masuda, T., Boku, S., Iwamoto, K., Ohtsuki, S., Uezono, Y., Aoki, J., Takebayashi, M. Abstract: Prototypic antidepressants, such as tricyclic/tetracyclic antidepressants (TCAs), have multiple pharmacological properties and have been considered to be more effective than newer antidepressants, such as selective serotonin reuptake inhibitors (SSRIs), in treating severe depression. However, the molecular mechanisms underlying the high efficacy of TCAs have not been completely understood. Herein, we found that lysophosphatidic acid receptor 1 (LPAR1), a G protein-coupled receptor, mediates the antidepressant effects of amitriptyline, a typical TCA. Amitriptyline directly bound to LPAR1 and activated downstream G protein signaling without affecting {beta}-arrestin signaling, which implied that amitriptyline could act as a G protein-biased agonist of LPAR1. This biased agonism is unique to TCAs and has not been observed in other antidepressants, such as SSRIs. Long-term infusion of mouse hippocampus with 1-oleoyl-2-O-methyl-glycerophosphothionate (OMPT), a potent G protein-biased LPAR1 agonist, induced behavior similar to that induced by antidepressants. In contrast, LPA, a non-biased agonist of LPAR1, induced anxious behavior, indicating that LPAR1 may regulate conflicting emotional behaviors because of the downstream signaling bias. Furthermore, RNA-seq analysis revealed that LPA and OMPT have opposite patterns of gene expression changes in hippocampus. Ingenuity pathway analysis indicated that chronic intrahippocampal administration of OMPT could activate LPAR1 downstream signaling (Rho and MAPK), whereas LPA suppressed LPAR1 signaling. The results reveal the unique antidepressant effects of TCAs and indicate the potential of G protein-biased agonists of LPAR1 as targets for novel antidepressants. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

PaperPlayer biorxiv neuroscience
A subpopulation of peripheral sensory neurons expressing the Mas-related G Protein-Coupled Receptor d (Mrgprd) generates pain hypersensitivity in painful diabetic neuropathy.

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Oct 28, 2022


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.10.27.514066v1?rss=1 Authors: George, D. S., Jayaraj, N. D., Ren, D., Miller, R. E., Malfait, A.-M., Miller, R. J., Menichella, D. M. Abstract: Painful diabetic neuropathy (PDN) is one of the most common and intractable complications of diabetes. PDN is characterized by neuropathic pain accompanied by dorsal root ganglion (DRG) nociceptor hyperexcitability, axonal degeneration, and loss of cutaneous innervation. However, the complete molecular profile underlying the hyper-excitable cellular phenotype of DRG nociceptors in PDN has not been elucidated. This gap in our knowledge is a critical barrier to developing effective, mechanism-based, and disease-modifying therapeutic approaches which are urgently needed to relieve the symptoms of PDN. Using single-cell RNA sequencing we demonstrated an increased expression of the Mas-related G Protein- Coupled Receptor d (Mrgprd) in a subpopulation of DRG neurons in the well-established High-Fat Diet (HFD) mouse model of PDN. In vivo calcium imaging allowed us to demonstrate that activation of Mrgprd receptors expressed by cutaneous afferents produced DRG neuron hyper-excitability and oscillatory calcium waves. Furthermore, Mrgprd-positive cutaneous afferents persist in diabetic mice skin. Importantly, limiting Mrgprd signaling or Mrgprd-positive DRG neuron excitability, reversed mechanical allodynia in the HFD mouse model of PDN. Taken together, our data highlights a key role of Mrgprd-mediated DRG neuron excitability in the generation and maintenance of neuropathic pain in a mouse model of PDN. Hence, we propose Mrgprd as a promising accessible target for developing effective therapeutics currently unavailable for treating neuropathic pain in PDN. Furthermore, understanding which DRG neurons cell type is mediating mechanical allodynia in PDN is of fundamental importance to our basic understanding of somatosensation and may provide an important way forward for identifying cell-type-specific therapeutics to optimize neuropathic pain treatment and nerve regeneration in PDN. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

PaperPlayer biorxiv cell biology
Structure of a G protein-coupled receptor with GRK2 and a biased ligand

PaperPlayer biorxiv cell biology

Play Episode Listen Later Oct 19, 2022


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.10.19.512855v1?rss=1 Authors: Duan, J., Liu, H., Ji, Y., Yuan, Q., Li, X., Wu, K., Gao, T., Zhu, S., Jiang, Y., Yin, W., Xu, H. E. Abstract: Phosphorylation of G protein-coupled receptors (GPCR) by GPCR kinases (GRKs) desensitizes G protein signaling and promotes arrestin signaling, which is also modulated by biased ligands. Molecular assembly of GRKs to GPCRs and the basis of GRK-mediated biased signaling remain largely unknown due to the weak GPCR-GRK interactions. Here we report the complex structure of neurotensin receptor 1 (NTSR1) bound to GRK2, Gaq, and an arrestin-biased ligand, SBI-553, at a resolution of 2.92 Angstrom. The high-quality density map reveals the clear arrangement of the intact GRK2 with the receptor, with the N-terminal helix of GRK2 docking into the open cytoplasmic pocket formed by the outward movement of the receptor TM6, analogous of the binding of G protein to the receptor. Strikingly, the arrestin-biased ligand is found at the interface between GRK2 and NTSR1 to enhance GRK2 binding. The binding mode of the biased ligand is compatible with arrestin binding but is clashed with the binding of a G protein, thus provide an unambiguous mechanism for its arrestin-biased signaling capability. Together, our structure provides a solid model for understanding the details of GPCR-GRK interactions and biased signaling. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Psychiatry.dev -  All Abstracts TTS
G protein-biased GPR3 signaling ameliorates amyloid pathology in a preclinical Alzheimer's disease mouse model – PubMed

Psychiatry.dev - All Abstracts TTS

Play Episode Listen Later Sep 28, 2022


https://psychiatry.dev/wp-content/uploads/speaker/post-10040.mp3?cb=1664385118.mp3 Playback speed: 0.8x 1x 1.3x 1.6x 2x Download: G protein-biased GPR3 signaling ameliorates amyloid pathology in a preclinical Alzheimer’s disease mouse model – PubMed Yunhong Huang et al. PNAS. 2022.Full EntryG protein-biased GPR3 signaling ameliorates amyloid pathology in a preclinical Alzheimer's disease mouse model – PubMed

PaperPlayer biorxiv neuroscience
The Tre1/S1pr1 phospholipid-binding G protein-coupled receptor signaling pathway is required for astrocyte morphogenesis

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Sep 17, 2022


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2022.09.15.508188v1?rss=1 Authors: Chen, J., Stork, T., Kang, Y., Sheehan, A., Paton, C., Monk, K., Freeman, M. Abstract: Astrocytes play crucial roles in regulating neural circuit function by forming a dense network of synapse-associated membrane specializations. Intimate astrocyte-synapse contact is thought to be crucial for astrocyte function, yet signaling pathways regulating astrocyte morphogenesis remain poorly defined. Here we show the Drosophila lipid-binding G protein-coupled receptor (GPCR) Tre1, likely acting through Rac1, is required for astrocytes to elaborate their complex morphologies. The lipid phosphate phosphatases Wunen/Wunen2, which process phospholipid ligands for Tre1, also regulate astrocyte growth, and act upstream of Tre1 as Wunen/Wunen2-mediated astrocyte overgrowth is suppressed by Tre1 loss. Loss of s1pr1, the functional analog of Tre1 in zebrafish, also leads to disruption of astrocyte morphogenesis. Live-imaging and pharmacology demonstrate that S1pr1 balances proper astrocyte process extension/retraction dynamics during morphogenesis, and that S1pr1 signaling is required throughout astrocyte development. Our work identifies lipid-binding GPCRs like Tre1 and S1pr1 as potent evolutionarily conserved regulators of astrocyte morphological complexity. Copy rights belong to original authors. Visit the link for more info Podcast created by PaperPlayer

IS PHARMACOLOGY DIFFICULT Podcast
Is Pharmacology Difficult Podcast G PROTEIN COUPLED RECEPTOR-2

IS PHARMACOLOGY DIFFICULT Podcast

Play Episode Listen Later Oct 17, 2021 13:19


Welcome all to IS PHARMACOLOGY DIFFICULT Podcast! I am Dr Radhika Vijay. As a continuation of yester episode, today i will be discussing the three major effector pathways of G Protein Coupled Receptor , firstly the Adenylyl cyclase enzyme-cAMP pathway, then IP3-DAG Pathway, Ion channel regulation,... All will be explained with apt examples and details! Termination of pathways forms a major notable point to discuss, never to be missed.. Wishing you all a very Happy Sunday!! For all the updates and latest episodes of my podcast, please visit www.ispharmacologydifficult.com where you can also sign up for a free monthly newsletter of mine. It actually contains lot of updates about the medical sciences, drug information and my podcast updates also. You can follow me on different social media handles like twitter, insta, facebook and linkedin. They all are with same name "IS PHARMACOLOGY DIFFICULT". If you are listening for the first time, do follow me here, whatever platform you are consuming this episode, stay tuned, do rate and review on ITunes, Apple podcasts, stay safe, stay happy, stay enlightened, Thank you!! You can access various links via - https://linktr.ee/ispharmacologydifficult

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IS PHARMACOLOGY DIFFICULT Podcast
Is Pharmacology Difficult Podcast G PROTEIN COUPLED RECEPTOR-2

IS PHARMACOLOGY DIFFICULT Podcast

Play Episode Listen Later Oct 17, 2021 13:19


Welcome all to IS PHARMACOLOGY DIFFICULT Podcast! I am Dr Radhika Vijay.As a continuation of yester episode, today i will be discussing the three major effector pathways of G Protein Coupled Receptor , firstly the Adenylyl cyclase enzyme-cAMP pathway,  then IP3-DAG Pathway, Ion channel regulation,...All will be explained with apt examples and details! Termination of pathways forms a major notable point to discuss, never to be missed.. Wishing you all a very Happy Sunday!! For all the updates and latest episodes of my podcast, please visit www.ispharmacologydifficult.com where you can also sign up for a free monthly newsletter of mine. It actually contains lot of updates about the medical sciences, drug information and my podcast updates also. You can follow me on different social media handles like twitter, insta, facebook and linkedin. They all are with same name "IS PHARMACOLOGY DIFFICULT". If you are listening for the first time, do follow me here, whatever platform you are consuming this episode, stay tuned, do rate and review on ITunes, Apple podcasts, stay safe, stay happy, stay enlightened, Thank you!! You can access various links via - https://linktr.ee/ispharmacologydifficult

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IS PHARMACOLOGY DIFFICULT Podcast
Is Pharmacology Difficult Podcast G PROTEIN COUPLED RECEPTOR-2

IS PHARMACOLOGY DIFFICULT Podcast

Play Episode Listen Later Oct 17, 2021 13:19


Welcome all to IS PHARMACOLOGY DIFFICULT Podcast! I am Dr Radhika Vijay.As a continuation of yester episode, today i will be discussing the three major effector pathways of G Protein Coupled Receptor , firstly the Adenylyl cyclase enzyme-cAMP pathway,  then IP3-DAG Pathway, Ion channel regulation,...All will be explained with apt examples and details! Termination of pathways forms a major notable point to discuss, never to be missed.. Wishing you all a very Happy Sunday!! For all the updates and latest episodes of my podcast, please visit www.ispharmacologydifficult.com where you can also sign up for a free monthly newsletter of mine. It actually contains lot of updates about the medical sciences, drug information and my podcast updates also. You can follow me on different social media handles like twitter, insta, facebook and linkedin. They all are with same name "IS PHARMACOLOGY DIFFICULT". If you are listening for the first time, do follow me here, whatever platform you are consuming this episode, stay tuned, do rate and review on ITunes, Apple podcasts, stay safe, stay happy, stay enlightened, Thank you!! You can access various links via - https://linktr.ee/ispharmacologydifficult

apple drugs camp difficult coupled termination pharmacology happy sunday ion receptor podcast g pharmacodynamics g protein gpcr radhika vijay is pharmacology difficult podcast is pharmacology difficult
IS PHARMACOLOGY DIFFICULT Podcast
Is Pharmacology Difficult Podcast G PROTEIN COUPLED RECEPTOR-1

IS PHARMACOLOGY DIFFICULT Podcast

Play Episode Listen Later Oct 9, 2021 11:42


Welcome all to IS PHARMACOLOGY DIFFICULT Podcast! I am Dr Radhika Vijay. In today's episode I will narrate the famous poem "Windows" by Wes Magee, communicate its amazing message and look into the window of today's topic i.e. G Protein coupled receptors. First and foremost a little word about the receptor nature and types, and then all about G protein coupled receptors, their structure, mechanism introduction and a teaser of upcoming episode too! All in all a good weekend diary to listen to! Hope you all like it........ For all the updates and latest episodes of my podcast, please visit www.ispharmacologydifficult.com where you can also sign up for a free monthly newsletter of mine. It actually contains lot of updates about the medical sciences, drug information and my podcast updates also. You can follow me on different social media handles like twitter, insta, facebook and linkedin. They all are with same name "IS PHARMACOLOGY DIFFICULT". If you are listening for the first time, do follow me here, whatever platform you are consuming this episode, stay tuned, do rate and review on ITunes, Apple podcasts, stay safe, stay happy, stay enlightened, Thank you!! You can access various links via https://linktr.ee/ispharmacologydifficult

apple difficult windows coupled pharmacology receptor podcast g g protein radhika vijay is pharmacology difficult is pharmacology difficult podcast
IS PHARMACOLOGY DIFFICULT Podcast
Is Pharmacology Difficult Podcast G PROTEIN COUPLED RECEPTOR-1

