AT1 calcium signaling in renal vascular smooth muscle cells.

Experiments were conducted to gain insight into calcium signaling mechanisms triggered by angiotensin II (AngII) stimulation in vascular smooth muscle cells (SMC) freshly isolated from preglomerular vessels of normotensive Wistar Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Cytosolic calcium concentration ([Ca2+]i) was measured using ratiometric Fura-2 fluorescence and a microscope-based photometer. Vascular SMC from preglomerular vessels were isolated and dispersed using an iron oxide-sieving method combined with collagenase treatment. AngII produced rapid increases in [Ca2+]i that remained elevated for the duration of continued stimulation. The same pattern of time response was observed in WKY and in SHR. AngII elicited dose-dependent increases in [Ca2+]i in groups of individual preglomerular arteriolar SMC from both strains. AngII (10(-10) M) induced an increase from baseline levels in WKY and SHR (37+/-9 and 32+/-13 nM; P < 0.05). In response to 10(-6) M AngII, steady-state responses were 165+/-30 and 170+/-35 nM (P < 0.01). The responses did not differ between strains (P > 0.4). The effects of AngII were inhibited by 88% by the AT1 receptor blocker candesartan in renal SMC. In SMC pretreated with calcium-free medium, baseline [Ca2+]i fell by about 60 nM. Thereafter, AngII did not elicit any [Ca2+]i response either in WKY or in SHR when calcium entry was prevented. Also, after prestimulation by AngII, a calcium-free solution completely reversed the effects of AngII. This study shows that AngII acts through AT1 receptors to stimulate [Ca2+]i by a predominant action on calcium entry with no evidence for calcium mobilization. Other studies have demonstrated that calcium entry in these SMC is mediated by voltage-gated, L-type entry channels sensitive to dihydropyridine agents. No strain differences were noted between the actions of AngII on individual renal SMC from SHR and normotensive control animals.