Signalling across the blood brain barrier by angiotensin II: novel implications for neurogenic hypertension

Angiotensin II (AngII) is a major culprit in essential hypertension. Based on a genetic rodent model of hypertension, we review here evidence that AngII may signal across the blood brain barrier to affect neuronal circuits within the nucleus tractus solitarii (NTS) of the brainstem, a pivotal region regulating both the baroreceptor reflex and set point control of arterial pressure. We have termed this form of signalling as vascular–neuronal signalling. We describe that the depressant action of AngII in NTS on the baroreceptor reflex is mediated via activation of endothelial nitric oxide synthase (eNOS) releasing NO that promotes release of the inhibitory transmitter—GABA. This could shunt the incoming excitatory baroreceptor afferent traffic impinging on NTS neurones. Chronic studies recording arterial pressure in conscious unrestrained rats using radio-telemetry have revealed that eNOS in NTS plays an endogenous physiological role in the homeostatic regulation of the gain of the cardiac baroreceptor reflex. However, in the spontaneously hypertensive rat, eNOS mRNA was higher (compared to normotensive rats), and its chronic blockade in NTS restored the abnormally depressed cardiac baroreceptor reflex to levels akin to normotensive rats, improved heart rate variability and lowered arterial pressure. Hence, it seems that excessive eNOS activity in NTS of the SHR contributes to the pathological state of this animal model’s cardiovascular autonomic nervous system. We speculate on why eNOS activity may be up regulated in the NTS of the SHR and propose that it is a consequence of high cerebral vascular resistance and inadequate blood perfusion of the brainstem.     

Altered cyclic AMP levels in specific cardiovascular brain centers of spontaneously hypertensive rats (SHR)

Summary The possible participation of cAMP in central regulation of arterial blood pressure was investigated in spontaneously hypertensive rats (SHR).Cyclic AMP concentrations of 36 microdissected individual brain areas — including primary and higher cardiovascular centers — were measured in adult SHR and compared with those of normotensive control rats of the same strain and age.In the adult SHR, elevated cAMP concentrations were found in brain areas which are in close connection with the central regulation of blood pressure: nucleus tractus solitarii, A1- and A2- catecholaminergic cell groups in the medulla oblongata, locus coeruleus, central grey matter (subnucleus medialis), and certain cortical areas (especially cingulate cortex and hippocampus).On the other hand, hypothalamic cell groups which have been also suggested to control blood pressure, such as paraventricular, anterior, ventromedial, dorsomedial and posterior hypothalamic nuclei, show lower concentrations of cAMP in the SHR than in normotensive controls.Cyclic AMP levels in 19 other brain areas of SHR which seem to be not involved in mechanisms of central blood pressure regulation practically do not differ from the values of normotensive rats.The results suggest that cAMP-dependent processe are involved in the regulatory mechanisms of central blood pressure control. The brain areas which show alterations of cAMP-levels are also distinguished by close neuronal connections to the baroreceptor reflex arc. It is supposed that these areas represent modulatory higher centers capable to affect baroreceptor reflex function.     

Hypotensive response in spontaneously hypertensive rats following microinjection of alpha-methylnoradrenaline in the caudal brain-stem.

Local microinjection of 1.25 nmol (-)-alpha-methylnoradrenaline in the A2-region of the nucleus tractus solitarii (NTS) caused a decrease of blood pressure and heart rate in both spontaneous hypertensive rats (SHR) and Wistar-Kyoto (W/K) rats. Although the maximal responses in both strains did not differ, the decrease in blood pressure lasted longer in the SHR. These results do not support the concept of a diminished sensitivity of catecholaminergic receptors in the NTS of SHR to alpha-methylnoradrenaline.     

