Search

It has been proposed that plasma cholesterol, a major risk factor for atherosclerosis, may modify cellular Ca2+. In particular, higher membrane cholesterol contents induce higher Ca2+ influx and decrease the activity of the Ca2+ pumps. Cellular Ca2+ may also control the number of accessible low density lipoprotein receptors. We investigated the question of whether plasma cholesterol influences cellular Ca2+ metabolism, by analysing the cytosolic free Ca2+ concentration [( Ca2+]i) in unstimulated platelets from 61 untreated hypertensive patients [systolic and diastolic arterial pressure: 157 +/- 3/95 +/- 2 mmHg (mean +/- s.e.m.), respectively, age 41.8 +/- 1.7 years, body mass index 24.7 +/- 0.6 kg/m2]. The subjects' plasma total cholesterol (5.5 +/- 0.5 mmol/l) and platelet [Ca2+]i concentration (228 +/- 7 nmol/l) were positively correlated (r = 0.375, P less than 0.003). This correlation persisted at constant age, arterial pressure or body mass index. Platelet [Ca2+]i tended to increase with plasma low-density lipoprotein concentration (n = 21, P = 0.01), and to decrease with the ratio of high-density lipoprotein cholesterol to total cholesterol (n = 21, P = 0.08). The observation that in normocholesterolaemic hypertensive patients [Ca2+]i concentration in unstimulated platelets was correlated with plasma cholesterol suggests that cell activation may be modulated by membrane fluidity or that cholesterol metabolism is influenced by cell Ca2+.

The digitalis-like activities of plasma extracts from 108 patients and normal subjects were measured by their ability to compete with ouabain for binding to the digitalis sites of the Na+-pump. High levels were found in 18 of 54 untreated patients with moderate hypertension, 10 of 14 patients with end-stage renal failure and six patients with active acromegaly. These levels returned to control values after dialysis in the patients with renal insufficiency and high levels of the inhibitor, and after successful surgery and cobalt therapy in seven acromegalic patients. An increase in circulating Na+, K+-ATPase inhibitor was also found in rats after chronic sodium loading. These results indicate that levels of the circulating compound with digitalis-like properties do not result from high blood pressure but, rather, are related to blood volume and Na+ balance.

It has been postulated that one or more plasma digitalis-like compounds may play an important role in body fluid regulation and in essential hypertension, although very little is known about their possible role in general haemodynamics. We therefore measured plasma inhibition of human kidney Na+,K+-ATPase and plasma cross-reactivity with digoxin antibodies in 11 normotensive cardiopathic subjects admitted to our clinic for heart catheterization. Possible correlations with haemodynamic parameters were studied. Plasma digoxin-like activity correlated directly with left atrial pressure and with pulmonary circulation data. The ability of the plasma to inhibit Na+,K+-ATPase showed an inverse correlation with cardiac output and cardiac index. No correlations were found with any of the other parameters measured, notably systemic resistance, blood pressure and natriuresis. These findings suggest the presence of more than one substance sharing chemical properties with digitalis: (1) a substance cross-reacting with digoxin antibodies and dependent on pulmonary vascular congestion; and (2) a substance capable of inhibiting the Na+-K+ pump and present in large amounts in heart diseases with a reduced cardiac index.

Active transport of Na+,K+ and Ca2+ was compared in heart plasma membranes from 3-week-old spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). ATP-dependent Ca2+ accumulation, which reflects Ca2+, Mg2+-ATPase activity, was higher in SHR than in WKY membranes. At a free calcium concentration of 4 X 10(-7)M, the addition of 2 X 10(-7)M calmodulin enhanced the active Ca2+-transport more in WKY than in SHR vesicles. Na+- and K+-dependent ATPase activity was two fold higher in SHR than in WKY. From ouabain binding studies this seemed to be due to an increased density of enzyme units. Physiological concentrations of calmodulin and calcium ions reduced Na+,K+-ATPase activity in the two strains but more in SHR than in WKY. This study demonstrates that active Na+ and Ca2+ transport is enhanced in young SHR; the Ca2+-calmodulin complex may regulate Na+,K+-ATPase activity and sensitivity of Na+,K+-ATPase and Ca2+,Mg2+-ATPase activities to Ca2+-calmodulin differs between SHR and WKY.

The presence of circulating Na+ pump inhibitors was investigated in hypertensive subjects using inhibition of ouabain binding to the pump and of Na+, K+-ATPase activity as tests. Plasma extracts from nearly half the normotensive subjects who were offspring of hypertensive parents as well as the essential hypertensive subjects were potent inhibitors. Three fractions, extracted from plasma, exhibited ouabain-like properties concerning competition for binding, inhibition of the Na+, K+-ATPase and of Na+-dependent serotonin uptake by platelets. When injected intracerebroventricularly, one of these fractions also induces a rise in blood pressure, as does ouabain. These results demonstrate the presence in some plasma of digitalis-like substances.