IS PHARMACOLOGY DIFFICULT Podcast

Play Episode Listen Later Oct 9, 2021 11:42


Welcome all to IS PHARMACOLOGY DIFFICULT Podcast! I am Dr Radhika Vijay.In today's episode I will narrate the famous poem "Windows" by Wes Magee, communicate its amazing message and look into the window of today's topic i.e. G Protein coupled receptors.First and foremost a little word about the receptor nature and types, and then all about G protein coupled receptors, their structure, mechanism introduction and a teaser of upcoming episode too! All in all a good weekend diary to listen to! Hope you all like it........ For all the updates and latest episodes of my podcast, please visit www.ispharmacologydifficult.com where you can also sign up for a free monthly newsletter of mine. It actually contains lot of updates about the medical sciences, drug information and my podcast updates also. You can follow me on different social media handles like twitter, insta, facebook and linkedin. They all are with same name "IS PHARMACOLOGY DIFFICULT". If you are listening for the first time, do follow me here, whatever platform you are consuming this episode, stay tuned, do rate and review on ITunes, Apple podcasts, stay safe, stay happy, stay enlightened, Thank you!!You can access various links viahttps://linktr.ee/ispharmacologydifficult

apple difficult windows coupled pharmacology receptor podcast g g protein radhika vijay is pharmacology difficult is pharmacology difficult podcast
IS PHARMACOLOGY DIFFICULT Podcast
Is Pharmacology Difficult Podcast G PROTEIN COUPLED RECEPTOR-1

IS PHARMACOLOGY DIFFICULT Podcast

Play Episode Listen Later Oct 9, 2021 11:42


Welcome all to IS PHARMACOLOGY DIFFICULT Podcast! I am Dr Radhika Vijay.In today's episode I will narrate the famous poem "Windows" by Wes Magee, communicate its amazing message and look into the window of today's topic i.e. G Protein coupled receptors.First and foremost a little word about the receptor nature and types, and then all about G protein coupled receptors, their structure, mechanism introduction and a teaser of upcoming episode too! All in all a good weekend diary to listen to! Hope you all like it........ For all the updates and latest episodes of my podcast, please visit www.ispharmacologydifficult.com where you can also sign up for a free monthly newsletter of mine. It actually contains lot of updates about the medical sciences, drug information and my podcast updates also. You can follow me on different social media handles like twitter, insta, facebook and linkedin. They all are with same name "IS PHARMACOLOGY DIFFICULT". If you are listening for the first time, do follow me here, whatever platform you are consuming this episode, stay tuned, do rate and review on ITunes, Apple podcasts, stay safe, stay happy, stay enlightened, Thank you!!You can access various links viahttps://linktr.ee/ispharmacologydifficult

apple drugs difficult windows coupled pharmacology receptor podcast g pharmacodynamics g protein gpcr radhika vijay is pharmacology difficult podcast is pharmacology difficult
Metabolism Made Easy
Signal Transduction II: G-protein coupled receptors (GPCR)

Metabolism Made Easy

Play Episode Listen Later Jun 3, 2021 34:42


This podcast covers thd mechanism of activation of two GPCRs: The beta- adrenergic and the alpha1-adrenergic receptors. This biochemistry content may be useful to premedical and medical students. --- Send in a voice message: https://anchor.fm/a-j-ghalayini/message

signal coupled receptors transduction g protein gpcr
My AP Biology Thoughts
Reception: G Protein Receptors, Tyrosine Kinase Receptors

My AP Biology Thoughts

Play Episode Listen Later May 25, 2021 5:12


My AP Biology Thoughts  Unit 4 Cell Communication and Cell CycleWelcome to My AP Biology Thoughts podcast, my name is Sid and I am your host for episode #86 called Unit 4 Cell Communication and Cell Cycle: G Protein Receptors and Tyrosine Kinase Receptors. Segment 1: Introduction to G protein receptors and tyrosine kinase receptors G Protein receptors and tyrosine kinase receptors both work to mediate cell communication by binding a signaling molecule, which is also called a ligand. Then this signal is sent through a transduction pathway where the last target protein causes some response. The response for both can be a variety of things such as gene expression, apoptosis, metabolic responses, cell division, or cell growth. Despite being similar in this way, g protein receptors and tyrosine kinase receptors work in very different ways Segment 2: More About G protein receptors Let's start by discussing g protein receptors. G proteins are very diverse and can bind to many different signals. One example is odorant (or scent) receptors. G proteins receptors are located in the cell membrane which is where an extracellular ligand binds to it. The signal is eventually sent to a g protein which is located on the membrane, but on the cytoplasmic side. Before the G protein is activated, GDP is bound to it which keeps it inactive. GDP is guanosine diphosphate. After the signal binds to the receptor, the receptor slightly changes shape and becomes active. Then, the GDP binds to the g protein receptor. Since the G protein no longer has a GDP bound to it, it frees it up to accept and bind to GTP. The GTP activates the G protein. The G protein is made up of three subunits: alpha, beta, and gamma. When the GTP is bound to the G protein and activates it, the alpha subunit detaches and moves away from the receptor. Now the G protein is split into two parts: one part is the single alpha subunit and the other is the beta and gamma subunits. These two parts can go on to interact with other proteins and cause a transduction pathway that results in one of many responses. Eventually, the alpha subunit comes back and hydrolyzes the GTP which keeps the G protein active and changes it back into GDP. At this point the G protein will once again become inactive. G proteins coupled receptors are very important in the human body. Disruptions can cause diseases like cystic fibrosis or cholera.  Now let's talk about tyrosine kinase receptors. Tyrosine kinase receptors are enzyme linked receptors. Enzyme linked receptors are receptors that are associated with an enzyme. A kinase is a protein that phosphorylates other proteins. For tyrosine kinase receptors, the kinase phosphorylates tyrosine. To start the process, a signalling molecule attaches to two tyrosine kinase receptors. These come together and form a dimer. Then, each tyrosine kinase receptor phosphorylates the domains of the tyrosine kinase receptor. Then, once the tyrosine is phosphorylated, it can send signals to other molecules Segment 3: Connection to the Course G protein receptors and tyrosine kinase receptors are very important to many species. Problems with g protein receptors can cause choler, cystic fibrosis, and some bacterial infections. Problems with tyrosine kinase receptors can also cause diseases and cancers. Both of these receptors play integral parts in many different species. This can be evidence of the endosymbiotic theory. Since so many species use these receptors, they likely came from a common ancestor and had an evolutionary advantage. Thank you for listening to this episode of My AP Biology Thoughts. For more student-ran podcasts and digital content, make sure that you visit http://www.hvspn.com (www.hvspn.com).  Music Credits: "Ice Flow" Kevin MacLeod (incompetech.com) Licensed under Creative Commons: By Attribution 4.0 License http://creativecommons.org/licenses/by/4.0/ Subscribe to our Podcast...

The Medbullets Step 1 Podcast
Pharmacology | G Protein-Coupled Receptors (GPCR)

The Medbullets Step 1 Podcast

Play Episode Listen Later Apr 30, 2021 23:24


In this episode, we review the high-yield topic of G Protein-Coupled Receptors (GPCR) from the Pharmacology section. --- Send in a voice message: https://anchor.fm/medbulletsstep1/message

BIOLOGO
#030 - Happy & High - durch Sport und Drogen? - Wirkung von Endorphinen und Opiaten

BIOLOGO

Play Episode Listen Later Apr 15, 2021 22:55


Unbedingt im Skript mitverfolgen (S. 29/30) Du erfährst... - wie Glücksgefühle beim Runners' High entstehen - wie es zu Abmilderung von Schmerzwahrnehmung kommt - wie Morphin, Opium und Heroin wirken! Fachbegriffe: präsynaptische Hemmung, postsynoptische Hemmung, Dopamin, Endorphin, Heroin, Opium, G-Protein, GTP, GDP, second messenger, cAMP, ATP, Calciumionenkanal, Adenylatzyklase, Silencer, Genexpression, Transkriptionsfaktor, Hypophyse, Nucleus accumbens, Area tegmentalis, sensorisches Neuron, Nozizeptoren, uvm.

BIOLOGO
#022 - Adrenalin-Kick oder Koffein-Boost? - Von Transmittern, Synapsen und second messenger-Systemen

BIOLOGO

Play Episode Listen Later Mar 3, 2021 15:50


HIER geht es zum ergänzenden Skript:) Du erfährst: 1) Was Botenstoffe sind... 2) wie sie an einer Synapse wirken... 3) wie Koffein und Adrenalin ein second messenger-System beeinflussen Schreibe gerne Fragen/Feedback/Wünsche an biologopodcast@googlemail.com oder ... @biologopodcast auf Instagram Fachbegriffe der Folge: Transmitter, Hormone, Serotonin, Dopamin, Glutamat, Acetylcholin, Aktionspotential, Endknöpfchen, Calciumkanäle, Transmitter, synaptische Vesikel, präsynaptische und postsynaptische Membran,  Acetylcholinesterase, Diffusion, cAMP, ATP, G-Protein, Adenosin, Adrenalin, Koffein, Adenylatcyclase. Viel Erfolg beim Lernen!

First Past the Post
G-Protein Linked Second Messangers

First Past the Post

Play Episode Listen Later Dec 28, 2020 3:34


This episode covers g-protein linked second messangers!

g protein
PaperPlayer biorxiv biochemistry
A Universal Allosteric Mechanism for G Protein Activation

PaperPlayer biorxiv biochemistry

Play Episode Listen Later Nov 16, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.15.383679v1?rss=1 Authors: Knight, K. M., Ghosh, S., Campbell, S. L., Lefevre, T. J., Olsen, R. H. J., Smrcka, A. V., Valentin, N. H., Yin, G., Vaidehi, N., Dohlman, H. G. Abstract: G proteins play a central role in signal transduction and pharmacology. Signaling is initiated by cell-surface receptors, which promote GTP binding and the dissociation of G from the G{beta}{gamma} subunits. Structural studies have revealed the molecular basis for subunit association with receptors, RGS proteins and downstream effectors. In contrast, the mechanism of subunit dissociation is poorly understood. We used cell signaling assays, MD simulations, biochemistry and structural analysis to identify a conserved network of amino acids that dictates subunit release. In the presence of the terminal phosphate of GTP, a glycine forms a polar network with an arginine and glutamate, putting torsional strain on the subunit binding interface. This "G-R-E motif" secures GTP and, through an allosteric link, discharges the G{beta}{gamma} dimer. Replacement of network residues prevents subunit dissociation, regardless of agonist or GTP binding. These findings reveal the molecular basis for the final committed step of G protein activation. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv neuroscience
The single pass membrane protein MRAP2 regulates energy homeostasis by promoting primary cilia localization of the G protein-coupled receptor MC4R

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Nov 15, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.13.382325v1?rss=1 Authors: Bernard, A. A., Ojeda Naharros, I., Bourgain-Guglielmetti, F., Ciprin, J., Yue, X., Zhang, S., McDaid, E., Nachury, M. V., Reiter, J., Vaisse, C. Abstract: The G protein-coupled receptor MC4R (Melanocortin-4 Receptor) and its associated protein MRAP2 (Melanocortin Receptor-Associated Protein 2) are both essential for the regulation of food intake and body weight in humans and mice. MC4R localizes and functions at the neuronal primary cilium, a microtubule-based organelle that senses and relays extracellular signals. Here, we demonstrate that MRAP2 is critical for the ciliary localization and weight-regulating function of MC4R. Our data reveal that GPCR localization to primary cilia can require specific accessory proteins that may not be present in heterologous cell systems. Our findings also demonstrate the essential role of neuronal primary cilia localization of MC4R for adequate control of energy homeostasis and the obesity-promoting effect of genetic disruption of this pathway. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv neuroscience
Synaptic integration by co-localized G protein-coupled receptors in presynaptic terminals

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Nov 12, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.12.362491v1?rss=1 Authors: Church, E., Hamid, E., Zurawski, Z. P., Alford, S. Abstract: Presynaptic terminals have been little studied as sites of synaptic integration because of inaccessibility. Gi/o-mediated presynaptic inhibition is ubiquitous and either reduces release probability (Pr) by inhibiting Ca2+ entry or inhibits directly at SNARE complexes. At CA1-subicular presynapses, 5-HT1B and GABAB receptors colocalize. GABABRs inhibit Ca2+ entry, whereas 5-HT1BRs Ca2+-dependently target SNARE complexes. We demonstrate that GABABRs, alter Pr, but 5-HT1BRs reduce cleft glutamate concentrations allowing strong inhibition of AMPA- but not NMDA-receptor responses. Simulations of glutamate release and receptor binding demonstrates that experimental effects on release and low affinity antagonism are well-fit by reduced release rates. Train-dependent presynaptic Ca2+ accumulation forces frequency-dependent recovery of neurotransmission during 5-HT1BR activation, consistent with competition between Ca2+-synaptotagmin and G{beta}{gamma} at SNARE complexes. Thus, stimulus trains in 5-HT unveil dynamic synaptic modulation and a sophisticated hippocampal output filter, that itself is modulated by colocalized GABABRs which alter presynaptic Ca2+ allowing complex presynaptic integration. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv neuroscience
The G protein-Coupled Metabotropic Glutamate Receptor 1 controls neuronal macroautophagy