Reflex inhibition of sympathetic activity during volume load in awake normotensive and spontaneously hypertensive rats

The reflex inhibition of the sympathetic activity in the splanchnic nerves was recorded upon volume expansion with blood in awake spontaneously hypertensive rats (SHR) and in normotensive Wistar-Kyoto rats (WKR) at an age of 16–20 weeks. At 10% blood volume expansion SHR showed a significantly greater nerve inhibition (43 %) in comparison with WKR (33 %). This augmented reflex response was not caused by the arterial baroreceptors, because the sensitivity of the arterial baroreceptor reflex arch, if anything, tended to be lower in SHR and the increase in arterial blood pressure upon volume load was also lower in SHR. It is suggested that the reason for this increased reflex inhibition in SHR is an augmented low pressure receptor response. The mechanism behind this is discussed. The most likely explanation is a decreased distensibility of the venous system, the systemic andlor the pulmonary veins.     

Dynamic exercise attenuates spontaneous baroreceptor reflex sensitivity in conscious rats

In humans, it has been reported that dynamic exercise attenuates spontaneous baroreceptor reflex sensitivity (sBRS), which is an index of the gain of the baroreceptor-cardiac reflex. We demonstrated previously that endogenously produced NO from endothelial nitric oxide synthase (eNOS) within the nucleus tractus solitarii (NTS), the central terminal site of baroreceptor afferents, depressed sBRS. In this study, we investigated whether eNOS activity within the NTS plays any role in down-modulating the sBRS during dynamic exercise. In conscious Wistar rats arterial pressure and heart rate (HR) were monitored continuously and chronically using radiotelemetry before and during wheel cage running at 6 m min(-1) for 10 min. sBRS was determined by a time-series method. During dynamic exercise systolic blood pressure (SBP) and HR were significantly increased (SBP: 138 +/- 2 vs. 125 +/- 2 mmHg, P < 0.001; HR: 447 +/- 6 vs. 362 +/- 8 beats min(-1), P < 0.001) while sBRS was significantly decreased (0.53 +/- 0.03 vs. 1.08 +/- 0.08 ms mmHg(-1), P < 0.001). In sino-aortic denervated rats the change in SBP in response to dynamic exercise was significantly larger than that in baroreceptor-intact rats (denervated: 21.6 +/- 2.5 mmHg; intact: 12.0 +/- 2.8 mmHg, P < 0.05). In contrast, denervation made no difference to the change in HR. Although disabling eNOS activity in the NTS by adenoviral-directed expression of a dominant negative mutant form of eNOS increased resting sBRS (1.48 +/- 0.20 vs. 1.09 +/- 0.15 ms mmHg(-1), P < 0.05), the absolute level reached during dynamic exercise was identical to control. These results demonstrate that during dynamic exercise (i) the sBRS decreases around the operating point of the baroreceptor-cardiac reflex function curve in normotensive rats, (ii) the baroreceptor reflex operates to limit the rise in arterial pressure, and (iii) the attenuation of sBRS is not mediated by changes in eNOS activity within the NTS.     

Substance P neurones in medullary baroreflex areas and baroreflex function of capsaicin-treated rats. Comparison with other primary afferent systems

Adult rats (age 13–17 weeks) treated 20 h after birth with capsaicin (50 mg/kg s.c.) showed a 50% loss of substance P-immunoreactive nerve terminals in the thoracic substantia gelatinosa as revealed by quantitative immunohistochemistry and radioimmunoassay. Other spinal substance P systems were unchanged. A similar decrease of immunoreactive substance P axons and nerve terminals was observed in medullary primary afferent systems, namely in IX and X rootlets, tractus spinalis nervi V, tractus solitarius and substantia gelatinosa nervi V. Other medullary substance P fibres, e.g. in the reticular formation, nucleus motorius nervi X and XII, inferior olive and area postrema, were not obviously changed. The nucleus tractus solitarii receiving, among others, IX and X primary afferents, e.g. from baroreceptors and chemoreceptors, showed a particular pattern of depletion: although the substance P axons penetrating from the tractus solitarius mainly into the intermediate and caudal nucleus tractus solitarii were markedly reduced in capsaicin-treated rats, the total number of substance P nerve terminals in the intermediate and caudal nucleus tractus solitarii was only moderately decreased and appeared normal in the cranial portion of the nucleus. The number of substance P-immunoreactive cells in the nucleus tractus solitarii was unchanged. Radioimmunoassay of substance P levels showed a 60% decrease in the microdissected substantia gelatinosa nervi V, but only insignificant decreases (by 20%) in microdissected divisions of the nucleus tractus solitarii and reticular formation.Thus, capsaicin destroyed a considerable part of primary spinal and medullary substance P afferents. In the entire nucleus tractus solitarii most of substance P is contained in a capsaicin-insensitive, probably intrinsic system. However, the intermediate and caudal divisions contain, in addition, primary afferent IX and X substance P terminals which were partly destroyed by capsaicin. In view of a recent proposal that substance P is the transmitter released from baroreceptor afferents, blood pressure and baroreflex function were measured in capsaicin-treated (50 and up to 600 mg/kg) rats; neither differed from controls. While, at first sight, this finding militates against a transmitter role of substance P in the baroreceptor reflex pathway, incomplete destruction of primary substance P afferents in the nucleus tractus solitarii by capsaicin and the possible development of compensatory mechanisms during growth of the animals have to be considered as alternative explanations. Furthermore, in the same rats, tests on the function of other primary afferents yielded, in part, incongruent results. The threshold for nociceptive chemical stimuli, as determined with capsaicin and formalin, was increased. On the other hand, threshold to nociceptive heat was unchanged in the tail-flick, but increased in the hot plate test.     

