Function In Normotensive Rats Research Articles

A16408 THE LACK OF ALAMANDINE EFFECTS ON ISCHEMIA/REPERFUSION IN TG (mREN-2)27 RATS HEARTS IS ASSOCIATED TO BLUNTED EXPRESSION OF MrgD RECEPTOR

Objectives: Our previous studies showed that Alamandine improved post-ischemic cardiac function in normotensive rats. Here we assessed Alamandine effects in a more challenging condition, using hypertensive rats. Methods: Hearts of Sprague Dawley (SD) or TGR (mREN2)27 rats, 12 weeks old, were placed on a Langendorff apparatus to evaluate the cardiac parameters. Hearts were submitted to 30 min of stabilization, 30 min of partial ischemia and 30 min of reperfusion. Alamandine (ALA) was used at 20pM concentration. 2,3,5-trypheniltetrazolium chloride (1%) was used to evaluate the extension of infarcted area. In additional hearts MrgD mRNA was determined by RT-PCR in each period. Results: Hypertensive rats showed an impairment of heart function at the baseline period. A decrease of left ventricular systolic pressure (LVSP), Maximum and minimum dP/dt (35.57 ± 5.839 mmHg, 493,6 ± 51,92mmHg/s and 270,7 ± 28,81mmHg/s, respectively) was observed. This impairment was attenuated by ALA (54.91 ± 6,304mmHg, 859,4 ± 80,85mmHg/s and 478,5 ± 32,94mmHg/s, respectively). Alamandine was unable to prevent the ischemia-induced decrease in LVSP, max and min dP/dt in hearts from mREN rats as it does in hearts from SD rats. ALA treatment decreased arrhythmia severity index and infarcted area extension in SD hearts. No improvement was observed in mREN hearts. MrgD mRNA expression which was already lower in mREN hearts in the baseline period, decreased to negligible values during ischemia/reperfusion. Conclusion: These data suggest that MrgD plays an essential role in heart function of TGR (mREN2)27 rats.

Abstract P190: Alamandine Improves Cardiac Post-Ischemic Function in Isolated Hearts of TGR(mREN2)27

The renin-angiotensin system (RAS) is an important regulator of cardiovascular function. Over the past years, new peptides have been described in this system. Among these new peptides, Alamandine was recently described by Santos and colleagues as an important participant in the control of cardiovascular function through its interaction with MrgD receptors. Our previous studies showed that Alamandine improved post-ischemic cardiac function in normotensive rats. Here we assessed the effects of Alamandine in a more challenging condition using hypertensive rats. TGR(mREN2)27 rats weighing between 250-300g were euthanized and their hearts were placed on Langendorff apparatus to evaluate the cardiac parameters. Hearts were submitted to 30min of stabilization, 30min of partial ischemia by occlusion of the left descending coronary artery and 30min of reperfusion. Alamandine (ALA, 22pM) was added to the perfusion setting from the beginning of the experiment until the end. 2,3,5-trypheniltetrazolium chloride were used to evaluate the extension of infarcted area. In hypertensive (mREN) animals there was an impairment in the baseline (35.57 ± 5.839 mmHg) of left ventricular systolic pressure (LVSP), and this impairment was attenuated by ALA (54.91 ± 6,304mmHg). Maximum and minimum dP/dt in mREN presented a significant reduction in the ischemic period (926.5 ± 172.3 mmHg/s and 682.7 ± 106.1 mmHg/s, respectively) compared to baseline (1761 ± 219.7 mmHg/s and 1242 ± 150.9 mmHg/s, respectively). Ischemia/reperfusion induced an arrhythmia severity index (ASI) in mREN hearts higher than in hearts treated with ALA (4,9 ± 1,26) vs (1,10 ± 0,58). ALA also reduced infarcted area compared to mREN (19,64 ± 2,61%) vs (33,85 ± 4,55%). In conclusion, our data shown that Alamandine exert cardioprotective effects in post-ischemic function in hypertensive rat isolated hearts by preventing LVSP, dP/dt max and min decrease. Furthermore, it reduced the infarcted area and I/R arrhythmias.

Flow mediated dilation variation based on normotensive rat strain

Previous studies from this lab have suggested that nitric oxide (NO) plays a significant role in the maintenance of arterial pressure in the same strain of rats obtained from different animal vendors. Subsequently, we hypothesized that similar variations in NO and thus, endothelial function, could be found in different strains of rat. Flow mediated dilation (FMD) is traditionally a non‐invasive technique used clinically to assess endothelial function. The hyperemic response elicited using this in vivo technique is thought to be mediated by NO release from the endothelium. The goal of the present study was to determine whether the FMD hyperemic response was different between two normotensive strains from the same supplier, Sprague‐Dawley (SD) and Wistar‐Kyoto rats (WKY) from Charles River (CR). In anesthetized rats, the femoral artery was isolated and occluded for 5 minutes, with femoral blood flow measured continuously before, during and after occlusion by use of an ultrasonic perivascular flow probe. Arterial pressure was measured directly via a catheter inserted into the contralateral femoral artery. The clamp procedure was repeated four times in each rat and then averaged. The average area under the FMD response curve (3 minute duration) for the CR SD rats was 79.0 ± 22.9 mL/min·s (n=6) and 371.0 ± 67.4 mL/min·s (n=5) for the CR WKY rats. These results suggest differences in endothelial function in normotensive rats and could have broad implications for research involving these strains of rat.

