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TheNa-K-2Clcotransporter2(NKCC2)wasthoughttobekidneyspecific.Hereweshowexpressioninthebrainhypothalamo-neurohypophyseal system (HNS), wherein upregulation follows osmotic stress. The HNS controls osmotic stability through the synthesis and release of the neuropeptide hormone, arginine vasopressin (AVP). AVP travels through the bloodstream to the kidney, where it promotes water conservation. Knockdown of HNS NKCC2 elicited profound effects on fluid balance following ingestion of a high-salt solution-rats produced significantly more urine, concomitant with increases in fluid intake and plasma osmolality. Since NKCC2 is the molecular target of the loop diuretics bumetanide and furosemide, we asked about their effects on HNS function following disturbed water balance. Dehydration-evoked GABAmediated excitation of AVP neurons was reversed by bumetanide, and furosemide blocked AVP release, both in vivo and in hypothalamic explants. Thus, NKCC2-dependent brain mechanisms that regulate osmotic stability are disrupted by loop diuretics in rats.

Experiments were performed in conscious male Wistar rats equipped with a radiotelemetric device. The PVN was unilaterally co-transfected with an adenoviral vector (Ad), engineered to overexpress OT receptors, and an enhanced green fluorescent protein (eGFP) tag. Control groups: PVN was transfected with an Ad expressing eGFP alone or untransfected, sham rats (Wt). Recordings were obtained without and with selective blockade of OT receptors (OTX), during both baseline and stressful conditions. Baroreceptor reflex sensitivity (BRS) and cardiovascular short-term variability were evaluated using the sequence method and spectral methodology respectively. KEY RESULTSUnder baseline conditions, rats overexpressing OT receptors (OTR) exhibited enhanced BRS and reduced BP variability compared to control groups. Exposure to stress increased BP, BP variability and HR in all rats. In control groups, but not in OTR rats, BRS decreased during stress. Pretreatment of OTR rats with OTX reduced BRS and enhanced BP and HR variability under baseline and stressful conditions. Pretreatment of Wt rats with OTX, reduced BRS and increased BP variability under baseline and stressful conditions, but only increased HR variability during stress. CONCLUSIONS AND IMPLICATIONSOT receptors in PVN are involved in tonic neural control of BRS and cardiovascular short-term variability. The failure of this mechanism could critically contribute to the loss of autonomic control in cardiovascular disease.

The hypothalamic paraventricular nucleus (PVN) is a key integrative site for the neuroendocrine control of the circulation and of the stress response. It is also a major source of the neuropeptide hormone vasopressin (VP), and co-expresses V1a receptors (V1aR). We thus sought to investigate the role of V1aR in PVN in cardiovascular control in response to stress.Experiments were performed in male Wistar rats equipped with radiotelemetric device. The right PVN was transfected with adenoviral vectors (Ads) engineered to over-express V1aR along with an enhanced green fluorescent protein (eGFP) tag. Control groups were PVN transfected with Ads expressing eGFP alone, or wild-type rats (Wt). Rats were recorded with and without selective blockade of V1aR (V1aRX) in PVN under both baseline and stressed conditions. Blood pressure (BP), heart rate (HR), their short-term variabilities, and baroreflex sensitivity (BRS) were evaluated using spectral analysis and the sequence method, respectively. Under baseline physiological conditions,V1aR rats exhibited reduced BRS and a marked increase of BP and HR variability during exposure to stress. These effects were all prevented by V1aRX pretreatment. In Wt rats, V1aRX did not modify cardiovascular parameters under baseline conditions, and prevented BP variability increase by stress.However, V1aRX pretreatment did not modify baroreflex desensitization by stress in either rat strain. It follows that increased expression of V1aR in PVN influences autonomic cardiovascular regulation and demarcates vulnerability to stress. We thus suggest a possible role of hypothalamic V1aR in cardiovascular pathology. KeywordsVasopressin, V1a receptor, paraventricular nucleus, adenoviral vector, baroreflex, blood pressure variability, heart rate variability Abbreviations BRS, baroreflex sensitivity; VLF, very low frequency short-term variability; LF, low frequency short-term variability; HF, high frequency short-term variability; VP, vasopressin;OT, oxytocin; OTR, oxytocin receptor; V1aR, vasopressin V1a receptor; V1bR, vasopressin V1b receptor; V2R, vasopressin V2 receptor; V1aRX, vasopressin V1a receptor antagonist, PVN, paraventricular nucleus; NTS, nucleus of the solitary tract; RVLM; rostroventrolateral medulla; IML, intermediolateral column of the spinal cord.

Vasopressin (VP) is a neurohypophyseal peptide best known for its role in maintaining osmotic and cardiovascular homeostasis. The main sources of VP are the supraoptic and paraventricular (PVN) nuclei of the hypothalamus, which co-express the vasopressin V1a and V1b receptors (V1aR and V1bR). Here we investigate the level of expression of VP and VP receptors in the PVN of borderline hypertensive rats (BHRs), a key integrative nucleus in neuroendocrine cardiovascular control. Experiments were performed in male BHRs and Wistar rats (WR) equipped with radiotelemetry device for continuous hemodynamic recordings, under baseline conditions and after saline load without or with stress. The autonomic control of the circulation was evaluated by spectral analysis of blood pressure (BP) and heart rate (HR) variability and baro-receptor reflex sensitivity (BRS) using the sequence method. Plasma VP was determined by radioimmunoassay, and VP, V1aR and V1bR gene expression by RT-qPCR. Under baseline conditions, BHRs had higher BP, lower HR and enhanced BRS in respect to WRs. BP and HR variability was unchanged. In the PVN, overexpression of VP and V1bRs genes were found and plasma VP was increased. Saline load downregulated V1bR mRNA without affecting VP mRNA expression, plasma VP and the BP. Adding stress increased BP, BP and HR low frequency sympathetic spectral markers, decreased plasma VP without altering the level of expression of VP and VP receptors in PVN. It follows that overexpression of VP and V1bR in the PVN is a characteristic trait of BHR, and that sympathetic hyperactivity underlies stressinduced hypertension.