mmHg In Sham Research Articles

Acid ceramidase gene therapy ameliorates pulmonary arterial hypertension with right heart dysfunction

BackgroundUp-regulation of ceramides in pulmonary hypertension (PH), contributing to perturbations in sphingolipid homeostasis and the transition of cells to a senescence state. We assessed the safety, feasibility, and efficiency of acid ceramidase gene transfer in a rodent PH model.MethodsA model of PH was established by the combination of left pneumonectomy and injection of Sugen toxin. Magnetic resonance imaging and right heart catheterization confirmed development of PH. Animals were subjected to intratracheal administration of synthetic adeno-associated viral vector (Anc80L65) carrying the acid ceramidase (Anc80L65.AC), an empty capsid vector, or saline. Therapeutic efficacy was evaluated 8 weeks after gene delivery.ResultsHemodynamic assessment 4 weeks after PH model the development demonstrated an increase in the mean pulmonary artery pressure to 30.4 ± 2.13 mmHg versus 10.4 ± 1.65 mmHg in sham (p < 0.001), which was consistent with the definition of PH. We documented a significant increase in pulmonary vascular resistance in the saline-treated (6.79 ± 0.85 mm Hg) and empty capsid (6.94 ± 0.47 mm Hg) groups, but not in animals receiving Anc80L65.AC (4.44 ± 0.71 mm Hg, p < 0.001). Morphometric analysis demonstrated an increase in medial wall thickness in control groups in comparison to those treated with acid ceramidase. After acid ceramidase gene delivery, a significant decrease of pro-inflammatory factors, interleukins, and senescence markers was observed.ConclusionGene delivery of acid ceramidase provided tropism to pulmonary tissue and ameliorated vascular remodeling with right ventricular dysfunction in pulmonary hypertension.Graphical

Abstract 10270: High Shear Stress Decreases ERG Causing Endothelial to Mesenchymal Transition and Pulmonary Arterial Hypertension

Introduction: Computational modeling studies indicated that pathological high shear stress (HSS) of 100 dynes/cm 2 is generated in pulmonary arteries (PA) (100-500μM) in patients with a ventricular septal defect or idiopathic pulmonary arterial hypertension (PAH) and occlusive vascular remodeling. Endothelial-to-mesenchymal transition (EndMT) is a feature of PAH. Hypothesis: Pathological HSS induces EndMT, which contributes to the initiation and progression of PAH. Methods: We apply the Ibidi perfusion system to human PA endothelial cells (EC), to determine whether HSS (100 dynes/cm 2 ) induces EndMT, when compared to normal laminar shear stress (LSS) (15 dynes/cm 2 ). We assessed the mechanism and targeted it to prevent PAH in a mouse with HSS resulting from an aortocaval (AV) shunt. Results: HSS induced EndMT, as assessed by an increase in transcription factors, SNAI1 and SNAI2, reduced BMPR2 (previously shown to inhibit EndMT), decreased EC markers PECAM1 and CDH5, and increased mesenchymal markers, ACTA2 and FSP-1. While the flow-induced transcription factors, KLF2 and KLF4 were similar in LSS and HSS, the co-transcription factor ERG was reduced with HSS. Indeed ERG siRNA under LSS caused EndMT whereas under HSS, transfection of ERG prevented EndMT. To address the significance of our observations in an animal model we created an AV shunt in mice and compared PAH in those sham operated vs transfected with an adeno-associated viral (AAV2- ESGHGYF) vector selectively targeting PAEC with a luciferase (control) or an ERG expressing construct (N=10 per group). Eight weeks after AV shunt, right ventricular systolic pressures was 21.9 ±0.6 mmHg in sham, 37.2±1.0 mmHg in AV shunt with luciferase vector (p&lt;0.01 vs sham) and 29.2±0.8 mmHg in ERG-vector (p&lt;0.01 vs both sham and luciferase). Muscularization of peripheral PA was similarly reduced in ERG-vector (p&lt;0.01 vs luciferase) but not normalized (p&lt;0.01 vs sham) as was immunohistochemical evidence of EndMT (p&lt;0.01 vs luciferase). Conclusions: Our study is the first to document pathological HSS as an inducer of EndMT and PAH and to show that this results from reduced PAEC ERG. Agents that upregulate ERG could reverse HSS mediated PAH and occlusive vascular remodeling as a consequence of high flow or narrowed PAs.

