Rapid desensitization of adrenaline- and neuropeptide Y-stimulated Ca2+ mobilization in HEL-cells.
1. Desensitization of Gs-coupled receptors, the beta 2-adrenoceptor for example, involves rapid and slower components but little is known regarding the existence of rapid desensitization of Gi-coupled receptors and its possible mechanisms. In HEL-cells stimulation of alpha 2A-adrenoceptors by adrenaline or Y1-like neuropeptide Y receptors by neuropeptide Y, transiently mobilizes Ca2+ from intracellular stores via a Gi-protein. We have used this model to study the existence and possible mechanisms of rapid desensitization of a Gi-mediated cellular response. 2. Following stimulation by adrenaline or neuropeptide Y Ca2+ levels returned towards baseline a few minutes after agonist addition and were refractory to a second agonist exposure demonstrating rapid desensitization. Cross-desensitization experiments with neuropeptide Y, adrenaline and moxonidine demonstrated the presence of homologous (both receptors) and heterologous desensitization (neuropeptide Y receptors only), and that the alpha 2A-adrenoceptor desensitization was not specific for phenylethylamine (adrenaline) or imidazoline agonists (moxonidine). 3. The protein kinase C activator, phorbol ester, rapidly desensitized the hormonal Ca2+ responses and inhibitors of protein kinase C enhanced the hormonal responses inconsistently. The tyrosine kinase inhibitor, herbimycin, enhanced Ca2+ mobilization by adrenaline and neuropeptide Y, whereas the protein phosphatase inhibitor, okdadaic acid, did not affect Ca2+ mobilization or its desensitization. 4. In the absence of extracellular Ca2+ the endoplasmic reticulum Ca(2+)-ATPase inhibitor, thapsigargin, reduced hormone-stimulated Ca2+ elevations, demonstrating that mobilization occurs from a thapsigargin-sensitive pool in the endoplasmic reticulum. The inositol phosphate-independent Ca2+release modulator, ryanodine, significantly enhanced adrenaline- and neuropeptide Y-stimulated Ca2+elevations. Blockade of the endoplasmic reticulum Ca2+-ATPase by thapsigargin in the presence of extracellular Ca2+ enhanced hormone-stimulated Ca2+ increases, demonstrating the importance of this enzyme for the termination of the Ca2+ signal.5. It is concluded that adrenaline and neuropeptide Y-stimulated Ca2+ mobilization in HEL-cells occurs from a thapsigargin- and ryanodine-sensitive store in the endoplasmic reticulum and desensitizes rapidly;this appears to involve multiple mechanisms including protein kinases, possibly acting on receptors, and Ca2+ release and sequestration mechanisms.
Michel
M C
MC
Dept. Medicine, University of Essen, Germany.
D058669:Adrenergic alpha-2 Receptor Antagonists; D000959:Antihypertensive Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002460:Cell Line; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D004837:Epinephrine / Q000494:pharmacology*; D016257:Fura-2; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D009478:Neuropeptide Y / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D011189:Potassium Chloride / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D018341:Receptors, Adrenergic, alpha-2 / Q000187:drug effects; D017476:Receptors, Neuropeptide Y / Q000037:antagonists & inhibitors / Q000187:drug effects; D012433:Ryanodine / Q000494:pharmacology; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin
D058669:Adrenergic alpha-2 Receptor Antagonists; D000959:Antihypertensive Agents; D007093:Imidazoles; D009478:Neuropeptide Y; D018341:Receptors, Adrenergic, alpha-2; D017476:Receptors, Neuropeptide Y; D013729:Terpenes; D012433:Ryanodine; D011189:Potassium Chloride; D019284:Thapsigargin; C043482:moxonidine; D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D019204:GTP-Binding Proteins; D000252:Calcium-Transporting ATPases; D002118:Calcium; D016257:Fura-2; D004837:Epinephrine