Effects of glycerol treatment and maintained depolarization on charge movement in skeletal muscle

Abstract

1. Voltage-clamp experiments were carried out using the techniques described in the preceding paper. 2. In one series of experiments an attempt was made to disrupt the T-system with glycerol treatment. Muscles were soaked in Ringer + 400 mM glycerol for 1 hr at room temperature, transferred to Ringer + 5 mM calcium + 5 mM magnesium for 20-30 min, and then cooled to around 2 degrees C and placed in an isosmotic test solution containing tetrodotoxin for electrical measurements. 3. The density of charge seen in isosmotic tetraethylammonium (TEA) solution with strong depolarization, normalized according to fibre capacitance, was decreased by glycerol treatment to about one third the amount seen in untreated hypertonic fibres. 4. An analysis of fibre capacitance revealed that only 0-4 of the tubular capacitance was removed by this particular glycerol procedure. If the density of charge with respect to capacitance is corrected for this decrease in capacitance, the results indicate that glycerol treatment removed or immobilized 0-77 of the charge initially present. Thus the effect of glycerol treatment to reduce charge does not depend entirely on disrupting the electrical continuity of the T-system. 5. The effects of maintained depolarization were studied using a TEA Ringer made hypertonic with sucrose. When the voltage was changed from -80 to -21 mV the measurable charge movement declined exponentially to zero with a time constant of 13-24 sec. On repolarization the process recovered exponentially to the initial level with a time constant of 21-53 sec. 6. Experiments were also carried out using a sodium Ringer made hypertonic with sucrose. For small depolarizations only charge movement currents were seen, whereas for large depolarizations large delayed ionic currents, presumably carried by potassium, were observed. With moderate depolarizations in the range V = -40 to -30 mV, both components were of comparable magnitude. 7. A plot of the fractional charge movement (Q/Qmax) vs. V fitted at moderate depolarizations is similar to that of n infinity vs. V fitted at larger depolarizations. Values of n infinity were obtained by fitting the delayed ionic current to n4(V - VK). For voltages between -40 and -30 mV the time constant for charge tauQ was always less than taun; values of taun/tauQ varied from 1-6 to 4-3. 8. Glycerol treatment had little if any effect on the kinetics of delayed rectifier currents. Values of gK measured in isosmotic solution following glycerol were about one third the values obtained in untreated fibres in a hypertonic solution (osmolality three times normal). The threefold difference in gK is probably due to a similar difference in internal potassium concentration. 9. These results and others are difficult to reconcile with the idea that the charge movement process acts as a gate for potassium channels. It seems more likely that charge movement is a step in the activation of contraction.