Exaggerated Activities of P2X3 and ASIC3 Signaling Pathway in Muscle Afferent Following Himblimb Muscle Ischemia‐Reperfusion

BackgroundPeripheral artery disease (PAD) is one of the major cardiovascular concerns and dramatically reduces the blood flow supply in the affected limbs. A combination of revascularization surgery in the femoral artery and supervised exercise intervention is among the most general approaches for the treatment of PAD. The approach of revascularization inevitably induces ischemia‐reperfusion (IR) injury in the tissues with the reperfusion of blood flow. Notably, following muscle IR in rats, we have observed an exaggerated exercise pressor reflex (EPR) indicating a risk of cardiovascular diseases, as the increased blood pressure (BP) response during exercise. In specific, a time‐course study (18, 66 and 114 hours post‐IR) showed that an increment in BP response was most profound in 18 hours post‐IR (IR 18h) among all the time points. However the underlying molecular mechanism leading to the IR‐induced exaggerated EPR has yet poorly understood.AimsWe targeted if the hindlimb muscle IR alters the activities of two key signaling pathway in muscle afferent nerves in regulating the EPR, namely P2X3 and ASIC3.MethodsWestern blot technique was used to determine the protein expression of P2X3 and ASIC3 in L4‐6 dorsal root ganglion (DRG) of sham rats and IR 18h rats. Meanwhile, the mean arterial pressure (MAP) responses to α‐β‐me‐ATP and lactic acid intra‐arterially injected into the hindlimb were examined to assess the P2X3 and ASIC3‐mediated EPR, respectively. Whole‐cell patch clamp was applied to evaluate the P2X and ASIC3 currents in the isolated muscle DRG neurons in both sham and IR 18h rats. All data were presented as mean ± standard deviation.ResultsCompared with sham rats, there were significant increases in both P2X3 and ASIC3 protein expression in L4‐6 DRGs in IR 18h rats (P2X3: 1.56±0.10 in IR 18h rats/n=4 vs. 1.00±0.10 in sham rats/n=6; P&lt;0.05; and ASIC3: 1.42±0.47 in IR 18h rats/n=8 vs. 1.00±0.07 in sham rats/n=10; P&lt;0.05). In addition, compared with sham rats, the MAP responses following the injection of α‐β‐me‐ATP (0.125 mM) and lactic acid (2 and 4 µmol/kg) were significantly amplified in IR 18h rats (α‐β‐me‐ATP: 29±8 mmHg in IR 18h rats/n=5 vs. 20±5 mmHg in sham rats/n=2; P&lt;0.05; Lactic acid: 2 µmol/kg, 28±8 mmHg in IR 18h rats/n=15 vs. 21±7 mmH in sham rats/n=8, P&lt;0.05; 4 µmol/kg, 37±10 mmHg in IR 18h rats/n=15 vs. 25±6 mmH in sham rats/n=9, P&lt;0.01). In the isolated DRG neurons, amplitude of both P2X and ASIC3 currents was significantly higher in IR 18h rats than that in sham rats. i.e., P2X transient currents: 1281.64±776.74 pA in IR 18h rats/n=25, vs. 717.91±556.2 pA in sham rats/n=9, (P&lt;0.05); and sustain currents: 466.94±168.68 pA in IR 18h rats/n=11, vs. 271.49±200.90 pA in sham rats/n=8 (P&lt;0.05).ConclusionThe activities of the P2X3 and ASIC3 signaling pathways in the muscle afferents are exaggerated following muscle IR. Results of this study shed light upon the future potential intervention strategy to minimize the IR‐induced exaggerated EPR response in the post‐revascularization PAD patients.

New Histopathologic Evidence for the Parasympathetic Innervation of the Kidney and the Mechanism of Hypertension Following Subarachnoid Hemorrhage.

The Cushing response was first described in 1901. One of its components is elevated systemic blood pressure secondary to raised intracranial pressure. However, controversy still exists in its pathophysiologic mechanism. Hypertension is attributed to sympathetic overactivity and vagotomy increased renal-based hypertension. However, the role of the parasympathetic system in hypertension has not been investigated. This subject was investigated following subarachnoid hemorrhage (SAH). A total of 24 rabbits were used: control group (n = 5), SHAM group (n = 5), and an SAH group (n = 14; bolus injection of blood into the cisterna magna). Blood pressures were examined before, during, and after the experiment. After 3 weeks, animals were decapitated under general anesthesia. Vagal nodose ganglion, axonal degeneration, and renal artery vasospasm (RAV) indexes of all animals were determined histopathologically. Significant degenerative changes were detected in the vagal axons and nodose ganglia following SAH in animals with severe hypertension. The mean degenerated neuron density of nodose ganglions, vasospasm index (VSI) values of renal arteries of control, SHAM, and study groups were estimated as 9.0 ± 2.0 mm, 1.87 ± 0.19; 65.0 ± 12.0 mm, 1.91 ± 0.34; and 986.0 ± 112.0 mm, 2.32 ± 0.89, consecutively. Blood pressure was measured as 94.0 ± 10.0 mmHg in control group, 102.0 ± 12.0 mmHg in SHAM; 112.0 ± 14.0 mmHg in middle (n = 9); and >122.0 ± 10.0 mmHg in severe RAV-developed animals (n = 5). Differences VSI values and blood pressure between groups were statistically significant (P < 0.05). The degeneration of vagal nodose ganglion has an important role in RAV and the development of RAV and hypertension following SAH.

Renal nerves contribute to hypertension in Schlager BPH/2J mice.

