title: Effect of mibefradil on voltage-dependent gating and kinetics of T-type Ca(2+) channels in cortisol-secreting cells.
abstract: We have studied the effect of the Ca(2+) antagonist mibefradil on low voltage-activated T-type Ca(2+) channels in whole-cell patch clamp recordings from bovine adrenal zona fasciculata (AZF) cells. AZF cells are distinctive in expressing only T-type Ca(2+) channels, allowing the mechanism of pharmacological agents to be explored without interference from other Ca(2+) channels. The inhibition of T-type Ca(2+) channels by mibefradil was voltage- and use-dependent. When Ca(2+) currents were activated from holding potentials of -80 and -60 mV, mibefradil inhibited currents with IC(50) values of 1.0 and 0.17 microM, respectively. When T-type Ca(2+) current (I(T)) was activated from a holding potential of -90 mV in the presence of 2 microM mibefradil, a single voltage step to -10 mV inhibited I(T) by 16.2% +/- 2.9% (n = 10). With subsequent voltage steps, applied at 10-s intervals, block reached a steady-state value of 51.9% +/- 5.0% (n = 5). Mibefradil (1 microM) produced a leftward shift of 5.7 mV (n = 4) in the voltage-dependent steady-state availability curve such that T-type Ca(2+) channels inactivated at more negative potentials, but this drug did not change the voltage-dependence of T channel opening. Mibefradil failed to alter the kinetics of T channel activation, inactivation, or deactivation, but markedly slowed T channel recovery following an inactivating prepulse. Mibefradil inhibited adrenocorticotropin-stimulated cortisol secretion from AZF cells with an IC(50) value of 3.5 microM. These results show that mibefradil is a relatively potent antagonist of T-type Ca(2+) channels in cortisol-secreting cells. The enhanced potency of mibefradil with sustained or repetitive depolarizations, its shifting of the steady-state inactivation curve, and its slowing of recovery all indicate that this drug preferentially interacts with Ca(2+) channels in the open or inactivated state. The inhibition of cortisol secretion by mibefradil at concentrations similar to those that block I(T) is consistent with a requirement for these channels in corticosteroidogenesis.
authors.affiliation: Department of Pharmacology, The Ohio State University, College of Medicine and Public Health, Columbus, OH 43210-1239, USA.
authors.forename: J C
authors.initials: JC
authors.lastname: Gomora
authors.forename: L
authors.initials: L
authors.lastname: Xu
authors.forename: J A
authors.initials: JA
authors.lastname: Enyeart
authors.forename: J J
authors.initials: JJ
authors.lastname: Enyeart
mesh_terms: D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015220:Calcium Channels / Q000145:classification / Q000187:drug effects*; D002417:Cattle; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D006854:Hydrocortisone / Q000378:metabolism*; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects*; D020748:Mibefradil / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D015383:Zona Fasciculata / Q000187:drug effects / Q000378:metabolism
chemical_list: D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020748:Mibefradil; D000324:Adrenocorticotropic Hormone; D006854:Hydrocortisone