Excitability And Regulation Of Rat Retinal Ganglion Cells

Cannabinoid CB1receptors, extensively distributed in the vertebrate retina, regulate a variety of physiological functions in the retina through modulation of various ion channels. In the present study, we investigated the effect of this receptor signaling on cell excitability of rat retinal ganglion cells (RGCs) by using current-clamp model of whole-cell patch-clamp techniques. Two methods, current injection and fluorescent dye staining, were used to identify ganglion cells’ subtype. In current-clamped condition, both ON-and OFF-type RGCs showed spontaneous firings, which were eliminated by perfusing cocktail synaptic blockers, including CNQX, D-APV, bicuculline and strychnine. In the presence of cocktail synaptic blockers, perfusion of CB1receptor agonist WIN55212-2(WIN,5μM) did not change the firing rate of the RGCs evoked by positive current injection (from+10to+100pA). The firing frequency was ranging from5.8±1.9Hz to67.6±3.3Hz (n=17) and from7.3±2.5Hz to65.5±3.3Hz (n=17) with injecting positive current of+10pA to+100pA before and after WIN was applied (P all>0.05). Phase-plane plot analysis showed the threshold and the time to first action potential were also not affected by WIN. However, the+dv/dt max (control:from185.5±16.6mV/ms to192.4±13.8mV/ms, WIN55212-2:from175.7±16.0mV/ms to183.0±13.1mV/ms, n=17, P<0.05to0.01) and-dv/dt max (control:from-159.4±17.7mV/ms to-163.5±13.0mV/ms, WIN55212-2:from-142.8±15.2mV/ms to-149.3±11.1mV/ms, n=17, P<0.05to0.001) of action potentials were significantly decreased in almost all the current injections, suggesting that WIN reduced the rising and descending velocities of the action potentials. Our results suggest that activation of cannabinoid receptors by WIN may regulate excitability of the rat RGCs through modulating some ion channels in the cells. We are investigating the underlying mechanisms of such modulation. Retinal ganglion cells (RGCs), output neurons of retina, receive excitatory input from bipolar cells and inhibitory input from amacrine cells. The excitability of these cells influences visual signal, which is regulated by inwardly rectifying potassium channels (Kir) and hyperpolarization-activated cation currents (Ih). However, expression, function and regulatory mechanisms of these channels in rat RGCs are still controversial. In the present work we used whole-cell recordings performed on rat RGCs in retinal slice preparations to investigate the regulation of Kir and Ih on cell excitability. Maintained activities were recorded under current-clamp mode. We found that:(1) For the majority of the RGCs, spontaneous firings of both ON-and OFF-type cells were dependent on synaptic inputs, while a small percentage of cells firing not only remained but strengthened significantly. These cells may be ipRGCs. Excitatory inputs enhanced firing frequency, shortened interspike interval and firing patterns became much more regular, while inhibitory synaptic inputs blocked most of cells firing, but increased26%of cells firing rate;(2) With the application of synaptic blockers and Kir blocker Ba2+, firing frequencies of both ON-and OFF-type RGCs were increased, interspike intervals shortened but firing patterns kept unchanged;(3)With the application of synaptic blockers and unspecific Ih blocker Cs+, firing frequencies of both ON-and OFF-type RGCs were increased, membrane potentials increased slightly, interspike intervals hardly changed and firing patterns became irregular. With the application of synaptic blockers and specific Ih blocker ZD7288,71%cells fired in a form of burst after a significant membrane potential decrease. These results suggest that spontaneous firings of both ON-and OFF-type RGCs in rat retina are dependent on synaptic inputs. Blocking inhibitory inputs increases the firing rate and shortens the intervals of spikes, while blocking excitatory inputs almost eliminates the firing. Kir and Ih, regulate spontaneous firing of rat RGCs.