Mechanisms of GABA-mediated inhibition of the angiotensin II-induced cytosolic Ca increase in rat subfornical organ neurons.

Neurons in the subfornical organ (SFO) sense both neurotransmitters and circulating humoral factors such as angiotensin II (AII) and atrial natriuretic peptide (ANP), and regulate multiple physiological functions including drinking behavior. We recently reported that AII at nanomolar concentrations induced a persistent [Ca] increase in acutely dissociated SFO neurons and that this effect of AII was reversibly inhibited by GABA. In the present study, we studied the inhibitory mechanism of GABA using Ca imaging and patch-clamp electrophysiology. The AII-induced persistent [Ca] increase was inhibited by GABA in more than 90% of AII-responsive neurons and by other two SFO inhibitory ligands, ANP and galanin, in about 60 and 30% of neurons respectively. The inhibition by GABA was mimicked by the GABA and GABA receptor agonists muscimol and baclofen. The involvement of both GABA receptor subtypes was confirmed by reversal of the GABA-mediated inhibition only when the GABA and GABA receptors antagonists bicuculline methiodide and CGP55845 were both present. The GABA agonist baclofen rapidly and reversibly inhibited voltage-gated Ca channel (VGCC) currents recorded in response to depolarizing pulses in voltage-clamp electrophysiology using Ba as a charge carrier (I). Baclofen inhibition of I was antagonized by CGP55845, confirming GABA receptor involvement; was reduced by N-ethylmaleimide, suggesting downstream Gi-mediated actions; and was partially removed by a large prepulse, indicating voltage-dependency. The magnitude of I inhibition by baclofen was reduced by the application of selective blockers for N-, P/Q-, and L-type VGCCs (ω-conotoxin GVIA, ω-agatoxin IVA, and nifedipine respectively). Overall, our study indicates that GABA inhibition of the AII-induced [Ca] increase is mediated by both GABA and GABA receptors, and that GABA receptors associated with Gi proteins suppress Ca entry through VGCCs in SFO neurons.