THIOPENTAL ALTERS CONTRACTION, INTRACELLULAR CA2+, AND PH IN RAT VENTRICULAR MYOCYTES

Background: Myocardial contractility is regulated by intracellular con centration of free Ca2+ ([Ca2+](i)) and myofilament Ca2+ sensitivity. The objective of this study was to elucidate the direct effects of thi opental on cardiac excitation-contraction coupling using individual, f ield-stimulated ventricular myocytes, Methods: Freshly isolated rat ve ntricular myocytes were loaded with the Ca2+ indicator, fura-2, and pl aced on the stage of an inverted fluorescence microscope in a temperat ure-regulated bath. [Ca2+](i) (340/380 ratio) and myocyte shortening ( video-edge detection) were monitored simultaneously in individual cell s field-stimulated at 0.3 Hz. Amplitude and timing of myocyte shorteni ng and [Ca2+](i) were compared before and after addition of thiopental . Intracellular pH was measured with the pH indicator, BCECF (500/440 ratio). Real-time uptake of Ca2+ into isolated sarcoplasmic reticulum vesicles was measured using fura-2 free acid in the extravesicular com partment. One hundred thirty-two cells were studied. Results: Field st imulation increased [Ca2+](i) from 85 +/- 10 nM to 355 +/- 22 nM (mean +/- SEM), Myocytes shortened by 10% of resting cell length (127 +/- 5 mu m). Times to peak [Ca2+](i) and shortening were 139 +/- 6 and 173 +/- 7 msec, respectively, Times to 50% recovery for [Ca2+](i) and shor tening were 296 +/- 6 and 290 +/- 6 ms, respectively. Addition of thio pental (30-1,000 mu M) resulted in dose-dependent decreases In peak [C a2+](i) and myocyte shortening. Thiopental altered time to peak and ti me to 50% recovery for [Ca2+](i) and myocyte shortening and inhibited the rate of uptake of Ca2+ into isolated sarcoplasmic reticulum vesicl es. Thiopental did not, however, alter the amount of Ca2+ released in response to caffeine in sarcoplasmic reticulum vesicles or intact cell s. Thiopental (100 mu M) increased intracellular pH and caused an upwa rd shift in the dose-response curve to extracellular Ca2+ for shorteni ng, with no concomitant effect on peak [Ca2+](i). These effects were a bolished by ethylisopropyl amiloride, an inhibitor of Na+-H+ exchange. Conclusion: Thiopental has a direct negative inotropic effect on card iac excitation-contraction coupling at the cellular level, which is me diated by a decrease in [Ca2+](i). Thiopental also increases myofilame nt Ca2+ sensitivity via alkalinization of the cell, which may partiall y offset its negative inotropic effect.