Cation-chloride cotransporters and the polarity of GABA signaling in mouse hippocampal parvalbumin interneurons

Abstract

Transmembrane chloride gradients govern the efficacy and polarity of GABA signaling in neurons and are usually maintained by the activity of cation chloride cotransporters, such as KCC2 and NKCC1. Whereas their role is well established in cortical principal neurons, it remains poorly documented in GABAergic interneurons. We used complementary electrophysiological approaches to compare the effects of GABAAR activation in adult mouse hippocampal parvalbumin interneurons (PV INs) and pyramidal cells (PCs). Loose cell attached, tight-seal and gramicidin-perforated patch recordings all show GABAAR-mediated transmission is slightly depolarizing and yet inhibitory in both PV INs and PCs. Focal GABA uncaging in whole-cell recordings reveal that KCC2 and NKCC1 are functional in both PV INs and PCs but differentially contribute to transmembrane chloride gradients in their soma and dendrites. Blocking KCC2 function depolarizes the reversal potential of GABAAR-mediated currents in PV INs and PCs, often beyond firing threshold, showing KCC2 is essential to maintain the inhibitory effect of GABAARs. Finally, we show that repetitive 10 Hz activation of GABAARs in both PV INs and PCs leads to a progressive decline of the postsynaptic response independently of the ion flux direction or KCC2 function. This suggests intraneuronal chloride buildup may not predominantly contribute to activity-dependent plasticity of GABAergic synapses in this frequency range. Altogether our data demonstrate similar mechanisms of chloride regulation in mouse hippocampal PV INs and PCs and suggest KCC2 downregulation in the pathology may affect the valence of GABA signaling in both cell types.Key point summaryCation-chloride cotransporters (CCCs) play a critical role in controlling the efficacy and polarity of GABAA receptor (GABAAR)-mediated transmission in the brain, yet their expression and function in GABAergic interneurons has been overlooked.We compared the polarity of GABA signaling and the function of CCCs in mouse hippocampal pyramidal neurons and parvalbumin-expressing interneurons.Under resting conditions, GABAAR activation was mostly depolarizing and yet inhibitory in both cell types. KCC2 blockade further depolarized the reversal potential of GABAAR-mediated currents often above action potential threshold.However, during repetitive GABAAR activation, the postsynaptic response declined independently of the ion flux direction or KCC2 function, suggesting intracellular chloride buildup is not responsible for this form of plasticity.Our data demonstrate similar mechanisms of chloride regulation in mouse hippocampal pyramidal neurons and parvalbumin interneurons.

Lateral diffusion of NKCC1 contributes to neuronal chloride homeostasis and is rapidly regulated by the WNK signaling pathway

“…This indicates that the recombinant NKCC1 transporter is functional in neurons and that the influx of chloride through NKCC1 in neurons in the absence of chloride ion extrusion capacity is toxic. In agreement with previous works [31-32], this result also means that in mature neurons KCC2 is the major regulator of [Cl - ] i . However, no effect of the endogenous NKCC1 or of the recombinant NKCC1-WT or NKCC1-T203/T207/T212/T217/T230A was observed on the YFP/CFP ratio, nor on bumetanide sensitivity in conditions of low KCC2 activity (Fig.…”

“…7C). These results are in agreement with data suggesting that at rest, endogenous NKCC1 do not significantly influence [Cl - ] i in mature hippocampal neurons [31-32]. Similarly, expression of the recombinant NKCC1-WT in mature neurons did not increase [Cl - ] i compared to neurons expressing the chloride probe alone, nor did it increase their sensitivity to bumetanide (Fig.…”

“…This indicates that the recombinant NKCC1 transporter is functional in neurons and that the influx of chloride through NKCC1 in neurons in the absence of chloride ion extrusion capacity is toxic. In agreement with previous works [31][32], this result also means that in mature neurons KCC2 is the major regulator of [Cl -]i.…”

“…7C). However, since NKCC1 plays an important role on [Cl - ] i in mature neurons under conditions where KCC2 is down-regulated [4, 9, 18, 32], we performed additional analyses on neurons co-transfected with the mutant transporter KCC2-T906/1007E, which has a reduced chloride extrusion capacity [24] and Fig. 7A).…”