Nitric oxide pathway is an important modulator of heat loss in rats during exercise

To assess the role of nitric oxide (NO) in central thermoregulatory mechanisms during exercise, 1.43 micromol (2 microL) of N(omega)-nitro-L-arginine methyl ester (L-NAME, n=6), a NO synthase inhibitor, or 2 microL of 0.15M NaCl (SAL, n=6) was injected into the lateral cerebral ventricle of male Wistar rats immediately before the animals started running (18 m min(-1), 5% inclination). Core (Tb) and skin tail (Ttail) temperatures were measured. Body heating rate (BHR), threshold Tb for tail vasodilation (TTbV), and workload (W) were calculated. During the first 11 min of exercise, there was a greater increase in Tb in the L-NAME group than in the SAL group (BRH=0.17+/-0.02 degrees C min(-1), L-NAME, versus 0.09+/-0.01 degrees C min(-1), SAL, p<0.05). Following the first 11 min until approximately 40 min of exercise, Tb levels remained stable in both groups, but levels remained higher in the L-NAME group than in the SAL group (39.16+/-0.04 degrees C, L-NAME, versus 38.33+/-0.02 degrees C, SAL, p<0.01). However, exercise went on to induce an additional rise in Tb in the SAL group prior to fatigue. These results suggest that the reduced W observed in L-NAME-treated rats (10.8+/-2.0 kg m, L-NAME, versus 25.0+/-2.1 kg m, SAL, p<0.01) was related to the increased BHR in L-NAME-treated animals observed during the first 11 min of exercise (r=0.74, p<0.01) due to the change in TTbV (39.12+/-0.24 degrees C, L-NAME, versus 38.27+/-0.10 degrees C, SAL, p<0.05). Finally, our data suggest that the central nitric oxide pathway modulates mechanisms of heat dissipation during exercise through an inhibitory mechanism.     

Muscarinic cholinoceptors in the ventromedial hypothalamic nucleus facilitate tail heat loss during physical exercise

The aim of this study was to evaluate the participation of ventromedial hypothalamic nucleus (VMH) muscarinic cholinoceptors in heat balance and central fatigue during treadmill exercise (24 m min(-1), 5% inclination). The animals were anesthetized with pentobarbital sodium (50 mg/kg body weight i.p.) and fitted with bilateral cannulae into the VMH 1 week prior to the experiments. Tail skin (T(tail)) and core body temperatures (T(b)) were measured after the injection of 0.2 microL of 5 x 10(-9) mol methylatropine (Matr) or 0.15 M NaCl solution (Sal) into the hypothalamus. Methylatropine injection into the VMH greatly increased heat storage rate (HSR) measured until fatigue (19.7+/-4.6 cal min(-1) Matr versus 9.7+/-3.3 cal min(-1) Sal; P<0.05) and attenuated the exercise-induced tail vasodilation as seen by T(tail) (23.98+/-0.43 degrees C Matr versus 25.52+/-0.85 degrees C Sal; at 6.5 min; P<0.05), indicating inhibition of the heat loss process. The 2 min delay and the increased DeltaT(b), which triggered the heat loss mechanisms observed in Matr-treated rats, are associated with increased HSR and may be responsible for the decreased running performance of these animals (21.0+/-2.9 min Matr versus 33.5+/-3.4 min Sal; P<0.001). In fact, a close negative correlation was observed between HSR and time to fatigue (r=-0.61; P<0.01). In conclusion, VMH muscarinic cholinoceptors facilitate tail heat loss mechanisms, and a delay in this adjustment would lead to a decrease in physical exercise performance due to excess heat storage.     

Physiological responses to maximal exercise on arm cranking and wheelchair ergometer with paraplegics.