Activation of brainstem endorphinergic neurons causes cardiovascular depression and facilitates baroreflex bradycardia

The effects of electrical stimulation of the arcuate nucleus on blood pressure, heart rate and baroreflex sensitivity were studied in urethane-anesthetized Sprague-Dawley rats. Stimulation of the mid-anterior parts of the arcuate nucleus at 80 Hz, 0.8 ms and 50-200 microA caused a biphasic, depressor/pressor, response and moderate bradycardia. Intravenous administration of a vasopressin V1-receptor antagonist eliminated the pressor component and unmasked a pure depressor response. This depressor response could be inhibited by naltrexone, 2 mg/kg i.v., by an antiserum against beta-endorphin, 100 nl injected directly into the ipsilateral nucleus tractus solitarii, or by deafferentation of the dorsal vagal complex (nucleus tractus solitarii and dorsal vagal nucleus) by an ipsilateral, dorsolateral knife-cut of the medulla oblongata. Stimulation of the arcuate nucleus at currents of 20-40 microA did not influence basal blood pressure or heart rate but potentiated the reflex bradycardia induced by phenylephrine, and this effect was completely blocked by naltrexone. It is concluded that a beta-endorphin-containing pathway projecting from the arcuate nucleus to the ipsilateral dorsal vagal complex is involved in depressor cardiovascular regulation and in the facilitation of baroreflex bradycardia.     

Low-Intensity physical activity beneficially alters the ultrastructural renal morphology of spontaneously hypertensive rats

INTRODUCTION AND OBJECTIVE:

Kidney disorders can cause essential hypertension, which can subsequently cause renal disease. High blood pressure is also common among those with chronic kidney disease; moreover, it is a well-known risk factor for a more rapid progression to kidney failure. Because hypertension and kidney function are closely linked, the present study aimed to observe the beneficial effects of low-intensity physical activity on structural and ultrastructural renal morphology and blood pressure in normotensive and spontaneously hypertensive rats.

METHOD:

Male Wistar-Kyoto rats and spontaneously hypertensive rats were randomly allocated into four groups: sedentary or exercised Wistar-Kyoto and sedentary or exercised spontaneously hypertensive rats. The exercise lasted 20 weeks and consisted of treadmill training for 1 hour/day, 5 days/week.

RESULTS:

The exercised, spontaneously hypertensive rats showed a significant blood pressure reduction of 26%. The body masses of the Wistar-Kyoto and spontaneously hypertensive strains were significantly different. There were improvements in some of the renal structures of the animals treated with physical activity: (i) the interdigitations of the proximal and distal convoluted tubules; (ii) the basal membrane of the proximal and distal convoluted tubules; and (iii) in the basal membrane, slit diaphragm and pedicels of the glomerular filtration barrier. The spontaneously hypertensive rats also showed a decreased expression of connexin-43.

