Abstract

1. We tested the hypothesis that the sarcoplasmic reticulum (SR) buffers (attenuates) the increase in averaged myoplasmic free [Ca2+] (Ca(im)) resulting from Ca2+ influx. 2. Fura-2 measurements of Ca(im) were obtained in single smooth muscle cells freshly dispersed from bovine coronary artery. 3. Caffeine (5 x 10(-3) M) elicited a transient increase in Ca(im) and depleted the SR Ca2+ store. In the continued presence of caffeine or 10(-5) M-ryanodine SR buffering of Ca(im) was inhibited. Subsequent exposure to high extracellular [K+] (greater than 30 mM, equimolar Na+ removal) elicited a 2-fold more rapid and 2-fold greater peak increase in Ca(im) than high K+ elicited when SR buffering of Ca(im) was normal. The augmented increase in Ca(im) was inhibited 35% by 10(-5) M-diltiazem, 65% by 2 x 10(-4) M-LaCl3, and 87% in Ca(2+)-free external solution. 4. When Ca(im) buffering capacity was increased by partially depleting the SR with a transient (1 min) exposure to caffeine, subsequent exposure to 80 nM-K+ solution increased Ca(im) almost 2-fold more slowly than 80 mM-K+ before depletion of Ca2+ from the SR. However, the influxing Ca2+ was sequestered by the SR and refilled it, as evident by the subsequent caffeine-induced Ca(im) transient being identical to the first. Increasing extracellular [K+] (thus, increasing depolarization and Na+ removal) caused proportional increases in Ca(im) and the subsequent caffeine-induced Ca(im) transients were proportionally larger, indicating a graded filling of the SR by Ca2+ influx. 5. Diltiazem (10(-5) M) inhibited the refilling of the SR achieved by 80 mM-K+, by 26%. Refilling was inhibited 76% by 80 mM-K+, Ca(2+)-free solution, indicating the fraction of refilling dependent on influx of Ca2+ through voltage-gated Ca2+ channels, leak channels, and other influx pathways. Mild depolarization with 35 mM-K+ (no Na+ removal) often caused no increase in Ca(im), but influx through voltage-gated Ca2+ channels occurred because the SR Ca2+ store was refilled. Also, 10(-5) M-diltiazem or 10(-6) M-TA3090 inhibited the refilling to levels attributable only to leak influx of Ca2+. 6. All data support our hypothesis that the SR significantly attenuates the amount of Ca2+ influx that accumulates to increase Ca(im).(ABSTRACT TRUNCATED AT 400 WORDS)

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