Structural integration and cardiovascular effects following transplantation of endothelial progenitor cells (EPCs) into the brainstem of rats

Our evidence indicates that brainstem endothelium is a viable target for controlling arterial pressure. Thus, we have used EPCs to target the brainstem microvasculature in vivo and assessed any effect on cardiovascular function. Bone marrow mononuclear cells from male Wistar rats were expanded for 7–15 days in conditions appropriate for the endothelial lineage. EPCs were characterized by immunofluorescence staining, flow cytometry, and using the Matrigel assay for network formation. Fluorescently tagged EPCs were injected intraparenchymally into the nucleus tractus solitarii (NTS), a region regulating arterial pressure, of male Wistar rats. Experiments were terminated 16 days post‐injection. EPCs remained at the site of injection until animals were sacrificed. During this time, EPCs migrated away from the injection site becoming closely associated with the vasculature. Peak measurements occurred between 10 to 15 days post injection and included a reduction in: systolic pressure (SP)(−7±0.5 mmHg), low frequency spectra of SP (−19%±1.0‐ indicating reduced sympathetic vasomotor tone) and heart rate (−18±4.3 beats/min). Cardiac baroreflex sensitivity increased (22%±0.8). In conclusion, EPCs remain viable for at least 16 days when transplanted into the NTS in vivo, integrate into the vascular wall and alter cardiovascular autonomic function in normotensive rats.MRC & Pfizer funded research.

Run training ameliorates the established erectile dysfunction in rats under long-term nitric oxide (NO) blockade

Introduction Stimulation of nitrergic neurons and endothelial cells in the erectile tissue results in release of NO that diffuse to surrounding smooth muscle cells where it activates the soluble guanylate cyclase (sGC), facilitating the conversion of GTP to cGMP. This second messenger diminishes the intracellular levels of calcium thereby causing penile smooth muscle relaxation and penile erection [1]. Epidemiological studies have shown a strong association between erectile dysfunction (ED) and arterial hypertension [2], where the deficiency of the NO-cGMP pathway seems to greatly contribute to such association [3]. Regular physical exercise has been shown to increase the NO production thus ameliorating cardiovascular diseases [2,4]. Recently, we have shown that prior physical conditioning improves the erectile function in normotensive rats [5] and prevents the impaired corpus cavernosum relaxation secondary to chronic NO blockade in rats [2].

Enhanced Superoxide Activity Modulates Renal Function in NO-Deficient Hypertensive Rats

An enhancement of superoxide (O2-) activity was shown to contribute to the development of hypertension induced by NO deficiency. To better understand the mechanistic role of O2- in this NO-deficient hypertension, we evaluated the renal responses to acute intraarterial administration of an O2- scavenger, tempol (50 microg/min per 100 g of body weight) in anesthetized male Sprague-Dawley rats treated with NO synthase inhibitor nitro-L-arginine methyl ester (15 mg/kg per day in drinking water, n=7) for 4 weeks, which caused increases in mean arterial pressure (146+/-3 versus 124+/-2 mm Hg) compared with normotensive control rats (n=6). Hypertensive rats had higher renal vascular resistance (29+/-2 versus 20+/-1 mm Hg/mL per minute per gram), as well as lower renal blood flow (5.2+/-0.3 versus 6.3+/-0.2 mL/min per gram; cortical blood flow, 153+/-13 versus 191+/-8 perfusion units; medullary blood flow, 43+/-2 versus 51+/-3 perfusion units) and glomerular filtration rate (0.69+/-0.04 versus 0.90+/-0.05 mL/min per gram) without a significant difference in urinary sodium excretion (0.81+/-0.07 versus 0.86+/-0.12 micromol/min per gram) compared with normotensive rats. Urinary 8-isoprostane excretion rate (6.8+/-0.7 versus 4.5+/-0.3 pg/min per gram) was higher in hypertensive than normotensive rats. Intraarterial infusion of tempol did not alter renal function in normotensive rats. However, tempol significantly decreased renal vascular resistance by 12+/-2% and urinary 8-isoprostane excretion rate by 24+/-4% and increased renal blood flow by 10+/-2%, cortical blood flow by 9+/-2%, medullary blood flow by 15+/-6%, glomerular filtration rate by 11+/-3%, and urinary sodium excretion by 19+/-5% in hypertensive rats. These data indicate that enhanced O2- activity modulates renal hemodynamics and excretory function during reduced NO production and, thus, contributes to the pathophysiology of the NO-deficient form of hypertension.

