The role of a humoral sodium-potassium pump inhibitor in low-renin hypertension

We have recently concentrated our efforts on bioassay of plasma supernatant from animals with experimental low-renin hypertension (one-kidney, one-wrapped in dogs, and one-kidney, one-clip, and reduced renal mass in rats) for sodium-potassium pump inhibiting activity. We have observed changes compatible with inhibitory activity by using three different in vitro bioassays: 1) ouabain-sensitive 86Rb uptake by the normal rat tail artery, 2) short-circuit current in the toad bladder, and 3) membrane potential in the rat tail artery. We have also generated evidence suggesting that the humoral pump inhibitor(s) comes from or is influenced by the anteroventral third ventricle area of the brain and that it acts on the vascular smooth muscle cell at least in part by depolarizing the membrane. These findings are compatible with our 1976 hypothesis in which we proposed that in volume-expanded hypertension there is a circulating agent that suppresses cardiovascular membrane Na+,K+-ATPase, which results in reduced activity of the Na+-K+ pump and hence increased contractility of heart, arteries, and veins and that in blood vessels the increased contractility may be secondary to depolarization. We attempt to relate these findings to those in the literature on monovalent ion transport in blood cells of hypertensive subjects.     

Pharmacologic agents for the in vivo detection of vascular sodium transport defects in hypertension

Anatagonists to angiotensin, catecholamines, aldosterone, and vasopressin have long been used to help determine agonist roles in hypertension. We here call attention to a possible extension of this approach to detect, evaluate, and treat vascular sodium transport defects in hypertension. Two basic types of transport defects have been identified in the blood vessels of hypertensive animals, increased sodium permeability and decreased sodium pump activity. Intravenous injection of 6-iodo-amiloride, a sodium channel blocker and vasodilator, produces an immediate and sustained decrease in blood pressure in two genetic models of hypertension characterized by increased permeability of the vascular smooth muscle cell membrane to sodium (Okamoto spontaneously hypertensive rat, Dahl salt sensitive rat), whereas it produces only a transient fall in arterial pressure in two renal models of hypertension having normal sodium permeability in vascular smooth muscle cells (reduced renal mass-saline rat, one-kidney, one clip rat). Canrenone, a metabolic product of spironolactone which can compete with oubain for binding to Na+,K+-ATPase at the digitalis receptor site, decreases blood pressure in a low renin, volume expanded model of hypertension which has been shown to have depressed sodium pump activity in arteries and increased sodium pump inhibitor in plasma (reduced renal mass-saline rat) but has no effect on blood pressure in a genetic model of hypertension which has been shown to have increased sodium pump activity secondary to increased sodium permeability (spontaneously hypertensive rat). Thus, a sodium channel blocker and a competitor to ouabain binding can detect and determine the functional significance of sodium transport defects in the blood vessels of intact hypertensive animals. Studies in red and white blood cells suggest that similar defects may exist in the blood vessels of hypertensive humans. Thus, this approach, probing for vascular transport defects in the intact animal, may ultimately also be useful in the clinical setting.     

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.     

Effects of rat atrial extract on sodium transport and blood pressure in the rat

Atrial cardiocytes contain specific atrial granules ( SAGs ) which are the storage site of atrial natriuretic factor (ANF). The purpose of the present study was to determine whether ANF produces natriuresis by inhibiting Na+-K+ pump activity and whether this factor is similar to the humoral sodium transport inhibiting factor ( HSTIF ) previously demonstrated in acutely volume expanded animals and humans as well as in experimental and human essential hypertension. Our results indicate that, in contrast to the HSTIF , ANF does not inhibit membrane Na+,K+-ATPase, vascular smooth muscle cell Na+-K+ pump activity, or sodium transport in the toad bladder. Intravenous infusion of ANF in the bilaterally nephrectomized, hexamethonium-treated rat produces only a small transient pressor response, probably due to potentiation of endogenous norepinephrine. These findings strongly suggest that the ANF is not the same as the HSTIF detected on acute volume expansion and in some forms of hypertension. They also suggest that the diuretic and natriuretic effects of ANF are due to mechanism(s) other than blood pressure elevation and inhibition of Na+-K+ pump activity.     