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Nov 2, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.02.365783v1?rss=1 Authors: Donoso, M., Speranza, L., Kalinowska, M., Castillo, C., De Sanctis, C., Francesconi, A. Abstract: Autophagy is an evolutionarily conserved, highly regulated catabolic process critical to neuronal homeostasis, function and survival throughout organismal lifespan. However, the external factors and signals that control autophagy in neurons are still poorly understood. Here we report that the G protein-coupled metabotropic glutamate receptor 1 (mGlu1) contributes to control basal autophagy in the brain. Autophagy is upregulated in the brain of adult mGlu1 knockout mice and genetic deletion or pharmacological inhibition of native mGlu1 receptors enhances autophagy flux in neurons. The evolutionarily conserved adaptor protein FEZ1, identified by a genome-wide screen as mGlu1 receptor interacting partner, was found to participate in the regulation of neuronal autophagy and to be required for repression of autophagy flux by the mGlu1 receptor. Furthermore, FEZ1 appears to enable association of mGlu1 with Ulk1, a core component of the autophagy pathway. Thus, we propose that the mGlu1 receptor contributes to restrain constitutive autophagy in neurons. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv bioinformatics
Functional Microswitches of Mammalian G Protein-Coupled Bitter-Taste Receptors

PaperPlayer biorxiv bioinformatics

Play Episode Listen Later Oct 23, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.10.23.348706v1?rss=1 Authors: Topin, J., Bouysset, C., Kim, Y., Rhyu, M., Fiorucci, S., Golebiowski, J. Abstract: Bitter taste receptors (TAS2Rs) are a poorly understood subgroup of G protein-coupled receptors (GPCR). No experimental structure of these receptors is available and key-residues controlling their function remain mostly unknown. Here, we have identified the functional microswitches that encode agonist sensing and downstream signaling mechanisms within TAS2Rs sequences. We thoroughly re-aligned the amino-acid sequences of the 25 human TAS2Rs considering residue conservations and all the experimental data from the literature as constraints. As a test case, an accurate homology model of TAS2R16 was constructed and examined by site-directed mutagenesis and in vitro functional assays. Conserved motifs acting as microswitches during agonist-sensing and receptor activation were pinpointed by comparison with the current knowledge on class A GPCRs. Unravelling these sequence-function relationships is of utmost importance to streamline how TAS2Rs functions are encrypted in their sequence. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv biophysics
Activation of G-protein coupled receptors is thermodynamically linked to lipid solvation

PaperPlayer biorxiv biophysics

Play Episode Listen Later Aug 21, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.08.20.259077v1?rss=1 Authors: Leonard, A. N., Lyman, E. Abstract: Preferential lipid solvation of the G-protein coupled A2A adenosine receptor (A2AR) is evaluated from 35 sec of all-atom molecular dynamics simulation. A coarse-grained transition matrix algorithm is developed to overcome slow equilibration of the first solvation shell, obtaining statistically robust estimates of the free energy of solvation by different lipids for the receptor in different activation states. Results indicate preference for solvation by unsaturated chains, which favors the active receptor. A model for lipid-dependent GPCR activity is proposed in which the chemical potential of lipids in the bulk membrane modulates receptor activity. The enthalpy and entropy associated with moving saturated vs. unsaturated lipids from bulk to A2AR's first solvation shell are compared. In the simulated mixture, saturated chains are disordered (i.e., obtain a favorable entropic contribution) when partitioning to the receptor surface, but this is outweighed by a favorable enthalpic contribution for unsaturated chains to occupy the first solvation shell. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv biophysics
Routine sub-2.5 A cryo-EM structure determination of B-family G protein-coupled receptors

PaperPlayer biorxiv biophysics

Play Episode Listen Later Aug 21, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.08.21.260851v1?rss=1 Authors: Danev, R., Belousoff, M., Liang, Y.-L., Zhang, X., Wootten, D., Sexton, P. M. Abstract: Cryo-electron microscopy (cryo-EM) experienced game-changing hardware and software advances about a decade ago. Since then, there have been gradual and steady improvements in experimental and data analysis methods. Nonetheless, structural analysis of nonsymmetric membrane proteins, such as G protein-coupled receptors (GPCRs), remains challenging. Their relatively low molecular weight and obstruction by the micelle/nanodisc result in marginal signal levels, which combined with the intrinsic flexibility of such complexes creates difficult structural study scenarios. Pushing the performance limits of cryo-EM requires careful optimization of all experimental aspects. To this end, it is necessary to build quantitative knowledge of the effect each parameter has on the outcome. Here, we present in-depth analysis of the influence of the main cryo-EM experimental factors on the performance for GPCR structure determination. We used a tandem experiment approach that combined real-world structural studies with parameter testing. We quantified the effects of using a Volta phase plate, zero-loss energy filtering, objective lens aperture, defocus magnitude, total exposure, and grid type. Through such systematic optimization of the experimental conditions, it has been possible to routinely determine class B1 GPCR structures at resolutions better than 2.5 A. The improved fidelity of such maps helps to build higher confidence atomic models and will be crucial for the future expansion of cryo-EM into the structure-based drug design domain. The optimization guidelines drafted here are not limited to GPCRs and can be applied directly for the study of other challenging membrane protein targets. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv genetics
Distinct specificity of two pheromone G-protein coupled receptors, Map3 and Mam2, in fission yeast species.

PaperPlayer biorxiv genetics

Play Episode Listen Later Jul 22, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.07.22.215566v1?rss=1 Authors: Seike, T., Sakata, N., Shimoda, C., Niki, H., Furusawa, C. Abstract: Most sexually reproducing organisms have the ability to recognize individuals of the same species. In ascomycetes including yeasts, potential mating between cells of opposite mating-type depends on the molecular recognition of two peptidyl mating pheromones by their corresponding G-protein coupled receptors (GPCRs). Although such pheromone/receptor systems are likely to function in both mate choice and prezygotic isolation, very few studies have focus on the differences in pheromone/receptor system between mating types. The fission yeast Schizosaccharomyces pombe has two mating types (sexes), Plus (P) and Minus (M). Here we investigated the specificity of the two GPCRs, Mam2 and Map3, for their respective pheromones, P-factor and M-factor, in fission yeast. First, we switched GPCRs between S. pombe and the closely related species Schizosaccharomyces octosporus, which showed that SoMam2 (Mam2 of S. octosporus) is partially functional in S. pombe, whereas SoMap3 (Map3 of S. octosporus) is not interchangeable. Next, we swapped individual domains of Mam2 and Map3 with the respective domains in SoMam2 and SoMap3, which revealed differences between the receptors both in the intracellular regions that regulate the downstream signaling of pheromones and in the molecular recognition for pheromone binding. In particular, we demonstrated that two amino acid residues of Map3, F214 and F215, are essential for the specificity of M-factor recognition. Thus, the differences in these two GPCRs are likely to reflect the significantly distinct specificities of their respective pheromone/receptor systems; that is, the specificity of Map3 is more stringent than that of Mam2. We speculate that this sexual asymmetry might allow ascomycete fungi to generate novel prezygotic barriers within a population, while maintaining strong mate choice. Our genetic analyses also contribute to our understanding of the receptors that comprise the Class D GPCRs belonging to the fungal pheromone receptor family. Author summaryCourtship signals play a key role in the fertilization processes of living beings from animals to microorganisms and, in particular, sex pheromones are involved in differentiating among species. On the other hand, changes in a pheromone/receptor system might also alter species-specificity during the selection of a mating partner, which is likely to facilitate prezygotic isolation. Here we demonstrate a distinct difference in the specificity of GPCRs for pheromones between cells of opposite mating-type in the fission yeast Schizosaccharomyces pombe, by conducting a comprehensive genetic analysis and comparison of these proteins between closely related fission yeast species. Our finding of sexual asymmetry due to the relative strictness of receptors in S. pombe suggests that it might be one of the driving forces behind the formation of new species. Such differences in pheromone/receptor interactions between the sexes may occur in a variety of life-forms such as insects and amphibians; hence, the findings of this study might be extended to other organisms. In addition, this study of S. pombe pheromone receptors sheds light on the functions of the unique Class D GPCRs. Copy rights belong to original authors. Visit the link for more info

PaperPlayer biorxiv neuroscience
Suppression of pyramidal neuron G protein-gated inwardly rectifying K+ channel signaling impairs prelimbic cortical function and underlies stress-induced deficits in cognitive flexibility

PaperPlayer biorxiv neuroscience

Play Episode Listen Later Jun 8, 2020


Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.06.08.139725v1?rss=1 Authors: Anderson, E., Loke, S., Wrucke, B., Engelhardt, A., Hess, E., Hearing, M. Abstract: Background: Imbalance in prefrontal cortical (PFC) pyramidal neuron excitation:inhibition is thought to underlie symptomologies shared across stress-related disorders and neuropsychiatric disease, including dysregulation of emotion and cognitive function. G protein-gated inwardly rectifying K+ (GIRK/Kir3) channels mediate excitability of medial PFC pyramidal neurons, however the functional role of these channels in mPFC-dependent regulation of affect, cognition, and cortical dynamics is unknown. Methods: In mice harboring a floxed version of the kcnj3 (Girk1) gene, we used a viral-cre approach to disrupt GIRK1-containing channel expression in pyramidal neurons within the prelimbic (PL) or infralimbic (IL) cortices. Additional studies used a novel model of chronic unpredictable stress (CUS) to determine the impact on PL GIRK-dependent signaling and cognitive function. Results: In males, loss of pyramidal GIRK-dependent signaling in the PL, but not IL, differentially impacted measures of affect and motivation, and impaired working memory and cognitive flexibility. CUS produced similar deficits in affect and cognition that paralleled a reduction in PL pyramidal GIRK-dependent signaling akin to viral approaches. Viral- and stress-induced behavioral deficits were rescued by systemic injection of a novel, GIRK1-selective agonist, ML-297. Unexpectedly, neither ablation of PL GIRK-dependent signaling or exposure to the CUS regimen impacted affect or cognition in female mice. Conclusions: GIRK-dependent signaling in male mice, but not females, is critical for maintaining optimal PL function and behavioral control. Disruption of this inhibition may underlie stress-related dysfunction of the PL and represent a therapeutic target for treating stress-induced deficits in affect regulation and impaired cognition that reduce quality of life. Copy rights belong to original authors. Visit the link for more info

MedFlashGo | 4 Minutes Or Less Daily Rapid Review Of USMLE, COMLEX, And Shelf For Medical Students
#12 G protein receptors and Bladder Dysfunction | Biochemistry | MedFlashGo Question Of The Day

MedFlashGo | 4 Minutes Or Less Daily Rapid Review Of USMLE, COMLEX, And Shelf For Medical Students

Play Episode Listen Later Feb 12, 2020 4:01


Welcome To The MedFlashGo Podcast. This Is Your Daily 4 Minutes Or Less Rapid Review For USMLE, COMLEX, And Shelf Exams. We release a new episode every weekday! In this question of the day, Sean asks students to identify the correct G protein receptor based on the given characteristics described. These questions are powered by MedFlashGo The First Voice-based interactive medical question bank currently available on Alexa. This tool allows medical students to study medical topics and be interactively tested without the use of a screen. You can study on your couch, in your car, and on the move without the use of a screen. To get access to your 50 free questions go on to your Alexa application and search medflashgo In the skills section. To learn more details go to medflashgo.com and check out our frequently asked questions section. Please know that these questions were creatively designed by medical students and physicians for the purpose of education and do not replace health information given from your health professionals. We have tried our best to make sure the information is accurate please, so please let us know if you find any errors and we will be sure to correct them. --- Send in a voice message: https://anchor.fm/medflashgo/message