CONCLUSION:

Physical exercise could be a therapeutic tool for improving kidney ultrastructure and, consequently, renal function in hypertensive individuals.     

Adenylate cyclase-cyclic AMP-phosphodiesterase system in microdissected brain areas of normotensive and spontaneously hypertensive rats

The topographical distribution of the adenylate-cyclase-cyclic adenosine monophosphate-phosphodiesterase system was investigated in specific brain areas of rats, and compared with spontaneously hypertensive rats. In normotensive animals, brain cyclic adenosine monophosphate levels were quite uniform (between 7.14 and 13.04 pmol/mg protein) with highest concentrations in catecholamine-containing cell groups. In contrast, the distribution of basal adenylate cyclase and phosphodiesterase activity is not uniform. The unstimulated adenylate cyclase activity was very low in the striatum and catecholamine-containing cell groups in the medulla oblongata (A1C1-cell groups) and very high in the central gray matter and the cerebellum, where the lowest phosphodiesterase activities were measured. In spontaneously hypertensive rats, altered cyclic adenosine monophosphate levels were found in 17 of 36 brain areas investigated in comparison to those of normotensive rats. Increased concentrations were found in regions which are known to participate in the central regulation of blood pressure (nucleus of the solitary tract, A1C1 catecholaminergic cell groups in the ventrolateral medulla oblongata, locus coeruleus) and in the periaqueductal central gray matter, the hippocampus and the cingulate cortex. Lower levels were measured only in hypothalamic nuclei, especially in the paraventricular and dorsomedial nucleus. No significant differences in basal adenylate cyclase activity were found in spontaneously hypertensive rats compared with Wistar-Kyoto control rats, while phosphodiesterase activity was generally higher in spontaneously hypertensive rats, most significantly in the medulla oblongata. Present data show that characterization of the adenylate cyclase-cyclic adenosine monophosphate-phosphodiesterase system helps to localize structural and/or functional differences between the spontaneously hypertensive rat and its normotensive control rat and indicate that more than one functional system is affected in spontaneous hypertension.     

Evidence for gamma-aminobutyric acid receptor-mediated modulation of the aortic baroreceptor reflex in the nucleus tractus solitarii of the rat

Microinjections of the gamma-aminobutyric acid (GABA) receptor antagonist bicuculline into the medial area of the nucleus tractus solitarii of the rat enhanced the depressor and bradycardiac responses to aortic nerve stimulation whereas the glycine receptor antagonist strychnine did not affect them. The GABA receptor agonist muscimol and the GABA uptake inhibitor nipecotic acid reduced the responses to aortic nerve stimulation. These results provide evidence suggesting GABA receptor-mediated modulation of the aortic baroreceptor reflex in the nucleus tractus solitarii of the rat.     

Comparative effects of hydralazine and captopril on the cardiovascular changes in spontaneously hypertensive rats.

Vascular changes that develop during the course of blood pressure rise in spontaneously hypertensive rats (SHRs) can be modified by antihypertensive therapy. It is not known, however, whether there is selectivity in the structural response to specific antihypertensive drugs. This issue was examined by comparing the effects of a direct vasodilator (hydralazine) and a converting enzyme inhibitor (captopril) on morphologic aspects of the cardiovascular system. Male SHRs, 21 weeks of age, were given either hydralazine (HCl plus hydrochlorothiazide, captopril plus hydrochlorothiazide, or hydrochlorothiazide alone. Untreated age-matched SHRs and Wistar-Kyoto normotensive rats (WKYs) were used as controls. Animals were sacrificed at 27-28 weeks of age. Both hydralazine and captopril lowered significantly the blood pressure of SHRs, whereas hydrochlorothiazide alone was ineffective. The heart/body weight ratios were dramatically reduced in captopril-treated SHRs to below the level of WKYs; hydralazine induced only a very modest (5%) reduction, whereas the diuretic alone was ineffective. The morphology of the aortic intima improved dramatically in response to captopril and hydralazine and, to a lesser extent, hydrochlorothiazide alone. This effect becomes apparent within 6 weeks of treatment, and the only evidence of preexisting disease is the persistence of collagen in the subendothelium. Captopril and hydralazine, but not hydrochlorothiazide alone, reduce the thickness of the aortic media below that of normotensive controls. In addition, captopril and hydralazine improve the structure of small intrarenal vessels. There was a strong correlation between the relative effectiveness of the three pharmacologic agents in lowering blood pressure and in improving the changes of intrarenal vessels. These results highlight the capacity of antihypertensive therapy to arrest or reverse the structural sequelae of hypertension. In addition, they underscore the heterogeneity in the response of different components of the cardiovascular system, which, in part, reflects selectivity in the action of antihypertensive agents.     

