Intracellular chloride ion concentration in differentiating neuronal cell and its role in growing neurite.

Chloride ion (Cl) is one of the most abundant anions in our body. Increasing evidence suggests that Cl plays fundamental roles in various cellular functions. We have previously reported that electroneutral cation-chloride cotransporters, such as Na-K-2Cl cotransporter 1 (NKCC1) and K-Cl cotransporter 1 (KCC1), are involved in neurite outgrowth during neuronal differentiation. In the present study, we studied if there is correlation between intracellular Cl concentrations ([Cl]) and the length of growing neurites. We measured [Cl] in the cell body and growing neurite tips using halide-sensitive fluorescent dye N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE), revealing that [Cl] in the tip of growing neurite was higher than that in cell body in a single cell. Importantly, there was a significant positive correlation between the length of growing neurite and [Cl] in neurite tip. Bumtanide (BMT), an inhibitor of NKCC1, significantly inhibited neurite outgrowth and decreased [Cl] in neurite tip. The results obtained in the present study and our previous studies together strongly suggest that high [Cl] in neurite tip region is crucial for efficient neurite outgrowth.