Circulation on the Run
Circulation February 28, 2017

Circulation on the Run

Play Episode Listen Later Feb 27, 2017


Dr. Carolyn L.:                    Welcome to Circulation on the Run, your weekly podcast, summary, and backstage pass to the journal and its editors. I'm Dr. Carolyn Lam, associate editor from the National Heart Center and Duke National University of Singapore. On our podcast today we are discussing the role of diastolic stress testing and the evaluation of heart failure with preserved dejection fraction, a really hot topic indeed, but first here's your summary of this week's issue.                                                 The first study tackles the obesity paradox in cardiac surgery, where morbidity and mortality are lower in obese patients. This study sought to ask the question, "Is this due to reverse epidemiology, bias, or confounding?" To answer this question, Dr. Maris Kelko and colleagues from University Leicester in United Kingdom used two separate analysis. One, registry data from the National Adult Cardiac Surgery Registry and two, a systematic review in meta-analysis of studies. Of more than 400,000 patients in the cohort study and more than 550,000 patients in the systematic review, the authors found a U shape association between mortality and body mass index classes, where lower mortality was observed in overweight and obese class one and two patients, relative to normal weight patients, and mortality was increased in underweight individuals.                                                 Now, the obesity paradox has been attributed to reverse epidemiology where the survival benefit associated with obesity is thought to actually reflect worse outcomes in the underweight patients who also had frailty, cachexia, or severe chronic disease. However, in the current study, counter to the reverse epidemiology hypothesis, the protective effects of obesity were less in patients with chronic renal, lung, or cardiac disease and greater in older patients as well as in those with complications of obesity, such as metabolic syndrome and atherosclerosis. Furthermore, adjustments for important confounders did not alter the results. The authors therefore concluded that obesity is associated with lower risks after cardiac surgery with consistent effects noted in multiple analysis even after attempting to address residual confounding and reverse causation.                                                 The authors even went as far as to suggest that their findings do not support common practice where weight loss is recommended prior to surgery or where very obese patients are refused surgery in the morbidly obese. These provocative findings are discussed in an accompanying editorial by Doctor's Carnethon and Kahn from Northwestern University. While the editorialists agree that this well-designed study highlights an important knowledge gap, they pointed out that the obese class two patients had nearly five times greater risk for deep sternal wound infection and 25% higher likelihood of needing renal replacement therapy.                                                 In such patients additional intervention in the perioperative period may still be indicated and include weight loss recommendations and postoperative surveillance for complications. Thus, a more cautious final recommendation may be for future studies to prospectively assess weight loss interventions prior to elective surgery in the context of overall surgical risk as assessed by the EuroSCORE or STS models.                                                 The next paper describes mechanistic studies showing for the first time that nucleoside diphosphate kinase suppresses cyclic-AMP formation in human heart failure. In this paper by First Authors, Dr. Abu Taha and Hagemann, corresponding authors Dr.'s Tobref and Weilend from the Heidelberg University in Germany, the authors performed biochemical studies of nucleoside diphosphate kinase and G Protein signaling in human and rat tissue samples, assessed the functional impact of nucleoside diphosphate kinase C on cyclic-AMP levels and contractility and isolated red cardiomyocytes and determined that in vitro effects of these nucleoside diphosphate kinases on contractility in zebra, fish and mice.                                                 They identified nucleoside diphosphate kinase as the critical isoform for the regulation of G Protein function and cyclic-AMP levels in the heart with important consequences for cardiac contractility. The increased nucleoside diphosphate kinase membrane content in human heart failure could potentially counteract a fading beta adrenoceptor response in the early stages of heart failure by increasing the amount of G Alpha stimulatory proteins in the plasma membrane. However, by switching from stimulatory to G Alpha inhibitory to activation, nucleoside diphosphate kinase may play a role in heart failure progression by reducing cyclic-AMP levels, typical for end-stage human heart failure.                                                 The study, therefore contributes to a better understanding of the molecular processes, underlying alter G Protein signaling in heart failure, and may help to develop new heart failure therapies.                                                 The next study tested the hypothesis, that high intensity interval training is superior to moderate continuous training in reversing cardiac remodeling and increasing aerobic capacity in patients with heart failure and reduced ejection fraction.                                                 Doctor Ellingson and colleagues from the Norwegian University of Science and Technology, performed a multicenter trial, comparing twelve weeks of supervised interventions of high-intensity interval training at 90 to 95% maximal heart rate, moderate continuous training at 60 to 70% maximal heart rate, or a recommendation of regular exercise in 261 patients with heart failure and ejection fraction less than 35%, in New York Heart Association class II or III status.                                                 The primary end point of change in left ventricular end-diastolic diameter from baseline to twelve weeks was not different between the high-intensity and moderate continuous groups. There was also no difference between the high-intensity and moderate groups in peak oxygen uptake, although both were superior to the recommendation for regular exercise. None of these changes were maintained at follow up after 52 weeks. Serious adverse events were not statistically different. However, training records showed that 51% of patients exercised below the prescribed target, during supervised high-intensity interval training, and 80% above the recommended target in those with moderate continuous training. Given that high-intensity interval training was not superior to moderate continuous training, in reversing remodeling or improving secondary end points, and considering that adherence to the prescribed exercise intensity based on heart rate was difficult to achieve even in the supervised setting.                                                 The authors concluded that moderate continuous training remains the standard exercise modality for patients with chronic heart failure.                                                 The final paper tells us, that brain emboli after left ventricular endocardial ablation may be more common than we knew. First author Doctor Whitman, corresponding author Doctor Marcus and colleagues from University of California studied eighteen consecutive patients, scheduled for ventricular tachycardia or premature ventricular contraction ablation, over a nine month period. Twelve patients undergoing left ventricular ablation were compared to a control group of six patients, undergoing right ventricular ablation only. Heparin was administrated with a goal activated clotting time of 300 to 400 seconds for all left ventricular procedures. Pre impulse procedural brain magnetic resonance imaging was performed on each patient within a week of the ablation procedure. The authors found that seven of the twelve patients, or 58% undergoing left ventricular ablation, experienced a total of sixteen cerebral emboli, compared with none among patients undergoing right ventricular ablation. Seven of the eleven patients undergoing a retrograde approach to the left ventricle, developed at least one new brain lesion. Thus, more than half of patients undergoing routine left ventricular ablation procedures, experienced new brain emboli after the procedure, even in the absence of clinically apparent stroke.                                                 Future research is critical to understanding the long-term consequences of these lesions and to determine optimal strategies to avoid them. This is further discussed in an editorial entitled "The Sound of Silence". How much noise should we make about post ablation silence strokes? By Doctor Z and Vora from Stanford University. Well, those were your summaries, now for our featured discussion.                                                 I am so thrilled to have with me two special guests to discuss the topic of the diagnosis of heart failure preserved ejection fraction or HFpEF. As you all know, that's my favorite topic and I have favorite people with me today. First, the corresponding author of our feature paper, Doctor Barry Borlaug from Mayo Clinic, Rochester, Minnesota. And, for the first time on the podcast, Doctor Mark Drazner, Senior Associate Editor from UT Southwestern. So, welcome Barry and Mark. Barry Borlaug:                    Thank you Carolyn. Mark Drazner:                   Thank you, great to be here. Dr. Carolyn L.:                    So, Barry, you talked about the role of stress diastolic testing, shall I call it, in the  diagnosis of HFpEF in your paper. Could you tell us why you looked at it and what you found? Barry Borlaug:                    Sure, Carolyn. When you have dyspnea and fatigue and you got a low EF, it's pretty easy to make the diagnosis of heart failure reduced EF, but we've been struggling for years with making the diagnosis of dyspnea, whether it's HFpEF or not in people with normal ejection fraction. And that's because physical and laboratory and clinical signs of high filling pressures and congestion, are either difficult to see or only present during stress, like physical exercise, in patients. So that's really what motivated us to pursue this study.                                                 We took patients, that were referred to our cath lab for invasive hemodynamic exercise testing, so we directly measured filling pressures, PA pressures and cardiac output reserve, to get a gold standard assessment, whether people have heart failure or not. And then we performed simultaneous echocardiography and blood testing to measure NT-proBNP levels, and then we just looked at what we could figure out. Can you accurately discern HFpEF patients from patients without cardiac dyspnea, using these non invasive estimates.                                                 We saw that a lot of people, with, for example, NT-proBNP levels that are low enough to be where most would consider HFpEF excluded, actually had HFpEF. And we saw that there were modest correlations between non-invasive echocardiographic estimates of filling pressures, specifically the E to E Prime ratio, and directly measured left heart filling pressures. But when we applied the criteria that had been initially proposed, we saw poor sensitivity to make the diagnosis with exercise. And this was largely related to the difficulty with getting all of the different echocardiographic indices, that are currently examined as part of the diastolic stress testing non-invasively. Next, we looked at just adding the exercise E to E Prime, which is an estimate of filling pressures, and when we used the cut-point, that's already been proposed, according to contemporary data, we found that this substantially improved the sensitivity to identify HFpEF, but there was a bit of a trade-off in that specificity decrease. Dr. Carolyn L.:                    That's so cool. So let me summarize some of these take-home messages here. First of all, using just rest echo. I was really impressed to see that rest echo indices alone only identifies a third to maybe up to 60% of the patients you found with invasively proven HFpEF. So, we may be specific, but we're really missing quite a number of patients. And then if you exercise them, what your data is really showing is that it's better to exercise them and use this data for the negative predictive value, isn't that what you're saying? Barry Borlaug:                    You know, the exercise is really the gold standard, so it gives you both, the negative and positive. With the echocardiography, relying on the exercise E to E Prime ratio, that was really helping us, as you say, Carolyn, with the higher negative predictive value. So most people, that had HFpEF, in this series, where we could get adequate, highly controlled environments, adequate diagnostic echocardiographic data, most people that ended up having HFpEF fit those criteria, we could see an elevation in this E to E Prime on exercise, so it did provide good negative predictive value. Dr. Carolyn L.:                    These are just such important data, because I think we're all still struggling with how to make this diagnosis of HFpEF. Mark, could you just share some thoughts on whether you think this is going to really change practice, even change guidelines? Mark Drazner:                   I think, if you read this paper, you would recognize it, that it's certainly a critical question that we're all facing, how to make the diagnosis of HFpEF. And all of their guidelines that have been advocated, there really wasn't much data, and these really are the best data out there. So, certainly, it's [inaudible 00:15:41] me a direction of changing practices. Barry says, certainly, the approach will need to be validated, I think, before it reaches high level guidelines, but certainly I think it's a step in the right direction, and points the way towards the future in terms of improving our ability to diagnose HFpEF. And really, that's why both reviewers and [inaudible 00:15:59] this is such an important paper. Dr. Carolyn L.:                    Right. Barry, I have a quick question for you though. Doing exercise echo, not easy. E to E Primes are all over the place usually. How easy was it? How feasible was this test? Barry Borlaug:                    So, first I'd like to say that we have outstanding, very well-trained, very highly skilled research scenographers, here at Mayo doing this. In very controlled environment, we're providing plenty of time for them to obtain images and that's going to be a question moving forward, because not everybody in clinical practice has that capability. But with that said, in this very controlled environments, skilled scenographers, we were able to measure the exercise data during low level exercise about 85 to 90% of the time and at peak exercise about 75 to 80% of the time. So, it's fairly feasible, but even in this best case scenario, we can't get it on everybody. Mark Drazner:                   Even in the [inaudible 00:19:49] echo lab, the recommended approach by the ASE with the four measures. How many times they were not able to acquire all those images, are necessary for those four techniques and so, here you have a [inaudible 00:20:03] of AS echo lab not being able to do that, and being transparent about that, and [inaudible 00:20:08] to the community, saying that, although these are ideal measures, even the [inaudible 00:20:12] perhaps you can't acquire them. I think that was another important point that came out of this and then lead to the focus on the E to E Prime. Barry Borlaug:                    I couldn't agree more. You got one of the world renowned labs, very skilled scenographers doing imaging, and we're still not able to get it all in each patient, and that just points to the difficulty of getting really high quality diagnostic images, and a lot of time you need the next level test, when that happens. And invasive exercise testing is really that test, the gold standard. Mark Drazner:                   When you get echos from the outside and you look at the E to E Primes, are you confident that the data, that's generally acquired, is gonna be acceptable for this [inaudible 00:20:50]? Barry Borlaug:                    Yes and no, I mean I'm always a little bit concerned, but it's not just being a control freak, you know, wanting to see everything, but I think that if it's a still frame doppler, tissue doppler spectrum, you can see that the sample volume is in the right place, and it's really unequivocally normal or abnormal, I feel pretty good about that. Not as good as when they get a full dedicated study here. Mark Drazner:                   Of course, the gold standard is also difficult. The invasive measurement. Barry Borlaug:                    Yes it is, I didn't [inaudible 00:21:18] that, but we've been doing a lot of invasive exercise tests for the last ten years now. And we do like 250 a year here, so we're really quite [inaudible 00:21:28] but we have all hands on deck in the lab. We have a couple technicians running gas samples around, all over the place. Somebody is on the medgraphics card, measuring oxygen consumption. We've got a nurse in there, that's helping out, so it's complicated, and of course we're using the micromanometer catheters for the pressure assessments, because you get so much more artifacts from width and under damping and over damping with the pressure tracing, so that's also not easy to do if you say. Mark Drazner:                   So maybe for practicing cardiologists it's gonna be hard to duplicate that and perhaps spend the energy in terms of doing the exercise echo techniques off the speed, for example. Perhaps, it's another message. Barry Borlaug:                    I would agree completely. And I think that again, when you do that, if you do a really high quality exercise echo and it's still not quite definitive, then you can refer on to a center that does have that capability, because obviously it's just reality, not everybody is going to be able to do this. Not every place has the size and resources to be able to do these really advanced tasks.   Dr. Carolyn L.:                    And do you apply exercise echo now in making your diagnosis? How do you use this data, for yourself, clinically? Barry Borlaug:                    We started to think about this, and I think that the best case scenario where the  people, that really have an intermediate pretest probability, based on their clinical characteristics. Somebody has jugular distension and a very high NT-proBNP level, and edema, you really don't need further testing, you know that that's going to be HFpEF. And if somebody has no risk factors, and everything is stone cold normal, they don't.                                                 But in some of these people that have some signals, but they don't quite meet criteria, we are doing this, again, if they have adequate echocardiographic images at rest. And then we're looking really carefully at the exercise echocardiography data, one concern from this data, I want to make sure people are very circumspect and really critically looking at the quality of their data, because we shouldn't over-interpret equivocal findings. And as you said earlier, E to E Primes can be all over the map, they're very difficult to obtain during exercise. But I think that if everything looks very high quality and is definitely abnormal or definitely normal, that can be helpful. More so, if it's normal. We     did see more false positive, so if it is abnormal, we did suggest that you may want to perform further confirmatory testing, because of the higher false positive rate with exercise echo. Mark Drazner:                                                                   I would say for the listeners, they should take a look at his figure six, which really is a nice diagnostic algorithm, where Barry shows, or advocates, for taking patients with intermediate probability and then using this to restratify that, using [inaudible 00:19:40] approach. I know that, that figure resonated with the editors and the reviewers dramatically, so I'd encourage listeners to take a look at that. Dr. Carolyn L.:                    Listeners, you heard it right. [inaudible 00:22:36] Circulation on the Run. Don't forget to tell all your friends about this podcast and tune in next week.  

