Effects of water deprivation on inhibitory GABAergic synaptic transmission in the hypothalamic paraventricular nucleus (PVN)

Abstract

Presympathetic neurons (PSNs) of the hypothalamic PVN mostly lack basal firing in unstressed animals due to the dominance of GABAergic inhibition, which our findings indicate is actively promoted and defended by the coupling of synaptic glutamate (Glu) release to heightened synthesis and packaging of GABA into synaptic vesicles. Our findings indicate that the latter process, referred to as Glu‐GABA strengthening (GGS), depends on EAAT3‐mediated uptake of Glu into PVN GABA terminals. To investigate the impact of PVN GGS on PSN responses to short‐term exposure to a physiological challenge that increases glutamatergic excitation of the PVN, male C57Bl/6 mice were either euhydrated (EUH) or water deprived (WD) for 48 h. Western blot analysis revealed significantly increased (P<0.05) PVN EAAT3 in WD (132 ± 1.2%) compared to EUH (100 ± 9.5 %) mice (n=18/group). Together with our previous report of reduced PVN expression of the glial Glu transporter GLT‐1 (EAAT2) in WD rats, the present findings suggest that Glu clearance in the PVN is shift during dehydration to favor uptake by neuronal EAAT3. If true, this could indicate that PVN GGS becomes enhanced during dehydration. To explore this further, we determined in brain slices from WD mice (n=4) that the amplitude of evoked IPSCs at baseline (−397 ± 102 pA) was unaffected by bath exposure to Glu (100 μM, 10 min) (−352 ± 89 pA), indicating either that dehydration disrupts PVN GGS or enhances it nearly to its maximum. To distinguish between these possibilities, we generated a cumulative eIPSC amplitude curve from 1000 stimuli delivered at a frequency of 5 Hz to the GABAergic peri‐nuclear zone (PNZ) that surrounds the PVN. The slope (‐pA/stimulus) of this relation in WD mice early during the stimulus train (stimuli 1–100) was similar to that of mice lacking GGS due to EAAT3 knockdown (KD) in PNZ GABA neurons (WD: 186 ± 0.9, EAAT3 KD: 180 ± 0.9). By contrast, the late phase slope (stimuli 901–1000) was 3 time higher for PSNs from WD mice than from the PNZ EAAT3 KD group (WD: 52 ± 0.2, EAAT3 KD: 17 ± 0.1). The latter indicates that eIPSCs in the WD group are resistant to rundown/depression and suggests that PVN GGS is likely near maximally enhanced, not disrupted, during dehydration. We posit that enhanced PVN GGS may be critical for preserving synaptic homeostasis and preventing excessive PSN activation and sympathoexcitation during short‐term dehydration stress.Support or Funding InformationNIH HL088052 (GMT)