Effects of thapsigargin in normal and pretreated with ryanodine guinea pig cardiomyocytes

Abstract

We compared the effects of thapsigargin (TG), a selective blocker of Ca(2+)-adenosinetriphosphatase of sarcoplasmic reticulum (SR), and ryanodine (Ry) in the single isolated myocytes of guinea pig ventricular myocardium loaded with indo 1 acetoxymethyl ester (AM). TG (2 x 10(-7) M) inhibited the rapid phase of Ca2+ transient, increased time to peak intracellular Ca2+ concentration ([Ca2+]i) from 158 +/- 12 to 391 +/- 60 ms and decreased the total amplitude of the transient to 89 +/- 4% of the pre-TG control. Time to peak of contractions increased from 350 +/- 47 to 410 +/- 37 ms and total duration from 666 +/- 62 to 850 +/- 198 ms. Total amplitude of contractions was hardly affected. In the cells not loaded with indo 1-AM TG decreased the amplitude of contractions to 71 +/- 3% of control. When the effects of TG were fully developed, the cells ceased to respond to 1 s of superfusion with 15.0 mM caffeine with transient elevation of [Ca2+]i and/or transient contracture. TG did not affect the amplitude or time course of Ca2+ current (ICa) or the current-voltage relation. We propose that Ca2+ transients and contractions in the cells treated with TG were initiated by sarcolemmal Ca2+ influx. Ry (1.0 microM) initiated similar changes in the time course of Ca2+ transients and contractions as TG; however, total amplitude of the transients and contractions was reduced to 78 +/- 5 and 55 +/- 7% of the control, respectively. The SR Ca2+ was also depleted by Ry. TG superfused over the cells pretreated with Ry increased the amplitude of Ca2+ transients and respective contractions to the pre-Ry level. TG did not affect the ICa in the cells pretreated with Ry nor did it change configuration of action potentials to increase the Ca2+ influx. We propose that the effect of Ry on amplitude of Ca2+ transients and contractions results from the trapping of a fraction of sarcolemmal Ca2+ influx by the SR and its rapid release into subsarcolemmal space. From there it is extruded out of the cell by Na(+)-Ca2+ exchange before ever reaching the contractile system.

Effects of inhibition of sarcoplasmic reticulum calcium uptake on contraction in myocytes isolated from failing human ventricle

“…This is consistent with results in guinea-pig myocytes, where it has been shown that ryanodine (which increases SR Ca2+ loss), but not thapsigargin, can reduce the size of the beat [27]. [t has been suggested that Ca.z+ uptake by the SR curtails the rise in Ca2+ at the contractile apparatus.…”

Alterations in the force‐frequency relationship by tert‐ butylbenzohydroquinone, a putative SR Ca2+ pump inhibitor, in rabbit and rat ventricular muscle

“…TBQ effects on AP duration are more probably explained by a smaller amount of SR released Ca2+ that would decrease the Na+/Ca2+ exchange current, a depolarizing current in rabbit ventricular cells (Bers, 1991 Another argument relies on the observation that in the rabbit and rat ventricle, TBQ, like CPA or TG, exerts a negative inotropic effect that was steady. We did not observe any reversal of the negative inotropic effect and we are not aware of such an effect with other inhibitors in multicellular preparations, whereas it has been reported in isolated guinea-pig (Lewartowski et al, 1994) or rabbit (Bassani et al, 1993; ventricular myocytes. As such a reversal is currently explained by abolition of trans-sarcolemmal Ca2" buffering by the SR, it is likely that a fraction of pumps unblocked by 30 uM TBQ is still able to buffer incoming Ca2".…”

“…In fact, it has been shown that both compounds had a negative inotropic effect in most animal species, which could be expected on the basis of a decreased SR Ca2" content (Bassani et al, 1993;Lewartowski et al, 1994). However, paradoxical results have also been reported which showed that the shortening of TG-or CPA-treated ventricular myocytes from rabbit or guinea-pig had similar amplitude but a longer time course compared to control, despite an empty SR (Bassani et al, 1994;Lewartowski et al, 1994). This effect has been interpreted as the result of the suppression of the rapid SR buffering of incoming Ca2+which could then directly activate the contractile proteins, although at a slower rate compared to the rate at which Ca2" binds to the myofibrillar proteins when it is released from the SR (Janczewski & Lakatta, 1993).…”

“…In fact, it has been shown that both compounds had a negative inotropic effect in most animal species, which could be expected on the basis of a decreased SR Ca2" content (Bassani et al, 1993;Lewartowski et al, 1994). However, paradoxical results have also been reported which showed that the shortening of TG-or CPA-treated ventricular myocytes from rabbit or guinea-pig had similar amplitude but a longer time course compared to control, despite an empty SR (Bassani et al, 1994;Lewartowski et al, 1994).…”

Effects of cyclopiazonic acid and thapsigargin on electromechanical activities and intracellular Ca2+ in smooth muscle of carotid artery of hypertensive rats

“…CPA and TG, which have been shown to inhibit sarcoplasmic reticular Ca2+-ATPase in skeletal and caridac muscles, are used as a tool for the study of excitation-contraction coupling in these muscles (Janczewski & Lakatta, 1993;Lewartowski et al, 1994;Chiesi et al, 1994;Yard et al, 1994). These drugs are also used in smooth muscle to deplete sarcoplasmic reticular Ca2+ or to inhibit Ca21 uptake by the sarcoplasmic reticulum (Sill et al, 1991;Low et al, 1991;Uyama et al, 1992;Low et al, 1992;Luo et al, 1993;Alexander & Cheung, 1994;Kwan et al, 1994;Naganobu et al, 1994).…”