Biochemistry (BIO/CHEM 4362) - Winter 2016
2b. How Adrenaline Signaling Works (G-Protein Coupled Receptors Part 1)

Biochemistry (BIO/CHEM 4362) - Winter 2016

Play Episode Listen Later Jan 8, 2016 23:55


Biochemistry (BIO/CHEM 4362) - Winter 2016
2b. How Adrenaline Signaling Works (G-Protein Coupled Receptors Part 1)

Biochemistry (BIO/CHEM 4362) - Winter 2016

Play Episode Listen Later Jan 8, 2016 23:56


Description Not Provided.

Biochemistry (BIO/CHEM 4362) - Winter 2016
2c. How Sight, Smell, and Taste Work (G-Protein Coupled Receptors Part 2)

Biochemistry (BIO/CHEM 4362) - Winter 2016

Play Episode Listen Later Jan 8, 2016 24:23


Biochemistry (BIO/CHEM 4362) - Winter 2016
2c. How Sight, Smell, and Taste Work (G-Protein Coupled Receptors Part 2)

Biochemistry (BIO/CHEM 4362) - Winter 2016

Play Episode Listen Later Jan 8, 2016 24:23


Description Not Provided.

Movement Disorders Journal Podcasts 2012-2015
Dopaminergic regulation of olfactory type G-protein α subunit expression in the striatum

Movement Disorders Journal Podcasts 2012-2015

Play Episode Listen Later Aug 27, 2015 2:58


regulation expression olfactory subunit g protein striatum
OIRT Undergraduate Research Spotlight
G-Protein Couple Receptor Expression in Tumors

OIRT Undergraduate Research Spotlight

Play Episode Listen Later Feb 24, 2015 2:23


Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 17/19
Der Stammzellmarker LGR5 (Leucine-Rich Repeat G-protein Coupled Receptor 5) übernimmt keine tumortreibende Funktion in der kolorektalen Karzinogenese

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 17/19

Play Episode Listen Later Dec 4, 2014


Thu, 4 Dec 2014 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/17726/ https://edoc.ub.uni-muenchen.de/17726/1/Blumrath_Kira.pdf Blumrath, Kira

keine funktion coupled receptor leucine g protein ddc:600 karzinogenese rich g
Medizin - Open Access LMU - Teil 22/22
The G-Protein-Coupled Estrogen Receptor (GPER/GPR30) in Ovarian Granulosa Cell Tumors

Medizin - Open Access LMU - Teil 22/22

Play Episode Listen Later Sep 1, 2014


Ovarian granulosa cell tumors (GCTs) are thought to arise from cells of the ovarian follicle and comprise a rare entity of ovarian masses. We recently identified the G-protein-coupled estrogen receptor (GPER/GPR30) to be present in granulosa cells, to be regulated by gonadotropins in epithelial ovarian cancer and to be differentially expressed throughout folliculogenesis. Thus, supposing a possible role of GPER in GCTs, this study aimed to analyze GPER in GCTs. GPER immunoreactivity in GCTs (n = 26; n (primary diagnosis) = 15, n (recurrence) = 11) was studied and correlated with the main clinicopathological variables. Positive GPER staining was identified in 53.8% (14/26) of GCTs and there was no significant relation of GPER with tumor size or lymph node status. Those cases presenting with strong GPER intensity at primary diagnosis showed a significant reduced overall survival (p = 0.002). Due to the fact that GPER is regulated by estrogens, as well as gonadotropins, GPER may also be affected by endocrine therapies applied to GCT patients. Moreover, with our data supposing GPER to be associated with GCT prognosis, GPER might be considered as a possible confounder when assessing the efficacy of hormone-based therapeutic approaches in GCTs.

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 17/19
Charakterisierung cAMP-unabhängiger Effektoren des Thyreotropin-Rezeptors in humanen Schilddrüsenkarzinom-Zelllinien

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 17/19

Play Episode Listen Later Jun 24, 2014


Der humane Thyreotropin-Rezeptor (TSH-R) steuert die zentralen Funktionen der Schilddrüse und ist der wichtigste Regulator für deren Wachstum und Differenzierung. Er gehört zur Superfamilie der G-Protein-gekoppelten Rezeptoren (GPCR) und kann nach einer Stimulation mit TSH die G-Proteine aller vier Familien (Gs, Gi/o, Gq/11 und G12/13) aktivieren. Dabei werden die meisten zellulären Reaktionen wie die Proliferation der Schilddrüsenepithelzellen und die Bereitstellung der Schilddrüsenhormone einer Aktivierung von Gs zugeordnet. Zu Gs-unabhängigen Signalwegen des TSH-R war dagegen erst wenig bekannt. Da der Gq/11-vermittelte Signalweg durch die Aktivierung der Phospholipase C und Proteinkinase C in maligne Prozesse von Schilddrüsenzellen involviert sein könnte, sollten in der vorliegenden Arbeit Gq/11-abhängige Effektoren des TSH-R in humanen Schilddrüsenkarzinomzellen identifiziert und näher charakterisiert werden. Als Modellsystem wurden FTC 133 wt TSH-R Zellen verwendet, eine follikuläre Schilddrüsenkarzinom-Zelllinie, die den humanen TSH-R überexprimiert. In diesen wurde die Aktivierung des Calcium/Calcineurin-abhängigen Transkriptionsfaktors NFAT nach TSH-Stimulation erstmalig beschrieben. Bei einer anschließenden Reihenuntersuchung der NFAT-abhängigen Zielgene Autotaxin, VEGF, c-Myc, Regulator von Calcineurin 1 (RCAN1) und Cyclooxygenase-2 (Cox-2) wurden c-Myc, RCAN1 und Cox-2 als TSH-regulierte Gene identifiziert. Die Induktion von c-Myc war unabhängig von NFAT, dagegen bestätigten Expressionsstudien mit Calcineurin-Inhibitoren und dem spezifischen NFAT-Inhibitor INCA-6, dass RCAN1 und Cox-2 durch eine NFAT-Aktivierung induziert wurden. Diese Aktivierung wurde durch Gq/11-Proteine vermittelt, denn nach spezifischer Herunterregulation der Gq- und G11-α-Untereinheiten mittels siRNA konnten die Zielgene nicht mehr TSH-abhängig induziert werden. Weitere Analysen zum Mechanismus der NFAT-Aktivierung zeigten, dass eine Erhöhung der intrazellulären Calciumionenkonzentration ([Ca2+]i) allein über intrazelluläre Speicher nicht ausreichend war. Um NFAT zu aktivieren, mussten zusätzlich Calciumionen aus dem Extrazellulärraum einströmen. Untersuchungen mit dem STIM1-Inhibitor SKF-96365 wiesen dabei auf einen Calciumioneneinstrom über Speicher-operierte Ionenkanäle hin. Zusätzlich zur NFAT-regulierten Genexpression wurde in dieser Arbeit die TSH-induzierte Expression des Metallothioneins MT1X in FTC 133 wt TSH-R Zellen und in primären Thyreozyten analysiert. Die mRNA Induktion dieses Cystein-reichen und zytoprotektiven Proteins war ebenfalls abhängig von einer Expression der Gq/11-Proteine. Eine Erhöhung der [Ca2+]i reichte jedoch nicht aus, um MT1X signifikant zu induzieren. Zur gesteigerten Expression war darüber hinaus auch die Aktivierung der Proteinkinase C notwendig. In der vorliegenden Dissertation konnten somit RCAN1, Cox-2 und MT1X als Gq/11-regulierte Zielgene des humanen TSH-R charakterisiert werden. Dabei wurde eine NFAT-regulierte Genexpression nach einer TSH-Stimulation erstmalig gezeigt. Die präsentierten Ergebnisse weisen damit auf eine bisher unbeachtete biologische Rolle von Gq/11-abhängigen Signalwegen des humanen TSH-R in der Schilddrüse hin.

er camp arbeit dabei rolle bei expression gi gq ergebnisse zur wachstum ftc cox prozesse zus reaktionen regulators unabh funktionen gs proteins stimulation erh dissertation untersuchungen tsh speicher proliferation proteine aktivierung mechanismus schilddr differenzierung bereitstellung vegf myc sirna charakterisierung ca2 g12 g protein genexpression zelllinien rezeptors eine erh ddc:600 extrazellul signalweg zelllinie die induktion cystein nfat cyclooxygenase calcineurin protein kinase c effektoren signalwegen zielgene ionenkan weitere analysen expressionsstudien g proteine herunterregulation
Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 16/19
Akzessorische Bindungspartner G-Protein-gekoppelter Rezeptoren: Untersuchungen zur Protein-Expression in der nicht-malignen humanen Prostata

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 16/19

Play Episode Listen Later Feb 20, 2014


Thu, 20 Feb 2014 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/16734/ https://edoc.ub.uni-muenchen.de/16734/1/Miersch_Josef.pdf Miersch, Josef

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 16/19
Liganden-abhängige Desensitisierung und pro-algetische Signalwege des hMrgX1-Rezeptors

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 16/19

Play Episode Listen Later Dec 10, 2013


Der humane Mas-related gene X1 (hMrgX1)-Rezeptor ist ein G-Protein-gekoppelter Rezeptor, der selektiv in nozizeptiven Spinalganglienneuronen exprimiert wird. Eine spezifische Aktivierung des Rezeptors durch das Proenkephalin-Spaltprodukt BAM8-22 (bovine adrenal medulla 8-22) wird als schmerzhaft wahrgenommen. Damit stellt der hMrgX1-Rezeptor eine neue molekulare Zielstruktur für eine potentiell nebenwirkungsarme, analgetische Therapie dar. Trotz dieses Potentials sind die pro-algetischen Signalwege des hMrgX1-Rezeptors bislang nicht verstanden. Der MrgX1-Rezeptor entwickelte sich unter hohem positivem Selektionsdruck und kommt nur in Primaten vor. Trotzdem wurden die nicht-homologen MrgC-Rezeptoren der Nagetiere zur Analyse des hMrgX1-Rezeptors verwendet. Im Rahmen dieser Arbeit wurden daher zunächst vergleichende Ligandenprofile des hMrgX1- und der MrgC-Rezeptoren aus Maus und Ratte erstellt. Dabei wurden deutliche Unterschiede offensichtlich, da der hMrgX1-Rezeptor exklusiv von BAM8-22 aktiviert wurde, während die MrgC-Rezeptoren durch weitere Liganden, u. a. Spaltprodukte des Proopiomelanocortins, z. T. sogar effizienter aktiviert wurden. Zudem konnte gezeigt werden, dass die MrgC-vermittelte Ca2+-Mobilisation auf Grund einer β-Arrestin-abhängigen Rezeptorendozytose deutlich desensitisierte, während der hMrgX1-Rezeptor resistent gegenüber dieser Liganden-induzierten Regulation war. Daher können die MrgC-Rezeptoren der Nagetiere nicht als Modellsytem für den hMrgX1-Rezeptor verwendet werden, so dass in dieser Arbeit weiterführend Signalwege des hMrgX1-Rezeptors in Spinalganglienneuronen-ähnlichen F11-Zellen und primären Spinalganglienneuronen untersucht wurden. Dabei zeigte sich eine duale funktionelle Regulation des etablierten pro-algetischen TRPV1 (transient receptor potential cation channel vanilloid 1)-Ionenkanals. Zum einen sensitisierte der hMrgX1-Rezeptor den TRPV1 über einen etablierten, Proteinkinase C-abhängigen Signalweg. Zum anderen zeigte sich eine direkte hMrgX1-mediierte Aktivierung des TRPV1. Dieser Regulationsmechanismus wurde durch eine Phospholipase C (PLC)-β-induzierte Produktion des endogenen TRPV1-Liganden Diacylglycerol und durch die Degradation des tonisch TRPV1-inhibierenden PLC-β-Substrates Phosphatidylinositol-4,5-bisphosphat vermittelt. Neben der TRPV1-Modulation induzierte der hMrgX1-Rezeptor die Expression verschiedener Gene, deren zentrale Bedeutung bei der inflammatorischen und neuropathischen Schmerzchronifizierung etabliert ist. Einerseits wurde eine hMrgX1-induzierte Phosphorylierung der extracellular signal-regulated kinases 1/2 beobachtet, die in einer Aktivierung von serum response factor-abhängigen Reportergenkonstrukten und in der Induktion von c-Fos auf mRNA- und von early growth response protein 1 auf mRNA- und Proteinebene resultierte. Andererseits zeigte sich die transkriptionelle Ca2+/Calcineurin-abhängige Aktivierung des nuclear factor of activated t cells, die in der Induktion des CCR2 (chemokine receptor 2) auf mRNA- und Proteinebene resultierte. Somit konnte erstmalig ein physiologischer Induktor des CCR2 in Spinalganglienneuronen beschrieben werden. Weiterhin wurde nach der Etablierung der endogenen Proteinexpression des hMrgX1-Rezeptors in LAD2-Mastzellen eine BAM8-22-induzierte Freisetzung des CCR2-Agonisten chemokine ligand 2 ermittelt, so dass der hMrgX1-Rezeptor die parakrine Stimulation von nozizeptiven Spinalganglienneuronen durch Mastzellen fördern könnte. Diese Dissertation trägt somit zum besseren molekularen Verständnis akuter und chronischer pro-algetischer Funktionen des hMrgX1-Rezeptors bei und könnte damit die Entwicklung neuer analgetischer Wirkstoffe ermöglichen.

