Abstract P198: Biphasic And Sex-dependent Roles Of The Prorenin Receptor In The Rostral Ventrolateral Nucleus In Mice Subjected To Deoxycorticosterone Acetate-salt Hypertension

The brain renin angiotensin system (RAS) regulates blood pressure (BP) and autonomic function. However, it remains unclear how and where angiotensin II (Ang II) is generated in conditions eliciting brain RAS overactivation including deoxycorticosterone acetate (DOCA)-salt hypertension (HT). In several tissues, the activation of prorenin requires its binding to the prorenin receptor (PRR). New evidence from this study indicates that prorenin and PRR are co-expressed in the proximity to the rostral ventrolateral nucleus (RVL), an anatomical brain region that controls sympathetic nerve activity. Therefore, we hypothesized that selective ablation of PRR targeting the RVL attenuates BP increase due to DOCA-salt. PRR ablation was targeted to the RVL by stereotactic microinjections of adeno-associated virus (AAV) expressing Cre recombinase-mCherry in PRR-flox mice (PRR RVL-KO ). AAV mCherry was used as control virus (WT). A pressor response to L-glutamate in the injection site served as confirmatory stereotactic target hit. RVL-targeted ablation of PRR resulted in lower BP responses to DOCA-salt in females (WT=115±3 vs KO=104±4 mmHg; p <0.05; n=8), but not males (n=5-8), only during the first 3 days of DOCA-salt treatment. However, at day 13 of DOCA-salt treatment, female PRR RVL-KO unexpectedly exhibited exaggerated increase in systolic BP (WT=149±3 vs KO=163±3 mmHg; p =0.004; n=8) and pulse pressure (WT=31±4 vs KO=45±4 mmHg; p =0.02; n=8) when compared to control. Next, mice were challenged with an intraperitoneal hypertonic saline injection equivalent to 10% of their body weight followed by 4 hours of urine collection. Urinary sodium excretion in female PRR RVL-KO was significantly lower when compared to WT ( p <0.05). These data indicate that the role of PRR in the RVL is sex-dependent and biphasic. That is, PRR contributes to the pressor response during the initial stage of DOCA-salt HT in females, presumably by facilitating the generation of angiotensin peptides in the RVL, while it plays a protective role by promoting renal sodium excretion and preventing elevation of systolic BP during the maintenance stage of DOCA-salt HT. This study suggests that distinct PRR expressing cell populations might elicit diverging physiological functions within the RVL.

Renal Denervation by Noninvasive Stereotactic Radiotherapy Induces Persistent Reduction of Sympathetic Activity in a Hypertensive Swine Model

Background We have previously reported the feasibility of noninvasive stereotactic body radiotherapy (SBRT) as a novel approach for renal denervation. Methods and Results Herein, from a translational point of view, we assessed the antihypertensive effect and chronological evolution of SBRT‐induced renal nerve injury within 6 months in a hypertensive swine model. Hypertension was induced in swine by subcutaneous implantation of deoxycorticosterone acetate pellets in combination with a high‐salt diet. A single dose of 25 Gy with SBRT was delivered for renal denervation in 9 swine within 3.4±1.0 minutes. Blood pressure levels at baseline and 1 and 6 months post‐SBRT were comparable to control (n=5), whereas renal norepinephrine was significantly lower at 6 months ( P <0.05). Abdominal computed tomography, performed before euthanasia and renal function assessment, remained normal. Standard semiquantitative histological assessment showed that compared with control (1.4±0.4), renal nerve injury was greater at 1 month post‐SBRT (2.3±0.3) and peaked at 6 months post‐SBRT (3.2±0.8) ( P <0.05), along with a higher proportion of active caspase‐3–positive nerves ( P <0.05). Moreover, SBRT resulted in continuous dysfunction of renal sympathetic nerves and low level of nerve regeneration in 6 months by immunohistochemistry analysis. Conclusions SBRT delivering 25 Gy for renal denervation was safe and related to sustained reduction of sympathetic activity by aggravating nerve damage and inhibiting nerve regeneration up to 6 months; however, its translation to clinical trial should be cautious because of the negative blood pressure response in the deoxycorticosterone acetate–salt hypertensive swine model.