Schlager mice (BPH/2J) are hypertensive due to a greater contribution of the sympathetic nervous system (SNS) and renin-angiotensin system (RAS). The kidneys of BPH/2J are hyper-innervated suggesting renal nerves may contribute to the hypertension. We therefore determined the effect of bilateral renal denervation (RD) on hypertension in BPH/2J. Mean arterial pressure (MAP) was measured by radiotelemetry before and for 3 weeks after RD in BPH/2J and BPN/3J. The effects of pentolinium and enalaprilat were examined to determine the contribution of the SNS and RAS, respectively.After 3 weeks, MAP was -10.9 ± 2.1 mmHg lower in RD BPH/2J compared to baseline and -2.1 ± 2.2 mmHg in sham BPH/2J (P < 0.001, n = 8-10). RD had no effect in BPN/3J (P > 0.1). The depressor response to pentolinium was greater in BPH/2J than BPN/3J, but in both cases the response in RD mice was similar to sham. Enalaprilat decreased MAP more in RD BPH/2J compared to sham (-12 vs -3 mmHg, P < 0.001) but had no effect in BPN/3J. RD reduced renal noradrenaline in both strains but more so in BPH/2J. RD reduced renin mRNA and protein, but not plasma renin in BPH/2J to levels comparable with BPN/3J mice.We conclude that renal nerves contribute to hypertension in BPH mice as RD induced a sustained fall in MAP, which was associated with a reduction of intrarenal renin expression. The lack of inhibition of the depressor effects of pentolinium and enalaprilat by RD suggests that vasoconstrictor effects of the SNS or RAS are not involved.

Abstract 12265: Normalization of Renal AT1 Receptor Expression is Observed With Enalapril But Not Diltiazem in a Rat Model of Chronic Kidney Disease

Introduction: Use of ACE inhibitors (ACEi) improves outcomes in patients with proteinuric chronic kidney disease (CKD) and is considered first line of treatment in recent guidelines. In addition to renin angiotensin (Ang) system blockers, other antihypertensive agents such as calcium channel blockers (CCBs) achieve similar therapeutic effectiveness in renoprotection. Our objective was to determine the impact of antihypertensive therapy on renal Ang type 1 receptor (AT 1 R) in 5/6 subtotal nephrectomy (Nx) rat model of CKD using PET imaging. Methods: Ten weeks after Nx, Sprague-Dawley rats (n=6-8/group) were administered 10mg/kg/d enalapril (NxE), 30 mg/kg/d diltiazem (NxD) or left untreated (Nx) for an additional ∼10 weeks. Systolic blood pressure (SBP), plasma creatinine, urine albumin, plasma and tissue Ang II were measured at the end. AT 1 R was assessed using in vivo [ 18 F]FPyKYNE-losartan PET and in vitro autoradiography. Heart function was evaluated with echocardiography. Results: Compared to shams, Nx rats exhibited higher SBP that was reduced by both enalapril and diltiazem (142.7±2, 188.8±17.3, 152±8.7 and 145.8±5.8 mm Hg in sham, Nx, NxE and NxD, respectively, p &lt;0.05). At ∼20 weeks, plasma creatinine and albuminuria were significantly increased in Nx rats, while no changes occurred in NxE rats compared to shams. By contrast, NxD rats demonstrated further increases in these parameters, compared to Nx alone ( p &lt;0.05). Plasma and kidney Ang II were raised in Nx rats, lowered by enalapril and increased significantly in NxD rats ( p &lt;0.05). PET kidney AT 1 R level decreased in Nx rats compared to shams and only enalapril treatment normalized AT 1 R expression (2.8±0.5, 2±0.3, 2.9±0.6, 1.9±0.5 ml/cm 3 in sham, Nx, NxE and NxD respectively, p &lt;0.05) and was confirmed by in vitro results. Enhanced echo LV mass in Nx rats was not reversed by enalapril but was further increased with diltiazem ( p &lt;0.001). Conclusions: In rats with CKD, delayed treatment with the ACEi enalapril normalized AT 1 R expression and reduced progression of the disease, whereas the CCB diltiazem exacerbated renal and cardiac dysfunctions. [ 18 F]FPyKYNE-Losartan PET allows longitudinal determination of AT 1 R abnormalities with progression of cardiorenal failure and monitoring of therapy.

105 - Oxidative Stress Blunts the Vasodilator Effects of Sodium Nitrite Mediated by Xanthine Oxidoreductase in 2K1C Hypertension

