Mechanisms that regulate [Ca2+](i) following depolarization in rat systemic arterial smooth muscle cells

Abstract

1. We have used the patch-clamp technique in combination with fluorimetric recording to study the mechanisms that regulate intracellular Ca2+, [Ca2+](i), following depolarization in cells isolated from the rat femoral artery. 2. Depolarization to 0 mV from a holding potential of -70 mV increased [Ca2+](i). Little Ca2+ release from sarcoplasmic reticulum, SR, was detected during depolarization since application of 30 mM ryanodine, a Ca2+-release inhibitor, had no significant effect on total Ca2+ buffering power. 3. Upon repolarization to -70 mV, 7 out of 13 cells showed three phases of Ca2+ removal; an initial rapid first phase, a slow second phase, and a faster third phase. Six cells, in which Ca2+ recovered quickly lacked the third phase. The third phase was also absent in cells treated with a SR Ca2+-pump inhibitor, cyclopiazonic acid. 4. The peak first-phase Ca2+ removal rate observed upon repolarization to -70 mV was significantly reduced in cells treated with a mitochondrial Ca2+ uptake inhibitor, carbonyl cyanide m-chlorophenylhydrazone. However, an ATP-synthase inhibitor, oligomycin B, had no significant effect. The Ca2+ removal rate was little affected by clamping the cell at +120 mV rather than -70 mV, suggesting that Ca2+ removal processes are largely voltage independent. Also, little inward current was associated with Ca2+ clearance, indicating that Ca2+ removal does not involve an electrogenic process. Our results suggest that Ca2+-induced Ca2+ release contributes little to the elevation of Ca2+ in these cells. The SR Ca2+ pump may contribute to Ca2+ removal over a low [Ca2+](i) range in cells where [Ca2+](i) remains high for long enough, while mitochondrial Ca2+ uptake may be important when [Ca2+](i) is high.