arbeit dabei grund entwicklung damit bedeutung regulation neben verst expression analyse trotzdem zudem trotz zum unterschiede daher mrna produktion therapie maus funktionen somit weiterhin stimulation einerseits andererseits plc potentials degradation aktivierung fos etablierung ratte wirkstoffe primaten freisetzung induktion rezeptor nagetiere g protein signalwege trpv1 liganden rezeptors phosphorylierung ddc:600 ccr2 proteinebene signalweg mastzellen proteinexpression diese dissertation protein kinase c selektionsdruck induktor zielstruktur phospholipase c plc
Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 16/19
Die Bradykinin B1 und B2 Rezeptoren als Modell für die Untersuchung der Regulation G-Protein-gekoppelter Rezeptoren

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 16/19

Play Episode Listen Later Dec 5, 2013


Die Familie A der G-Protein-gekoppelten Rezeptoren (GPCRs) bildet die größte und vielfältigste aller Transmembranrezeptorfamilien. Ihre Mitglieder spielen eine wesentliche Rolle in fast allen (patho)physiologischen Prozessen. Nach Agonistenbindung aktivieren GPCRs, wie ihr Name andeutet, heterotrimere G-Proteine aber auch G-Protein-unabhängige Signalwege. Die verschiedenen aktiven G-Proteinuntereinheiten (Gα-GTP und βγ) induzieren nach Dissoziation vom Rezeptor entsprechende Signalkaskaden z.B. über Phospholipase A und Cβ. Um eine Fehlregulation zellulärer Prozesse z.B. durch „Überstimulation“ zu verhindern, unterliegen GPCRs strengen Regulationsmechanismen, die ihre Fähigkeit zur Signaltransduktion und ihre Verfügbarkeit an der Zelloberfläche bestimmen. Die Bradykininrezeptoren B1 und B2 (B1R, B2R) gehören zur Familie A der GPCRs, also zu den Rhodopsin-ähnlichen GPCRs, und werden durch die pro-inflammatorischen Peptide desArg9-Bradykinin/desArg10-Kallidin (DABK/DAK) bzw. Bradykinin (BK)/Kallidin aktiviert. Im Gegensatz zum konstitutiv exprimierten B2R, der nach Stimulation schnell desensitisiert und internalisiert wird, erfolgt eine B1R-Expression fast ausschließlich unter pathophysiologischen Bedingungen über Induktion durch Zytokine. Nach Stimulation wird der B1R nicht internalisiert, sondern verbleibt an der Zelloberfläche. Beide Rezeptoren koppeln sowohl an Gαq/11 als auch an Gαi und aktivieren somit weitgehend identische Signalwege [vor allem Phospholipase Cβ (PLCβ) und „mitogen activated protein kinase“ (MAPK)-Kaskaden]. Durch ihre - besonders im Hinblick auf ihre Internalisierungs-eigenschaften - konträre Regulation, stellen die Bradykininrezeptoren ein interessantes Modell zur Untersuchung regulatorischer Mechanismen und deren Einflüsse auf die Signalübertragung von GPCRs dar. Beide Bradykininrezeptoren spielen bei inflammatorischen Prozessen eine Rolle. Sie fördern die Ausschüttung pro-inflammatorischer Zytokine und rekrutieren Immunzellen. Während entzündlicher Ereignisse kommt es zu erhöhter Zytokinfreisetzung z.B. von Interleukin-1β (IL-1β) und dadurch zur de novo Synthese von B1R. Pro-inflammatorische Zytokine wie IL-1β, die zur B1R-Expression führen, induzieren unter anderem aber auch einen Anstieg der Körpertemperatur (Fieber), eine häufige Begleiterscheinung inflammatorischer Vorgänge. Trotz des bekannten Zusammenhangs zwischen Inflammation und erhöhter Temperatur war über den Einfluss eines Temperaturanstiegs auf Membranrezeptoren und ihre Signalvermittlung auf zellulärer Ebene bisher nur sehr wenig bekannt. In dieser Arbeit wurde - unseres Wissens nach - erstmals auf die Temperatur als regulatorische Komponente für GPCR-vermittelte Signalübertragung eingegangen. Am Beispiel der Bradykininrezeptoren wurde gezeigt, dass die Stärke der Signalübertragung von GPCRs signifikant durch eine Temperaturerhöhung von 37°C auf 41°C beeinflusst werden kann. Hierbei war jedoch zwischen einer Temperaturabhängigkeit des Signalwegs selbst und einer rezeptorspezifischen Temperatursensitivität zu unterscheiden. So wurde die Aktivierung von ERK1/2 unter pathophysiologisch erhöhter Temperatur (41°C; normale Körpertemperatur: 37°C) signifikant gesteigert, unabhängig davon ob sie durch B1 oder B2 Rezeptoren stimuliert wurde. Die gesteigerte Aktivität PLCβ-vermittelter Signalkaskaden bei 41°C konnte hingegen auf eine nur für den B1R spezifische Temperaturabhängigkeit zurückgeführt werden. Diese Beobachtung zusammen mit der Tatsache, dass die B1R-Expression unter pathophysiologischen Bedingungen besonders induziert wird, deutet darauf hin, dass der B1R in Kombination mit Fieber eine verstärkte Wirkung im Organismus haben könnte. Ob diese Heilungs-fördernd oder -abträglich wirkt, müsste noch genauer untersucht werden. Neben dem Einfluss der Temperatur wird die Signalübertragung der GPCRs durch die jeweiligen Rezeptorkonformationen und die sich daraus ergebenden Funktionsunterschiede bestimmt. Die Familie A der GPCRs wird durch einige hoch konservierte Strukturmerkmale wie die E/DRY-Sequenz mit R3.50 in der dritten Transmembrandomäne (TM) oder die NPXXY-Sequenz am Ende der siebten TM charakterisiert. Publizierte Ergebnisse deuten darauf hin, dass bovines Rhodopsin durch eine Salzbrücke zwischen R3.50135 (TM3) und E6.30247 (TM6), auch „ionic lock“ genannt, im inaktiven Zustand gehalten wird. Der B2R ist einer der wenigen Peptid-GPCRs, der ein Glutamat an Position 6.30 (E6.30238) trägt, und eignete sich daher zur Untersuchung der Anwesenheit und Funktion eines möglichen „ionic lock“ auch in „nicht-Rhodopsin“-GPCRs. Für alle bisher entsprechend untersuchten GPCRs ist bekannt, dass R3.50 für eine effiziente G-Protein-Aktivierung unabdingbar ist (selbiges wurde in der vorliegenden Arbeit auch für den B2R bestätigt). Die funktionelle Analyse eines „ionic lock“ anhand einer R3.50 Mutation und G-Protein-abhängiger Kaskaden ist deshalb nicht möglich. Die Rolle eines „ionic lock“ im Hinblick auf G-Protein-unabhängige Mechanismen wie die Rezeptorinternalisierung, einem wichtigen Regulationsschritt für die meisten GPCRs, wurde bisher jedoch noch nicht untersucht. In der vorliegenden Arbeit wurde erstmals gezeigt, dass die Rezeptorendozytose durch Mutation von R3.50128 zu Alanin (R3.50128A), im Gegensatz zur G-Protein-Aktivierung, nicht zum Erliegen kommt. Die mutierten Rezeptorkonstrukte wiesen sogar ein konstitutives Internalisierungsverhalten auf. Dies verwies auf unterschiedliche Funktionen dieser Aminosäure bei der G-Protein-vermittelten Signaltransduktion und bei der Rezeptorinternalisierung. Ein Aufbrechen des möglichen „ionic lock“ durch Mutation von E6.30238 zu Alanin oder Arginin resultierte ebenfalls in konstitutiv internalisierenden Rezeptorkonstrukten. Im Gegensatz zur Endozytose zeigten diese Mutanten zwar keine konstitutive Signalübertragung, wurden aber auch durch prinzipiell als Antagonisten klassifizierte Verbindungen effizient aktiviert. Diese Ergebnisse legen einen mehrstufigen Aktivierungsprozess nahe, dessen Stufen sich durch verschiedene Rezeptorkonformationen mit unterschiedlichen Interaktionsmöglichkeiten für die G-Protein-Rekrutierung/Aktivierung oder mit der Internalisierungsmaschinerie [GPCR-Kinasen (GRKs), Arrestine] auszeichnen. Der wechselseitige Austausch der beiden hoch konservierten Aminosäuren R3.50128 und E6.30238 ermöglichte wahrscheinlich die Bildung eines inversen „ionic lock“, wodurch normales B2R-Verhalten wieder hergestellt wurde. Diese Arbeit zeigt somit erstmals, dass ein Aufbrechen eines möglichen „ionic lock“ in einem Peptidrezeptor unterschiedliche Auswirkungen für die Prozesse der G-Protein-Aktivierung und der Rezeptorendozytose haben kann. Dadurch wird die Annahme bestärkt, dass es bei einem GPCR mehrere aktive Konformationen geben kann, die unterschiedliche Affinitäten gegenüber regulatorischen Proteinen (GRKs, Arrestinen) oder Effektoren (G-Proteinen, Arrestinen) aufweisen und dadurch differenziert zelluläre Signale auslösen können. Die Aufklärung der unterschiedlichen Aktivierungsmechanismen von GPCRs in Kombination mit der Herstellung von Verbindungen z.B. sogenannten „small molecule compounds“, die bestimmte Rezeptorkonformationen mit ihren signalspezifischen Eigenschaften stabilisieren können, wäre möglicherweise für die Entwicklung von Agonisten oder Antagonisten, die nur ganz bestimmte Signalwege modulieren und so eine optimierte therapeutische Anwendung erlauben, hilfreich.

Medizin - Open Access LMU - Teil 21/22
Human Mas-related G protein-coupled receptors-X1 induce chemokine receptor 2 expression in rat dorsal root ganglia neurons and release of chemokine ligand 2 from the human LAD-2 mast cell line

Medizin - Open Access LMU - Teil 21/22

Play Episode Listen Later Mar 7, 2013


Primate-specific Mas-related G protein-coupled receptors-X1 (MRGPR-X1) are highly enriched in dorsal root ganglia (DRG) neurons and induce acute pain. Herein, we analyzed effects of MRGPR-X1 on serum response factors (SRF) or nuclear factors of activated T cells (NFAT), which control expression of various markers of chronic pain. Using HEK293, DRG neuron-derived F11 cells and cultured rat DRG neurons recombinantly expressing human MRGPR-X1, we found activation of a SRF reporter gene construct and induction of the early growth response protein-1 via extracellular signal-regulated kinases-1/2 known to play a significant role in the development of inflammatory pain. Furthermore, we observed MRGPR-X1-induced up-regulation of the chemokine receptor 2 (CCR2) via NFAT, which is considered as a key event in the onset of neuropathic pain and, so far, has not yet been described for any endogenous neuropeptide. Up-regulation of CCR2 is often associated with increased release of its endogenous agonist chemokine ligand 2 (CCL2). We also found MRGPR-X1-promoted release of CCL2 in a human connective tissue mast cell line endogenously expressing MRGPR-X1. Thus, we provide first evidence to suggest that MRGPR-X1 induce expression of chronic pain markers in DRG neurons and propose a so far unidentified signaling circuit that enhances chemokine signaling by acting on two distinct yet functionally co-operating cell types. Given the important role of chemokine signaling in pain chronification, we propose that interruption of this signaling circuit might be a promising new strategy to alleviate chemokine-promoted pain.

IFOM Podcasts
Rap G Protein Signaling in Hematologic Malignancy

IFOM Podcasts

Play Episode Listen Later Feb 5, 2013 27:04


IFOM Podcasts
Rap G Protein Signaling in Hematologic Malignancy

IFOM Podcasts

Play Episode Listen Later Feb 5, 2013 27:04


Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 04/06
Biomimetic screening of class B G protein-coupled receptors

Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 04/06

Play Episode Listen Later Jul 12, 2012


Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis is a hallmark of complex and multifactorial psychiatric diseases such as anxiety and mood disorders. The 41-amino acid neuropeptide Corticotropin Releasing Hormone (CRH) is a major regulator of the mammalian stress response. Upon stressful stimuli, it binds to the Corticotropin Releasing Hormone Receptor 1 (CRHR1), a typical member of the class B GPCRs and a potential novel target for the therapeutic intervention in major depressive disorder. A precise understanding of the peptide-receptor interactions is an essential prerequisite towards the development of efficient CRHR1 specific antagonists. To chemically probe the molecular interaction of CRH with its cognate receptor, a high-throughput conjugation approach which mimics the natural activation mechanism for class B GPCRs was developed. Acetylene-tagged peptide libraries were synthesized and conjugated to high-affinity azide-modified carrier peptides using copper-catalyzed dipolar cycloaddition. The resulting conjugates reconstitute potent ligands and were tested in situ for modulation of the CRHR1 activity in a cell-based assay. This approach allows to (i) define the sequence motifs which are required for receptor activation or inhibition, (ii) identify the critical functional groups and investigate structure-activity-relationships, and (iii) develop novel optimized, highly potent peptide probes which are specific for the transmembrane domain of the receptor. The membrane recruitment by a high-affinity carrier peptide enhances the potency of tethered peptides and allows the initial testing of weak fragments that otherwise would be inactive. The biomimetic screening led to the discovery of transtressin, a highly modified and potent CRHR1 transmembrane domain-specific optimized agonist (EC50 = 4 nM). Beyond its intrinsic agonistic activity, transtressin is an essential tool for the pharmacological characterization of CRHR1 antagonists in competition assays.