Influence of sodium intake on the cardiovascular and renal effects of brain mineralocorticoid receptor blockade in normotensive rats.

We have previously shown that brain mineralocorticoid receptors (MR) participate in the control of arterial pressure and renal excretory function in normotensive rats. In the present study, we evaluate the influence of sodium intake on the control of cardiovascular and renal function by brain MR in normotensive conscious rats. We hypothesize that modulation of sodium intake affects the cardiovascular and renal effects of brain MR blockade. We examined the effect of intracerebroventricular (ICV) administration of MR antagonist (RU28318) on systolic blood pressure (SBP), heart rate, and urinary excretion of water and electrolytes in normotensive Wistar rats subjected to different dietary sodium content. Rats were fed high (8%), normal (0.4%), or depleted (0%) sodium for 3 weeks. SBP and heart rate measurements were performed using an indirect tail cuff method. Metabolic cages were used to assess renal function. ICV injection of 100 ng RU28318 induced a long-lasting decrease ( 0.01) in SBP in rats submitted to different sodium intake. At 8 h, the decrease in SBP did not differ between rats on high (30 +/- 5 mmHg), normal (28 +/- 6 mmHg), and low (21 +/- 3 mmHg) sodium diets. At 24 h, the decrease in SBP was also comparable between rats on different sodium diets. Increased diuresis was observed during the period 0-8 h after ICV injection of RU28318; this was less pronounced in rats on the low sodium diet. Urinary excretions of potassium and chloride were also increased during this period, particularly in rats on the high and normal sodium diets compared with rats with low sodium intake. Urinary excretion of sodium was increased only in the rats fed high and normal sodium diets. Measurement of plasma renin activity, which was suppressed and stimulated, respectively, by high and low sodium intake, indicated that the effects on SBP and renal function induced by ICV RU28318 were independent from the level of activation of the renin-angiotensin system. In contrast to our hypothesis, in normotensive Wistar rats, sodium intake does not affect the hypotension induced by brain MR blockade. However, sodium depletion attenuated the renal effects of brain MR blockade.

Influence of age on cardiovascular effects of increased dietary sodium and angiotensin-converting enzyme inhibition in normotensive Wistar rats.

Recent studies have shown that increased intake of dietary sodium chloride produces blood pressure-independent increase in cardiac and renal mass even in young normotensive rats. With advancing age the harmful cardiovascular effects of increased dietary sodium are not so well known. In the present study the influence of advancing age on the cardiovascular effects of increased intake of sodium (control diet, 0.3% and high-sodium diet, 2.6% sodium in the chow) were examined in young and aged (3 and 18 months old, respectively, at the beginning of the experiment) male normotensive Wistar rats in a six-week study. Moreover, the potential role of renin-angiotensin system in ageing during normal and a high-sodium intake was studied using a pharmacological tool, angiotensin converting enzyme (ACE) inhibitor ramipril. Ageing did not significantly modify basal systolic blood pressure measured by the tail cuff method. A high intake of sodium chloride increased blood pressure significantly only in aged rats, while in young rats it increased renal weight. Left ventricular weight was not affected by high-sodium diet in either age group. The ACE inhibition during control diet lowered blood pressure and decreased left ventricular weight in young rats only and these effects were completely blocked by a high-sodium diet. The maximal vascular contraction force of mesenteric arterial rings to noradrenaline was decreased with ageing while endothelium-dependent and -independent relaxation responses were unaltered with ageing. The sensitivity to sodium nitroprusside was impaired by the high-sodium diet in young rats. In both age groups the urinary excretion of calcium was increased during the high-sodium diet. In conclusion, the increased intake of sodium produced different changes in cardiovascular function in normotensive rats depending on age. With advancing age, the sensitivity to sodium-induced increase in blood pressure was increased. In aged rats a high intake of dietary sodium elevated blood pressure, while in young rats it increased renal mass without increase in blood pressure. In both age groups sodium did not affect left ventricular hypertrophy. Both high-sodium intake and ageing attenuated or even abolished the cardiovascular effects of ACF inhibition.