Chronic kidney diseases lead to severe cardiovascular consequences such as hypertension and cardiac failure. Apelin, along with its receptor APJ have been shown to involve in cardiovascular functions including blood pressure (BP) lowering effect and also a positive inotropic effect on failing hearts. Therefore we investigated the effect of apelin on BP and cardiac contractility in chronic two-kidney–one-clip (2K1C) hypertension, a kidney disease hypertension model. The changes in the level of apelin and some other hemodynamically effective hormones in serum and apelin receptor gene expression in nonischemic and ischemic kidneys were also assessed.2K1C was produced by placing a Plexiglas clip around the left renal artery. 16 weeks later, BP and cardiac indices of contractility were measured by power lab system. The mRNA and protein level of APJ were determined by RT-PCR and Western blot methods respectively.2K1C increased BP from 115/75 mm Hg in sham to 180/120 mm Hg in test group. Hypertensive rats had about ten times higher basal left ventricular end-diastolic pressure (LVEDP) (P < 0.001) and higher basal LV systolic pressure (LVSP) (P < 0.01). Apelin-13 (20 μg/kg, iv) significantly decreased LVEDP and LVSP (P < 0.001). Furthermore, apelin in 20 μg/kg dose significantly decreased systolic (SBP) and diastolic (DBP) blood pressures in hypertensive rats. This reduction was persistent and prominent in 40 μg/kg dose. 20 μg/kg apelin increased + LVdp/dt max and − LV dp/dt max. However in the dose of 40 μg/kg SBP, DBP, + LVdp/dt max and − LV dp/dt max greatly decreased. All of these effects were completely blocked by apelin antagonist F13A. 2K1C decreased serum apelin (P < 0.05), did not change ang II and arginine-vasopressin levels, and slightly increased serum aldosterone. Apelin receptor mRNA and protein expression significantly decreased in both ischemic and non-ischemic kidneys.Overall, chronic 2K1C rats showed hypertension and signs of cardiac failure. Apelin in medium doses induced antihypertensive and positive myocardial inotropic effects. Reduction of serum apelin and down regulation of apelin receptors in kidneys of hypertensive rats may play a role in pathophysiology of cardiovascular complications.

Abstract 519: Increased Renal Angiotensinogen Expression in Non-clipped Kidneys of 2-Kidney 1-Clip Hypertensive Rats

In 2-kidney 1-clip (2K1C) hypertension, intrarenal angiotensin II (Ang II) levels are increased in both kidneys but the mechanisms for augmentation of Ang II may be different. We recently reported that urinary angiotensinogen (AGT) is increased only in the non-clipped kidneys. However, it has not been determined if these changes are accompanied by augmentation of intrarenal AGT mRNA synthesis in the non-clipped kidneys of 2K1C hypertensive rats. Experiments were performed on male Sprague-Dawley rats (n=11) subjected to left renal arterial clipping (.25 mm gap) and followed for 18-21 days prior to anesthesia and separate measurements of renal function. Systolic arterial pressure increased to 180±3 mmHg compared to 126±4 mmHg in sham operated rats. There were no significant differences in water intake, body weights, and 24 hour urine volume and sodium excretion in awake rats. Separate measurements of renal function showed that renal plasma flow and glomerular filtration rate were similar in clipped and non-clipped kidneys and not different from those in sham rats. However, urine flow, sodium excretion and urinary AGT (uAGT) excretion were significantly greater in non-clipped kidneys compared to clipped and sham kidneys. While kidney AGT protein levels were not increased significantly, AGT transcript measured by real time RT-PCR revealed that the AGT mRNA levels in the cortex were 2.15 fold significantly greater in the non-clipped kidneys than in sham (1.0±.1) or clipped kidneys (.98±.15). The results support the hypothesis that in the non-clipped kidneys of 2K1C rats there is an augmentation of intrarenal AGT mRNA synthesis which explains increased uAGT excretion rates and intrarenal Ang II levels.

Role of afferent and efferent renal nerves in the antihypertensive effect of renal denervation in Ang II‐salt hypertension (856.4)

Renal denervation (RDNX) attenuates drug resistant hypertension (HT) in humans, but it is unclear whether this is due to ablation of afferent or efferent renal nerves. We have previously shown that RDNX attenuates the intravenous (IV) AngII‐salt model of HT in the rat. The present study tested the hypothesis that the antihypertensive effect of RDNX in the IV AngII‐salt model is due to ablation of afferent renal nerves, specifically. Male Sprague Dawley rats underwent SHAM surgery, RDNX or selective ablation of afferent renal nerves by periaxonal capsaicin (renal‐CAP). Rats were instrumented with an IV catheter for delivery of AngII and a radio‐telemeter to measure mean arterial pressure (MAP) and heart rate (HR). Rats were placed on a 4% NaCl diet and, after recovery and a baseline period, AngII was infused IV (10 ng/kg/min) for 16 days. Rats were then sacrificed and kidneys were harvested and assayed for the efferent nerve marker, norepinephrine and the afferent nerve marker, CGRP. Initial results suggest that RDNX and renal‐CAP have similar antihypertensive effects in AngII‐salt rats. On the last day of AngII treatment, MAP was 161 ± 2.8 mmHg in SHAM (n = 4) rats, compared to 133 ± 11.5 mmHg in RDNX (n = 5) and 140 ± 5.2 mmHg in renal‐CAP (n = 4) rats. These preliminary results support the hypothesis that the antihypertensive effect of RDNX in the IV AngII‐salt model is primarily due to the ablation of afferent renal nerves.

