A role for the cystic fibrosis transmembrane conductance regulator in the nitric oxide-dependent release of Cl⁻ from acidic organelles in amacrine cells

Abstract

γ-Amino butyric acid (GABA) and glycine typically mediate synaptic inhibition because their ligand-gated ion channels support the influx of Cl^- However, the electrochemical gradient for Cl^- across the postsynaptic plasma membrane determines the voltage response of the postsynaptic cell. Typically, low cytosolic Cl^- levels support inhibition, whereas higher levels of cytosolic Cl^- can suppress inhibition or promote depolarization. We previously reported that nitric oxide (NO) releases Cl^- from acidic organelles and transiently elevates cytosolic Cl^-, making the response to GABA and glycine excitatory. In this study, we test the hypothesis that the cystic fibrosis transmembrane conductance regulator (CFTR) is involved in the NO-dependent efflux of organellar Cl^- We first establish the mRNA and protein expression of CFTR in our model system, cultured chick retinal amacrine cells. Using whole cell voltage-clamp recordings of currents through GABA-gated Cl^- channels, we examine the effects of pharmacological inhibition of CFTR on the NO-dependent release of internal Cl^- To interfere with the expression of CFTR, we used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 genome editing. We find that both pharmacological inhibition and CRISPR/Cas9-mediated knockdown of CFTR block the ability of NO to release Cl^- from internal stores. These results demonstrate that CFTR is required for the NO-dependent efflux of Cl^- from acidic organelles.<b>NEW & NOTEWORTHY</b> Although CFTR function has been studied extensively in the context of epithelia, relatively little is known about its function in neurons. We show that CFTR is involved in an NO-dependent release of Cl^- from acidic organelles. This internal function of CFTR is particularly relevant to neuronal physiology because postsynaptic cytosolic Cl^- levels determine the outcome of GABA- and glycinergic synaptic signaling. Thus the CFTR may play a role in regulating synaptic transmission.