Do canrenone and 6,7-dihydroxylated derivatives compete with ouabain at the same site on Na,K-ATPase?

Canrenone is the major metabolic product of the synthetic steroids spironolactone and K+-canrenoate, used in antihypertensive therapy. Canrenone can competitively displace [3H]ouabain from Na,K-ATPase [Na+- and K+-activated, Mg2+-dependent adenosine triphosphatase (E.C.3.6.1.3)] and partially inhibit enzymatic activity. These features have led to a hypothesis that competition between canrenone and endogenous digitalis-like materials, suggested to be involved in etiology of essential hypertension, could underly the antihypertensive effect of canrenone. Surprisingly, three derivatives of canrenone (6 beta,7 alpha-,6 beta,7 beta-, and 6 alpha,7 alpha-dihydroxy-6,7-dihydrocanrenone) reportedly occur in normal human and rat urine. This paper characterizes the interactions with partially purified renal Na,K-ATPase of canrenone, the three 6,7-dihydroxylated derivatives, and one epoxide intermediate, synthesized from K+-canrenoate. Canrenone and all the 6,7-substituted derivatives partially inhibited Na,K-ATPase activity (39-45%), with approximately the same apparent affinity, 100-200 microM. Canrenone displaced [3H]ouabain in an apparently competitive fashion (Ki = 100-300 microM) but none of the tested derivatives significantly displaced ouabain even at very high concentrations. The ability to differentiate the ATPase inhibition and [3H]ouabain displacement by modification of the 6,7-double bond indicates that inhibition of ATPase activity does not occur from within the ouabain binding site. We suggest that neither canrenone nor the 6,7-derivatives bind to the ouabain site, but rather interact with it 'allosterically.' Our findings do not support the proposed mechanisms for the antihypertensive action of canrenone.     

Digitalis-like compounds in animal tissues.

The Na+, K+ activated adenosine triphosphatase is present in the membrane of eukaryotic cells and represents a major pathway for Na+ and K+ transport across the plasma membrane. Cardiac glycosides such as ouabain or digoxin suppress this enzyme activity by binding to a specific receptor on the membrane. Studies conducted in this and other laboratories have proven the existence of digitalis-like compounds in animal tissues which may serve as in vivo regulators of the Na+, K(+)-pump activity. This review summarizes the attempts to identify these compounds from animal tissues and examines the potential physiological role of some of the identified compounds.     

Inhibition of the erythrocyte Na+, K+-pump by mammalian lignans

Several mammalian lignans, particularly enterolactone, 3-oxy-methyl enterolactone and prestegane B are able to inhibit Na+, K+-pump activity in human red cells with IC50 of about 1 mM. The inhibition of Na+, K+-pump activity by mammalian lignans have the following properties: the IC50 for ouabain remains unchanged suggesting a noncompetitive inhibition. The apparent affinity for internal Na+ and maximal rate of cation translocation are both diminished. The above inhibition of the Na+, K+-pump was obtained at doses 2-3 orders of magnitude higher than those required for ouabain. However we cannot exclude that a glycosyl- (and/or butenolide)-derivative of enterolactone could be one endogenous ouabain-like factor.     

Quantitative changes in cardiac Na+, K+ -adenosine triphosphatase of spontaneously hypertensive rats

Sodium pumps of cardiac plasma membranes were studied in young, spontaneously hypertensive rats (SHR) and in their normotensive controls (Wistar-Kyoto; WKY) using the two following methods. The enzymatic activity and its sensitivity to ouabain were measured as the Na+, K+ -dependent ATP hydrolysis, and the number of pumps was estimated by [3H] ouabain binding. The main results of this study were the observations that (a) concentrations of ouabain as low as 10(-10) M inhibited 10-15% of the enzyme activity in both strains; (b) Na+, K+- adenosine triphosphatase (ATPase) activity in membranes from SHR was double that in membranes from WKY (16.5 +/- 3.2 mumol Pi/h/mg protein vs. 8.2 +/- 1.2 mumol Pi/h/mg protein for 10(-7) M ouabain; p less than 0.01); (c) sensitivity to three different cardiac glycosides, ouabain, digoxin, and digitoxigenin, was identical in SHR and WKY vesicles; and (d) the binding capacity of [3H] ouabain was significantly higher in SHR than in WKY vesicles, but the dissociation constant (KD) did not appear to differ between the two substrains. These studies, performed on 3-week-old rats before the appearance of hypertension, showed, on the one hand, the existence of a Na+, K+ -ATPase of very high affinity in the rat heart, and, on the other, that cardiac sarcolemmal membranes from SHR had a greater number of sodium pumps than those from WKY and thus a greater ability to extrude sodium.     

