Lowering of blood pressure in chronic aortic coarctate hypertensive rats with anti-digoxin antiserum

A marked lowering of arterial blood pressure was observed in chronic aortic coarctate (CAC) hypertensive rats after intravenous administration of anti-digoxin antiserum. The hypotensive effect lasted for about 30 min after dosing. In contrast to the pronounced changes observed following treatment with anti-digoxin antiserum in CAC rats, only a transient pressor effect was observed in both water-and saline-drinking CAC rats following injection of normal goat serum. In addition, a transient depressor effect was observed in normotensive rats after injection of anti-digoxin antiserum. The results of this study provide additional evidence indicating that an endogenous digoxin-like substance may play an important role in the maintenance of chronic low-renin hypertension induced by aortic coarctation.     

Cardiovascular investigations of an endogenous digoxin-like factor

A circulating factor with digoxin immunoreactivity has been demonstrated. Elevated levels of this substance appear to be present after volume expansion and salt loading, and in some forms of hypertension. The potentially causative role for this factor in hypertension can be demonstrated by the normalization of blood pressure after antidigoxin antibody infusions in low-renin and sodium-dependent hypertension. The possibility that renal excretory defects may be the initiating event to elevate endogenous digoxin is suggested by studies with normotensive humans and monkeys with renal disease. In the latter case cardiovascular deficits were noted that were analogous to those detected in renal hypertensive monkeys with elevated endogenous digoxin. Considered together, these results suggest the existence of a natriuretic and hypertensive substance that plays a role in body fluid homeostasis and blood pressure regulation.     

Determination of bufalin-like immunoreactivity in serum of humans and rats by time-resolved fluoroimmunoassay for using a monoclonal antibody

The existence of a mammalian natriuretic substance or endogenous digitalis-like factor, which inhibits Na+,K+-ATPase and thereby regulates body fluid volume, has been speculated for a long time but has yet to be defined. We established in the present study a simple and highly sensitive procedure to measure bufalin, a constituent of toad venom preparation and a specific inhibitor of Na+,K+-ATPase by time-resolved fluoroimmunoassay (TR-FIA) and using a monoclonal antibody. The antibody was specific to bufalin and resembled bufadienolides but showed no cross-reactivity with digitoxin and ouabain. A bufalin-like immunoreactivity was detectable in serum of humans and rats by the proposed TR-FIA. The levels of bufalin-like immunoreactivity in serum of healthy volunteers were significantly correlated with their systolic blood pressure. Moreover, bufalin-like immunoreactivity in serum of Dahl-S rats increased in parallel with a period of high-salt diet. These results suggest that increased bufalin-like immunoreactivity may be associated with certain types of hypertension.     

Humoral factors and the sodium-potassium pump in volume expanded hypertension.

Bioassay studies in the old and recent literature suggest the presence of an unknown slowly acting pressor agent in the blood of animals and man with volume expanded (low renin) hypertension. Recent studies in our laboratories suggest that the sodium-potassium pump activity of blood vessels is suppressed in animals with one-kidney, one wrapped, one-kidney, one clip, and one-kidney, DOCA, salt hypertension. Similar reduction of Na⁺, K⁺-ATPase activity has been observed in the left ventricle of animals with one-kidney, one clip and one-kidney, DOCA salt hypertension. The changes do not appear to result from increased pressure since they have also been observed in veins and right ventricle. Acute volume expansion of the normal rat with saline suppresses pump activity in the tail artery and plasma from these animals suppresses pump activity when applied to a tail artery from another rat. Data in the literature indicate that the adrenergic nerve terminals are depleted of norepinephrine. Suppression of pump activity, with ouabain for example, is known to activate cardiovascular muscle and reduce norepinephrine uptake by nerve terminals. These observations suggest a role for a slowly acting ouabain-like humoral agent, which acts directly on cardiovascular muscle to increase contractility and on nerve endings to reduce reflex compensation, in the genesis of volume expanded hypertension.     

