Chloride channels with ClC-1-like properties differentially regulate the excitability of dopamine receptor D1- and...

Dynamic chloride (Cl^-) regulation is critical for synaptic inhibition. In mature neurons, Cl^- influx and extrusion are primarily controlled by ligand-gated anion channels (GABA_A and glycine receptors) and the potassium chloride cotransporter K⁺-Cl^- cotransporter 2 (KCC2), respectively. Here, we report for the first time, to our knowledge, a presence of a new source of Cl^- influx in striatal neurons with properties similar to chloride voltage-gated channel 1 (ClC-1). Using whole cell patch-clamp recordings, we detected an outwardly rectifying voltage-dependent current that was impermeable to the large anion methanesulfonate (MsO^-). The anionic current was sensitive to the ClC-1 inhibitor 9-anthracenecarboxylic acid (9-AC) and the nonspecific blocker phloretin. The mean fractions of anionic current inhibition by MsO^-, 9-AC, and phloretin were not significantly different, indicating that anionic current was caused by active ClC-1-like channels. In addition, we found that Cl^- current was not sensitive to the transmembrane protein 16A (TMEM16A; Ano1) inhibitor Ani9 and that the outward Cl^- rectification was preserved even at a very high intracellular Ca²⁺ concentration (2 mM), indicating that TMEM16B (Ano2) did not contribute to the total current. Western blotting and immunohistochemical analyses confirmed the presence of ClC-1 channels in the striatum mainly localized to the somata of striatal neurons. Finally, we found that 9-AC decreased action potential firing frequencies and increased excitability in medium spiny neurons (MSNs) expressing dopamine type 1 (D1) and type 2 (D2) receptors in the brain slices, respectively. We conclude that ClC-1-like channels are preferentially located at the somata of MSNs, are functional, and can modulate neuronal excitability.