High sensitivity of the Na+, K+-pump of human red blood cells to genins of cardiac glycosides.

1. Four different cardiac glycosides (ouabain, digitoxin, digoxin and gitoxin) and their corresponding genins were tested on Na+, K+-pump fluxes measured under steady-state and initial rate conditions (non equilibrium conditions) in human and rat erythrocytes and in mouse macrophages. 2. In human red cells, Na+, K+-pump fluxes exhibited up to 8 fold higher sensitivity to genins than to glycosides. In addition genins, but not the corresponding glycosides, exhibited double reactivity with regard to the erythrocyte Na+, K+-pump (with the exception of gitoxigenin). A weak reactivity component was similar to the one of the corresponding glycosides (IC50 of about 10(-6) M) and a high reactivity component exhibited IC50 values varying from 0.1 to 0.5 X 10(-6) M for digitoxigenin and ouabagenin respectively. 3. In contrast with human red cells, the initial rate of Na+, K+-pump fluxes in rat erythrocytes and mouse macrophages was less sensitive to genins than to the corresponding cardiac glycosides. 4. Dihydroouabain was 3, 10 and 75 times less active than ouabain in inhibiting the initial rate of Na+, K+-pump fluxes in human and rat erythrocytes and in mouse macrophages respectively. 5. In conclusion, Na+, K+-pump fluxes measured under initial rate conditions in human erythrocytes exhibit an unusually high sensitivity to genins of cardiac glycosides. This property probably results from the fast binding rate constants of genins and the slow association rates of glycosides to human red cells.     

Beta3-adrenoceptor agonist stimulation of the Na+, K+ -pump in rat skeletal muscle is mediated by beta2- rather than beta3-adrenoceptors

BACKGROUND AND PURPOSE: In cardiac muscle, BRL 37344, a selective beta3-adrenoceptor agonist, activates the Na+, K+ -pump via NO signalling. This study investigated whether BRL 37344 also activates the Na+, K+ -pump via beta3-adrenoceptors in skeletal muscle. EXPERIMENTAL APPROACH: Isolated rat soleus muscles were incubated between 1 and 60 min in buffer. Intracellular Na+, K+ content and Na+, K+ -pump activity were measured using flame photometry and ouabain-suppressible 86Rb+ uptake, respectively. Additional muscles were mounted on force transducers and stimulated (60 Hz for 2 s) every 10 min. KEY RESULTS: BRL 37344 (10(-8) -10(-5) M) induced a concentration- and time-dependent reduction in intracellular Na+, and increased ouabain-suppressible 86Rb+ uptake by up to 112%. BRL 37344-induced reductions in intracellular Na+ were blocked by the beta1/beta2-adrenoceptor antagonist, nadolol (10(-7) M), and the beta2-adrenoceptor antagonist, ICI 118,551 (10(-7) -10(-5) M), but not by beta3- or beta1-adrenoceptor antagonists, SR 59230A (10(-7) M) and CGP 20712A (10(-7) -10(-5) M), respectively. Another beta3-adrenoceptor agonist, CL 316,243, did not alter intracellular Na+. BRL 37344-induced reductions in intracellular Na+ were not blocked by L-NAME, an NOS inhibitor, or ODQ, a guanylyl cyclase inhibitor. The NO donors, SNP and SNAP, did not alter intracellular Na+. BRL 37344 rapidly recovered force in muscles depressed by high [K+]o, an effect that was blocked by nadolol, but not L-NAME. CONCLUSIONS AND IMPLICATIONS: In rat soleus muscle, the beta3-adrenoceptor agonist BRL 37344 stimulated the Na+, K+ -pump via beta2-adrenoceptors. A more selective beta3-adrenoceptor agonist did not affect Na+, K+ homeostasis in skeletal muscle. NO did not seem to mediate Na+, K+ -pump stimulation in skeletal muscle.     