This study describes the responses of 20 paraplegic athletes (mean age: 26.8 +/- 1.6 years) to a continuous incremental workload test until exhaustion on an arm cranking ergometer (ACE) and on a wheelchair ergometer (WCE). Both ergometers used the same electromagnetic braking device allowing a fair comparison between results. Tests were conducted at a 24 hour interval at the same time of the day. Oxygen uptake (VO2), heart rate (HR), workload (W), blood pressure (BP), Borg index, and mechanical efficiency (ME) were measured at every minute during the effort and the cool down periods of both tests. The purpose of this study was to analyse the different responses obtained on ACE and on WCE during maximal effort by paraplegics, and also to determine which ergometer permits the higher ME. Results indicate that paraplegics reached the same max HR on ACE and on WCE (97% of the predicted max HR). The lack of significant difference (p less than 0.05) between ACE and WCE in terms of maximal values of VO2, VE and HR suggests that the subjects reached their maximal capacity on each test regardless of the type of ergometer. Nevertheless, W max (in Watts) was 26% higher on ACE than on WCE. Maximal ME values were respectively 16% and 11.6% on ACE and WCE. Results suggest that ergometers and protocol used in this study are appropriate to measure physiological responses of paraplegic athletes during arm cranking and wheelchair exercise without excessive or early arm fatigue.     

Angiotensin AT(2) receptor stimulates ERK1 and ERK2 in quiescent but inhibits ERK in NGF-stimulated PC12W cells

To investigate the influence of AT(2) receptor stimulation on the ERK pathway and elucidate potential mechanisms of angiotensin II (ANG II)-mediated neuronal differentiation, we analysed tyrosine phosphorylation and activity of ERK after ANG II treatment of both quiescent and NGF-treated PC12W cells. Tyrosine phosphorylation of ERK1 and ERK2 corresponded with the activity of ERK. While ANG II induced an initial activation of ERK in quiescent cells, the NGF-mediated plateau of ERK-stimulation was lowered by costimulation with ANG II. All effects of ANG II were sensitive to AT(2) - but not AT(1) receptor blockade. Ang II-mediated neurite outgrowth in PC12W cells was inhibited by co-treatment with the MEK inhibitor PD 098059. These findings demonstrate that the AT(2) receptor modulates ERK activity depending on the overall cellular input. The distinct regulation of ERK by ANG II and NGF further indicates basic differences in AT(2) receptor- and NGF-induced neuronal differentiation.     

The differential effect of angiotensin II and angiotensin 1-7 on norepinephrine, epinephrine, and dopamine concentrations in rat hypothalamus: the involvement of angiotensin receptors

Angiotensin 1-7 has been recently claimed the active member of the angiotensins' family. In the present study we compared the effect of angiotensin II and angiotensin 1-7 on the concentration of dopamine, serotonin, epinephrine, and norepinephrine and some of their metabolites in the rat hypothalamus, where the levels of angiotensins are particularly high. Intracerebroventricular injection of angiotensin II, but not angiotensin 1-7, time-dependently elevated the levels of both epinephrine (p < 0.05) and norepinephrine (p < 0.05) in the hypothalamus and both effects could be prevented by intracerebroventricular injection of either AT(1) (candesartan), AT(2) (PD123319) or AT(1-7) (A-779) receptor antagonist. Neither angiotensin II nor angiotensin 1-7 produced any changes in the level of dopamine, dihydroxyphenylacetic acid, homovanilic acid, serotonin, 5-hydroxyindoleacetic acid, or tryptophan at any time point in comparison with the control groups. However, AT(1) but not AT(2) receptor blockade, unmasked the stimulatory effect of angiotensin 1-7 on dopamine concentration in the hypothalamus. Thus, angiotensin II and its active metabolite angiotensin 1-7 regulate selectively, albeit differentially, adrenergic, noradrenergic and dopaminergic systems in the hypothalamus, the effects that involve AT(1), AT(2) and AT(1-7) angiotensin receptors.     

Endurance training in MS: short-term immune responses and their relation to cardiorespiratory fitness, health-related quality of life, and fatigue