454: Biliverdin-Supplementation into the Preservation Solution Prevents Lung Cold Ischemia/Reperfusion Injury

Purpose: Biliverdin (BV), one of the byproducts of heme catalysis through the heme oxygenase system, is known to be a scavenger of reactive oxygen species. We hypothesized that the addition of BV to the perfusate and cold storage solution could protect rat lung grafts from oxidative injury via its anti-oxidant properties. Methods and Materials: Orthotopic left lung transplantation was performed in a syngeneic Lewis to Lewis rat combination under 100% oxygen mixed with isoflurane. Grafts were preserved in low potassium dextran solution at 4C for 6 hrs. The lung graft function was assessed by graft pulmonary vein (PV) blood gas analysis. Results: Prolonged cold storage and reperfusion impaired graft function and resulted in a decrease of graft PV PaO2 to 195 mmHg at 2 hrs after reperfusion from 297 mmHg in sham operated animals. BV treatment significantly improved lung graft function (graft PV PaO2 of 244.8 mmHg). Graft MDA level, a marker of lipid peroxidation, increased to 0.38 M 2 hrs after reperfusion compared to those of sham controls (0.25 M). BV reduced MDA levels to 0.28 M, demonstrating that BV attenuated graft oxidative injury. The effect of BV on proinflammatory cytokine production was also evaluated. Realtime RT-PCR revealed that intragraft mRNA for IL-6 and TNFwere rapidly upregulated within 2 hrs after reperfusion and elevated to 693 fold and 12 fold from base line of sham-operated controls. BV markedly inhibited the increase of these inflammatory mediators to 417 fold and 8.4 fold. IRI resulted in recruitment of polymorphonuclear cells (PMN) into the grafts, but BV-treatment did not reduce the number of infiltrating PMN cells (control 47/0.1mm vs BV 44/ 0.1mm). Conclusions: This study demonstrates that exposure of the lung grafts to BV during cold storage can impart potent cytoprotective effects in lung I/R injury and significantly improve function of lung grafts following extended cold preservation and transplantation by a mechanism involving amelioration of oxidative injury and subsequent inflammatory events.

EARLY RENAL DENERVATION PREVENTS DEVELOPMENT OF HYPERTENSION IN GROWTH‐RESTRICTED OFFSPRING

1. Low birth weight is associated with an increased risk for the development of hypertension. Our laboratory uses a model of reduced uterine perfusion in the pregnant rat that results in intrauterine growth-restricted (IUGR) offspring that develop hypertension at a prepubertal age. Although hypertension develops in both prepubertal male and female IUGR offspring, only male IUGR offspring remain hypertensive after puberty. We reported previously that bilateral renal denervation abolishes hypertension in adult male IUGR offspring, indicating an important role for the renal nerves in the maintenance of established IUGR-induced hypertension. We also reported that angiotensin-converting enzyme inhibition abolishes hypertension in adult male IUGR offspring. However, activation of the renin-angiotensin system does not occur in male IUGR offspring until after puberty, or after the development of established IUGR-induced hypertension. Therefore, the mechanisms involved in the development of IUGR-induced hypertension may differ from those involved in the maintenance of established IUGR-induced hypertension. Thus, the purpose of the present study was to determine whether the renal nerves play a causative role in the early development of IUGR-induced hypertension in prepubertal IUGR offspring. 2. Intrauterine growth-restricted and control offspring were subjected to either bilateral renal denervation or sham denervation, respectively, at 4 weeks of age. Mean arterial pressure (MAP) was determined at 6 weeks of age in conscious, chronically instrumented animals. Adequacy of renal denervation was verified by renal noradrenaline content. 3. Whereas renal denervation had no effect on MAP in control offspring (103 +/- 2 vs 102 +/- 3 mmHg for sham vs denervated, respectively), it reduced blood pressure in growth-restricted offspring (114 +/- 3 vs 104 +/- 1 mmHg for sham vs denervated, respectively; P < 0.01). Renal noradrenaline content was significantly reduced in denervated animals relative to sham operated rats. 4. Thus, the data indicate a role for the renal nerves in the aetiology of IUGR-induced hypertension and suggest that the renal nerves may participate in the early development of hypertension in IUGR offspring in addition to established hypertension observed in adult male IUGR offspring.

Alterations in the Brain–Gut Axis Underlying Visceral Chemosensitivity in Nippostrongylus brasiliensis-Infected Mice

Visceral hypersensitivity, a hallmark of irritable bowel syndrome, is generally considered to be mechanosensitive in nature and mediated via spinal afferents. Both stress and inflammation are implicated in visceral hypersensitivity, but the underlying molecular mechanisms of visceral hypersensitivity are unknown. Mice were infected with Nippostrongylus brasiliensis (Nb) larvae, exposed to environmental stress and the following separate studies performed 3-4 weeks later. Mesenteric afferent nerve activity was recorded in response to either ramp balloon distention (60 mm Hg), or to an intraluminal perfusion of hydrochloric acid (50 mmol/L), or to octreotide administration (2 micromol/L). Intraperitoneal injection of cholera toxin B-488 identified neurons projecting to the abdominal viscera. Fluorescent neurons in dorsal root and nodose ganglia were isolated using laser-capture microdissection. RNA was hybridized to Affymetrix Mouse whole genome arrays for analysis to evaluate the effects of stress and infection. In mice previously infected with Nb, there was no change in intestinal afferent mechanosensitivity, but there was an increase in chemosensitive responses to intraluminal hydrochloric acid when compared with control animals. Gene expression profiles in vagal but not spinal visceral sensory neurons were significantly altered in stressed Nb-infected mice. Decreased afferent responses to somatostatin receptor 2 stimulation correlated with lower expression of vagal somatostatin receptor 2 in stressed Nb-infected mice, confirming a link between molecular data and functional sequelae. Alterations in the intestinal brain-gut axis, in chemosensitivity but not mechanosensitivity, and through vagal rather than spinal pathways, are implicated in stress-induced postinflammatory visceral hypersensitivity.

