Separation of charge movement components in mammalian skeletal muscle fibres

1. Intramembrane charge movement, I-ICM, were measured in rat skeletal musc le fibres in response to voltage steps from a -90 mV holding potential to a wide test voltage range (-85 to 30 mV), using a double Vaseline-gap voltag e-clamp technique. Solutions were designed to minimise ionic currents. Ca2 current was blocked by adding Cd2+ (0.8 mM) to the external solution, In a subset of experiments Cd2+ was emitted to determine which components of th e charge movement best correlated with L-type Ca2+ channel gating. 2. Detailed kinetic analysis of I-ICM identified two major groups of charge s. The first two components, designated Q(a) and Q(b) were the only charges moved by small depolarising steps. The second group of components, Q(c) an d Q(d), showed a more positive voltage threshold, -35.6 +/- 2.0 mV, (n = 6) in external solution with Cd2+, and -41.1 +/- 2.0 mV (n = 12) in external solution without Cd2+. Notably, in external solution without Cd2+ the volta ge threshold of Ca2+ current, activation had a similar value, being -38.1 /- 2.4 mV. 3. The sum of three Boltzmann functions, Q(1), Q(2), and Q(3), showing prog ressively more positive transition voltages, could be fitted to charge vers us voltage, Q(ICM)-V, plots. The three Boltzmann terms identified three cha rge components: Q(1) described the shallow voltage-dependent Q(a) and Q(b), charges, Q(2), and Q(3) described the steep voltage-dependent Q(c) and Q(d ) charges. 4. In external solution without Cd2+ the charge kinetics changed: a slow de caying phase was replaced by a pronounced delayed hump. Moreover, the trans ition voltages of the individual steady-state charge components were shifte d towards negative potentials (front 6.3 to 8.2 mV). Nevertheless, the over all charge and steepness factors were conserved. 5. In conclusion, these experiments allowed a clear separation of four Comp onents of intramembrane charge movements in rat skeletal muscle, showing th at there are no fundamental differences with respect to charge movement com ponents between amphibian and mammalian twitch muscle. Moreover, Q(c) and Q (d) charge are correlated with L-type Ca2+ channel gating.