The influences of exercise on cytokine response, health-related quality of life (HR-QoL), and fatigue are important aspects of MS rehabilitation. Physical exercises performed within these programs are often practiced in water, but the effects of immersion have not been investigated. To investigate the influences of short-term immune responses and cardiorespiratory fitness on HR-QoL and fatigue during 3 weeks endurance training conducted on a cycle-ergometer or an aquatic-bike. Randomized controlled clinical trial in 60 MS patients. HR-QoL, fatigue, cardiorespiratory fitness, and short-term immune changes (serum concentrations in response to cardiopulmonary exercise test) of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), interleukin-6, and the soluble receptor of IL-6 (sIL-6R) were determined at the beginning and end of 3 weeks of training intervention. Subjects performed daily 30 min training at 60 % of their VO2peak. SF-36 total (p = 0.031), physical (p = 0.004), and mental health (p = 0.057) scores show time effects within both groups. Between-group effects were shown for FSMC total (p = 0.040) and motor function score (p = 0.041). MFIS physical fatigue showed time effects (p = 0.008) for both groups. Linear regression models showed relationships between short-term immune responses and cardiorespiratory fitness with HR-QoL and fatigue after the intervention. This study indicates beneficial effects of endurance training independent of the training setting. Short-term immune adaptations and cardiorespiratory fitness have the potential to influence HR-QoL and fatigue in persons with MS. The specific immune responses of immersion to exercise need further clarification.     

An exercise test to evaluate fitness for wheelchair activity.

The purpose of this study was to develop a wheelchair ergometer (WERG) test to evaluate fitness for manual wheelchair activity. Thirty able-bodied females participated in a progressive intensity, discontinuous test where exercise bouts were 4 min in duration interspersed with 5-min rest periods. Physiological responses of oxygen uptake (VO2), respiratory exchange ratio (R), net mechanical efficiency (ME), pulmonary ventilation (VO) and heart rate (HR) were determined during the final minute of exercise at power output (PO) levels of 30, 60, 90, 120 and 150 kpm/min. These responses were generally found to be linearly related to PO, however, net ME initially increased with PO and plateaued at approximately 11 per cent at 90 kpm/min. Criteria for fitness evaluation were based upon: (1) magnitude of physiological responses at each PO level; and (2) the maximal PO level completed.     

AT(2) but not AT(1) receptor antagonism abolishes angiotensin II increase of the acquisition of conditioned avoidance responses in rats

In this study we attempted to determine behavioural, including cognitive, consequences of the brain AT(1) (losartan, 2 nmol), AT(2) (PD 123319, 1.5 nmol), and joint AT(1)/AT(2) angiotensin receptors blockade. Male Wistar rats (160-180 g) were injected into the left cerebral ventricle with the above doses of the blockers dissolved in 0.9% NaCl solution (vehicle) or with the vehicle alone. Five minutes later they received, to the right cerebral ventricle, 1 nmol of angiotensin II (Ang II) dissolved in vehicle or the vehicle alone. Ang II consistently increased rate of acquisition of conditioned avoidance response (CARs) and facilitated recall of the passive avoidance behaviour. In one out of the three series of experiments in open field Ang II stimulated rats locomotor activity. Losartan and PD 123319, both ineffective alone, given prior to Ang II abolished all the behavioural changes produced by the peptide except for the Ang II facilitation of CARs acquisition, which was unchanged by losartan. Interestingly, joint injection of losartan and PD 123319 significantly decreased the rate of CARs acquisition both in control and Ang II treated animals. In conclusion, the present data suggest significant though different involvement of both AT(1) and AT(2) angiotensin receptors in cognitive processes.     

Candesartan prevents angiotensin II-induced facilitation of hypoxic neuronal damage through PKCdelta inhibition

To investigate the role of protein kinase Cdelta (PKCdelta) in angiotensin II-induced facilitation mechanisms of hypoxic neuronal damage and whether candesartan, an AT1 receptor antagonist, can suppress these mechanisms, we performed in vitro experiments which were free from vascular components using PC12 cells under hypoxic (12 h)/reoxygenation (0-48 h) conditions. Angiotensin II apparently increased the basal expression level of PKCdelta phosphorylated at Ser(643) before hypoxia, promoted the cleavage of PKCdelta to its catalytic fragment, and fostered the progression of DNA fragmentation after hypoxia. Candesartan inhibited both phosphorylation and cleavage of PKCdelta and suppressed the angiotensin II-induced facilitation of DNA fragmentation under hypoxic/reoxygenation conditions. However, PD123319, an AT2 receptor antagonist, influenced neither PKCdelta nor the angiotensin II-induced facilitation of DNA fragmentation. Furthermore, in PC12 cells expressing the ATP-binding mutant of PKCdelta (PKCdelta(K376R)) acting as a dominant-negative protein, both phosphorylation and cleavage of PKCdelta were attenuated and DNA fragmentation was markedly suppressed regardless of the presence of angiotensin II. These findings suggest that angiotensin II-induced facilitation of DNA fragmentation under hypoxic conditions is mediated by PKCdelta, and the mechanisms can be suppressed by the candesartan mediated blockade of the AT1 receptor.     

