Orexin facilitates GABAergic IPSCs via postsynaptic OX ₁ receptors coupling to the intracellular PKC signalling cascade in the rat cerebral cortex

Orexin has multiple physiological functions including wakefulness, appetite, nicotine intake, and nociception. The cerebral cortex receives abundant orexinergic projections and expresses both orexinergic receptor 1 (OX ₁ R) and 2 (OX ₂ R). However, little is known about orexinergic regulation of GABA-mediated inhibitory synaptic transmission. In the cerebral cortex, there are multiple GABAergic neural subtypes, each of which has its own morphological and physiological characteristics. Therefore, identification of presynaptic GABAergic neural subtypes is critical to understand orexinergic effects on GABAergic connections. We focused on inhibitory synapses at pyramidal neurons (PNs) from fast-spiking GABAergic neurons (FSNs) in the insular cortex by a paired whole-cell patch-clamp technique, and elucidated the mechanisms of orexin-induced IPSC regulation. We found that both orexin A and orexin B enhanced unitary IPSC (uIPSC) amplitude in FSN→PN connections without changing the paired-pulse ratio or failure rate. These effects were blocked by SB-334867, an OX ₁ receptor (OX ₁ R) antagonist, but not by TCS-OX2-29, an OX ₂ R antagonist. [Ala ¹¹ , D-Leu ¹⁵ ]-orexin B, a selective OX ₂ R agonist, had little effect on uIPSCs. Variance-mean analysis demonstrated an increase in quantal content without a change in release probability or the number of readily releasable pools. Laser photolysis of caged GABA revealed that orexin A enhanced GABA-mediated currents in PNs. Downstream blockade of G _q/11 protein-coupled OX ₁ Rs by IP ₃ receptor or protein kinase C (PKC) blockers and BAPTA injection into postsynaptic PNs diminished the orexin A-induced uIPSC enhancement. These results suggest that the orexinergic uIPSC enhancement is mediated via postsynaptic OX ₁ Rs, which potentiate GABA _A receptors through PKC activation.