Effects of expression of a mouse brain L-type calcium channel alpha 1 subunit on secretion from bovine adrenal chromaffin cells.
Regulated exocytosis from bovine chromaffin cells is stimulated by the influx of Ca2+ through plasma membrane ion channels that are opened by nicotinic stimulation and/or depolarization. Recently, we developed a novel method that enabled us to investigate the function of a cloned Ca2+ channel type C alpha 1 subunit in forming channels that stimulate exocytosis. In the present study, we demonstrate by immunocytochemistry that bovine chromaffin cells normally express an epitope specific for the type C alpha 1 subunit. We investigated the effects of expression of additional class C alpha 1 subunits (mouse brain clone) on various aspects of secretory function in bovine chromaffin cells by measuring secretion of cotransfected human growth hormone (GH, a reporter for the regulated secretory pathway in the transfected cells). New channels were activated in response to depolarization by both elevated K+ and nicotinic cholinergic agonist. The new channels had their greatest effects when secretion was stimulated suboptimally. Secretion was enhanced only after the first 30 sec of stimulation, and the enhancement extended beyond 5 min of continuous stimulation. In contrast to the endogenous L-type Ca2+ channels, the latency was not decreased by the dihydropyridine L-type Ca2+ channel agonist, Bay K 8644. The findings suggest that (i) the Ca(2+)-sensitive mechanism for triggering or maintaining exocytosis is capable of being saturated by high levels of Ca2+, (ii) secretion caused by nicotinic agonist stimulation can be significantly enhanced by activation of voltage-sensitive Ca2+ channels, and (iii) the effects on secretion of the L-type Ca2+ channels formed on expression of the mouse brain class C alpha 1 subunit are distinct from those of endogenous L-type Ca2+ channels.
Wick
P F
PF
Department of Pharmacology, University of Michigan Medical School, Ann Arbor 48109-0632, USA.
Westenbroek
R E
RE
Holz
R W
RW
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000313:Adrenal Medulla / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000096:biosynthesis / Q000737:chemistry / Q000502:physiology*; D020746:Calcium Channels, L-Type; D002395:Catecholamines / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D003001:Cloning, Molecular; D013006:Growth Hormone / Q000096:biosynthesis / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007700:Kinetics; D046911:Macromolecular Substances; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D018722:Nicotinic Agonists / Q000494:pharmacology; D010957:Plasmids; D011950:Receptors, Cholinergic / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D002395:Catecholamines; D046911:Macromolecular Substances; D018722:Nicotinic Agonists; D011950:Receptors, Cholinergic; D011994:Recombinant Proteins; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D013006:Growth Hormone; D002118:Calcium