Effects Of Renal-artery Denervation On Renal Sympathetic Nerve Activity And Sodium Handling In Hypertensive Rats

Hypertension has become a global health problem. Excessive sympatheticdrive is involved in the pathophysiology of hypertension. Clinical and animalexperiments show that renal sympathetic denervation can reduce the peripheryand the central sympathetic activity, increase renal sodium excretion, and lead tolower blood pressure. There has been some controversy on renal sympatheticnerve in regulation of blood pressure. The mechanism of how renal sympatheticdenervation to reduce peripheral and central sympathetic activity, and increaserenal sodium excretion is still unclear.The sodium retention induces the secretion of endogenous ouabain (EO),resulting in reduced activity of Na＋-K＋-ATPase and expression of renal sodiumhydrogen exchange protein (NHE), leads to stimulation of the RAAS; on theother hand, increased central sympathoexcitation, and enhanced peripheralsympathetic nerve activity. The purpose of the present study was to investigatethe effects of renal denervation on RAAS and EO of spontaneously hypertensiverats (SHR), and to investigate the potential role of renal sympathetic nerve onthe expression of NHE and NHE regulatory protein family in the kidneys ofSHR. Our studies demonstrate regulation mechanism of renal denervation on renal sodium/water handling, and confirm an essential role of sympatheticexcessive activation in maintenance and progression of the essentialhypertension. In the current studies, the renal nerves were denervated byapplying a solution of phenol in95%alcohol on renal vessels, and theresearches were carried out as the following.Part1: To establish an animal model of renal-artery denervationObjective: To confirmed the effectiveness of the animal model of renalsympathetic denervation by detecting NE content of serum and kidney tissues,and TH staining of kidney.Methods: Twelve-week-old male SHR were divided into2groups, therenal denervated (RDNX) or Sham groups, using Wistar-Kyoto (WKY) rats asnormotensive controls(n=8). Renal denervation was induced by painting therenal vessels with10%phenol, and the sympathetic nervous activity wasassessed2weeks later. The expressions of tyrosine hydroxylase (TH) andnorepinephrine (NE) content were determined in kidney, hypothalamus andadrenal. Serum NE was also examined in rats.Results: Following the renal sympathetic denervation, the BP in RDNXwas normalized to the level found in WKY rats, and serum NE content wassignificantly lower in RDNX than in Sham, and was not different between WKYrats and RDNX. The NE contents and TH stainings of kidney, hypothalamus andadrenal medulla were significantly lower in RDNX than in Sham, and were notdifferent between WKY rats and RDNX.Conclusion: Our results suggest that it is a successful method for the renalsympathetic denervation by applying a solution of phenol in95%alcohol onrenal vessels, and resulted in decreased sympathetic nerve activity of peripheral and central sympathetic nervous system. The renal sympathoinhibition canlower blood pressure, and reduce serum NE content, and cause the inhibition ofsympathoadrenal system.Part2: Effects of renal denervation on the renal sodium handlingand renal sympathetic nervous activity in SHRObjective: To evaluate the potential role of the adrenergic system in theregulation of solute and fluid secretion, and investigated the effect of renaldenervation on renal function.Methods: The rats were divided into RDNX, Sham and WKY groups. The renalfunction was assessed2weeks later by Kw/Bw, serum creatinine, urinaryprotein, Ccr, CNa, FELi, FDRNaand Ucr/Scr. The sections of the kidney werestained with PAS and Masson, and assessed glomerular and arterialmorphological characteristics by glomerular injury score, IF score and themedial thickness-to-lumen ratio.Results: No significant differences in Kw/Bw, urinary protein, FDRNaandCcr were found among the three groups. Ucr/Scr was significantly elevated afterrenal denervation. CNaand FELiwere increased in RDNX compared with Shamgroup. Glomerulosclerosis score and medial thickness-to-lumen diameter ratioof small arteries were significantly decreased in the RDNX group comparedwith the Sham group (0.41±0.09vs.0.83±0.16,0.28±0.07vs.0.54±0.05; p＜0.05for both). Interstitial fibrosis score was similar between RDNX andSham.Conclusion: Renal sympathetic nerve activity has important effects onsodium/water handling, and renal function of concentration and dilution.Decreased renal sympathetic activity may prevent the progression of renal injury caused by hypertension.Part3: Effects of renal denervation on RAAS in SHRObjective: To investigate the alteration of RAAS, so that to discuss thepossible mechanism of renal sympathetic denervation on treatment ofhypertension.Methods: The rats were divided into RDNX, Sham and WKY groups. Theexpression of the three systems was examined2weeks later. ALD contents ofserum and tissue were measured by RIA. The expressions of AngⅡand AT1Rprotein were determined by immunohistochemical staining. Relative qRT-PCRwas performed to detect the expressions of AT1RmRNA in kidney.Results: Renal denervation not only decreased peripheral contents of ALD,but also reduced both of these hormones in kidney and hypothalamus of SHR.AT1R mRNA expression was significantly decreased in the kidney of RDNXcompared with Sham (P＜0.05), and not difference between RDNX and WKYrats. Immunohistochemistry of AT1R confirmed their decreases in thedenervated kidney, and ANGⅡprotein was also lower in RDNX than in Sham.Conclusion: Decreased renal sympathetic activity result in decreases inALD contents of serum and kidney, AT1R and ANGⅡexpression in kidney.Part5: Effects of renal denervation on Na+/H+exchanger proteinin the kidney of SHRObjective: To investigate the potential role of renal sympathetic nerve onthe expression of NHE and NHE regulatory protein family in the kidneys ofSHR.Methods: The rats were divided into RDNX, Sham and WKY groups. Theexpression of NHE and NHE regulatory protein family was examined2weeks later. Relative qRT-PCR was performed to detect the expressions of NHE1,NHE3, NHERF1and NHERF2mRNA in kidney. The expressions of NHE1,NHE3, NHERF1and NHERF2protein were determined byimmunofluorescence histochemistry and Western-blot.Results: Following the renal denervation, NHE1and NHE3mRNA weresignificantly decreased in the kidney compared with Sham (-1.84fold and-4.01fold, respectively). NHERF1mRNA expression was significantly increased inRDNX compared with Sham, whereas NHERF2mRNA expression wassignificantly decreased after renal denervation. Immunohistochemistry (FITCsetting) and western blot analysis of NHE1, NHE3, NHERF1and NHERF2confirmed their changes in the denervated kidney.Conclusion: It is suggested that the sympathetic nerve should play thespecific excitatory role in the regulation of NHE in the kidney. Moreover,NHERF1is likely to be the relevant biological regulator of NHE3in theproximal tubule.Taken together with our previous observations, we propose that：(1) It is asuccessful method for the renal sympathetic denervation by applying a solutionof phenol in95%alcohol on renal vessels, and resulted in decreased sympatheticnerve activity of peripheral and central sympathetic nervous system.(2) Renalsympathetic nerve activity has important effects on sodium/water handling, andrenal function of concentration and dilution. Decreased renal sympatheticactivity may prevent the progression of renal injury caused by hypertension.(3)RAAS and sympathetic nervous system may be relevant to the regulation ofNHE activity and sodium/water handling by direct and/or indirect effects, andplay an important role in the regulation of Na-K-ATPase activity in the kidney.