Effects of a short-term exercise training program on aerobic fitness, fatigue, health perception and activity level of subjects with multiple sclerosis

Multiple sclerosis (MS) patients of an inpatient rehabilitation program have been randomly assigned to an exercise training (MS-ET) or nontraining group (MS-NI). Before and after 4 weeks of aerobic exercise training, a graded maximal exercise test with measurement of gas exchange and a lung function test was administered to all 26 patients fulfilling the inclusion criteria. Activity level, fatigue and health perception were measured by means of questionnaires. Twenty-six healthy persons served as control group and were matched in respect of age, gender and activity level. Training intervention consisted of 5x30 min sessions per week of bicycle exercise with individualised intensity. Compared with baseline, the MS training group demonstrated a significant rightward placement of the aerobic threshold (AT) (VO2+13%; work rate [WR])+11%), an improvement of health perception (vitality+46%; social interaction+36%), an increase of activity level (+17%) and a tendency to less fatigue. No changes were observed for the MS-NI group and the control groups. Maximal aerobic capacity and lung function were not changed by either training or nontraining in all four groups. Overall compliance to the training program was quite low (65%), whereas incidence of symptom exacerbation by physical activity has been lower than expected (6%).     

Angiotensin protects cortical neurons from hypoxic-induced apoptosis via the angiotensin type 2 receptor

The effects of angiotensin on mouse cortical neuronal cultures exposed to chemical-induced hypoxia was investigated. Cultures exposed to 10 mM sodium azide for 5 min showed a 17% increase in apoptosis when assayed 24 h postinsult. The N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 blocked sodium azide-induced cell death suggesting that the NMDA receptor contributes to the mediated cell death. Pretreatment of cultured neurons with angiotensin decreased sodium azide-induced apoptosis by 94%. When the AT(1) receptor was blocked by its receptor antagonist, losartan, angiotensin activation of the AT(2) receptor completely inhibited sodium azide-induced apoptosis. Pretreatment of neurons with the AT(2) receptor antagonist PD123319 resulted in angiotensin reducing sodium azide-induced apoptosis by 48%. These results demonstrate that angiotensin can significantly attenuate sodium azide-induced apoptosis primarily through activation of the AT(2) receptor and suggests that angiotensin may have a protective role in neurons undergoing ischemic injury.     

Effects of angiotensin II on visual evoked potentials in the superior colliculus of juvenile rats

There are age-related changes in the relative expression of the AT(1)and AT(2)receptors of angiotensin II (Ang II) in brain regions such as the superior colliculus, a midbrain visual structure where both receptor subtypes are found. We investigated the effects of Ang II on gross visual activity in the colliculus of anesthetized rats aged between 15 and 35 post-natal days. Microinjection of Ang II in the superficial layers yielded a strong reduction in the amplitude of visual evoked potentials in a dose-related manner. Injection of the peptide in more ventral collicular layers did not modify the potential confirming the discrete localization of the angiotensinergic receptors in the superficial layers. Preliminary data indicated that the co-injection of Ang II with Losartan or PD 123319 yielded a partial blockade of Ang II suppressive effects, indicating that both AT(1)and AT(2)receptors are likely to be involved in mediating these responses. Overall, this study shows that the inhibitory nature of Ang II action is similar in juvenile and adult animals (Merabet et al. 1994 and Merabet et al. 1997) Copyright 2000 Harcourt Publishers Ltd.     

The expression of human brain vascular smooth muscle cell AT receptor after the UL83 gene of HCMV inhibition by small interfering RNAs

Abstract

Objectives: To explore the role of angiotensin receptors (AT1 and AT2) and human cytomegalovirus (HCMV) infection in arthrosclerosis, and to observe effect of HCMV UL83 gene on angiotensin receptor AT receptors.