Cytosolic free Ca2+ ([Ca2+]i) concentrations were measured in platelets from hypertensive and normotensive man and rat with the fluorescent indicator Quin-2/AM, taking into account the signal of the free chelator. In the absence of added external Ca2+, no difference in [Ca2+]i was observed between platelets of hypertensive patients and those of normotensive subjects or between platelets of spontaneously hypertensive rats (SHR; Okamoto-Aoki strain) and those of normotensive Wistar-Kyoto (WKY) rats. In the presence of 0.5-1 mmol/l external Ca2+, [Ca2+]i was higher in patients with essential hypertension than in their normotensive controls (250 +/- 14 versus 198 +/- 10 nmol/l; n = 30 and 36, P < 0.01). In SHR, platelet [Ca2+]i was higher than in WKY rats but did not change with age and blood pressure. Removal of external K+ or addition of 10(-4) mol/l ouabain were used to inhibit the Na+K(+)-pump. Whereas an increase in [Ca2+]i was observed in platelets from normotensives in the absence of external K+ (273 +/- 29 versus 197 +/- 9 nmol/l; n = 6; P < 0.05), no significant change in [Ca2+]i was observed after ouabain treatment (220 +/- 2 versus 203 +/- 22 nmol/l, n = 8). These results suggest that primary hypertension is accompanied by a disequilibrium between cellular Ca2+ influx, storage and extrusion. Such a characteristic, if present in other excitable cells and in particular in vascular smooth muscle cells, may play a major role in the increase in peripheral resistance. However, the relationship between Na(+)-pump inhibition and the rise in the intracellular calcium remains unclear.

Plasma membrane properties of 3 to 4-week-old spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats were investigated in both excitable cells, i.e. cardiomyocytes and platelets, and non-excitable cells, i.e. erythrocytes. Cardiac sarcolemma from SHR exhibited: lowered capacity of high-affinity Ca2+-binding sites; higher Ca2+ pump activity; higher Na+K+-ATPase activity due to increased density of Na+ pump units; suppression of the above three effects by the Ca2+-calmodulin complex, and increased Na+Ca2+ exchange. No difference in platelet cytosolic free Ca2+ concentration was observed between SHR and WKY. In both substrains, erythrocyte intracellular Na+ content was similar in spite of reduced Na+ and K+ net fluxes. Isolated membranes from SHR erythrocytes were also characterized by: lowered phosphoinositide turnover; decreased ATP-dependent Ca2+-transport, and lowered capacity of high affinity Ca2+-binding sites. Structural alterations detected by fluorescence polarization of diphenylhexatriene were observed in SHR cardiac sarcolemma, erythrocyte and brain synaptosomal membranes. Membrane organization and activity of transport systems controlling the intracellular Na+ and Ca2+ level were thus already modified in the prehypertensive animals whereas the resulting intracellular ion contents were still unaltered.

It has been proposed that peripheral resistance can be increased by ouabain-like factors that are able to increase cell sodium and thereby cell calcium. Canrenone has been reported to be a partial agonist of ouabain. The effect of canrenone was investigated in rats with reduced renal mass (RRM) showing evidence of excess circulating ouabain-like factors. Wistar rats were uninephrectomized, 30% of the other kidney was removed, and they were given a 0.8% NaCl solution to drink. Half of them received 60 mg/kg per day of canrenone orally for 26 days. In RRM, the following indices of a ouabain-like activity were found: erythrocyte Na+K(+)-pump activity was decreased by 39% (P < 0.001), sodium content increased by 12% (P < 0.01), net erythrocyte sodium extrusion in plasma decreased by 20% (P < 0.01), and plasma digoxin equivalents increased by 62% (P < 0.02). Canrenone increased the IC50 for ouabain from 1.05 to 2.16 x 10(-4) mol/l (P < 0.05) in erythrocytes. In RRM with systolic blood pressure of 165 mmHg, acute administration of canrenone decreased blood pressure by 36 mmHg. Chronic administration blunted the blood pressure rise by 12, 26 and 21 mmHg at days 5, 14 and 26, respectively (P < 0.05). Haematocrit was markedly reduced in RRM (33%) and much less when treated with canrenone (37.5%). In conclusion, in contrast with spontaneously hypertensive rats, RRM hypertension is a model where a ouabain-like factor is present and in which canrenone reduces blood pressure. The hypotensive effect of canrenone may be related to a competition with ouabain-like factors.

Essential hypertension is accompanied by several modifications to platelet metabolism suggesting hyper-reactivity to various aggregating agents. As the platelet response is mediated by both cytosolic free calcium, which is stimulatory, and cyclic (c)AMP, which is inhibitory, this hyper-reactivity may be caused by a modification in cAMP metabolism. We therefore determined cAMP in unstimulated platelets from 19 patients with essential hypertension and 27 age-matched normotensive subjects, nine with and 19 without a family history of hypertension. The platelet cAMP content was reduced in the essential hypertensives and in the normotensives with a positive family history by 37.5% and 42%, respectively (P less than 0.001 for both). Platelet cAMP was inversely correlated with diastolic blood pressure (P = 0.036). After prostaglandin (PG) E1 stimulation, the platelet cAMP content remained lower in the patients with essential hypertension than in the normotensive subjects, whatever their hypertensive heredity. The rises in cAMP caused by inhibition of phosphodiesterase by 7-bromo-1,5-dihydro-3,6-dimethylimidazo-[2,1-b]quinazolin-2[ 3H]-one (Ro 15-2041) were similar in the three groups. These results indicate that cAMP, the platelet inhibitory messenger, is reduced in hypertensive patients and in their normotensive offspring and may affect the various platelet abnormalities previously described in this disease.