Endogenous ouabain-like substances in animal tissues

Methods of purification and properties of the digitalis-like substances from different animal-origin sources are considered. Possible mechanisms of the cardiac glycoside-effect on the cardiac muscle and endogenous digitalis-like factors' effect on the smooth muscles of the blood vessels are discussed. Criteria that may be used to characterize the digitalis-like activity in samples and extracts obtained from different tissues are presented. The role of these substances in some types of pathologies, especially in the development of hypertension, is considered.     

Circulating digitalis-like substance is increased in DOCA-salt hypertension

Blood pressure and digitalis-like substance were measured in the plasma of control, salt-treated, and DOCA-salt treated rats. Blood pressure in DOCA-salt treated rats was significantly higher than that of either control or salt-treated animals. Digitalis-like activity was measured by two methods, radioimmunoassay for digoxin, and a receptor binding assay employing a rat brain synaptosomal membrane fraction. Digoxin-like immunoreactivity in plasma was not detected in either control or salt-treated rats, but was detected in DOCA-salt treated rats. Receptor binding activity in salt-treated rats was slightly but significantly higher than that of control rats. In DOCA-salt treated rats, receptor binding activity was significantly higher than that of salt-treated rats. Partial purification of the digitalis-like substance in plasma was performed by gel filtration using Sephadex G-25. Two peaks containing digoxin-like immunoreactivity were observed. Receptor binding activity, as well as Na+-K+ ATPase inhibitory activity, was detected only in the second peak, in which approximately 70% of the digoxin-like immunoreactivity was eluted. These results indicate that a circulating digitalis-like substance is increased in DOCA-salt hypertension.     

Critical opening pressure and reactivity of tail vessels in conscious hypertensive rats.

The possible role of increased vascular reactivity in the mechanism of experimental hypertension was studied by measurements of the critical opening pressure (COP) of tail vessels in conscious rats. In hypertension induced by administration of desoxycorticosterone acetate (DOCA) and replacement of the drinking water by 1% NaCl solution (DOCA-NaCl hypertension), and in one-kidney Goldblatt renovascular hypertension, the raised level of blood pressure was associated with an increased COP of the tail vessels when measured both before and after ganglionic blockade. In rats treated with either DOCA alone or 1% NaCl alone there was no significant increase in systolic blood pressure (SBP) or COP relative to the corresponding controls. In all four experimental series intravenous infusion of angiotensin or norepinephrine in conscious ganglion-blocked rats produced dose-dependent increases in SBP and COP. In DOCA-NaCl hypertensive rats but not in renovascular hypertensives, nor in rats treated with DOCA alone or 1% NaCl alone, the increase in COP for a given increment in dose of angiotensin or norepinephrine was significantly greater than in the control rats. It is concluded that in DOCA-NaCl hypertension there is a true increase in the reactivity of the smooth muscle of the resistance vessels to angiotensin and norepinephrine. In renovascular hypertension this is not the case and other factors must therefore be involved in causing the increased blood pressure and COP.     

Enhanced depressor effect of muscimol in the DOCA/NaCl hypertensive rat: evidence for altered GABAergic activity in brain

To elucidate the role of the central gamma-aminobutyric acid (GABA) system in the maintenance of deoxycorticosterone (DOCA)NaCl hypertension, the responses of mean arterial pressure (MAP), plasma norepinephrine (NE), and epinephrine (EP) to intracerebroventricular (ICV) administration of muscimol, a GABA agonist, and the responses of MAP to bicuculline, a GABA antagonist, and to clonidine, an alpha 2-adrenoceptor agonist known to lower blood pressure by inhibiting sympathetic tone, were examined in conscious, unrestrained 4 week DOCA/NaCl hypertensive rats and age-matched uninephrectomized control rats. Muscimol (50-1000 ng/300 g, ICV) caused dose-dependent decreases in MAP which were greater in DOCA/NaCl rats than in controls. Basal plasma NE and EP were significantly higher in DOCA/NaCl rats than in controls. Muscimol (1000 ng/300 g, ICV) induced decreases in plasma EP which were greater in DOCA/NaCl rats than in controls without changing NE levels in either group. Bicuculline (3 micrograms/300 g, ICV) caused increases in MAP which were the same in both groups. The depressor response to clonidine (5 micrograms/300 g) was greater in DOCA/NaCl rats than in controls. These results suggest that the activity of the central GABAergic system is altered in the rat with established DOCA/NaCl hypertension and that the alteration in central GABAergic function may be related to the increased sympathoadrenal activity and the maintenance of hypertension in this model.     

