A MACRO CELL-ATTACHED PATCH-CLAMP STUDY OF THE PROPERTIES OF THE NA CURRENT IN THE VICINITY OF THE MOTOR END-PLATE REGION OF FROG SINGLE INTEROSSEAL SKELETAL-MUSCLE FIBERS

A macro cell-attached patch-clamp technique using large (16-mu-m) electrodes was developed to study the properties of the Na current (I(Na)) 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, I(Na) 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 V(rev) equal to + 63.2 +/- 1.9 mV. There is no evidence for the presence of a tubular I(Na). When the [Na] in the recording pipette is changed, V(rev) 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 I(Na) is maximum, the half-time of activation is 223.9 +/- 10.5-mu-s, the time to peak 365.7 +/- 13.5-mu-s and the time constant of inactivation 260.7 +/- 11.2-mu-s. The time constant of reactivation at - 100 mV is 1.44 +/- 0.19 ms. These and other results show that I(Na) can be adequately studied with this technique.