title: Regulation by L channels of Ca(2+)-evoked secretory responses in ouabain-treated chromaffin cells.
abstract: It is known that the sustained depolarisation of adrenal medullary bovine chromaffin cells (BCCs) with high K(+) concentrations produces an initial sharp catecholamine release that subsequently fades off in spite depolarisation persists. Here, we have recreated a sustained depolarisation condition of BCCs by treating them with the Na(+)/K(+) ATPase blocker ouabain; in doing so, we searched experimental conditions that permitted the development of a sustained long-term catecholamine release response that could be relevant during prolonged stress. BCCs were perifused with nominal 0Ca(2+) solution, and secretion responses were elicited by intermittent application of short 2Ca(2+) pulses (Krebs-HEPES containing 2 mM Ca(2+)). These pulses elicited a biphasic secretory pattern with an initial 30-min period with secretory responses of increasing amplitude and a second 30-min period with steady-state, non-inactivating responses. The initial phase was not due to gradual depolarisation neither to gradual increases of the cytosolic calcium transients ([Ca(2+)]c) elicited by 2Ca(2+) pulses in BBCs exposed to ouabain; both parameters increased soon after ouabain addition. Νifedipine blocked these responses, and FPL64176 potentiated them, suggesting that they were triggered by Ca(2+) entry through non-inactivating L-type calcium channels. This was corroborated by nifedipine-evoked blockade of the L-type Ca(2+) channel current and the [Ca(2+)]c transients elicited by 2Ca(2+) pulses. Furthermore, the plasmalemmal Na(+)/Ca(2+) exchanger (NCX) blocker SEA0400 caused a mild inhibition followed by a large rebound increase of the steady-state secretory responses. We conclude that these two phases of secretion are mostly contributed by Ca(2+) entry through L calcium channels, with a minor contribution of Ca(2+) entry through the reverse mode of the NCX.
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.|Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, C/ Arzobispo Morcillo, 4, 28029, Madrid, Spain.
authors: Ricardo
authors: R
authors: De Pascual
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.|Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, C/ Arzobispo Morcillo, 4, 28029, Madrid, Spain.
authors: Inés
authors: I
authors: Colmena
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
authors: Lucía
authors: L
authors: Ruiz-Pascual
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
authors: Andrés Mateo
authors: AM
authors: Baraibar
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.
authors: Javier
authors: J
authors: Egea
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain.|Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, C/ Arzobispo Morcillo, 4, 28029, Madrid, Spain.
authors: Luis
authors: L
authors: Gandía
authors: Instituto Teófilo Hernando, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo, 4, 28029, Madrid, Spain. agg@uam.es.|Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, C/ Arzobispo Morcillo, 4, 28029, Madrid, Spain. agg@uam.es.|Servicio de Farmacología Clínica, Instituto de Investigación Sanitaria, Hospital Universitario de la Princesa, Universidad Autónoma de Madrid, c/ Diego de León, 62, 28006, Madrid, Spain. agg@uam.es.
authors: Antonio G
authors: AG
authors: García
mesh_terms: D000814:Aniline Compounds / Q000494:pharmacology; D000818:Animals; D020746:Calcium Channels, L-Type / Q000378:metabolism*; D020013:Calcium Signaling; D002395:Catecholamines / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D010042:Ouabain / Q000494:pharmacology*; D010647:Phenyl Ethers / Q000494:pharmacology
chemical_list: D000814:Aniline Compounds; D020746:Calcium Channels, L-Type; D002395:Catecholamines; D004791:Enzyme Inhibitors; D010647:Phenyl Ethers; C430918:SEA 0400; D010042:Ouabain
keywords: Amperometry; Catecholamine release; Chromaffin cell; L-type Ca2+ channels; N-type Ca2+ channels; NCX; Ouabain; P/Q-type Ca2+ channels