Effect of a potassium-deficient diet on arterial blood pressure, plasma and tissue cations, and tissue norepinephrine in the hypertensive dog

Chronic potassium deficiency in one-kidney one-clip hypertensive dogs significantly reduces blood pressure and plasma potassium, with a simultaneous increase in plasma renin activity. Tissue potassium concentration was decreased and tissue sodium concentration was increased in striated muscle and adrenal glands, which may suggest that the sodium-potassium pump was inhibited. In myocardium the sodium concentration was higher but the potassium concentration was not significantly lower than in control hypertensive dogs on normal diets. Arterial cation concentrations in the potassium-deficient group were not significantly different from those in the control group. Tissue norepinephrine concentration was higher in arteries from potassium-deficient animals, significantly so in the mesenteric and femoral arteries. The conclusion is that potassium deficiency may decrease blood pressure in the one-kidney one-clip hypertensive dogs by impairing the release of norepinephrine.     

Atrial natriuretic factor inhibits the sympathetic nervous activity in one-kidney, one-clip hypertension in the rat

The one-kidney, one-clip model of rat hypertension was found to have an increased natriuresis following chronic infusion of atrial natriuretic factor (ANF). We have now found that this natriuretic effect of ANF is associated with a suppression of the initially elevated urinary excretion of norepinephrine and epinephrine and increase of the excretion of the main dopamine metabolite-dihydroxyphenylacetic acid as well as of the urinary dopamine to norepinephrine ratio. These data are compatible with the hypothesis that ANF suppresses the increased sympathetic activity in this model of hypertension and this action combined with opposite changes of dopamine may contribute to the natriuretic effect of ANF.     

The role of humoral agents in volume expanded hypertension.

It has long been known that increased salt intake or decreased salt excretion leads to elevated blood pressure and increased extracellular salt and volume. The blood pressure rises slowly and for this reason and others cannot be explained by the increased salt or volume per se. Rather it appears that pressure rises as a result of some indirect effect of the increased volume. Autoregulation has been implicated, but reasons are presented which make this an unlikely possibility. Recent evidence suggests that some humoral agent or agents, operating through the Na⁺?K⁺ pump in cardiovascular muscle, participate in the genesis of the volume expanded hypertensions.     

Central neural peptides and catecholamines in spontaneous and DOCA/salt hypertension

Hypothalamic and neurophypophyseal levels of catecholamines and peptides were measured in spontaneous and deoxycorticosterone (DOCA)/salt hypertension. Catecholamines, norepinephrine, epinephrine and dopamine were measured by electrochemical detection while the peptides, vasopressin, oxytocin, luteinizing hormone-releasing hormone (LHRH), the enkephalins and somatostatin (SRIF) were measured by radioimmunoassay. Blood pressure was significantly elevated in both groups as compared to their controls. Marked changes in central neural peptides were observed in the SHR, while no differences were seen in DOCA/salt hypertension. Hypothalamic vasopressin, oxytocin, LHRH and SRIF were significantly decreased. In the posterior pituitary, enkephalins were increased twofold in the SHR. With regard to catecholamines, there was no change in hypothalamic content. However, a dramatic decrease in neurohypophyseal dopamine was observed in SHR. Plasma levels of vasopressin were significantly elevated in both types of hypertension while oxytocin was increased only in the DOCA/salt model. These result show that (1) a wide spectrum of neuroendocrine changes are associated with genetic hypertension, (2) there are CNS differences between DOCA/salt and spontaneous hypertension, and (3) central aminergic changes may be involved in th neuroendocrine alterations seen in the SHR.     

Passive transfer of pressor hyperresponsiveness from renal-hypertensive to normotensive rats

The role of serum factors in the pathogenesis of pressor hyperresponsiveness in hypertension was investigated by the passive transfer of serum from donor rats with chronic one-kidney, one clip hypertension into syngeneic normotensive recipient rats (0.25 ml iv, bid) for 3 weeks. Rats injected twice daily with the serum of normotensive rats served as controls. In rats injected with the serum of hypertensive rats there was a gradual increase in pressor responses to norepinephrine and angiotensin II and, at the end of the study, increased water content of the aorta and sodium content of the myocardium. In volume-expanded renal hypertension unidentified serum factors contribute to pressor hyperresponsiveness and increased sodium content of cardiovascular tissue.     

Correlation of the fibrinogen level with blood pressure and plasma renin activity in rats with early Goldblatt hypertension

Goldblatt hypertension was induced in rats by constricting the renal artery on one side. In one group of animals the contralateral kidney remained untouched (two-kidney hypertension), while in the other it was removed (one-kidney hypertension). In the two-kidney hypertension group, renin activity was higher than in the control animals, the fibrinogen was normal both in arterial and venous blood while in one-kidney hypertension the PRA was normal, but the fibrinogen was increased. A close significant correlation could be demonstrated between blood pressure and fibrinogen.     