RESETTING OF AORTIC BARORECEPTORS IN RESPONSE TO HYPOTENSION DOES NOT ALTER GAIN SENSITIVITY

1. In chronic hypertension, the baroreceptors reset to hypertensive levels with a decrease in gain sensitivity, but only a few studies have evaluated baroreceptor resetting during chronic hypotension and, under these conditions, no consistent information is available concerning changes in baroreceptor gain sensitivity. Therefore, in the present study, the aortic baroreceptor function curve and the baroreflex control of heart rate (HR) were evaluated in chronic hypotension produced by myocardial infarction (MI) with no heart failure. 2. Aortic baroreceptor function curves were studied in anaesthetized three groups of rats: (i) MI-7, six rats 7 days after MI; (ii) MI-30, nine rats 30 days after MI; and (iii) five control animals (SHAM). The pressure-nerve activity relationship was measured during rapid changes in blood pressure by integrating the whole-nerve activity of the baroreceptors in a computerized beat-to-beat analysis. 3. Both long-term periods (7 or 30 days) of hypotension were accompanied by complete resetting of the baroreceptor in rats (the leftward displacement of the baroreceptor curve matched the decrease in blood pressure). Moreover, the resetting of the baroreceptor function curve was not accompanied by changes in gain sensitivity (1.47, 1.64 and 1.67%/mmHg for SHAM, MI-7 and MI-30 groups, respectively) and the baroreflex control of HR was normal comparing SHAM and MI-30 groups (bradycardic 1.62 +/- 0.18 vs 1.99 +/- 0.52 b.p.m./mmHg, respectively; tachycardic 3.6 +/- 0.5 vs 4.1 +/- 0.4 b.p.m./mmHg for, respectively). 4. The data indicate that the resetting of baroreceptors in chronic hypotension is stable and is not accompanied by changes in gain sensitivity, as observed in hypertension. This may account for the normal baroreflex control of HR observed in non-anaesthetized rats.

Increased c‐Fos in medullary cardiovascular nuclei in acute and chronic renal wrap hypertension

Renal wrap hypertension is characterized by sodium sensitivity, increased sympathetic outflow, and altered baroreflex function. The present study used c-Fos staining to determine if more neurons are active in brainstem cardiovascular regions in this form of hypertension. Hypertension was produced using a unilateral nephrectomy, renal wrap procedure. Sham operated controls and 1 wk and 4 wk hypertensive rats were perfused and brainstems sectioned coronally (40 um). The sections were processed for c-Fos immunocytochemistry (Oncogene AB-5). Mean arterial pressures were 103 ± 2 mmHg for sham (n = 6), 138 ± 4 for 1 wk renal wrap (n=6), and 159 ± 6 for 4 wk renal wrap (n=6). Renal wrap hypertension was associated with a significant increase in c-Fos staining in the NTS (sham 3 ± 1; 1 wk 12 ± 3; 4 wk 36 ± 3; P < 0.01) with 4 wk renal wrap significantly higher than 1 wk renal wrap. Similar increases were observed in the RVLM (sham 2 ± 1; 1 wk 9 ± 2; 4 wk 17 ± 2; P < 0.01). In the CVLM, c-Fos staining was significantly increased in rats with hypertension but there were no differences between 1wk and 4 wk groups (sham 3 ± 1; 1 wk 6 ± 0.4; 4 wk 7 ± 1; P < 0.01). Changes in c-Fos staining were not observed in the area postrema. Renal wrap hypertension was associated with significant increases in c-Fos staining in the NTS and the RVLM that were related to the level of blood pressure. The failure to observe similar changes in the CVLM could be related to baroreceptor resetting at this site or reflect convergence of NTS inputs to this region. Supported by HL067529 &HL56637

Acute Blood Pressure Effects and Chronic Hypotensive Action of Calcimimetics in Uremic Rats

