Differential contribution of the Na⁺-K⁺-2Cl⁻ cotransporter NKCC1 to chloride handling in rat embryonic dorsal root ganglion neurons and motor neurons

Plasma membrane chloride (Cl⁻) pathways play an important role in neuronal physiology. Here, we investigated the role of NKCC1 cotransporters (a secondary active Cl⁻ uptake mechanism) in Cl⁻ handling in cultured rat dorsal root ganglion neurons (DRGNs) and motor neurons (MNs) derived from fetal stage embryonic day 14. Gramicidin-perforated patch-clamp recordings revealed that DRGNs accumulate intracellular Cl⁻ through a bumetanide- and Na⁺-sensitive mechanism, indicative of the functional expression of NKCC1. Western blotting confirmed the expression of NKCC1 in both DRGNs and MNs, but immunocytochemistry experiments showed a restricted expression in dendrites of MNs, which contrasts with a homogeneous expression in DRGNs. Both MNs and DRGNs could be readily loaded with or depleted of Cl⁻ during GABAA receptor activation at depolarizing or hyperpolarizing membrane potentials. After loading, the rate of recovery to the resting Cl⁻ concentration (i.e., [Cl⁻]i decrease) was similar in both cell types and was unaffected by lowering the extracellular Na⁺ concentration. In contrast, the recovery on depletion (i.e., [Cl⁻]i increase) was significantly faster in DRGNs in control conditions but not in low extracellular Na⁺. The experimental observations could be reproduced by a mathematical model for intracellular Cl⁻ kinetics, in which DRGNs show higher NKCC1 activity and smaller Cl⁻-handling volume than MNs. On the basis of these results, we conclude that embryonic DRGNs show a higher somatic functional expression of NKCC1 than embryonic MNs. The high NKCC1 activity in DRGNs is important for maintaining high [Cl⁻]i, whereas lower NKCC1 activity in MNs allows large [Cl⁻]i variations during neuronal activity.--Chabwine, J. N., Talavera, K., Verbert, L., Eggermont, J., Vanderwinden, J.-M., De Smedt, H., Van Den Bosch, L., Robberecht, W., Callewaert, G. Differential contribution of the Na⁺-K⁺-2Cl⁻ cotransporter NKCC1 to chloride handling in rat embryonic dorsal root ganglion neurons and motor neurons.