Ionic plasticity diminishes GABAergic inhibition of pre‐sympathetic PVN neurons and facilitates AngII‐salt hypertension

Exaggerated sympathetic nerve activity (SNA) strongly contributes to treatment resistant forms of hypertension (HTN), but neural mechanisms are only partially delineated. Here, we investigated synaptic mechanisms governing hypothalamic paraventricular nucleus (PVN) pre‐sympathetic neurons (PSNs) and how their disruption contributes to heightened SNA in our established mouse model of angiotensin II (AngII)‐dependent salt‐sensitive (AngII‐salt) HTN. Previously, we determined that ongoing synaptic inhibition of PVN PSNs is actively maintained by EAAT3‐mediated uptake of L‐glutamate (Glu) into synaptic terminals of GABAergic neurons in the surrounding peri‐nuclear zone (PNZ). We further determined that activation of excitatory inputs from the forebrain median preoptic nucleus (MnPO), which strongly contribute to AngII‐salt HTN, can acutely strengthen GABAergic inhibition of PVN PSNs. This suggests that forebrain excitation in AngII‐salt HTN can strengthen GABAergic inhibitory tone, but it was previously reported that PVN inhibitory tone is diminished in AngII‐salt HTN, not strengthened. To reconcile these apparently disparate observations, we postulated that reduced PVN inhibition in AngII‐salt HTN results from increased, not decreased, synaptic GABA release that increases intracellular Cl‐ concentration ([Cl‐]i) amongst PVN PSNs, thereby diminishing the inhibitory efficacy of GABA‐A receptor activation. Using gramicidin perforated patch clamp recordings in brain slices, we determined that [Cl‐]i amongst PSNs from normotensive (NT) mice (N=3) and mice with AngII‐salt HTN (N=6) averaged 11.1 ± 2.5 mM (n=3 cells) and 43.5 ± 15.7 mM (n=6 cells), respectively (P=0.0033). The corresponding reversal potential of GABA‐AR current in the NT group was ‐67.2 ± 6.3 mV and was significantly depolarized in the AngII‐salt HTN group to ‐32.9 ± 8.8 mV (P<0.001). In slices prepared from vGAT‐ChR2(H134R) expressing mice, photo‐stimulation of PNZ GABA release rapidly reversed the polarity of evoked IPSCs (Vhold = ‐50 mV). Consistent with this observation, the polarity of postsynaptic current responses to puff application of the GABA‐AR agonist isoguvacine at baseline (+3.29 ± 0.47 pA) was also reversed after PNZ photo‐activation (‐7.47 ± 0.52 pA). Findings indicate that AngII‐salt HTN is accompanied by loss of GABA‐AR inhibitory efficacy reflecting increased [Cl‐]i and that the latter can occur rapidly in response to synaptic GABA release from local PNZ terminals. Findings indicate that inhibitory inputs to PVN from the PNZ can diminish GABAergic inhibition through excess Cl‐ influx via activation of GABA‐AR.