Methods: Cell model of RNA interference (RNAi) HCMV UL83 gene was set up by transfection RNAi into HCMV infected human cerebral artery vascular smooth muscle cells cultured in vitro by application of transfection technique and adoption of RT-PCR, etc. Effect of HCMV on gene expression of AT1 and AT2 receptors in human cerebral artery vascular smooth muscle cells cultured in vitro under the action of Ganciclovir (GCV), AT1 and AT2 receptor antagonists by immunofluorescence technique, RT-PCR and protein electrophoresis.

Results: The duplication and protein expression of HCMV UL83 gene mRNA was successfully interfered using RNAi technique. After silencing HCMV UL83 gene, gene expression of AT1 receptor was down-regulated while that of AT2 was up-regulated. AT1 and AT2 receptor antagonists and GCV, under certain condition, could inhibit the activation on AT1 and AT2 receptor in human cerebral artery vascular smooth muscle cells induced by HCMV infection.

Conclusion: Cell model of HCMV infected UL83 silence gene was successfully set up in human cerebral artery vascular smooth muscle cells cultured in vitro. HCMV UL83 might play a role by activating AT1 receptor and inhibiting expression of AT2 receptor.     

Normalization of endothelial and inducible nitric oxide synthase expression in brain microvessels of spontaneously hypertensive rats by angiotensin II AT1 receptor inhibition.

Inhibition of angiotensin II AT1 receptors protects against stroke, reducing the cerebral blood flow decrease in the periphery of the ischemic lesion. To clarify the mechanism, spontaneously hypertensive rats (SHR) and normotensive control Wistar Kyoto (WKY) rats were pretreated with the AT1 receptor antagonist candesartan (0.3 mg. kg.(-1) d(-1)) for 28 days, a treatment identical to that which protected SHR from brain ischemia, and the authors studied middle cerebral artery (MCA) and common carotid morphology, endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) messenger RNA (mRNA), and protein expression in cerebral microvessels, principal arteries of the Willis polygon, and common carotid artery. The MCA and common carotid artery of SHR exhibited inward eutrophic remodeling, with decreased lumen diameter and increased media thickness when compared with WKY rats. In addition, there was decreased eNOS and increased iNOS protein and mRNA in common carotid artery, circle of Willis, and brain microvessels of SHR when compared with WKY rats. Both remodeling and alterations in eNOS and iNOS expression in SHR were completely reversed by long-term AT1 receptor inhibition. The hemodynamic, morphologic, and biochemical alterations in hypertension associated with increased vulnerability to brain ischemia are fully reversed by AT1 receptor blockade, indicating that AT1 receptor activation is crucial for the maintenance of the pathologic alterations in cerebrovascular circulation during hypertension, and that their blockade may be of therapeutic advantage.     

Candesartan reduces superoxide production after global cerebral ischemia

Excessive superoxide production after cerebral ischemia is known to mediate neuronal injury. Angiotensin II type 1 receptor activation results in production of superoxide, but whether angiotensin II type 1 receptor blockade prevents production of superoxide and subsequent neuronal injury after ischemia remains unclear. Normotensive rats received the angiotensin II type 1 receptor blocker, candesartan or only vehicle before induction of global cerebral ischemia. Approximately 30% of the hippocampal CA1 neurons survived in candesartan-treated animals, whereas only 2% of neurons survived in vehicle-treated animals. Superoxide production was significantly less in these vulnerable neurons in candesartan-treated animals than in vehicle-treated animals. Angiotensin II type 1 receptor may have an essential role in superoxide production and subsequent injury in vulnerable neurons after global cerebral ischemia.     

