INTRACELLULAR CA2+ TRANSIENTS INDUCED BY HIGH EXTERNAL K+ AND TETRACAINE IN CULTURED RAT MYOTUBES

Cultured myotubes from rat neonatal skeletal muscle were used to measu re intracellular Ca2+ concentration ([Ca2+](i)) and membrane potential s (Vm) using the Indo-1 microfluorimetry method and the nystatin perfo rated membrane patch technique, respectively. Sudden increases in exte rnal [K+](o) from 5 mM to either 22, 42 or 84 mM elicited transient el evations in [Ca2+](i) from a resting level of 106.2 +/- 10.3 nM (n=41) to peak values of 297, 409 and 454 nM, respectively. V-m changes indu ced by elevated [K+](o) followed the Nernst equation for [K+](o). The complex Ca2+ release response induced by elevated [K+](o) can be descr ibed by a minimal model involving two components with different kineti cs. This analysis revealed that the extent of the Ca2+ release by the fast component bears a sigmoidal relationship with V-m (midpoint at -4 7.5 mV and an effective valence of 4). Furthermore, while the fast tra nsitory component was rather insensitive to [Ca2+](o) and nifedipine, the slow component was profoundly inhibited by the dihydropyridine (10 mu M) both in normal and in a Ca2+ deficient medium. Tetracaine (0.05 to 2 mM), a blocker of the charge movement associated with excitation -contraction (E-C) coupling, elicited a fast elevation in [Ca2+](i) fo llowed by a rise at a constant rate to levels as high as 1-2 mu M, and the changes in [Ca2+](i) were readily reversible. Simultaneous measur ements of V-m and [Ca2+](i) suggest that the fast component is coupled to the rapid depolarization of the membrane induced by the anesthetic . We concluded that tetracaine triggers the release of Ca2+ from inter nal stores by at least two different mechanisms, one of which is assoc iated with the depolarizing effects of the drug.