Effect of 1-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine inhibitor on the reduction of one-kidney, one clip hypertension after unclipping in the rat

The role of 1-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine in regulating blood pressure was studied in one-kidney, one clip hypertensive rats using 3-(N-n-octadecylcarbamoyloxy)-2-methoxypropyl-2-thiazolio ethylphosphate, which specifically inhibited the hypotensive activity of exogenously injected 1-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine. The blood pressure of rats with established hypertension produced by clipping one renal artery and contralateral nephrectomy normally decreases rapidly after unclipping the artery, but this rapid decrease was significantly inhibited by intravenous infusion of 3-(N-n-octadecylcarbamoyloxy)-2-methoxypropyl-2-thiazolio ethylphosphate. This shows that endogenous 1-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine participates in the rapid decrease of blood pressure after unclipping the kidney in one-kidney, one clip hypertensive rats.     

Reversal of chronic renal hypertension: role of salt and water excretion.

Rats with chronic one-kidney Goldlatt hypertension underwent an unclipping procedure with and without maintenance of body fluid volume through administration of iv salt solution. The blood pressure declined equally in the two groups. It is concluded that external loss of salt and water is not the mechanism for the reversal of this form of hypertension.     

Effects of acute and chronic administration of sotalol on the blood pressure and the sympathoadrenal activity of anesthetized deoxycorticosterone acetate-salt hypertensive rats

Using plasma catecholamine (CA) levels as an index of the sympathoadrenal activity, the effects of chronic and acute beta-blockade on the blood pressure and sympathetic activity were evaluated in deoxycorticosterone acetate (DOCA) - salt hypertensive (HT) rats. The acute administration of one beta-blocker (sotalol, 5 mg/kg) to intact of vagotomized anesthetized HT animals induced a significant decrease in plasma norepinephrine (NE) concentrations and mean arterial pressure (MAP). The amplitude of the decrease of the MAP or NE levels were linearly correlated with the basal NE levels, suggesting that sotalol reduced the blood pressure and sympathetic NE release more efficiently in rats with increased sympathetic activity. Similarly, chronic infusion of sotalol (1.5 mg X day-1 X rat-1) through an osmotic pump for 12 days in DOCA-salt HT rats significantly reduced NE and epinephrine (E) plasma levels compared with those observed in untreated DOCA-salt HT rats. Moreover, the chronic treatment with sotalol significantly reduced the plasma E elevation induced by bilateral carotid occlusion (CO) in vagotomized normotensive (NT) and HT rats. It therefore appears that acute administration of sotalol to HT rats causes a significant reduction in the sympathetic activity which is associated to a decrease in MAP. Although chronic sotalol treatment causes a significant reduction in the sympathoadrenal basal activity and in the adrenal reactivity, this treatment did not prevent the development of DOCA-salt hypertension.     

Vascular responses to sodium arachidonate in experimental hypertension

This study characterizes vascular responsiveness to sodium arachidonate (C 20:4) in four models of hypertension [deoxycorticosterone acetate (DOCA) hypertensive rats, two kidney-one clip (2K-1C) renal hypertensive rats, spontaneously hypertensive rats (SHR), and psychosocial hypertensive mice]. Isolated arterial strips (aorta, mesenteric artery, tail artery) were equilibrated under optimal resting tension in physiological salt solution for measurement of isometric force generation. Dose-response curves to arachidonate (10(-10) to 10(-4) g/ml) in arteries from DOCA and 2K-1C hypertensive rats were shifted to the left compared to those in arteries from control rats. In arteries from SHR and psychosocial hypertensive mice, the dose-response relationships were unchanged compared to normotensive values. Arteries from DOCA hypertensive and 2K-1C hypertensive rats developed greater maximal contractile responses to arachidonate than controls; maximal responses in arteries from SHR and psychosocial hypertensive mice were unchanged compared to normotensive values. Contractions to arachidonate were inhibited by indomethacin (0.5 and 5 micrograms/ml) and by aspirin (5 and 50 micrograms/ml). The fatty acid, oleate (C 18:1), had no effect on the contractile state of the arteries, whereas prostaglandin F2 alpha caused contraction. These results indicate altered responsiveness to exogenous arachidonate in arteries from DOCA and 2K-1C hypertensive rats, but not in arteries from SHR and psychosocial hypertensive mice.     

