title: Role of membrane depolarization and T-type Ca2+ channels in angiotensin II and K+ stimulated aldosterone secretion.
abstract: The hypothesis that Ca2+ influx necessary for angiotensin II (AngII) and K+ stimulation of aldosterone secretion is primarily mediated by membrane depolarization and activation of T-type Ca2+ channels was examined in isolated rat adrenal glomerulosa cells. Perforated-patch clamp recordings of membrane potential (Vm) demonstrated that AngII and K+ induce concentration-dependent depolarizations capable of activating T channels and, at high K+ and AngII concentrations, activating L channels and inactivating T channels. K+-induced depolarizations were stable and readily reversible. Vm was proportional to K+ concentration, exhibiting a linear slope of 53.7 mV per 10-fold increase in K+. AngII-induced depolarizations were complex, consisting of a slow maintained component superimposed with small amplitude depolarizing fluctuations. Slow oscillations in Vm were occasionally observed in response to 10(-9) M AngII or greater. The slow, maintained component of depolarization coincided with inhibition of K+ conductance. Neither rapid fluctuations nor slow oscillations in Vm were blocked by mibefradil or other treatments that inhibit voltage-gated Ca2+ channels. Perforated-patch clamp experiments also demonstrated that AngII (10(-8) M) inhibited L channels by 45.6% without affecting T channels. Thus AngII activates T channels by depolarization rather than T channel modulation in rat cells. The concentration dependencies of mibefradil inhibition of T channels and AngII- and K+-induced aldosterone secretion were compared. Under whole-cell patch clamp mibefradil induced a concentration-dependent inhibition of T channels, exhibiting a K(app) of 0.62 microM. Mibefradil inhibition was use-dependent but mibefradil neither acted as an open channel blocker nor significantly affected T channel inactivation or activation. Mibefradil inhibited K+- and AngII-induced secretion at concentrations similar to that for T channel inhibition; at high concentrations (10 microM) mibefradil inhibited AngII-induced secretion by 88% and completely inhibited K+-induced secretion. The IC50 for K+-induced secretion was dependent on K+ concentration, increasing from 0.2 microM for 6 mM K+ to 2.5 microM for 10 mM K+ or greater. Mibefradil exhibited an IC50 of 1.1 microM for inhibition of secretion at all AngII concentrations examined (0.1, 1.0, and 10 nM). Mibefradil also exhibited multiple nonspecific effects, which complicated the assessment of T channel function, including; inhibition of leak and voltage-dependent K+ conductances, inhibition of Ca2+-independent aldosterone secretion, and inhibition of secretion under conditions expected to completely inactivate T channels (10 nM AngII or 20 mM K+). In summary, these results indicate that voltage-gated T channels represent the primary Ca2+ influx pathway activated by physiological concentrations of AngII and K+ but other Ca2+ influx pathways must mediate aldosterone secretion induced by high K+ or AngII concentrations.
authors.affiliation: Department of Biological Sciences, Northern Illinois University, DeKalb, IL 60115, USA. dlotshaw@niu.edu
authors.forename: D P
authors.initials: DP
authors.lastname: Lotshaw
mesh_terms: D000450:Aldosterone / Q000378:metabolism*; D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020747:Calcium Channels, T-Type / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D018929:Cell Culture Techniques; D004305:Dose-Response Relationship, Drug; D005260:Female; D007700:Kinetics; D008564:Membrane Potentials / Q000502:physiology; D020748:Mibefradil / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015384:Zona Glomerulosa / Q000166:cytology
chemical_list: D002121:Calcium Channel Blockers; D020747:Calcium Channels, T-Type; D000804:Angiotensin II; D020748:Mibefradil; D000450:Aldosterone; D011188:Potassium; D002118:Calcium