The hippocampus in spontaneously hypertensive rats: a quantitative microanatomical study

The influence of hypertension on the morphology of hippocampus was assessed in spontaneously hypertensive rats of two, four and six months and in age-matched normotensive Wistar-Kyoto rats. Values of systolic pressure were slightly increased in two-month-old spontaneously hypertensive rats in comparison with age-matched Wistar-Kyoto rats and augmented progressively with age in spontaneously hypertensive rats. No microanatomical changes were observed in the hippocampus of spontaneously hypertensive rats of two months in comparison with age-matched Wistar-Kyoto rats, whereas a decrease of white matter volume was observed in the CA(1) subfield and in the dentate gyrus of four-month-old spontaneously hypertensive rats. In the hippocampus of six-month-old spontaneously hypertensive rats a reduction of grey matter volume both in the CA(1) subfield and in the dentate gyrus, a loss of neurons affecting to a greater extent the CA(1) subfield and an increase of glial fibrillary acid protein-immunoreactive astrocytes was found. The occurrence of apoptosis and/or necrosis identified using the terminal deoxyribonucleotidyl transferase-mediated biotin-16-dUTP nick end labelling technique was also observed in the CA(1) subfield and to a lesser extent in the dentate gyrus. The only change noticeable in the CA(3) subfield of six-month-old spontaneously hypertensive rats was a slight increase in the number of glial fibrillary acid protein-immunoreactive astrocytes.These findings indicate the occurrence of neuronal loss and of astrocyte changes in the hippocampus of spontaneously hypertensive rats of six months, being the CA(1) subfield the area most affected. The relevance of these neurodegenerative changes in hypertension and the possible occurrence of apoptosis and/or necrosis as expression of hypertensive brain damage is discussed.     

Effects of antihypertensive agents on blood pressure and the progress of renal failure in partially nephrectomized spontaneously hypertensive rats

The purpose of this study was to investigate the effects of antihypertensive agents on blood pressure and the development of glomerular changes in salt-loaded, 5/6 nephrectomized spontaneously hypertensive rats (SHR). Thirty-two spontaneously hypertensive rats with 5/6 nephrectomy were divided into 4 groups: a control group (N = 8), and group treated with 10 mg/kg/day trichlormethiazide. (N = 8), 30 mg/kg/day captopril (N = 8), and 200 mg/kg/day nicardipine (N = 8). Each of these antihypertensive drugs was added to the drinking water for 10 weeks and the rats were given the drugs and a high-salt diet (5% NaCl). During the experiment, body weight and systolic blood pressure were measured every 2 weeks. At the end of the study, blood was collected for determination of serum creatinine, blood urea nitrogen, serum protein, serum sodium and potassium, and plasma renin activity and aldosterone concentration. Also renal tissues were obtained for light and electron microscopic examination at the end of the study. Systolic blood pressure in 5/6 nephrectomized SHR loaded with a high salt was significantly reduced by administration of trichlormethiazide (155 +/- 12 versus 204 +/- 12 mm Hg), but not by administration of either captopril or nicardipine. However, levels of serum creatinine were not significantly elevated in rats treated with captopril and nicardipine (control: 0.93 +/- 0.11 mg/dl, captopril: 0.62 +/- 0.01 mg/dl, nicardipine: 0.55 +/- 0.05 mg/dl). In contrast to changes in blood pressure, marked glomerular sclerosis with hyalinosis, which was found in the control group was not ameliorated by treatment with trichlormethiazide. However, these changes were not observed in rats treated with either captopril or nicardipine in spite of the absence of a prominent fall in blood pressure. These data suggest that captopril and nicardipine ameliorated glomerular injury regardless of the level of systolic blood pressure through the direct and/or indirect actions on the glomerulus.     

