Distinct functions of cardiac β-adrenergic receptors in the T-tubule vs. outer surface membrane
Uloženo v:
| Název: | Distinct functions of cardiac β-adrenergic receptors in the T-tubule vs. outer surface membrane |
|---|---|
| Autoři: | George WP Madders, Marion Barthé, Flora Lefebvre, Emilie Langlois, Florence Lefebvre, Patrick Lechêne, Maya Dia, Xavier Iturrioz, Catherine Llorens-Cortes, Tâp Ha-Duong, Laurence Moine, Nicolas Tsapis, Rodolphe Fischmeister |
| Zdroj: | eLife, Vol 14 (2025) |
| Informace o vydavateli: | eLife Sciences Publications Ltd, 2025. |
| Rok vydání: | 2025 |
| Sbírka: | LCC:Medicine LCC:Science LCC:Biology (General) |
| Témata: | T-tubules, membrane compartmentation, β-adrenergic receptors, PEGylation, cardiomyocyte, cyclic AMP, Medicine, Science, Biology (General), QH301-705.5 |
| Popis: | β-Adrenoceptors (β-ARs) regulate cardiac function during sympathetic nerve stimulation. β-ARs are present in both the cardiac T-tubule (TTM) and outer surface membrane (OSM), but how their location impacts their function is unknown. Here, we developed a technology based on size exclusion to explore the function of β-ARs located in the OSM. We synthesized a PEG-Iso molecule by covalently linking isoprenaline (Iso) to a 5000 Da PolyEthylene-Glycol (PEG) chain to increase the size of the β-AR agonist and prevent it from accessing the T-tubule network. The affinity of PEG-Iso and Iso on β1- and β2-ARs was measured using radioligand binding. Molecular dynamics simulation was used to assess PEG-Iso conformation and visualize the accessibility of the Iso moiety to water. Using confocal microscopy, we show that PEGylation constrains molecules outside the T-tubule network of adult rat ventricular myocytes (ARVMs) due to the presence of the extracellular glycocalyx. β-AR activation in OSM with PEG-Iso produced a lower stimulation of [cAMP]i than Iso but a larger stimulation of cytosolic PKA at equivalent levels of [cAMP]I and similar effects on excitation–contraction coupling parameters. However, PEG-Iso produced a much lower stimulation of nuclear cAMP and PKA than Iso. Thus, OSM β-ARs in ARVMs control mainly cytosolic cAMP/PKA pathway and contractility, while TTM β-ARs control mainly nuclear cAMP, PKA, and consequent nuclear protein phosphorylation. Size exclusion strategy using ligand PEGylation provides a unique approach to evaluate the respective contribution of T-tubule vs. OSM proteins in cardiac cells. |
| Druh dokumentu: | article |
| Popis souboru: | electronic resource |
| Jazyk: | English |
| ISSN: | 2050-084X |
| Relation: | https://elifesciences.org/articles/84243; https://doaj.org/toc/2050-084X |
| DOI: | 10.7554/eLife.84243 |
| Přístupová URL adresa: | https://doaj.org/article/98886af2e72f4fbd9b1f52a583f2021d |
| Přístupové číslo: | edsdoj.98886af2e72f4fbd9b1f52a583f2021d |
| Databáze: | Directory of Open Access Journals |
Full Text 
Full Text Finder 
Nájsť tento článok vo Web of Science | Abstrakt: | β-Adrenoceptors (β-ARs) regulate cardiac function during sympathetic nerve stimulation. β-ARs are present in both the cardiac T-tubule (TTM) and outer surface membrane (OSM), but how their location impacts their function is unknown. Here, we developed a technology based on size exclusion to explore the function of β-ARs located in the OSM. We synthesized a PEG-Iso molecule by covalently linking isoprenaline (Iso) to a 5000 Da PolyEthylene-Glycol (PEG) chain to increase the size of the β-AR agonist and prevent it from accessing the T-tubule network. The affinity of PEG-Iso and Iso on β1- and β2-ARs was measured using radioligand binding. Molecular dynamics simulation was used to assess PEG-Iso conformation and visualize the accessibility of the Iso moiety to water. Using confocal microscopy, we show that PEGylation constrains molecules outside the T-tubule network of adult rat ventricular myocytes (ARVMs) due to the presence of the extracellular glycocalyx. β-AR activation in OSM with PEG-Iso produced a lower stimulation of [cAMP]i than Iso but a larger stimulation of cytosolic PKA at equivalent levels of [cAMP]I and similar effects on excitation–contraction coupling parameters. However, PEG-Iso produced a much lower stimulation of nuclear cAMP and PKA than Iso. Thus, OSM β-ARs in ARVMs control mainly cytosolic cAMP/PKA pathway and contractility, while TTM β-ARs control mainly nuclear cAMP, PKA, and consequent nuclear protein phosphorylation. Size exclusion strategy using ligand PEGylation provides a unique approach to evaluate the respective contribution of T-tubule vs. OSM proteins in cardiac cells. |
|---|---|
| ISSN: | 2050084X |
| DOI: | 10.7554/eLife.84243 |