Isolation of Somatic Na+Currents by Selective Inactivation of Axonal Channels with a Voltage Prepulse

Abstract

We present a simple and effective method for isolating the somatic Na(+) current recorded under voltage clamp from neurons in brain slices. The principle is to convert the axon from an active compartment capable of generating uncontrolled axonal spikes into a passive structure by selectively inactivating axonal Na(+) channels. Typically, whole-cell currents from intact neurons under somatic voltage clamp contain a mixture of Na(+) current and axial current caused by escaped axonal spikes. We found that a brief prepulse to voltages near spike threshold evokes the axonal spike, which inactivates axonal but not somatic channels. A subsequent voltage step then evokes only somatic Na(+) current from electrotonically proximal sodium channels under good voltage-clamp control. Simulations using a neuron compartmental model support the idea that the prepulse effectively inactivates currents from the axon and isolates well controlled somatic currents. Na(+) currents recorded from cortical pyramidal neurons in slices, using the prepulse, were found to have voltage dependence nearly identical to that of currents recorded from acutely dissociated pyramidal neurons. In addition, studies in dissociated neurons show that the prepulse has no visible effect on the voltage dependence and kinetics of Na(+) currents elicited by the subsequent voltage step, only decreasing the amplitude of the currents by 10-20%. The technique was effective in several neuronal types in brain slices from male and female neonatal rats and mice, including raphé neurons, cortical pyramidal neurons, inferior olivary neurons, and hypoglossal motoneurons.