Science Signaling Podcast
Science Signaling Podcast, 27 March 2012

Science Signaling Podcast

Play Episode Listen Later Mar 26, 2012 14:37


Regulated protein synthesis and degradation mediate skeletal muscle homeostasis.

Science Signaling Podcast
Science Signaling Podcast, 20 September 2011

Science Signaling Podcast

Play Episode Listen Later Sep 19, 2011 13:43


Activation of the small GTPase Rho downstream of a chemokine receptor is required for metastasis of some breast cancer cells.

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 06/19
Identifizierung von extrazellulären Bindungspartnern des Tumorendothelmarkers (TEM) 5 und Charakterisierung seiner Funktion bei der Angiogenese

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 06/19

Play Episode Listen Later Feb 26, 2007


Tumorendothelmarker (TEM) 5 gehört zu den Adhäsions-G-Protein-gekoppelten Rezeptoren und wird in Endothelzellen während der physiologischen Angiogenese und Tumorangiogenese exprimiert. Bisher wurden noch kein Ligand und keine Funktion von TEM5 beschrieben. In dieser Arbeit wurde gezeigt, dass TEM5 in Endothelzellen während der In Vitro-Angiogenese intrazellulär an einer konservierten Proteolysestelle (GPS) gespalten wird. Diese Proteolyse führte einerseits zur Translokation der nicht-kovalent verbunden TEM5-Fragmente an die Zelloberfläche und andererseits zur Freisetzung von löslichem TEM5 (sTEM5). Bindungsstudien haben ergeben, dass sTEM5 mit verschiedenen Glykosaminoglykanen interagiert. Sequenzanalysen und funktionelle und biochemische Studien haben gezeigt, dass sTEM5 eine kryptische RGD-Bindungsstelle für Integrin alpha(v)beta(3) enthält. Matrixmetalloprotease 9-prozessiertes, jedoch nicht unprozessiertes, sTEM5 vermittelte Endothelzelladhäsion durch direkte Interaktion mit Integrin alpha(v)beta(3). Die Interaktion von immobilisiertem proteolytisch prozessierten sTEM5 mit Integrin alpha(v)beta(3) vermittelte Überleben von Wachstumsfaktor-deprivierten Endothelzellen. Die Ergebnisse dieser Arbeit führen zu der Schlußfolgerung, dass sTEM5 während der Angiogenese von Endothelzellen freigesetzt wird und an Glykosaminoglykane in der extrazellulären Matrix und auf der Oberfläche von Zellen bindet. Proteolytische Prozessierung von sTEM5 führt zur Freilegung seines RGD-Motivs und vermittelt Überleben von Endothelzellen durch die Interaktion mit Integrin alpha(v)beta(3).

matrix arbeit funktion studien bisher oberfl interaktion seiner die ergebnisse adh identifizierung charakterisierung freisetzung ligand g protein die interaktion integrin endothelzellen translokation ddc:600 extrazellul angiogenese sequenzanalysen bindungsstudien
Fakultät für Biologie - Digitale Hochschulschriften der LMU - Teil 02/06
Proinflammatorische und prothrombotische Wirkung von MT-SP1/Matriptase über Aktivierung des Protease-aktivierten Rezeptors-2 in Endothelzellen

Fakultät für Biologie - Digitale Hochschulschriften der LMU - Teil 02/06

Play Episode Listen Later May 30, 2006


Atherosklerose wird heute als entzündliche Gefäßerkrankung verstanden, an deren Beginn ein Funktionsverlust des Endothels steht. Genablationsversuche zeigen, dass der Protease-aktivierte Rezeptor 2 (PAR-2) eine Rolle bei der Vermittlung inflammatorischer Reaktionen des Endothels spielt. PAR-2 gehört zur Familie der heptahelikalen G-Protein-gekoppelten Rezeptoren und wird durch proteolytische Spaltung seines N-Terminus aktiviert. Zusätzlich zu bekannten PAR-2-Liganden wie Trypsin und Gerinnungsfaktoren Xa und Tissue Factor/VIIa wurde mittels positional scanning synthetic combinatorial library die Typ II transmembrane Serinprotease Matriptase/MT-SP1 als PAR-2-aktivierende Protease identifiziert. MT-SP1/Matriptase wird bislang ausschließlich eine Rolle bei Tumorinvasion und Metastasierung zugeschrieben. In der vorliegenden Arbeit wurde eine entzündungsfördernde Wirkung der katalytischen Domäne von MT-SP1/Matriptase in primären Gefäßendothelzellen und der daran beteiligte Rezeptormechanismus untersucht. MT-SP1/Matriptase induzierte die de novo-Synthese der proinflammatorischen Mediatoren Interleukin-8 (IL-8), IL-6 und Monocyte Chemoattractant Protein (MCP)-1 abhängig von der katalytischen Aktivität und über die Aktivierung von PAR-2. Die MT-SP1/Matriptase-induzierten Signalwege beinhalteten die Aktivierung des Transkriptionsfaktors NF-kB in Abhängigkeit von der Aktivität der MAPK p38 und p42/44. Die IL-8-Induktion durch MT-SP1/Matriptase erforderte dabei lediglich die Aktivität von p38. Zusätzlich wurde ein zweiter, PKCalpha-abhängiger Signalweg zur MT-SP1/Matriptase-induzierten IL-8-Expression nachgewiesen, der unabhängig von p38, p42/44 und NF-kB war. Die endotheliale Dysfunktion in der Atherosklerose kennzeichnet sich nicht nur durch Inflammation, sondern auch durch prothrombotische Veränderungen. MT-SP1/Matriptase induzierte zusätzlich zu inflammatorischen Zytokinen die Neusynthese des Gerinnungsfaktors Tissue Factor und könnte dadurch proatherogen wirken. Tissue Factor selbst induzierte wiederum IL-8 unabhängig von seinem Liganden FVIIa, aber abhängig von den Serinresten 253 und 258 in der zytoplasmatischen Domäne des Rezeptors. Expressionsstudien zeigten die erhöhte Expression von MT-SP1/Matriptase in der endothelialen Innenwand atherosklerotischer Gefäße im Vergleich zu gesundem Gefäß. Auch am Endothel adhärierte Blutzellen wiesen MT-SP1/Matriptase-Expression auf. Die Basalexpression von MT-SP1/Matriptase war nicht in Endothelzellen, aber in Monozyten nachweisbar, die im atherosklerotischen Prozess mit dem Endothel interagieren können. MT-SP1/Matriptase könnte daher eine Rolle bei der PAR-2-vermittelten Entzündungsreaktion in der atherosklerotischen Gefäßwand spielen.

arbeit gef rolle familie dom beginn expression prozess wirkung vergleich inflammation aktivit zus reaktionen abh spaltung vermittlung entz aktivierung synthese dysfunktion rezeptoren induktion protease nf kb rezeptor mapk g protein ddc:500 blutzellen signalwege atherosklerose liganden metastasierung endothelzellen monozyten rezeptors trypsin endothel zytokinen signalweg ddc:570 typ ii n terminus endothels funktionsverlust expressionsstudien tissue factor neusynthese
Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 02/06
Expression, Isolierung und Funktionsanalyse des OR5 Geruchsrezeptors: einem Vertreter der G-Protein gekoppelten Rezeptoren

Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 02/06

Play Episode Listen Later Feb 13, 2006


Die Familie der Geruchsrezeptoren ist zwar ein zentrales Forschungsgebiet, dennoch ist wenig über sie bekannt. Im Folgenden wird dargestellt, worin die beiden Hauptursachen für die Problematik der Analyse von Geruchsrezeptoren bestehen und welche Strategien in dieser Arbeit gewählt wurden, um einen experimentellen Ansatz zur Untersuchung von Geruchsrezeptoren zu finden: 1. Erst wenigen Geruchsrezeptoren konnten Liganden zugeordnet werden . Da noch keine Struktur eines Geruchsrezeptors bekannt ist, können bislang Modellvorstellungen nur über Sequenzhomologien innerhalb der Rezeptorgruppe oder anhand verwandter Rezeptoren, z. B. dem Rinderopsin, erstellt werden. Offensichtlich reichen diese Modelle jedoch nicht aus, um effektiv Liganden zuzuordnen oder Struktur-Funktion-Zusammenhänge zu erkennen. Das HEK-293-Zellsystem erwies sich zwar als das bisher effektivste heterologe Expressionssystem für diese Art von Rezeptoren, doch auch hier ist die Zahl beschriebener, funktionell exprimierter ORs begrenzt . Es gibt also nur wenige Möglichkeiten, Geruchsrezeptoren funktionell zu exprimieren, und daher besteht ein Bedarf an weiteren Expressionssystemen, um diese Rezeptoren in vivo untersuchen zu können. 2. Neben der Problematik einer funktionellen Expression gibt es bislang kein Expressionssystem, welches die Herstellung geeigneter Mengen für eine Strukturaufklärung dieser Rezeptoren ermöglicht. Die Menge der synthetisierten Rezeptoren ist in der Regel zu gering oder die Zielproteinspezies liegt in Einschlusskörpern vor. Zu 1) Die funktionelle Expression mit korrekter Translokation des Membranproteins sollte in Kombination mit einer Semliki Forest Virus-basierten Infektion in Säugetier P19-Zellen durchgeführt werden. Ausgewählt wurde diese Zelllinie aufgrund folgender Tatsachen: es handelt sich bei der P19-Zelllinie um Teratokarzinom-Zellen, welche ursprünglich aus embryonalen Stammzellen, implantiert in Hodengewebe, generiert wurden. Einem Gewebe also, in dem in vivo eine Geruchsrezeptor-Expression nachgewiesen wurde . Ein weiterer vielversprechender Umstand war die Differenzierbarkeit dieser Fibroblasten-ähnlichen Zellen in neuronale Zellen, dem Zelltyp, der auch in vivo für olfaktorische Neuronen vorliegt. Dementsprechend lautet die Annahme, dass es sich um eine optimale Zelllinie für die Expression rekombinanter ORs handeln kann, die zur zeitgerechten Expression aller für die olfaktorische Signalkaskade notwendigen Bestandteile befähigt ist. Die Zelllinie sollte bezüglich ihrer Eignung als Expressionsplattform für Geruchsrezeptoren charakterisiert und Expressionsstudien am Beispiel des Rezeptors OR5 durchgeführt werden. Zu 2) Die Überexpression des Membranproteins zur quantitativen Isolierung erfolgte in Hefe. Gegenüber E. coli ist der Hefeorganismus zur Durchführung posttranslationaler Modifikationen fähig. Ein Vorteil im Vergleich zu Säugerzellen sind die hohen erreichbaren Zelldichten. Primärgewebe kam für diese Fragestellung nicht in Frage: eine Isolierung wäre mit sehr geringen Ausbeuten verbunden, eine Problematik, die durch die Verwendung von heterologen Expressionssystemen umgangen werden kann. Es wurde eine „unfolded protein response“ (UPR)-kontrollierte Expression in Saccharomyces cerevisiae ausgewählt, um zu gewährleisten, dass überwiegend korrekt gefaltetes Rezeptorprotein gebildet wird. Mit dieser Arbeit sollten erstmals durch eine optimierte Expression, Produktion und Isolierung ausreichende Proteinmengen des Geruchsrezeptors OR5 (aus R. norvegicus) mit einer Homogenität von >90% zur Verfügung gestellt werden, um biochemische und strukturelle Charakterisierungen durchzuführen. Zusätzlich sollten monoklonale OR5-Antikörper generiert werden, um eine spezifische Detektion und Immunopräzipitation des Geruchsrezeptors OR5 zu ermöglichen. Zusammengefasst tragen die etablierten Systeme dazu bei, die genaue Rolle der ORs in der Geruchswahrnehmung in Zukunft entschlüsseln zu können. Mit Hilfe der P19-Zelllinie wird die OR-Charakterisierung in einem heterologen System ermöglicht, und durch den Gebrauch des Hefeexpressionsystem und die optimierte Isolierungs-Strategie kann das Material für eine Strukturaufklärung bereitgestellt werden.