A previous study in subtotally nephrectomized (SNX) rats suggested beneficial effects of the calcimimetic R-568 beyond the control of mineral metabolism. This study analyzed potential blood pressure (BP)-lowering effects of R-568. Male Sprague-Dawley rats received two-stage subtotal nephrectomy or sham operation. Telemetry devices were inserted into the abdominal aorta, and BP was measured every 5 min. R-568 (20 mg/kg per d) or solvent was infused for 4 wk followed by once-daily subcutaneous injections for 2 wk. Total body sodium was measured by neutron activation analysis. The uremia-induced increase of mean arterial pressure from baseline to day 42 in SNX solvent rats (103+/-5 to 128+/-14 mmHg, P=0.006) was attenuated by R-568 (104+/-5 to 111+/-8 mmHg; P<0.0001 for difference of slopes). The circadian rhythm was abrogated in SNX rats and not restored by R-568. In sham-operated rats, R-568 had only a minor transient antihypertensive effect. R-568 injection induced a transient rise of mean arterial pressure by 23+/-4 and 26+/-10 mmHg in sham and SNX rats but only by 9+/-3 and 10+/-5 mmHg in solvent-treated rats (P<0.01 versus baseline and solvent versus R-568). Plasma angiotensin-converting enzyme activity and aldosterone levels were similar; food intake and physical activity did not differ throughout the study. In healthy rats, total body sodium was higher after 14 d of R-568 compared with solvent infusion (37.1+/-4 versus 32.5+/-1.4 mmol/kg; P=0.01). The calcimimetic R-568 causes an initial BP increase in sham-operated and uremic rats, which in uremic rats is followed by a marked and sustained antihypertensive effect.

Thyroid hormone receptors α1 and β1 are downregulated in the post-infarcted rat heart: consequences on the response to ischaemia-reperfusion

There is accumulating evidence that thyroid hormone metabolism is altered after myocardial infarction (AMI) but its physiological relevance remains largely unknown. The present study investigated the possible role of thyroid hormone signaling in the response of the post-infarcted heart to ischaemia-reperfusion. Wistar rats were subjected to left coronary artery ligation (AMI), or sham operation (SHAM). After 8 weeks, hearts from AMI and SHAM rats were perfused in Langendorff mode and subjected to 20 min of zero-flow global ischaemia (I) and 45 min of reperfusion (R); AMI(I/R), n = 7 and SHAM(I/R), n = 7. Basal left ventricular pressure (LVDP), +dp/dt, and -dp/dt were significantly reduced. Left ventricular weight of the viable myocardium was increased by 14% in the AMI as compared to SHAM hearts, P < 0.05. T(3) and T(4) plasma levels in nM were 1.83 (0.08) and 53.3 (2.9) for SHAM and 1.76 (0.06) and 59.4 (5.2) for AMI rats, respectively, P > 0.05. TRalpha1 and TRbeta1 expression levels were 1.3- and 1.8-fold less in AMI than in SHAM hearts, P < 0.05. Furthermore, SERCA and NHE1 expression levels were 2.1- and 1.8-fold less in AMI than in SHAM, P < 0.05. PKCepsilon was 1.35-fold more in AMI compared to SHAM, P < 0.05. Myocardial glycogen content (in micromol/g) was 7.8 (1.2) in AMI as compared to 4.4 (0.5) for SHAM hearts, P < 0.05. After I/R, left ventricular end-diastolic pressure at 45 min of R (LVEDP45 in mmHg) was 20.3 (3.2) for AMI(I/R) vs 50.6 (4.8) mmHg for SHAM(I/R), P < 0.05. LDH release per gram of tissue was 251 (103) for AMI(I/R) and 762 (74) for SHAM(I/R), P < 0.05. In conclusion, TRalpha1 and TRbeta1 are downregulated after myocardial infarction and this was associated with altered expression of thyroid hormone responsive genes and increased tolerance of the post-infarcted heart to ischaemia-reperfusion injury.

CHRONIC EFFECTS OF ANGIOTENSIN II and AT1 RECEPTOR ANTAGONISTS IN SUBFORNICAL ORGAN‐LESIONED RATS

1. Angiotensin (Ang) II is known to exert some of its effects centrally via circumventricular organs. These unique central nervous system areas lack the normal blood-brain barrier and, therefore, allow peptide hormones access to the brain. Of these, the subfornical organ (SFO) has been shown to be involved in many of the acute dipsogenic and pressor effects of AngII, but much less is known about the role of the SFO in the chronic effects of AngII. We hypothesized that the SFO is a central site involved in the chronic hypotensive effects of endogenous AT(1) receptor blockade, as well as the chronic hypertensive effects of exogenously administered AngII. 2. In order to test these hypotheses, SFO-lesioned (SFOx) or sham Sprague-Dawley rats were instrumented with venous catheters and radiotelemetric pressure transducers for intravenous administration of losartan or AngII and continuous measurement of blood pressure and heart rate. Rats were given 3 days of saline control infusion (7 mL/day of 0.9% NaCl) and were then infused with either losartan (10 mg/kg per day) or AngII (10 ng/kg per min) for 10 days. 3. By day 4 of losartan treatment, arterial pressure had decreased 24 +/- 2 and 18 +/- 2 mmHg in sham (n = 9) and SFOx (n = 10) rats, respectively. Furthermore, by day 5 of AngII infusion, arterial pressure had increased 12 +/- 3 mmHg in sham rats (n = 9), but only by 4 +/- 1 mmHg in SFOx rats (n = 9). In each treatment group, these attenuated pressure responses in SFOx rats continued through day 10 of treatment. 4. These results support the hypotheses that the SFO plays a role in both the hypotensive effects of chronic AT(1) receptor blockade and the chronic hypertensive phase of exogenously administered AngII.

Cardiac angiotensin-(1-7) in ischemic cardiomyopathy.

