Two inhibitors of store operated Ca2+ entry suppress excitation contraction coupling in frog skeletal muscle

摘要

Two drugs, 2-APB and SKF-96365, commonly used to block Store Operated Ca2+ Entry (SOCE) were found to have inhibitory effects at different levels of the Excitation Contraction Coupling (ECC) process in frog skeletal muscle fibers. Treatment with either drug suppressed Ca2+ release from the Sarcoplasmic Reticulum, but this effect was not due to inhibition of SOCE as it occurred in Ca2+-free conditions. 2-APB applied extracellularly at 100 μM, the usual concentration to suppress SOCE, reversibly reduced the charge movement elicited by pulses in the range between −45 and −35 mV from 7.99 ± 0.73 nC/μF (N = 17) before drug application to 6.27 ± 0.68 nC/μF in the presence of 2-APB. This effect was mostly on the delayed Q_γ component. In fibers treated with the SERCA ATPase inhibitor CPA the Q_γ component disappeared, under this condition the application of 2-APB did not suppress the remaining charge movement. Thus the effect of 2-APB on charge movement currents seemed to be secondary to the suppression of Ca2+ release, likely occurring directly on the release channels. No significant suppression of ECC was observed for concentration below 20 μM. 2-APB also inhibited the L-type Ca2+ current (20 ± 4%, N = 8). On the other hand SKF-96365 had a direct effect on the voltage sensor promoting its voltage dependent inactivation. Applied at 20 μM, a typical concentration used for inhibiting SOCE, to fibers held at −80 mV inhibited the charge moved in response to pulses ranging −45 to −30 mV from 7.95 ± 2.59 nC/μF to 3.48 ± 0.9 nC/μF (N = 12). A parallel reduction of Ca2+ release was observed. Wash out was drastically increased by hyperpolarization of the holding potential to −100 mV. SKF-96365 also inhibited the L-type Ca2+ current (41 ± 8%, N = 4) and increased its rate of inactivation.