F2-Isoprostane Levels in Deoxycorticosterone Acetate (DOCA)-Salt Induced Hypertensive Rats Administered with Coffee-Corn Mixture

The purpose of this study is to analyze F2-Isoprostane levels in Deoxycorticosterone Acetate (DOCA)–Salt induced hypertensive rats that is given Coffee-Corn mixture. Measurement of blood pressure was carried out by non-invasive methods using CODA instruments. The coffee-corn mixture was made from Robusta (Coffea canephora) obtained from Kaliwining, Jember Regency, and Yellow Corn (Zea Mays) bought from the local market in Jember, East Java, Indonesia. F2-Isoprostane levels were obtained by ELISA examination. The mice’s blood pressure was measured using a non-invasive CODA® blood pressure gauge. Serum F2-isoprostane levels were measured using an F2-isoprostane-ELISA kit that was read with an Elisa reader at a wavelength of 450nm. The results showed that there was a decrease in mean systolic blood pressure in each treatment group, with the most decrease by 41.88mmHg occured in the group that was given a coffee-corn mixture with a ratio of 50% Robusta coffee (Coffea canephora) Kaliwining and 50% yellow corn (Zea Mays). The highest decrease in diastolic blood pressure by 35.25mmHg was also attained in the same group. From the results of serum F2 isoprostrane level examination, there was a decrease in the average level of serum F2 isoprostrane in the treatment group with the most reduction by 51.34pg/mL occured in the group given a coffee-corn mixture with a ratio of 50% Robusta (Coffea canephora) Kaliwining and 50% yellow maize (Zea Mays). Coffee-corn mixture can reduce blood pressure in hypertensive mice through anti-oxidant effects.

Anti-Hypertensive Activity of Hydro Alcoholic Extract of Cìraka Cūraṇam on High Salt Loaded Wistar Albino Rats

Background: Hypertension is called as Silent killer. The most important risk factor for heart diseases and stroke and it may leads to premature death. Several medicinal plants have high effective in anti-hypertensive and anti-thrombotic activity without any side effects. Aim: To evaluate the anti-hypertensive activity of hydro alcoholic extract of Cìraka cūraṇam on deoxycorticosterone acetate (DOCA) salt induced wistar albino rats. Study design: Observational in-vivo study Place and duration of study: Animal house, Dept. of Pharmacology, Arulmigu Kalasalingam College of Pharmacy, Krishnankoil, Srivilliputtur,Tamilnadu. Materials and methods: Antihypertensive activity was conducted on wistar albino rats by determining serum Sodium and Potassium levels by using semi auto analyzer (RA-50, Bayer Diagnostics), using specific kits (Auto span, India) at 500 and 550 nm respectively and left carotid artery (for recording BP) was cannulated under aseptic conditions with polyethylene cannula filled with 1% heparin in normal saline. Rest procedure, which was stated under the 2K1C-model was followed and BP was observed in terms of mm of Hg. Results: The Cìraka cūraṇam possesses strong antihypertensive effect against DOCA-salt hypertensive rats, which is evidenced by a considerable decrease in blood pressures. Keywords: Anti-hypertensive activity, Cìraka cūraṇam, Wistar albino rats, Deoxycorticosterone acetate.

