The action of perchlorate (ClO4/-), an agonist of the voltage sensor in excitation-contraction (EC) coupling, has been examined using bundles of intact muscle cells, isolated membrane vesicles [sarcoplasmic reticulum (SR) and transverse tubule (TT)], and cultured myotubes. The effect of ClO4/- on mechanical parameters was investigated in isolated murine limb muscles. The presence of ClO4/- (5 or 10 mM) greatly increased twitch tension (>250%), slightly enhanced tetanic tension, and increased K contracture tension. K contracture thresholds of extensor digitorum longus (EDL, 40 mM K+) and soleus (30 mM K+) muscles were not altered by ClO4/-. However, in whole cell patch-clamp studies of mouse myotubes, contractile activation was shifted by approximately -10 mV by 10 mM ClO4/-. To further define the site of alteration of EC coupling by ClO4/-, studies were conducted with isolated porcine SR and TT vesicles and with cultured mouse myotubes. The rate constant of Ca-induced 45Ca release from SR vesicles was significantly increased by ClO4/-. However, neither the affinity nor level of [3H]PN200- 110 binding to TT vesicles was significantly affected by ClO4/- concentrations that increased twitch tension. Furthermore, slow plasmalemmal Ca currents of myotubes recorded in the whole cell patch-clamp mode were enhanced by 10 mM ClO4/-, and the current-voltage relationship was shifted approximately -7mV. Thus, in enhancing EC coupling in mammalian muscle, ClO4/- may act at multiple sites including the SR Ca release channel and the TT Ca channel-voltage sensor.
- calcium current
- contractile threshold
- dihydropyridine receptor
- potassium contracture
- sarcoplasmic reticular calcium release channel