Accumulating evidence suggests that angiotensin-(1-7) (Ang-[1-7]) may play an important role in counteracting the pressor, proliferative, and profibrotic actions of angiotensin II in the heart. Thus, we evaluated whether Ang-(1-7) is expressed in the myocardium of normal rats and those in which myocardial infarction was produced 4 weeks beforehand. The left coronary artery in 10-week-old Lewis rats was either ligated (n=5) or exposed but not occluded in age-matched controls (sham; n=5). Left ventricular end-diastolic pressures were significantly elevated 4 weeks after myocardial infarction (25+/-1 versus 5+/-1 mm Hg for sham; P<0.001), whereas left ventricular systolic pressures were significantly reduced (ligated 86+/-4 versus sham 110+/-5 mm Hg; P<0.01). Hemodynamic effects of coronary artery ligation were accompanied by significant cardiac hypertrophy (heart weight to body weight: ligated 4.3+/-0.1 versus sham 2.9+/-0.1 mg/g; P<0.001). In both ligated and sham rats, Ang-(1-7) immunoreactivity was limited to cardiac myocytes and absent in interstitial cells and coronary vessels. Ang-(1-7) immunoreactivity was significantly augmented in ventricular tissue surrounding the infarct area in the heart of rats with myocardial infarction. Development of heart failure subsequent to coronary artery ligation leads to increased expression of Ang-(1-7),which was restricted to myocytes.

Glucocorticoids act in the dorsal hindbrain to increase arterial pressure.

Glucocorticoid receptors (GRs) are present at a high density in the nucleus of the solitary tract (NTS), an area of the dorsal hindbrain (DHB) that is critical for blood pressure regulation. However, whether these receptors play any role in the regulation of blood pressure is unknown. We tested the hypothesis that glucocorticoids act in the DHB to increase arterial pressure using two experimental strategies. In one approach, we implanted pellets of corticosterone (Cort) or sham pellets onto the DHB over the NTS. Compared with rats with sham pellets, rats with DHB Cort pellets had an increased (P < 0.05) mean arterial pressure (111 +/- 2 vs. 104 +/- 1 mmHg) and heart rate (355 +/- 9 vs. 326 +/- 5 beats/min) after 4 days. In the second approach, we implanted subcutaneous Cort pellets to increase the systemic Cort concentration and then subsequently implanted pellets of the GR antagonist mifepristone (Mif; previously RU-38486) or sham pellets onto the DHB. Two days of DHB Mif treatment reduced (P < 0.05) mean arterial pressure in those rats with elevated plasma Cort levels (118 +/- 2 vs. 108 +/- 1 mmHg for sham vs. Mif DHB pellets). Cort and Mif pellets placed on the dura had no effects on arterial pressure or heart rate, ruling out systemic cardiovascular effects of the steroids. DHB Cort treatment had no effects on plasma Cort concentration or adrenal weight, indicating that the contents of the DHB Cort pellet did not diffuse into the systemic circulation or into the forebrain areas that regulate plasma Cort concentration in concentrations sufficient to produce physiological effects. Immunohistochemistry for the occupied GRs demonstrated that the Cort and Mif from the DHB pellets were delivered to the DHB with minimal diffusion to the ventral hindbrain or forebrain. We conclude that glucocorticoids act in the DHB to increase arterial pressure.

Estrogen or the AT1 antagonist olmesartan reverses the development of profound hypertension in the congenic mRen2. Lewis rat.

The influence of estrogen on the regulation of cardiovascular function remains a controversial and complex area of investigation. We assessed the effects of estrogen depletion in the congenic mRen(2). Lewis rat, established from the back-cross of the original (mRen2)-27 transgenic onto the Lewis inbred strain. Ovariectomy of heterozygous mRen(2). Lewis at 4 to 5 weeks resulted in a progressive increase in blood pressure compared with the sham surgery congenics at weeks 6 to 11. At 11 weeks, the ovariectomized mRen(2). Lewis (OVX) systolic blood pressure averaged 195+/-3.7 mm Hg versus 141+/-4.0 mm Hg for sham. Plasma Angiotensin (Ang) II, serum ACE activity, plasma renin concentration, as well as urinary excretion of Ang II, 8-isoprostane F2alpha, and endothelin-1 were elevated; however, renal mRNA levels of eNOS were suppressed after ovariectomy. Estrogen replacement reduced blood pressure below both the sham and OVX by 11 weeks (125+/-2.9 mm Hg, n=7, P<0.01 versus OVX and sham). Moreover, the AT1 receptor antagonist olmesartan (CS866; week 12 to 16) essentially normalized blood pressure to 113+/-5.4 mm Hg (n=6, P<0.01 versus OVX and sham). The attenuation of the hypertension was still evident 7 weeks after complete withdrawal of treatment (124+/-4.1 mm Hg at week 23). In summary, the OVX mRen.2. Lewis exhibited a rapid and sustained increase in blood pressure. Estrogen or olmesartan lowered pressure by a similar extent. We conclude that the ovary exerts considerable influence on the regulation of the blood pressure in the mRen2. Lewis strain, possibly by limiting activation of the renin-angiotensin system.