Hypothalamic SOCS-3 expression and the effect of intracerebroventricular angiotensin II injection on water intake and renal sodium handling in SHR

Abstract

In rats, the acute central dipsogenic and natriuretic action of angiotensin II (AngII) seems to be independent of the hemodynamic effects of the peptide; however, in genetically hypertensive models, this relationship has not yet been investigated. It has been demonstrated that AngII induces the suppressor of cytokine signaling (SOCS-3) expression in the brain that, in turn, modulates further activation of the pathway, leading to desensitization to AngII stimuli with regard to its dipsogenic effect. This study investigates age-related Janus kinase (JAK-2) and SOCS-3 hypothalamic expression, by immunoblotting, and the involvement of SOCS-3 expression in urinary sodium handling and dipsogenic response in spontaneously hypertensive rats (SHR), compared with age-matched Wistar-Kyoto (WKy) rats. The intracerebroventricular (i.c.v.) application of AngII significantly enhanced the dipsogenic response, reduced C(Cr), and reciprocally promoted increased absolute and fractional rates of excretion of sodium in WKy rats. The central AngII-induced dipsogenic effect in WKy and SHR was significantly attenuated by prior i.c.v. administration of DUP753. In addition, the magnitude of the dipsogenic and renal response to AngII was significantly attenuated in age-matched SHR. Blocking of hypothalamic SOCS-3 expression by an antisense oligonucleotide resulted in partial reversal of the refractory nature of AngII in thirst responses in SHR. The altered centrally applied AngII response in SHR associated with increased hypothalamic JAK-2/SOCS-3 expression may suggest that abnormal regulation of the central angiotensin pathways may contribute to dysfunction of water-electrolyte homeostasis in SHR.

Fig. 1

Effect of i.c.v. dose–response injection of AngII on drinking behavior in WKy and SHR rats. Rats were i.c.v. cannulated and received 3.0 μl saline, 3.0 μl of a solution containing AngII or 1 μl DUP753 before 3 μl AngII (amounts stated in the figure). In all groups, the volume of spontaneous water consumption was measured over the next 30 min. In all experiments, n = 5; *P ≤ 0.05 versus saline-treated (ANOVA)

Fig. 2

Effect of i.c.v. injection of 40 pmol AngII upon renal function behavior of rats. Rats were i.c.v. cannulated and received 3.0 μl saline or 3.0 μl of a solution containing AngII. Creatinine clearance (C_Cr), fractional sodium excretion (FE_Na), proximal (FEP_Na) and post-proximal (FEPP_Na) fractional sodium excretion, fractional potassium excretion (FE_K), and absolute urinary sodium excretion (UNaV) in 12-week-old SHR and WKy rats. The data are reported as the mean ± SD. *P ≤ 0.05 versus WKy (ANOVA and Bonferroni’s contrast test, n = 10). See “Results” for details of statistical analysis

Fig. 3

Age-related (at 4, 8, and 12-week-old) SOCS-3 expression and activation of JAK-2 and SOCS-3/JAK-2 association signal transduction in the hypothalami of SHR and WKy rats. After 4, 8, and 12 weeks, hypothalami were obtained and total protein was extracted, according to the protocols described in “Materials and methods”. Immunocomplexes were separated by SDS-PAGE, transferred to nitrocellulose membranes and blotted (IB) with anti-SOCS-3 or anti-phosphotyrosine (pY) antibodies. In all experiments, n = 5; *P ≤ 0.05 SHR versus WKy rats (ANOVA). At the bottom, we show representative densitometric expression of β-actin as the internal standard for protein loading

Fig. 4

Effect of SOCS-3 partial blockade upon AngII-elicited SOCS-3 drinking behavior of SHR. Two groups of rats (a, b) were i.c.v. cannulated and received either SOCS-3 antisense or SOCS-3 sense phosphorothioate-modified oligonucleotides (4.0 nmol in 3.0 μl solution). Thirty minutes after SOCS-3 antisense or sense, a dose of AngII (40 pmol) was injected and water intake was measured over the next 30 min. In all experiments n = 5; *P ≤ 0.05 versus No oligo (no oligonucleotides) (ANOVA and Bonferroni’s contrast test)