Angiotensin II generation in mesenteric arteries in rats: effects of nephrectomy, deoxycorticosterone and dexamethasone

Angiotensin II (ANG II) generation in the mesenteric arteries was studied in four groups of rats: deoxycorticosterone (DOCA)/salt treated, glucocorticoid treated, nephrectomized and control rats. Basal plasma renin activity (PRA) was undetectable in the nephrectomized group and suppressed in the DOCA/salt treated rats, but was increased in the rats treated with glucocorticoid. The Basal plasma ANG II concentration changed comparably with PRA in all four groups of rats. In the control rats, ANG II was released from the mesenteric arteries at a rate of 43.0 +/- 12.0 pg/h, and it was not decreased by nephrectomy. In DOCA/salt rats and glucocorticoid rats, ANG II release significantly decreased to 12.8 +/- 7.1 and 6.9 +/- 1.5 pg/h, respectively. Captopril treatment significantly reduced ANG II release from the mesenteric arteries in both controls and nephrectomized rats, but did not influence ANG II output in DOCA/salt rats or in glucocorticoid treated rats. In nephrectomized rats, captopril lowered blood pressure in association with a significant reduction in the mesenteric ANG II formation. These results indicate that the renal and vascular renin-angiotensin system (RAS) may be independently regulated, and in nephrectomized animals the vascular RAS contributes in part to the maintenance of blood pressure. The present results also suggest that volume expansion per se and/or pharmacological intervention by DOCA and glucocorticoid could modulate vascular ANG II generation.     

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.     

Enhanced Assymetrical Noradrenergic Transmission in the Olfactory Bulb of Deoxycorticosterone Acetate-Salt Hypertensive Rats

The ablation of olfactory bulb induces critical changes in dopamine, and monoamine oxidase activity in the brain stem. Growing evidence supports the participation of this telencephalic region in the regulation blood pressure and cardiovascular activity but little is known about its contribution to hypertension. We have previously reported that in the olfactory bulb of normotensive rats endothelins enhance noradrenergic activity by increasing tyrosine hydroxylase activity and norepinephrine release. In the present study we sought to establish the status of noradrenergic activity in the olfactory bulb of deoxycorticosterone acetate (DOCA)-salt hypertensive rats. Different steps in norepinephrine transmission including tyrosine hydroxylase activity, neuronal norepinephrine release and uptake were assessed in the left and right olfactory bulb of DOCA-salt hypertensive rats. Increased tyrosine hydroxylase activity, and decreased neuronal norepinephrine uptake were observed in the olfactory bulb of DOCA-salt hypertensive rats. Furthermore the expression of tyrosine hydroxylase and its phosphorylated forms were also augmented. Intriguingly, asymmetrical responses between the right and left olfactory bulb of normotensive and hypertensive rats were observed. Neuronal norepinephrine release was increased in the right but not in the left olfactory bulb of DOCA-salt hypertensive rats, whereas non asymmetrical differences were observed in normotensive animals. Present findings indicate that the olfactory bulb of hypertensive rats show an asymmetrical increase in norepinephrine activity. The observed changes in noradrenergic transmission may likely contribute to the onset and/or progression of hypertension in this animal model.     

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.     

Resistance of Fischer 344 rats to deoxycorticosterone hypertension

Implantation of one 40 mg pellet of DOCA causes hypertension in the majority of young female Sprague-Dawley rats within three weeks without removal of a kidney or adding salt to the diet. Similar identically-treated Fischer 344 rats remain normotensive. If one kidney is removed and 1% saline is given to drink, the hormone dosage causes hypertension in rats of both strains, although even here Fischer 344 rats develop the disorder more slowly and less severely. It is concluded that for rat strains resistant to mineralocorticoid hypertension, sensitization is necessary for its induction, whereas for susceptible strains it is not. Fischer 344 rats appear to have higher levels of resting serum renin activity than Sprague-Dawley rats, but the relationship that this bears to hypertension susceptibility is unknown.     

Evidence for a circulating endogenous Na+-K+ pump inhibitor in low-renin hypertension

It is now more than 10 years since we suggested that an endogenous Na+,K+-ATPase inhibitor might participate in the genesis of certain forms of ren hypertension. Although the question is not yet fully resolved, there has been much activity in the area. We here review that activity. In 1980 we reported that supernatant of boiled plasma from dogs with one-kidney, one wrapped hypertension reduces Na+-K+ pump activity when applied to an artery from another animal. Since then, we and a number of other investigators have described Na+-K+ pump inhibitory activity in the plasma of animals and humans with hypertension, particularly the low-renin varieties. The activity results from a heat-stable small molecule, but the chemical structure of the molecule is unknown. It appears to be released from the hypothalamus in response to pulmonary vascular distension and to act on blood vessels via electrogenic depolarization. Although it may be sufficient by itself to raise pressure, it may be most effective when superimposed on vascular smooth muscle cells that are abnormally permeable to Na+. Efforts to determine the chemical structure of the agent or agents should be intensified.