Expression of c-fos protein in the medulla oblongata of conscious rabbits in response to baroreceptor activation.

Neuronal expression of c-fos protein (Fos) in the medulla in response to baroreceptor activation was studied in conscious rabbits. Raising arterial pressure resulted in a marked increase, compared to control animals, in Fos immunoreactivity in the nucleus tractus solitarius, area postrema and ventrolateral medulla (VLM). Fos-immunoreactive neurons in the VLM extended from the level just rostral to the obex to 3 mm more caudal. Only a small proportion of these neurons showed tyrosine hydroxylase immunoreactivity. The results indicate that baroreceptor activation induces Fos expression in circumscribed medullary regions which have previously been shown to receive excitatory baroreceptor inputs.     

Effects of Aqueous Extract of Lycopersicum esculentum L. var. “Camone” Tomato on Blood Pressure,Behavior and Brain Susceptibility to Oxidative Stress in Spontaneously Hypertensive Rats

Behavioral disorders affect millions of people worldwide. Hypertension contributes to both the development and progression of brain damage and cognitive dysfunction and could represent the most powerful modifiable risk factor for cerebral vessel dysfunction and consequent behavioral impairment. Tomato contains antioxidants and bioactive molecules that might play an important role in the prevention of cardiovascular and brain diseases. The effects of the combined gel and serum from Lycopersicum esculentum L. var. “Camone” tomatoes and those of purified tomato glycoalkaloids (tomatine) and an antihypertensive drug (captopril) were investigated in male spontaneously hypertensive rats (SHRs) and compared with normotensive Wistar Kyoto (WKY) rats. Body weight, systolic blood pressure, behavioral parameters, as well as brain susceptibility to oxidative stress and brain cytokine contents, were assessed. Treating hypertensive rats with tomato gel/serum or captopril for four weeks caused a significant reduction in blood pressure, decreased locomotor activity and increased grooming behavior; the last two parameters were also significantly affected by tomatine treatment. Brain slices obtained from hypertensive rats treated with tomato gel/serum were more resistant to oxidative stress and contained lower levels of inflammatory cytokines than vehicle-treated ones. In contrast, tomatine treatment had no effect. In conclusion, the tomato-derived gel/serum can be considered a dietary supplement able to drive in vivo blood pressure towards healthier values and also control some central effects such as behavior and brain oxidative stress.     

Barosensory cells in the nucleus tractus solitarius receive convergent input from group III muscle afferents and central command

Some neural mechanism must prevent the full expression of the baroreceptor reflex during static exercise because arterial blood pressure increases even though the baroreceptors are functioning. Two likely candidates are central command and input from the thin fiber muscle afferents evoking the exercise pressor reflex. Recently, activation of the mesencephalic locomotor region, an anatomical locus for central command, was found to inhibit the discharge of nucleus tractus solitarius cells that were stimulated by arterial baroreceptors in decerebrated cats. In contrast, the effect of thin fiber muscle afferent input on the discharge of nucleus tractus solitarius cells stimulated by baroreceptors is not known. Consequently in decerebrated unanesthetized cats, we examined the responses of barosensory nucleus tractus solitarius cells to stimulation of thin fiber muscle afferents and to stimulation of the mesencephalic locomotor region, a maneuver which evoked fictive locomotion. We found that electrical stimulation of either the mesencephalic locomotor region or the gastrocnemius nerve at current intensities that recruited group III afferents inhibited the discharge of nucleus tractus solitarius cells receiving baroreceptor input. We also found that the inhibitory effects of both gastrocnemius nerve stimulation and mesencephalic locomotor region stimulation converged onto the same barosensory nucleus tractus solitarius cells. We conclude that the nucleus tractus solitarius is probably the site whereby input from both central command and thin fiber muscle afferents function to reset the baroreceptor reflex during exercise.