雌激素、褪黑素、红细胞源性降压因子降压机制的研究进展

雌激素(estrogen)作为一种重要的类固醇激素,对心血管系统发挥着重要作用,雌激素被认为对心血管系统起着有益的保护作用,临床流行病学研究显示,女性在绝经前或围绝经期的高血压患病率显著低于同龄男性。女性绝经后雌激素缺乏是高血压发病率增加的主要原因之一。褪黑素(Melatonin,Mel)主要是由哺乳动物和人类的松果体产生的一种胺类激素,它的合成受光周期的制约,可使正常动物和人的血压降低,也可对抗病理性的血压升高。红细胞源降压因子(erythrocyte-derived depressing factor,EDDF)是从人的红细胞中提取出一种能够显著降低血压的物质,它不同于迄今已发现的体液中的降压物质,很可能是一种新的内源性血压调节物质。体内物质雌激素、褪黑素、红细胞源性降压因子均可起到降压作用,将可能成为治疗高血压的新靶点。     

Arterial blood pressure and high calcium diet in normal and mineralcorticoid (DOCA and sodium chloride hypertensive rats]

High calcium diet induces an hypertension lasting one week in normal rats. In mineralocorticoid treated rats (DOCA + NaCl), the same diet prevents for 10 weeks the increase of arterial blood pressure. Parathyroid activity (estimated by urinary cAMP) is decreased after the high calcium diet. These results confirm the role of the parathyroid glands in mineralocorticoid hypertension in the rat.     

Splanchnic sympathetic nerves in the development of mild DOCA-salt hypertension

We previously reported that mild deoxycorticosterone acetate (DOCA)-salt hypertension develops in the absence of generalized sympathoexcitation. However, sympathetic nervous system activity (SNA) is regionally heterogeneous, so we began to investigate the role of sympathetic nerves to specific regions. Our first study on that possibility revealed no contribution of renal nerves to hypertension development. The splanchnic sympathetic nerves are implicated in blood pressure (BP) regulation because splanchnic denervation effectively lowers BP in human hypertension. Here we tested the hypothesis that splanchnic SNA contributes to the development of mild DOCA-salt hypertension. Splanchnic denervation was achieved by celiac ganglionectomy (CGX) in one group of rats while another group underwent sham surgery (SHAM-GX). After DOCA treatment (50 mg/kg) in rats with both kidneys intact, CGX rats exhibited a significantly attenuated increase in BP compared with SHAM-GX rats (15.6 ± 2.2 vs. 25.6 ± 2.2 mmHg, day 28 after DOCA treatment). In other rats, whole body norepinephrine (NE) spillover, measured to determine if CGX attenuated hypertension development by reducing global SNA, was not found to be different between SHAM-GX and CGX rats. In a third group, nonhepatic splanchnic NE spillover was measured as an index of splanchnic SNA, but this was not different between SHAM (non-DOCA-treated) and DOCA rats during hypertension development. In a final group, CGX effectively abolished nonhepatic splanchnic NE spillover. These data suggest that an intact splanchnic innervation is necessary for mild DOCA-salt hypertension development but not increased splanchnic SNA or NE release. Increased splanchnic vascular reactivity to NE during DOCA-salt treatment is one possible explanation.     

