A macro cell-attached patch-clamp study of the properties of the Na current in the vicinity of the motor endplate region of frog single interosseal skeletal muscle fibres.

Abstract

A macro cell-attached patch-clamp technique using large (16 microns) electrodes was developed to study the properties of the Na current (INa) in the motor endplate region of frog interosseal muscle fibres (resting potential approx.-99 mV). The fibre isolation procedure allows the formation of gigaOhm seals and thus a good voltage-clamp control of the patch. At 22 degrees C, in the presence of 110 mM external [Na] and of K channel blockers, INa activates at -49.6 +/- 1.9 mV, reaches a maximum of 4.33 +/- 0.66 mA/cm2 at -7.5 +/- 2.5 mV and reverses at Vrev equal to +63.2 +/- 1.9 mV. There is no evidence for the presence of a tubular INa. When the [Na] in the recording pipette is changed, Vrev exactly follows the Nernst equation for Na ions. An internal [Na] of 9.69 mM is determined. The Na conductance is maximum (63.56 +/- 7.03 mS/cm2) at +8.9 +/- 3.0 mV and markedly decreases for potentials positive to +25 mV. In contrast, the Na permeability (maximum of 6.13 +/- 0.95 x 10(-4) cm/s at +51.4 +/- 5.6 mV) remains more constant at positive potentials. The Na channels are half-inactivated at -68.9 mV and half-activated at -37.9 mV. At the potential at which INa is maximum, the half-time of activation is 223.9 +/- 10.5 microseconds, the time to peak 365.7 +/- 13.5 microseconds and the time constant of inactivation 260.7 +/- 11.2 microseconds. The time constant of reactivation at -100 mV is 1.44 +/- 0.19 ms. These and other results show that INa can be adequately studied with this technique.