Cellular and Molecular Properties of Neurons: A role for the cystic fibrosis transmembrane conductance regulator in the nitric oxide-dependent release of Cl− from acidic organelles in amacrine cells

摘要

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.>NEW & NOTEWORTHY 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.