Hypertension from chronic central sodium chloride in mice is mediated by the ouabain-binding site on the Na,K-ATPase α₂-isoform

A chronic increase in the concentration of sodium chloride in the cerebrospinal fluid (CSF) (↑CSF [NaCl]) appears to be critically important for the development of salt-dependent hypertension. In agreement with this concept, increasing CSF [NaCl] chronically by intracerebroventricular (icv) infusion of NaCl-rich artificial CSF (aCSF-HiNaCl) in rats produces hypertension by the same mechanisms (i.e., aldosterone-ouabain pathway in the brain) as that produced by dietary sodium in salt-sensitive strains. We first demonstrate here that icv aCSF-HiNaCl for 10 days also causes hypertension in wild-type (WT) mice. We then used both WT and gene-targeted mice to explore the mechanisms. In WT mice with a ouabain-sensitive Na,K-ATPase α(2)-isoform (α2(S/S)), mean arterial pressure rose by ~25 mmHg within 2 days of starting aCSF-HiNaCl (0.6 nmol Na/min) and remained elevated throughout the study. Ouabain (171 pmol/day icv) increased blood pressure to a similar extent. aCSF-HiNaCl or ouabain given at the same rates subcutaneously instead of intracerebroventricularly had no effect on blood pressure. The pressor response to icv aCSF-HiNaCl was abolished by an anti-ouabain antibody given intracerebroventricularly but not subcutaneously, indicating that it is mediated by an endogenous ouabain-like substance in the brain. We compared the effects of icv aCSF-HiNaCl or icv ouabain on blood pressure in α2(S/S) versus knockout/knockin mice with a ouabain-resistant endogenous α(2)-subunit (α2(R/R)). In α2(R/R), there was no pressor response to icv aCSF-HiNaCl in contrast to WT mice. The α2(R/R) genotype also lacked a pressor response to icv ouabain. These data demonstrate that chronic ↑CSF [NaCl] causes hypertension in mice and that the blood pressure response is mediated by the ouabain-like substance in the brain, specifically by its binding to the α(2)-isoform of the Na,K-ATPase.     

5α-Dihydro-11-deoxycorticosterone: Effect on blood pressure in the rat

The bioactivity of 5α-reduced sex steroids such as 5α-dihydrotestosterone has increased interest in an analagous role for 5α-reduced mineralocorticoids in hypertensive syndromes. In view of its relatively high mineralocorticoid receptor affinity despite relatively low electrolyte-altering effects, 5α-dihydro-11-deoxycorticosterone, or 5αDHDOC (2) was compared to 11-deoxycorticosterone acetate (DOCA) for blood pressure-altering ability by continuous subcutaneous infusion into uninephrectomized saline-drinking Sprague-Dawley male rats, at doses selected on the basis of relative mineralocorticoid receptor affinity. After three weeks of treatment, DOCA significantly raised blood pressure, body weight, heart and kidney weight, and produced a discernible increase in fluid intake; 5αDHDOC failed to affect any of these parameters commonly influenced by mineralocorticoids. We conclude that 1) the ability of 5αDHDOC to affect blood pressure was not predicted by its relatively high affinity for the mineralocorticoid receptor, and 2) these data do not support a role for 5αDHDOC in mineralocorticoid hypertension, although differences in protein binding and clearance could affect its blood pressure-altering activity.     

Endogenous digoxin-like factor: serum concentration and interrelations with the electrolyte picture in normal subjects

Endogenous Digitalis-Like Factor (DLF) is a putative hypothalamic Na+,K+-ATPase inhibitor that mediates natriuresis in response to intravascular volume expansion or sodium loading. The precise structure of this substance remains unknown; however, it cross-reacts with antibody to digoxin. Using a radioimmunoassay, we measured DLF concentrations in 26 normal subjects: mean value of this factor was 0.512 ng digoxin-equivalents/ml +/- 0.038 SEM; DLF correlated significantly with serum sodium levels (r = 0.59 - p less than 0.01) and daily urinary sodium excretion (r = 0.48 - p less than 0.05). Our results confirm that endogenous digitalis-like factor has a physiological role as regulator of natriuresis, in response to plasma sodium concentrations.     