Development of AT(1) and AT(2) receptors in the ovine fetal brain

This study determined the development of AT(1) and AT(2) receptors in the ovine fetal brain from preterm to term by utilizing Western blot for the receptor expression at the protein level, RT-PCR for the receptor mRNA, and immunostaining for the specific receptor immunoreactivity. The results demonstrated that AT(1) and AT(2) receptors developed in an increasing pattern from preterm to term gestational periods in the fetal sheep brain. Both AT(1) and AT(2) receptors have appeared in the major structures in the angiotensin-related central cardiovascular and body fluid controlling pathways at the 0.7 of the gestational age. Importantly, AT(1) receptors have been discovered in the supraoptic nuclei in the fetal hypothalamus, and in the lateral parabrachial nuclei and the ventrolateral medulla in the fetal hindbrain. This provides evidence of the anatomical existence of the angiotensin receptors in the brain areas that are critical for cardiovascular and fluid regulatory functions in utero. In addition, although the results demonstrated the predominance of AT(2) receptors in several regions such as the cerebellum in the ovine fetal brain, dominant occupation of AT(1) receptors in the hypothalamus have appeared early in the life of sheep animals before birth. Together, the data support the hypothesis that the central angiotensin receptors are well developed and established in the last third trimester of gestation. The brain receptors provide a pharmacological basis for the action of angiotensin in the maintenance of in utero fetal physiological functions, including cardiovascular and body fluid balance.     

AT1 receptor inhibition prevents astrocyte degeneration and restores vascular growth in oxygen-induced retinopathy

We investigated the effect of receptor blockade induced by an angiotensin II type-1 receptor antagonist (AT(1)-RB) on glial and vascular changes in oxygen-induced retinopathy (OIR), a model of retinopathy of prematurity (ROP). OIR was induced in Sprague-Dawley rats by exposure to 80% oxygen from postnatal (P) days 0-11, followed by 7 days in room air. Control animals were in room air for the entire duration. One cohort of OIR and control pups received the AT(1)-RB valsartan (40 mg/kg/day intraperitoneal) from P11 to P18. The vascular response was examined immunocytochemically using retinal wholemounts and vertical sections labeled with endothelial (Isolectin-B4) and pericyte (NG2, desmin) markers. Glial cell changes were assessed by measuring cell numbers and immunoreactivity (S100beta, connexin-26, and glial fibrillary acidic protein). OIR resulted in extensive intravitreal neovascularization and under-development of the outer vascular plexus. Pericyte numbers were not significantly affected in OIR, although pericyte-endothelial (desmin-IB4) interactions were impaired. Peripheral astrocyte degeneration occurred between P11 and P13 with prominent Müller cell reactivity at P18. Valsartan imparted a protective effect on glia and blood vessels in OIR. At P18, valsartan-treated OIR retinae showed significantly greater astrocyte survival, improved revascularization of the retina, and reduced preretinal neovascularization and Müller cell reactivity. This study identifies a glio-vascular protective effect with AT(1)-RB in OIR.     

Anxiety-like behavior in mice lacking the angiotensin II type-2 receptor

The main biological role of angiotensin II type 2 receptor (AT2) has not been established. We made use of targeted disruption of the mouse AT2 gene to examine the role of the AT2 receptor in the central nervous system (CNS). AT2-deficient mice displayed anxiety-like behavior compared with wild-type mice. However, AT2-deficient mice showed no depressant-like activity and no change in hexobarbital-induced sleeping time as compared with findings in wild-type mice. Both noradrenergic and corticotropin-releasing factor (CRF) neuronal systems appear to be involved in this anxiety-like behavior. Diazepam, captopril (angiotensin I converting enzyme inhibitor), prazosin (alpha1 antagonist) reversed the anxiety-like behavior in these AT2-deficient mice, whereas yohimbine (alpha2 antagonist), phenylephrine (alpha1 agonist), clonidine (alpha2 agonist), isoproterenol (beta1/beta2 agonist), propranolol (beta1/beta2 antagonist) and alpha-helical CRF9-41 (CRF receptor antagonist) has no apparent effects on anxiety-like behavior in AT2-deficient mice. In addition, concentrations of plasma adrenocorticotropic hormone (ACTH) and corticosterone in AT2-deficient mice did not differ from these in wild-type mice, hence, there are probably no endocrine abnormalities involving the hypothalamic-pituitary-adrenal axis (HPA). The amygdala appears to play an important role in many of the responses to fear and anxiety. The number of [3H]prazosin but not [125I]CRF binding sites in the amygdala was significantly reduced in AT2-deficient mice. These findings indicate that the noradrenergic system is involved in mediating the anxiety-like behavior in AT2-deficient mice.