Kv3 channels modulate calcium signals induced by fast firing patterns in the rat retinal ganglion cells

Abstract

Expression of non-inactivating Kv3.1/Kv3.2 potassium channels determines fast-spiking phenotype of many types of neurones including retinal ganglion cells (RGCs); furthermore Kv3 channels regulate neurotransmitter release from presynaptic terminals. In the present study we investigated how inhibition of Kv3 channel by low TEA concentrations modifies firing properties and Ca2+ influx in the rat RGCs. Experiments were performed on the whole-mount retinal preparations from 4 to 6 weeks old Wistar rats using simultaneous whole cell patch clamp and intracellular Ca2+ measurements in combination with single-cell RT-PCR. In response to 500-ms depolarization step the RGCs demonstrated fast firing tonic behaviour with a mean frequency of spiking 61±5Hz (n=28). All of the tonic cells tested (n=9) expressed specific mRNA for either Kv3.1 or Kv3.2 or for both channels. Bath applications of TEA (250μM, 500μM and 1mM) modified firing patterns dose-dependently as follows: firing frequency was decreased, mean action potential (AP) half-width increased and mean amplitude of after hyperpolarization was reduced. The amplitude of the Ca2+ signals induced by the cells firing was linearly dependent on number of APs with a mean slope of 7.3±0.9nM per one AP (n=8). APs widening by TEA increased the slope of the amplitude vs. AP number plots in a dose-dependent manner: 250μM of TEA increased the mean slope value to 9.5±1.2nM/AP, 500μM to 12.4±2.4nM/AP and 1mM to 13.2±2.9nM/AP (n=6). All these parameters, as well as the cells firing properties, were significantly different from controls and from each other except between 500μM and 1mM. This is consistent with the pharmacological properties of Kv3.1/Kv3.2 channels: the TEA IC50 is in the range 150–300μM with almost complete block at 1mM. This suggests that Kv3.1/Kv3.2 channels underlie the fast firing of the rat RGCs and provide at a given firing frequency 1.8-fold restriction Ca2+ influx, thus protecting the cells from its cytotoxic action.