Diurnal variation in neuronal chloride levels and seizure susceptibility, in neocortex, reflecting changes in activity of chloride-cation-cotransporters

Summary The main inhibitory synaptic currents, gated by gamma-aminobutyric acid (GABA), are mediated by Cl - -conducting channels 1–3 , and are therefore sensitive to changes in the chloride electrochemical gradient. GABAergic activity dictates the neuronal firing range 4,5 and timing 6–9 , which in turn influences the rhythms of the brain, synaptic plasticity, and flow of information in neuronal networks 7,10–12 . The intracellular chloride concentration [Cl - ] i is, therefore, ideally placed to be a regulator of neuronal activity. Chloride levels have been thought to be stable in adult cortical networks, except when associated with pathological activation 13–16 . Here, we used 2-photon LSSmClopHensor imaging, in anaesthetized young adult mice 13 , to show that [Cl - ] inside pyramidal cells shows a physiological diurnal rhythm, with an approximately 1.8-fold range, equating to an ~15mV positive shift in ECl at times when mice are typically awake (midnight), relative to when they are usually asleep (midday). This change of [Cl - ] i alters the stability of cortical networks, as demonstrated by a greater than 3-fold longer latency to seizures induced by 4-aminopyridine at midday, compared to midnight. Importantly, both [Cl - ] i and latency to seizure, in night-time experiments, were shifted in line with day-time measures, by inhibition of NKCC1. The redistribution of [Cl - ] i reflects diurnal changes in surface expression and phosphorylation states of the cation-chloride-co-transporters, KCC2 and NKCC1, leading to a greatly reduced chloride-extrusion capacity at night (awake period). Our data demonstrate a means by which changes in the biochemical state of neurons are transduced into altered brain states.