Tierärztliche Fakultät - Digitale Hochschulschriften der LMU - Teil 02/07
Zum vasodilatierenden Effekt des antidiabetisch wirkenden Hintonia-latiflora-Extraktes: In-vitro- und In-vivo-Messungen

Tierärztliche Fakultät - Digitale Hochschulschriften der LMU - Teil 02/07

Play Episode Listen Later Jul 15, 2005


ZUSAMMENFASSUNG In dieser Arbeit wird zum ersten Mal nachgewiesen, dass ein Extrakt aus Hintonia latiflora, der unter dem Namen Sucontral® als Phytoantidiabetikum Anwendung findet, neben seiner Blutzucker-senkenden Wirkung auch einen vasodilatierenden Effekt besitzt und deshalb möglicherweise Diabetes-assozierte Gefäßveränderungen günstig beeinflusst. Diese Gefäß-erweiternde Wirkung wird sowohl in vitro an der isolierten Aorta von Meerschweinchen als auch in vivo am wachen Kaninchen gezeigt. An isolierten Aortenringen können Noradrenalin-induzierte (also G-Protein-vermittelte) Kontraktionen durch den Hintonia-latiflora-Extrakt nahezu vollständig relaxiert werden (Relaxation um ca. 99 % bei einer Konzentration von 60 µg/ml). Die EC50 liegt bei 51,98 µg/ml. Kontraktionen, die dagegen durch Erhöhung der extrazellulären Kalium-Konzentration erzeugt werden, reagieren wesentlich schwächer auf die Zugabe des Extraktes. Selbst bei der hohen Konzentration von 300 µg/ml kommt es nur zu einer geringen Relaxation von ca. 19 %. Die Kalium-induzierte Kontraktion wird in erster Linie durch eine Kalium-induzierte Depolarisation, eine Aktivierung der membranären Calcium-Kanäle und einen daraus resultierenden erhöhten Calcium-Einstrom in die Zelle ausgelöst. Aufgrund dieser Ergebnisse kommt als Wirkungsmechanismus für die Gefäßrelaxation nicht eine Calcium-Kanal-Blockade, sondern primär eine Hemmung des G-Protein-vermittelten intrazellulären Calcium-Anstiegs in Frage. Auch ein Inhaltsstoff des Extraktes, das Neoflavonoid Coutareagenin, das hauptsächlich für die Blutzucker-senkende Wirkung des Extraktes verantwortlich gemacht wird, zeigt die vasodilatierende Wirkung (Relaxation bei Noradrenalin-induzierter Kontraktion um ca. 94 % bei einer Konzentration von 60 µg/ml) im In-vitro-Testsystem. Der vermutete Wirkmechanismus wird durch fluoreszenzmikroskopische Versuche an gezüchteten Aortenzellen der Ratte mit dem Calcium-Indikator Fura-2 bestätigt. In diesen Versuchen wird Vasopressin verwendet, das ebenfalls über G-Proteine auf die Zelle wirkt. Der dadurch ausgelöste Calcium-Transient wird durch Coutareagenin (60 µg/ml) stark reduziert (um ca. 72 %), was für eine Hemmung des G-Protein-vermittelten intrazellulären Calcium-Anstiegs spricht. Auch das Ergebnis, dass Hintonia latiflora keinen wesentlichen negativen Einfluss auf die Kontraktionskraft des isolierten Herzmuskels zeigt, spricht gegen eine Beeinflussung von Spannungs-abhängigen Calcium-Kanälen und stützt die G-Protein-Hypothese. Im In-vivo-Teil der Studie werden Untersuchungen an wachen Kaninchen durchgeführt, wobei eine Hintonia-latiflora-Gruppe und eine Kontroll-Gruppe (n=8) erstellt wird. Nach Bestimmung der Basalwerte erfolgt die einmalige orale Applikation des Hintonia-latiflora-Extraktes bzw. die Gabe der Kontrollsubstanz (Lösungsvermittler Macrogol 300®). Sowohl an der Aorta abdominalis wie auch an der A. carotis communis kann sonographisch eine Zunahme des Gefäßdurchmessers bei gleichzeitiger Abnahme der Blutflussgeschwindigkeit unter dem Einfluss des Extraktes nachgewiesen werden. Zusätzlich kann auch eine signifikante Blutzucker-senkende Wirkung in der Hintonia-latiflora-Gruppe gezeigt werden. Eine einmalige Applikation ruft also am gesunden und wachen Tier sowohl eine Blutzuckersenkung als auch eine tendenzielle Vasodilatation hervor. Diese Ergebnisse sind bemerkenswert, da in bisherigen Studien meist davon ausgegangen wurde, dass die Effekte des Extraktes erst bei länger dauernder wiederholter Anwendung sichtbar werden. Der gezeigte vasodilatierenden Effekt könnte möglicherweise nützlich sein, um Spätschäden des Diabetes mellitus wie Mikro- und Makroangiopathien zu mindern. Aus diesem Grund könnte es sinnvoll sein, den Hintonia-latiflora-Extrakt besonders bei Typ 2 Diabetikern einzusetzen, um Antidiabetika vom Sulfonylharnstoff-Typ einzusparen und gleichzeitig im Hinblick auf Mikro- und Makroangiopathien durch Vasodilatation günstige hämodynamische Bedingungen zu schaffen.

Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 02/06
Investigation on the EGFR transactivation by G protein coupled receptors in cancer cells

Fakultät für Chemie und Pharmazie - Digitale Hochschulschriften der LMU - Teil 02/06

Play Episode Listen Later May 3, 2005


Tue, 3 May 2005 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/3749/ https://edoc.ub.uni-muenchen.de/3749/1/Marg_Beatrice.pdf Marg, Beatrice ddc:540, ddc:500, Fakultät für Che

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 03/19
Untersuchungen über die Expression von Early-Response Genen und intrazelluläre Signalübertragungswege nach Verletzung humaner vaskulärer Endothelzellmonolayer in vitro

Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 03/19

Play Episode Listen Later Nov 11, 2004


Perkutane Interventionsverfahren stellen ein etabliertes und wichtiges Behandlungskonzept der Arteriosklerose dar. Ihr Ergebnis wird getrübt durch hohe Restenoseraten nach z.B. Ballon-Angioplastie. Wesentlich für die Ausbildung und den Grad einer postinterventionellen Restenose ist die Reendothelialisierung der zerstörten Intima, weshalb es von besonderem Interesse ist, intrazelluläre Signalübertragungswege, welche nach Zerstörung eines Endothelzellmonolayers in den überlebenden Zellen ablaufen, zu untersuchen, um potentielle Angriffspunkte z.B. für eine pharmakologische Modulation der Endothelzellantwort nach Angioplastie zu finden. In der vorliegenden Arbeit wurden diese Signalübertragungswege an einem Modell zur Verletzung vaskulärer Endothelzellmonolayer in vitro untersucht. Das Hauptaugenmerk lag dabei auf Untersuchung des an der Regulation der Zellproliferation beteiligten Transkriptionsfaktors und Proto-Oncogens AP-1 bzw. dessen mRNA-Vorstufe c-fos. An humanen vaskulären Endothelzellen (HUVEC) wurde in Zellkultur mittels Northern-Blot Analysen und Electrophoretic mobility shift assays untersucht, ob c-fos und AP-1 nach Endothelzellverletzung exprimiert bzw. aktiviert werden. Es konnte gezeigt werden, dass die Endothelzellverletzung im vorliegenden Modell eine starke Expression von c-fos sowie die Aktivierung des korrespondierenden Transkriptionsfaktors AP-1 bewirkt. Diese Aktivierung wird gesteuert durch einen Anstieg der intrazellulären Calcium-Konzentration sowie unter Beteiligung negativ regulierender G-Protein, der Proteinkinase C sowie von Protein-Tyrosinkinasen. Keine Beteiligung konnte nachgewiesen werden für MAPKinasen, Proteinkinase A sowie die CamKinasen. Die Ergebnisse dieser Arbeit liefern in Zusammenschau mit publizierten Ergebnissen anderer Arbeitsgruppen wichtige Erkenntnisse für eine weiterführende Untersuchung der Signalübertragungswege in huamnen Endothelzellen nach z.B. perkutaner Angioplastie und führen in Zukunft zu neuen Ansatzpunkten für die Pharmakotherapie vaskulärer Läsionen, z.b. durch drug-eluting Stents.

zukunft arbeit regulation signal expression erkenntnisse interesse grad ausbildung modell zerst ergebnissen verletzung beteiligung untersuchung untersuchungen anstieg zellen die ergebnisse vitro modulation aktivierung genen intima wesentlich stents arbeitsgruppen arteriosklerose das hauptaugenmerk g protein zellkultur endothelzellen die expression angriffspunkte vaskul ddc:600 behandlungskonzept zusammenschau pharmakotherapie zellproliferation ansatzpunkten protein kinase c angioplastie restenose calcium konzentration endothelzellen huvec
Medizinische Fakultät - Digitale Hochschulschriften der LMU - Teil 01/19

Reaktive – Sauerstoff - Spezies (ROS) spielen in der Physiologie und Pathophysiologie des vaskulären Systems eine wichtige Rolle. So kommt es z.B. bei Hypertonie, Atherosklerose, Ischämie / Reperfusion und weiteren Krankheiten und Stoffwechselstörungen, wie z.B. Hypercholesterinämie und Diabetes mellitus zu einem Ungleichgewicht zwischen Sauerstoffradikalbildung und anti - oxidativen Mechanismen. Superoxidanionen (O2 -) spielen insofern eine besondere Rolle, als sie durch direkte Interaktion endotheliales NO inaktivieren, so daß es seine vasodilatatorische, anti – proliferative und plättchenaggregationshemmende Funktion nicht mehr voll erfüllen kann. Damit ist O2 - maßgeblich an der Induktion der Endotheldysfunktion beteiligt. Bei Beginn dieser Arbeit gab es erste Hinweise, daß eine der leukozytären NAD(P)H - Oxidase ähnlichen Oxidase auch im Endothel existiert und wesentlich zur endothelialen O2 - - Bildung beiträgt. Wenig erforscht waren jedoch die Regulationsmechanismen dieser Oxidase. Ein bisher noch nicht bekannter Stimulus zur Steigerung der endothelialen O2 - - Bildung wurde 1996 beschrieben. In Endothelzellen aus bovinen Pulmonararterien führte eine Depolarisation zu einer gesteigerten O2 - - Bildung. Dies kann insofern von Bedeutung sein, als es sowohl unter physiologischen, als auch pathophysiologischen Bedingungen zu akuten oder chronischen Veränderungen des endothelialen Membranpotentials kommt. In dieser Arbeit wurde nun untersucht, ob eine NAD(P)H – Oxidase in der Tat auch in humanen Endothelzellen vorhanden ist, ob sie im Gegensatz zur leukozytären Form konstitutiv aktiv ist, und welchen Beitrag sie zur basalen endothelialen O2 - - Bildung leistet. Weitere Untersuchungen in HUVEC sollten zeigen, ob und wie sich sowohl De – als auch Hyperpolarisation der Zellmembran auf die O2 - - Bildung auswirken, welches Enzym hierbei eine Rolle spielt und welche Signaltransduktionsmechanismen beteiligt sind. Zur O2 - - Messung an vaskulären Zellen war die Verwendung der Lucigenin – Chemilumineszenz – Methode etabliert, so daß auch hier anfänglich mit dieser Methode gearbeitet wurde. Da jedoch dann Befunde veröffentlicht wurden, die zeigten, daß Lucigenin in Enzymsyste-men, die sonst kein oder nur wenig O2 - produzieren, zu einer erheblichen O2 - - Bildung führte, mußte mit weiteren Methoden der O2 - - Messung überprüft werden, ob diese Nachteile auch unter unseren Versuchsbedingungen auftraten. Verwendet wurden hierzu die MCLA – verstärkte Chemilumineszenz, die NBT – und Cytochrom C – Methode. Mit diesen verschiedenen, voneinander unabhängigen Methoden zeigte sich, daß in Anwesenheit von NADH Lucigenin selbst zu einer wesentlich gesteigerten O2 - - Bildung in Lysaten von humanen Umbilikalvenenendothelzellen (HUVEC) führt. Daher wurde zur Untersuchung der endothelialen O2 - - Bildung in dieser Arbeit schließlich nur die Cytochrom C Methode verwendet. Zur Überprüfung der Auswirkungen der verwendeten Substanzen auf das Membranpotential wurde die Membranpotentialmeßmethode mittels dem Potential – sensitiven Fluoreszenzfarbstoff Bis - oxonol aufgebaut und verwendet. Intakte HUVEC zeigten eine basale O2 - - Produktion, die durch bekannte Inhibitoren der leukozytären NAD(P)H – Oxidase, mit unterschiedlichen Wirkmechanismen signifikant gehemmt wurde (Diphenyleniodonium ca. 48%, Phenylarsenoxid ca. 34% ). Ebenso resultierte die Inaktivierung des GTP - bindenden - Proteins rac mit Clostridium difficile Toxin B in einer signifikanten Reduktion der basalen endothelialen O2 - - Produktion um ca. 30%. Weiterhin konnte gezeigt werden, daß nach Aufhebung der zellulären Integrität durch das Lysieren der HUVEC die Gabe von NADH eine um ca. 2.7 fach erhöhte O2 - - Produktion im Vergleich zu NADPH bewirkte. Mit Hilfe der Immunfluoreszenz bzw. rtPCR konnten außerdem zumindest ein Teil der leukozytären NAD(P)H – Oxidase Untereinheiten, p67phox und gp91phox auch in HUVEC nachgewiesen werden. Zur gezielten Depolarisation des Membranpotentials wurden ein Puffer mit erhöhter Kaliumkonzentration (90 mM), der nicht selektive Kalium – Kanal - Blocker Tetrabutylammonium Chlorid (1 mM) und das Kation – Ionophor Gramicidin (1 µM) verwendet. Die basale endotheliale O2 - - Produktion wurde durch diese Substanzen in ähnlichem Ausmaß (~ 60% ) signifikant gesteigert (n=23, p