Renal function in rats with angiotensin li-salt-induced hypertension: effect of thromboxane synthesis inhibition and receptor blockade

This study was designed to assess the contribution of thromboxane (Tx) A2 to the pathogenesis of renal dysfunction in rats with angiotensin II (Ang II)-salt hypertension. Hypertension was induced in rats drinking 0.15 mol/l NaCl by infusion of Ang II (125 ng/min, intraperitoneally) for 12 days. Relative to values in water- and saline-drinking rats without Ang II infusion, rats with Ang II-salt hypertension exhibited increased renal vascular resistance, decreased renal blood flow, and increased renal excretion and glomerular synthesis of TxB2. Treatment with an inhibitor of TxA2 synthesis, UK 38,485, had no effect on renal function in normotensive and hypertensive rats. Similarly, the TxA2 and prostaglandin endoperoxide antagonist SQ 29,548 did not affect renal function in normotensive rats. In contrast, in rats with Ang II-salt hypertension of 12 days' duration, SQ 29,548 caused a reduction in renal vascular resistance, allowing for maintenance of renal blood flow in the face of an accompanying reduction in blood pressure. A comparable reduction in renal perfusion pressure, produced by constriction of the abdominal aorta above the renal arteries, was not accompanied by a reduction in renal vascular resistance in Ang II-salt hypertensive rats. Therefore, the SQ 29,548-induced lowering of renal vascular resistance is attributable not to renal blood flow autoregulation, but to blockade of the renal vasoconstrictor actions of TxA2 and/or prostaglandin endoperoxides. This interpretation implies that pressor eicosanoids contribute to increase renal vascular resistance in rats with severe Ang II-salt hypertension.

Cardiovascular effects of chronic high-dose atriopeptin III infusion in normotensive rats

Seventy-eight Sprague-Dawley rats received continuous intravenous infusions of either atriopeptin III (APIII), 60 μg/kg/hr, or distilled water vehicle for a period of 7 days by means of osmotic minipumps. On Day 7 approximately one-half of the animals (20 vehicle-treated rats and 21 APIII-treated rats) were instrumented for evaluation of cardiac function and terminated for measurement of heart weight. The minipumps remained in place during the evaluation of cardiac function. Also on Day 7, the osmotic pumps were removed from the remaining animals and an additional 7 days were allowed to elapse before heart weight and cardiac function were evaluated. Mean arterial blood pressure (MAP) of rats receiving APIII for 7 days was significantly lower (−9%, p < 0.05) than that of rats receiving vehicle for 7 days. In addition, reductions ( p < 0.05) of total ventricular weight dry (−7%), left ventricular weight dry (−8%), and right ventricular weight dry (−9%) were observed in the APIII-treated rats (all ventricular weights are normalized for body weight). Hematocrit (HCT) was significantly higher (13%, p < 0.05) in the APIII-treated group. Chronic APIII infusion did not influence ventricular performance nor did it affect regional vascular resistances. Seven days after termination of the APIII infusion the differences in MAP and HCT between vehicle-treated and APIII-treated animals were no longer evident. Partial recovery of the effect on heart weights was apparent, with total ventricular weight dry and left ventricular weight dry remaining slightly reduced (−4 and −5%, respectively; p < 0.05). No differences were found between the two recovery groups for any index of cardiac function. In separate experiments, it was demonstrated that APIII, 60 μg/kg/hr iv, caused a significant increase in urine volume ( p < 0.05 relative to vehicle) during the initial 24 hr of infusion. The results indicate that chronic infusion of a large diuretic dose of APIII exerts relatively little influence on overall cardiovascular function in normotensive rats.

Thyrotropin releasing hormone: autonomic effects upon cardiorespiratory function in endotoxic shock

These studies define potential sites and mechanisms by which thyrotropin releasing hormone (TRH) stimulates cardiorespiratory function in normotensive rats as well as in rats subjected to endotoxic shock. Changes in mean arterial pressure, pulse pressure, heart rate, and respiratory rate were determined in conscious animals following injection of TRH into the lateral, third, or fourth ventricular spaces. Injections of TRH into the third ventricular space resulted in a greater increase in cardiorespiratory variables than did fourth ventricular injection. In endotoxin-treated rats, the cardiorespiratory effects of intracerebroventricular (icv) TRH and its analog MK 771 were assessed. TRH and MK 771 were shown to act within the brain to reverse endotoxic shock hypotension; at the doses used, the pressor effects of these two tripeptides were achieved through selectively different actions upon heart rate and pulse pressure. Adrenal demedullated and sham-operated control rats subjected to endotoxic shock were injected with icv and intravenous (iv) TRH in order to evaluate the potential involvement of sympatho-medullary function in cardiorespiratory responses. The cardiovascular effects of icv TRH were dependent upon adrenal medullary integrity; effects of iv TRH were not. Doses of iv TRH which effectively reverse shock neither altered nociceptive latencies nor interfered with analgesic responses to morphine. Collectively, these studies reinforce the potential therapeutic utility of TRH and its analogs in the treatment of shock and indicate potential sites and mechanisms which mediate these salutary effects.