Angiotensin ? And Superoxide Anion Production In Hypothalamic Paraventricular Nucleus Contribute To Capsaicin-induced Excitatory Renal Reflex And Sympathetic Activation

BackgroundSympathetic over-activity contributes to the development of chronic heart failure,hypertension,and chronic kidney disease.Kidneys play important roles in the sympathetic activation in these diseases.Excitatory renal reflex(ERR)is chemical stimulation of kidney with capsaicin,leading to increases in sympathetic activity and blood pressure.Afferent activity from kidneys are closely associated with several brain sites related to the modulation of cardiovascular and sympathetic activity,including nucleus solitary tract(NTS),rostral ventrolateral medulla(RVLM),paraventricular nucleus of hypothalamus(PVN).PVN receives various inputs from visceral receptors including arterial baroreceptors and pulmonary/cardiac vagal afferents,cardiac sympathetic afferents,adipose afferents,and renal afferents.The PVN is critical in the integration and regulation of sympathetic and cardiovascular activity.Renal afferent inputs increase sympathetic activity,and renal nerve stimulation induces excitation of some neurons in the PVN.We found that stimulation of renal afferents with capsaicin promoted c-Fos expression in bilateral PVN,while lesion of bilateral PVN with kainic acid abolished the renal infusion of capsaicin-induced ERR,indicate that the PVN is the important component of the ERR neurocircuitry,but the molecular signaling in the PVN in medating the capsaicin-induced ERR is still unknown.ObjectiveThis study was designed to determine the molecular signaling in the PVN in mediating the capsaicin-induced ERR and sympathetic activation.MethodsExperiments were carried out in 196 male Sprague–Dawley rats weighing 280-320 g.The animals were kept with free access of tap water and chow.The experiments were performed on rats anaesthetized by intraperitoneal injection of?-chloralose(40mg/kg)combined with urethane(800 mg/kg).A right flank incision was performed to expose right kidney.A stainless steel tube(0.31 mm OD)was horizontally inserted into the kidney from the right side to the left side for renal infusion,the insertion was stopped when the tube encountered a slight resistance,indicating that its tip had reached the cortico-medullary border.The outer end of the tube was connected to a programmable pressure injector through a PE50 polyethylene catheter.The ERR was elicited by infusion of capsaicin(1 nmol/?L)into the kidney at 1.0?L/min for 20 min.The ERR was evaluated by the capsaicin-induced RSNA(renal sympathetic nerve activity)and MAP(mean arterial pressure)responses.In anesthetized rats,RANA and MAP were recorded in vivo on a Power Lab data acquisition system.Each rat was fixed prone in a stereotaxic frame(Stoelting,Chicago,IL,USA).The coordinates for microinjections into the PVN were 1.8 mm caudal to bregma,0.4 mm lateral to the midline,and 7.9 mm below the dorsal surface.Glass micropipettes with tip size 50?m were used for bilateral PVN microinjections of 50 n L on each side completed in 1 min.Specific fluorogenic probe dihydroethidim(DHE)and lucigenin-derived chemiluminescence method was used to detect in situ superoxide anions in the PVN.Results1.The capsaicin-induced ERR was almost abolished by PVN pretreatment with losartan or captopril,but losartan or the angiotensin-converting enzyme inhibitor captopril had no significant effects on these baseline values.Microinjection of Ang II into the PVN increased the baseline RSNA and MAP,but failed to have a greater effect on the capsaicin-induced RSNA and MAP changes than those caused by microinjection of PBS.2.The capsaicin-induced ERR was almost abolished by pretreatment with PVN microinjection of tempol or NAC.Microinjection of DETC into the PVN failed to cause greater effects on the capsaicin-induced RSNA and MAP changes than those caused by microinjection of PBS.3.Renal infusion of capsaicin promoted superoxide anion production in the bilateral PVN,and this was prevented by bilateral losartan pretreatment and right renal denervation.4.Microinjection of the NAD(P)H oxidase inhibitor apocynin or DPI reduced the baseline RSNA and MAP,but the xanthine oxidase inhibitor allopurinol had no significant effects on these values.5.Renal infusion of capsaicin increased NAD(P)H oxidase activity in the bilateral PVN,and this was prevented by bilateral losartan pretreatment and right renal denervation.ConclusionCapsaicin-induced ERR is mediated by Ang II in the PVN.The Ang II acts on AT₁receptors in the PVN,and then causes NAD(P)H oxidase activation and superoxide anion production,which induces sympathetic activation and pressor responses.