Activation of Orexin 1 Receptors in the Paraventricular Nucleus Contributes to the Development of Deoxycorticosterone Acetate-Salt Hypertension Through Regulation of Vasopressin

Salt-sensitivity is a major factor in the development of hypertension. The brain orexin system has been observed to play a role in numerous hypertensive animal models. However, orexin’s role in the pathology of salt-sensitive hypertension (SSH) remains to be adequately explored. We assessed the impact of orexin hyperactivity in the pathogenesis of the deoxycorticosterone acetate (DOCA) – salt rat model, specifically through modulation of Arginine Vasopressin (AVP). Adult male rats were separated into three groups: vehicle control, DOCA-salt, and DOCA-salt+OX1R-shRNA. DOCA-salt rats received subcutaneous implantation of a 21-day release, 75 mg DOCA pellet in addition to saline drinking water (1% NaCl and 0.2% KCl). DOCA-salt+OX1R-shRNA rats received bilateral microinjection of AAV2-OX1R-shRNA into the paraventricular nucleus (PVN) to knockdown function of the Orexin 1-Receptor (OX1R) within that area. Following 2-week to allow full transgene expression, a DOCA pellet was administered in addition to saline drinking solution. Vehicle controls received sham DOCA implantation but were given normal water. During the 3-week DOCA-salt or sham treatment period, mean arterial pressure (MAP) and heart rate (HR) were monitored utilizing tail-cuff plethysmography. Following the 3-week period, rat brains were collected for either PCR mRNA analysis, as well as immunostaining. Plasma samples were collected and subjected to ELISA analysis. In line with our hypothesis, OX1R expression was elevated in the PVN of DOCA-salt treated rats when compared to controls. Furthermore, following chronic knockdown of OX1R, the hypertension development normally induced by DOCA-salt treatment was significantly diminished in the DOCA-salt+OX1R-shRNA group. A concurrent reduction in PVN OX1R and AVP mRNA was observed in concert with the reduced blood pressure following AAV2-OX1R-shRNA treatment. Similarly, plasma AVP concentrations appeared to be reduced in the DOCA-salt+OX1R-shRNA group when compared to DOCA-salt rats. These results indicate that orexin signaling, specifically through the OX1R in the PVN are critical for the onset and maintenance of hypertension in the DOCA-salt model. This relationship is mediated, at least in part, through orexin activation of AVP producing neurons, and the subsequent release of AVP into the periphery. Our results outline a promising mechanism underlying the development of SSH through interactions with the brain orexin system.

β-Arrestin–Biased Agonist Targeting the Brain AT 1 R (Angiotensin II Type 1 Receptor) Increases Aversion to Saline and Lowers Blood Pressure in Deoxycorticosterone Acetate–Salt Hypertension

Activation of central AT 1 Rs (angiotensin type 1 receptors) is required for the increased blood pressure, polydipsia, and salt intake in deoxycorticosterone acetate (DOCA)–salt hypertension. TRV120027 (TRV027) is an AT 1 R-biased agonist that selectively acts through β-arrestin. We hypothesized that intracerebroventricular administration of TRV027 would ameliorate the effects of DOCA-salt. In a neuronal cell line, TRV027 induced AT 1a R internalization through dynamin and clathrin-mediated endocytosis. We next evaluated the effect of chronic intracerebroventricular infusion of TRV027 on fluid intake. We measured the relative intake of water versus various saline solutions using a 2-bottle choice paradigm in mice subjected to DOCA with a concomitant intracerebroventricular infusion of either vehicle, TRV027, or losartan. Sham mice received intracerebroventricular vehicle without DOCA. TRV027 potentiated DOCA-induced water intake in the presence or absence of saline. TRV027 and losartan both increased the aversion for saline—an effect particularly pronounced for highly aversive saline solutions. Intracerebroventricular Ang (angiotensin) II, but not TRV027, increased water and saline intake in the absence of DOCA. In a separate cohort, blood pressure responses to acute intracerebroventricular injection of vehicle, TRV, or losartan were measured by radiotelemetry in mice with established DOCA-salt hypertension. Central administration of intracerebroventricular TRV027 or losartan each caused a significant and similar reduction of blood pressure and heart rate. We conclude that administration of TRV027 a selective β-arrestin biased agonist directly into the brain increases aversion to saline and lowers blood pressure in a model of salt-sensitive hypertension. These data suggest that selective activation of AT 1 R β-arrestin pathways may be exploitable therapeutically.