CA2+ CURRENT AND CHARGE MOVEMENT IN ADULT SINGLE HUMAN SKELETAL-MUSCLE FIBERS

1. The Vaseline-gap technique was used to record calcium currents (I(Ca)) and charge movement in single cut fibres from normal human muscle. Experiments were carried out in 2 or 10 mM-extracellular Ca2+ concentration ([Ca2+]o) and at 17 or 27-degrees-C. 2. The passive electrical properties of the fibres with this technique were: membrane resistance for unit length r(m) = 59.4 k-OMEGA cm; longitudinal resistance per unit length r(i) = 4.9 M-OMEGA/cm; longitudinal resistance per unit length under the Vaseline seals r(e) = 438 M-OMEGA/cm; specific membrane resistance R(m) = 1.176 k-OMEGA cm2; input capacitance = 5.53 nF; specific membrane capacitance = 8.9-mu-F/cm2. 3. The maximum amplitude of I(Ca) at 17-degrees-C was: in 2 mM [Ca2+]o, -0.42-mu-A/mu-F and in 10 mM [Ca2+]o, -1.44-mu-A/mu-F. At 27-degrees-C and in 10 mM [Ca2+]o, it increased to -3.04-mu-A/mu-F. The calculated temperature coefficient (Q10) for the increase in amplitude from 17 to 27-degrees-C was 2.1. 4. Ca2+ permeability (P(Ca)) was calculated using the Goldman-Katz relation; in 2 mM [Ca2+]o at 17-degrees-C, P(Ca) = 1.26 x 10(-6) cm/s; in 10 mM [Ca2+]o at 17-degrees-C, P(Ca) = 2.23 x 10(-6) cm/s; in 10 mM [Ca2+]o at 27-degrees-C, P(Ca) = 4.03 x 10(-6) cm/s. 5. The activation curve calculated from the P(Ca) was shifted by 10 mV to positive potentials when raising [Ca2+]o from 2 to 10 mM. Increasing the temperature did not change the curve. The mid-point potentials (V(a 1/2)) and steepness (k) of the activation curves were: at 17-degrees-C, in 2 mM [Ca2+]o, V(a 1/2) = -1.53 mV and k = 6.7 mV; in 10 mM [Ca2+]o, V(a 1/2) = 9.96 mV and k = 6.8 mV; at 27-degrees-C and 10 mM [Ca2+]o, V(a 1/2) = 11.3 mV and k = 7.7 mV. The activation time constant in 10 mM [Ca2+]o reached a plateau at potentials positive to 10 mV, with a value of 93.8 ms at 17-degrees-C and 17.4 ms at 27-degrees-C. The calculated Q10 was 4.5. 6. The deactivation of the current was studied from tail currents at different membrane potentials in 10 mM [Ca2+]o. For potentials from -120 to -50 mV, the deactivation time constant ranged between 5 and 7 ms at 17-degrees-C and between 7 and 14 ms at 27-degrees-C. For potentials positive to -50 mV the deactivation became much slower at 27-degrees-C. 7. I(Ca) was reversibly reduced by the addition of 0.2 or 0.5-mu-M-nifedipine and completely blocked by 2-mu-M-nifedipine. 8. Charge movement was recorded in 2 or 10 mM [Ca2+]o at 17-degrees-C. Increasing [Ca2+]o caused an increase of the maximum amount of charge moved (Q(max)) and a positive shift of the curve. The parameters (V(q 1/2) = mid-point activation and k = steepness) for the voltage-charge relationship were: 2 mM [Ca2+]o, Q(max) = 5.2 nC/mu-F, V(q 1/2) = -46.0 mV and k = 12.9 mV; 10 mM [Ca2+]o, Q(max) = 9.8 nC/mu-F, V(q 1/2) = -28.8 mV and k = 15.2 mV. 9. The fact that the activation curves of the I(Ca) and the charge movement differ widely in the mid-point of activation can be explained by the presence of several closed states before the channel opens. In addition, it is also possible that only part of the charge movement corresponds to the gating of the Ca2+ channel.