Regulation of chemoconvulsant-induced seizures by store-operated Orai1 channels.

KEY POINTS: Temporal lobe epilepsy is a complex neurological disease caused by imbalance of excitation and inhibition in the brain. Growing literature implicates altered Ca2+ signalling in many aspects of epilepsy but the diversity of Ca2+ channels that regulate this syndrome are not well-understood. Here, we report that mice lacking the store-operated Ca2+ channel, Orai1, in the brain show markedly stronger seizures in response to the chemoconvulsants, kainic acid and pilocarpine. Electrophysiological analysis reveals that selective deletion of Orai1 channels in inhibitory neurons disables chemoconvulsant-induced excitation of GABAergic neurons in the CA1 hippocampus. Likewise, deletion of Orai1 in GABAergic neurons abrogates the chemoconvulsant-induced burst of spontaneous inhibitory postsynaptic currents (sIPSCs) on CA1 pyramidal neurons in the hippocampus. This loss of chemoconvulsant inhibition likely aggravates status epilepticus in Orai1 KO mice. These results identify Orai1 channels as regulators of hippocampal interneuron excitability and seizures. ABSTRACT: Store-operated Orai1 channels are a major mechanism for Ca2+ entry in many cells and mediate numerous functions including gene expression, cytokine production and gliotransmitter release. Orai1 is expressed in many regions of the mammalian brain; however, its role in regulating neuronal excitability, synaptic function and brain disorders has only now begun to be investigated. To investigate a potential role of Orai1 channels in status epilepticus induced by chemoconvulsants, we examined acute seizures evoked by intraperitoneal injections of kainic acid (KA) and pilocarpine in mice with a conditional deletion of Orai1 (or its activator STIM1) in the brain. Brain-specific Orai1 and STIM1 knockout (KO) mice exhibited significantly stronger seizures (P = 0.00003 and P < 0.00001), and higher chemoconvulsant-induced mortality (P = 0.02) compared with wildtype (WT) littermates. Electrophysiological recordings in hippocampal brain slices revealed that KA stimulated the activity of inhibitory interneurons in the CA1 hippocampus (P = 0.04) which failed to occur in Orai1 KO mice. Further, KA and pilocarpine increased the frequency of spontaneous IPSCs in CA1 pyramidal neurons >twofold (KA: P = 0.04; pilocarpine: P = 0.0002) which was abolished in Orai1 KO mice. Mice with selective deletion of Orai1 in GABAergic neurons alone also showed stronger seizures to KA (P = 0.001) and pilocarpine (P < 0.00001) and loss of chemoconvulsant-induced increases in sIPSC responses compared with WT controls. We conclude that Orai1 channels regulate chemoconvulsant-induced excitation in GABAergic neurons and that destabilization of the excitatory/inhibitory balance in Orai1 KO mice aggravates chemoconvulsant-mediated seizures. These results identify Orai1 channels as novel molecular regulators of hippocampal neuronal excitability and seizures.

Astrocyte reactivity and inflammation-induced depression-like behaviors are regulated by Orai1 calcium channels.

Astrocytes contribute to brain inflammation in neurological disorders but the molecular mechanisms controlling astrocyte reactivity and their relationship to neuroinflammatory endpoints are complex and poorly understood. In this study, we assessed the role of the calcium channel, Orai1, for astrocyte reactivity and inflammation-evoked depression behaviors in mice. Transcriptomics and metabolomics analysis indicated that deletion of Orai1 in astrocytes downregulates genes in inflammation and immunity, metabolism, and cell cycle pathways, and reduces cellular metabolites and ATP production. Systemic inflammation by peripheral lipopolysaccharide (LPS) increases hippocampal inflammatory markers in WT but not in astrocyte Orai1 knockout mice. Loss of Orai1 also blunts inflammation-induced astrocyte Ca2+ signaling and inhibitory neurotransmission in the hippocampus. In line with these cellular changes, Orai1 knockout mice showed amelioration of LPS-evoked depression-like behaviors including anhedonia and helplessness. These findings identify Orai1 as an important signaling hub controlling astrocyte reactivity and astrocyte-mediated brain inflammation that is commonly observed in many neurological disorders.

CRAC channels regulate astrocyte Ca2+ signaling and gliotransmitter release to modulate hippocampal GABAergic transmission.

Astrocytes are the major glial subtype in the brain and mediate numerous functions ranging from metabolic support to gliotransmitter release through signaling mechanisms controlled by Ca2+ Despite intense interest, the Ca2+ influx pathways in astrocytes remain obscure, hindering mechanistic insights into how Ca2+ signaling is coupled to downstream astrocyte-mediated effector functions. Here, we identified store-operated Ca2+ release-activated Ca2+ (CRAC) channels encoded by Orai1 and STIM1 as a major route of Ca2+ entry for driving sustained and oscillatory Ca2+ signals in astrocytes after stimulation of metabotropic purinergic and protease-activated receptors. Using synaptopHluorin as an optical reporter, we showed that the opening of astrocyte CRAC channels stimulated vesicular exocytosis to mediate the release of gliotransmitters, including ATP. Furthermore, slice electrophysiological recordings showed that activation of astrocytes by protease-activated receptors stimulated interneurons in the CA1 hippocampus to increase inhibitory postsynaptic currents on CA1 pyramidal cells. These results reveal a central role for CRAC channels as regulators of astrocyte Ca2+ signaling, gliotransmitter release, and astrocyte-mediated tonic inhibition of CA1 pyramidal neurons.