Relationships among endogenous digitalis-like factors in essential hypertension

Elements of a hypothesis that relate endogenous digitalis-like factors to both natriuretic hormone and hypertension are briefly reviewed. The stimulus for secretion of these factors appears to involve a tendency toward a state of extracellular fluid volume expansion as a consequence of an inherited or an acquired defect in renal function. Several studies implicate the brain and, in particular, the hypothalamus in the control of the secretion. The digitalis-like factors are thought to act by partial inhibition of active sodium transport, thereby promoting increased intracellular levels of Na+ and Ca2+ in a variety of cell types. In the kidney, inhibition of sodium transport leads to a compensatory natriuresis to correct the tendency for volume overload. In smooth muscle, the inhibition of sodium transport will indirectly increase intracellular calcium levels. The increased availability of Ca2+ will elevate muscle tone and increase peripheral vascular resistance. Also presented are criteria that may be used to characterize digitalis-like activity in samples and extracts obtained from purification procedures. Finally, we review our measurements of the 6-h integrated plasma levels of digitalis-like factors and other hormones for normotensive subjects and patients with essential hypertension. The data indicate the presence of two classes of digitalis-like factors with potentially different roles in electrolyte metabolism and hypertension.     

Is the endogenous digitalis-like factor the link between hypertension and metabolic disorders as diabetes mellitus, obesity and acromegaly?

Endogenous factors cross-reacting with antidigoxin antibodies have been found in several tissues and body fluids of animals and humans, using commercially available digoxin radioimmunoassay or enzyme immunoassay methods. The chemical characteristics of these endogenous factors are, at present, unknown, although it has been suggested that they could be substances with low molecular weight. Experimental studies and theoretical considerations indicate that endogenous digitalis-like factors (DDLFs), in addition to the ability to react with antibodies, might also bind to the specific cellular receptor of the cardiac glycosides and thus inhibit the membrane Na+/K(+)-ATPase (sodium pump). Therefore, EDLF can be an endogenous modulator of the membrane sodium-potassium pump and several authors have suggested that EDLF could play a role in the regulation of fluids and electrolytes, muscular tone of myocardial and also in the pathogenesis of arterial hypertension. In this review, the authors discuss the hypothesis that, in metabolic diseases such as diabetes mellitus, obesity and acromegaly, the sodium retention and volume expansion, possibly due to exaggerated sodium intake, and/or exogenously induced peripheral hyperinsulinemia and high levels of growth hormone, could trigger a sustained release of EDLF, which in turn increases the blood pressure.     

Effect of prazosin and yohimbine on systolic blood pressure and on renal norepinephrine content in DOCA-salt rats

We studied the effect of alpha-1 and alpha-2 blockers (prazosin and yohimbine) on systolic blood pressure (SBP) and on renal norepinephrine (NE) content in Sprague-Dawley normotensive and DOCA-salt rats. The administration of desoxycorticosterone acetate (DOCA) to these rats for 6 weeks increased their SBP from 137 to 183 mmHg (p less than .001). This increase was prevented by simultaneous administration of prazosin (p less than .001), yohimbine (p less than .01), or prazosin + yohimbine (p less than .001). DOCA rats on saline and on yohimbine had lower renal NE content (p less than .05 and p less than .001, respectively) than normotensive rats. Renal NE content of DOCA rats on yohimbine decreased with respect to those treated with prazosin (p less than .001) or prazosin + yohimbine (p less than .05). Besides, renal NE content of DOCA rats on prazosin increased when compared to control DOCA rats (p less than .05). However, these drugs showed no effect on SBP and on renal NE content in normotensive rats. These findings further confirm that the alpha adrenoceptor blockade can prevent the hypertension of DOCA-salt rats in such a way that their blood pressure stabilizes at similar levels to those observed in normotensive treated animals.     

