Role of pH in a nitric oxide-dependent increase in cytosolic Cl−in retinal amacrine cells

Nitric oxide (NO) synthase-expressing neurons are found throughout the vertebrate retina. Previous work by our laboratory has shown that NO can transiently convert inhibitory GABAergic synapses onto cultured retinal amacrine cells into excitatory synapses by releasing Cl(-) from an internal store in the postsynaptic cell. The mechanism underlying this Cl(-) release is currently unknown. Because transport of Cl(-) across internal membranes can be coupled to proton flux, we asked whether protons could be involved in the NO-dependent release of internal Cl(-). Using pH imaging and whole cell voltage-clamp recording, we addressed the relationship between cytosolic pH and cytosolic Cl(-) in cultured retinal amacrine cells. We found that NO reliably produces a transient decrease in cytosolic pH. A physiological link between cytosolic pH and cytosolic Cl(-) was established by demonstrating that shifting cytosolic pH in the absence of NO altered cytosolic Cl(-) concentrations. Strong buffering of cytosolic pH limited the ability of NO to increase cytosolic Cl(-), suggesting that cytosolic acidification is involved in generating the NO-dependent elevation in cytosolic Cl(-). Furthermore, disruption of internal proton gradients also reduced the effects of NO on cytosolic Cl(-). Taken together, these results suggest a cytosolic environment where proton and Cl(-) fluxes are coupled in a dynamic and physiologically meaningful way.