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The suprachiasmatic nucleus is the circadian pacemaker of the mammalian brain. Suprachiasmatic nucleus neurons display synchronization of their firing frequency on a circadian timescale, which is required for the pacemaker function of the suprachiasmatic nucleus. However, the mechanisms by which suprachiasmatic nucleus neurons remain synchronized in vivo are poorly understood, although synaptic communication is considered indispensable. Suprachiasmatic nucleus neurons contain the neurotransmitter GABA and express GABA receptors. This has inspired the hypothesis that GABA signalling may play a central role in network synchronization, although this remains untested in vivo. Here, using local genetic deletion, we show that disruption of GABA synaptic transmission within the suprachiasmatic nucleus of adult mice results in the eventual deterioration of physiological and behavioural rhythmicity in vivo and concomitant cellular desynchrony in vitro. These findings suggest that intercellular GABA signalling is essential for behavioural rhythmicity and cellular synchrony of the suprachiasmatic nucleus neural network.

Cl-sensitive with-no-lysine kinase (WNK) plays a key role in regulating the thiazide-sensitive Na+-Cl- cotransporter (NCC) in the distal convoluted tubule (DCT). Cl- enters DCT cells through NCC and leaves the cell across the basolateral membrane via the Cl- channel ClC-K2 or K+-Cl- cotransporter (KCC). While KCC is electroneutral, Cl- exit via ClC-K2 is electrogenic. Therefore, an alteration in DCT basolateral K+ channel activity is expected to influence Cl- movement across the basolateral membrane. Although a role for intracellular Cl- in the regulation of WNK and NCC has been established, intracellular Cl- concentrations ([Cl-]i) have not been directly measured in the mammalian DCT. Therefore, to measure [Cl-]i in DCT cells, we generated a transgenic mouse model expressing an optogenetic kidney-specific Cl-Sensor and measured Cl- fluorescent imaging in the isolated DCT. Basal measurements indicated that the mean [Cl-]i was ~7 mM. Stimulation of Cl- exit with low-Cl- hypotonic solutions decreased [Cl-]i, whereas inhibition of KCC by DIOA or inhibition of ClC-K2 by NPPB increased [Cl-]i, suggesting roles for both KCC and ClC-K2 in the modulation of [Cl-]i. Blockade of basolateral K+ channels (Kir4.1/5.1) with barium significantly increased [Cl-]i. Finally, a decrease in extracellular K+ concentration transiently decreased [Cl-]i, whereas raising extracellular K+ transiently increased [Cl-]i, further suggesting a role for Kir4.1/5.1 in the regulation of [Cl-]i. We conclude that the alteration in ClC-K2, KCC, and Kir4.1/5.1 activity influences [Cl-]i in the DCT. [ABSTRACT FROM AUTHOR]