Effects of irbesartan and bosentan on the blood pressure and adrenal zona glomerulosa function in heterozygous transgenic TGR[mREN2]27 rats

The role of angiotensin-II (Ang-II) and endothelin-1 (ET-1) in the development of hypertension and zona glomerulosa (ZG) hyperfunction in the transgenic rat strain TGR[mREN2]27 (TGR) has been investigated. Male heterozygous TGR were given per os for 4 weeks the Ang-II ATI receptor antagonist irbesartan (50 mg/kg x day) or the mixed ETA/ETB receptor antagonist bosentan (100 mg/kg x day). A group of TGR received a placebo gavage. Irbesartan lowered blood pressure (BP), while bosentan was ineffective. Conversely, both antagonists decreased plasma aldosterone concentration, the volume of ZG and its parenchymal cells, and in vitro aldosterone secretion by capsule-ZG preparations. Collectively, our results allow us to conclude that (i) only Ang-II is involved in the genesis of hypertension in TGR, while both endogenous Ang-II and ET-1 play a role in the genesis of ZG hyperfunction; and (ii) hyperaldosteronism does not contribute significantly to the development of hypertension in TGR.     

Abnormal regulation of the sympathoadrenal system in deoxycorticosterone acetate-salt hypertensive rats

With the use of circulating norepinephrine (NE) and epinephrine (E) levels, the sympathoadrenal activity as well as its local modulation by adrenoceptors were studied in normotensive (NT) and DOCA-salt hypertensive (HT) rats. In anesthetized hypertensive rats, plasma NE levels were higher, whereas in conscious animals both NE and E levels were found to be increased, suggesting an increased basal sympathoadrenal tone in these animals. The finding of a close correlation between blood pressure levels and NE levels suggests that the elevation of blood pressure may be linked to sympathetic system activity in this experimental model of hypertension. The reactivity of the sympathoadrenal system was also found to be increased in DOCA HT rats. Following a bilateral carotid occlusion of 1 min, which specifically activates the adrenal medulla, the elevation of E levels was found to be potentiated in intact or vagotomized HT rats. Moreover, in response to prolonged or acute hypotension in anesthetized and conscious animals, the elevation in plasma NE and E levels was found to be markedly potentiated in DOCA HT rats. The local modulating adrenoceptor-mediated mechanisms of the sympathoadrenal system appeared to be altered in this model of hypertension. Although it was possible to demonstrate that the E response to carotid occlusion can be greatly potentiated by administration of an alpha2-antagonist (yohimbine) and completely abolished by an alpha2-agonist (clonidine) in NT rats, the E response was found to be unaffected by the same treatments in HT rats, suggesting a reduced sensitivity in the alpha2-mediated inhibitory modulation of the adrenal medulla. Moreover, the acute treatment with a beta-blocker (sotalol) lowered circulating NE levels and blood pressure only in HT rats, suggesting the possibility of a more sensitive beta-receptor-mediated presynaptic facilitatory mechanism on sympathetic fibers of these animals. Finally, it was observed that the functional balance which exists between the activities of sympathetic fibers and the adrenal medulla in normotensive animals appears to be impaired in DOCA HT rats. In conclusion, the present studies suggest that the increased sympathoadrenal tone and reactivity may be due, in part, to a variety of dysfunctions in local adrenoceptor modulatory mechanisms of the sympathoadrenal system in DOCA hypertensive rats.     

Antihypertensive effects of fluoxetine and L-5-hydroxytryptophan in rats.

The combination of fluoxetine (10 mg/kg) and L-5-hydroxytryptophan (5-HTP) (10 mg/kg) significantly lowered blood pressure in spontaneously hypertensive rats and in rats made hypertensive by treatment with deoxycorticosterone (DOCA) and saline. Fluoxetine alone also had a significant effect on blood pressure in DOCA hypertensive rats, but not as great an effect as the combination. Since fluoxetine is an inhibitor of serotonin reuptake and 5-HTP is the serotonin precursor, the antihypertensive effect of this drug combination strengthens previous evidence that serotonin neurons have a role in the central regulation of blood pressure.