Abstract
Determining the factors regulating cytosolic Cl− in neurons is fundamental to our understanding of the function of GABA- and glycinergic synapses. This is because the Cl− distribution across the postsynaptic plasma membrane determines the sign and strength of postsynaptic voltage responses. We have previously demonstrated that nitric oxide (NO) releases Cl− into the cytosol from an internal compartment in both retinal amacrine cells and hippocampal neurons. Furthermore, we have shown that the increase in cytosolic Cl− is dependent upon a decrease in cytosolic pH. Here, our goals were to confirm the compartmental nature of the internal Cl− store and to test the hypothesis that Cl− is being released from acidic organelles (AO) such as the Golgi, endosomes or lysosomes. To achieve this, we made whole cell voltage clamp recordings from cultured chick retinal amacrine cells and used GABA-gated currents to track changes in cytosolic Cl−. Our results demonstrate that intact internal proton gradients are required for the NO-dependent release of internal Cl−. Furthermore, we demonstrate that increasing the pH of AO leads to release of Cl− into the cytosol. Intriguingly, the elevation of organellar pH results in a reversal in the effects of NO. These results demonstrate that cytosolic Cl− is closely linked to the regulation and maintenance of organellar pH and provide evidence that acidic compartments are the target of NO.
Highlights
The regulation of cytosolic Cl− plays a key role in determining the inhibitory strength within neuronal circuits
GABA-gated current in cultured amacrine cells are known to be entirely due to the activation of GABAA receptors (GABAARs)
Inward GABA-gated currents were often observed in response to the first few GABA applications, but when observed, these currents typically dissipated as Cl− washed out of the cytosol
Summary
The regulation of cytosolic Cl− plays a key role in determining the inhibitory strength within neuronal circuits. Regulation of intracellular Cl− levels influences the sign and synaptic strength of GABAergic and glycinergic synapses by influencing the reversal potential of GABA- or glycine-gated synaptic currents. GABA and glycinergic amacrine cells are known to generate inhibitory output onto bipolar cells, ganglion cells and other amacrine cells. This signaling is known to be key in regulating the response properties of different classes of retinal ganglion cells (Zhou and Lee, 2008; Masland, 2012; Venkataramani et al, 2014). Transient and possibly localized modifications in cytosolic Cl− levels have the potential to alter synaptic signaling in the inner retina and the output of the retina
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