Clampfit 8

Calcium currents were recorded with the ruptured whole-cell patch-clamp technique in voltage-clamp mode using an Axopatch 200B amplifier (Axon Instruments). Patch pipettes (borosilicate glass; Harvard Apparatus) were filled with (mM) 145 Cs-aspartate, 2 MgCl₂, 10 HEPES, 0.1 Cs-EGTA, and 2 Mg-ATP (pH 7.4 with CsOH) and had a resistance between 1.5 and 3 MΩ. For the recording of the calcium currents, the myotubes were bathed in solution containing (mM) 10 CaCl₂, 145 tetraethylammonium chloride, and 10 HEPES (pH 7.4 with tetraethylammonium hydroxide). Data acquisition and command potentials were controlled by pCLAMP software (version 8.0; Axon Instruments); analysis was performed using Clampfit 8.0 (Axon Instruments) and SigmaPlot 8.0 (SPSS Science) software. The current-voltage dependence was fitted according to $I=G_{\max }\cdot \left(V-V_{rev}\right)/1\left(1+\exp \left(-\left(V-V_{1/2}\right)/k\right)\right),$ where, G_max is the maximum conductance of the L-type calcium currents, V_rev is the extrapolated reversal potential of the calcium current, V_1/2 is the potential for half maximal conductance, and k is the slope. The conductance was calculated using $G=\left(-I\ast 1000\right)/\left(V-V_{rev}\right),$ and its voltage dependence was fitted according to a Boltzmann distribution: $G=G_{\max }/\left(1+\exp \left(-\left(V-V_{1/2}\right)/k\right)\right).$ The statistical significance was calculated using the Mann–Whitney U test (see Tables 1 and 2).