Effects of fluvastatin treatment on red blood cell Na+ transport systems in hypercholesterolemic subjects

This study was performed to ascertain the effects of short-term cholesterol-lowering therapy with fluvastatin on red blood cells Na+ transport systems. Forty familial hypercholesterolemic subjects (FH; 19 men and 21 women) without hypertension or cardiovascular disease were given a placebo for 4 weeks, and then randomized in two groups. Twenty (fluvastatin group) were given fluvastatin (40 mg/day), and the other 20 (placebo group) continued placebo administration. After the placebo period and after 4 and 12 weeks of placebo or fluvastatin treatment, we measured Na+/K+ pump activity, Na+/K+ cotransport (Na+/K+ Ct), Na+/Li+ countertransport (Na+/Li+ Cnt), passive Na+ permeability (Na+PP), and internal Na+ content (Na+i). The same parameters were measured in 23 control subjects (C) with normal cholesterolemic values, who were matched for sex and age. FH had higher Na+/Li+ Cnt values than C (193.2 +/- 59.4 vs. 139.8 +/- 48.7 microM cells/h; p < 0.01), an increase in Na(+)PP (0.034 +/- 0.012/h vs. 0.018 +/- 0.004/h; p < 0.001), and higher Na(+)i (7.5 +/- 1.5 vs. 6.2 +/- 0.9 mM cells; p < 0.001). In hypercholesterolemic subjects, Na(+)i values were correlated with cholesterol (total and LDL) and apo B levels, whereas an inverse correlation was found for HDL-c and apo AI levels. Reduced total and LDL cholesterol and apo B levels after fluvastatin treatment caused a decrease in both Na(+)/Li(+) Cnt (from 186.1 +/- 60.5 to 125.1 +/- 34.0 microM cells/h; p < 0.001) and Na(+) PP (from 0.035 +/- 0.013/h to 0.02 +/- 0.016/h; p < 0.01), and an increase in Na+/K+ pump activity (from 1,549.0 +/- 507.7 to 1,894.2 +/- 536.2 microM cells/h; p < 0.04), with a significant reduction in the internal Na+ content (from 7.5 +/- 1.6 to 5.8 +/- 2.4 mM cells; p < 0.001). Our findings show that hypercholesterolemia affects red blood cell Na+ transport systems, with an increase in Na+/Li+Cnt, Na+PP, and the internal Na+ content. Cholesterol-lowering treatment with fluvastatin influences Na+ transport systems and reduces the internal Na+ content. This might also be responsible for the greater vascular reactivity observed in hypercholesterolemic patients, and its amelioration after a reduction in cholesterol levels.     

POPULATION-BASED SURVEY OF HUMAN SODIUM AND POTASSIUM EXCRETION

1. During the 1989 National Heart Foundation Risk Factor Prevalence Survey a subsample in Hobart collected 24 h urine samples to measure electrolyte excretion. 2. The ranges were 30-344 mmol/24 h for Na+ excretion (mean 160 mmol/24 h for men, 124 mmol/24 h for women), and 25-119 mmol/24 h for K+ excretion (mean 77 mmol/24 h for men, 68 mmol/24 h for women). 3. As in other surveys, women excreted about 20-25% less Na+ and K+ than men, although there was no significant sex difference in the ratio of Na+/K+ excretion. 4. The recommended dietary intake (RDI) for Na+ and K+ was followed simultaneously by 19% of subjects, and 13% had a 24 h urinary Na+/K+ ratio less than or equal to 1.0. 5. Observance of the RDI limited the value of iodized salt for goitre prophylaxis. 6. Sodium excretion rates were outside the therapeutic range of thiazide diuretics in 22% of subjects. 7. Diet groups for long-term prospective cohort studies to test the prophylactic value of avoiding salt could apparently be recruited from existing subsamples of the population.     

Cardenolide analogues. 7. Synthesis and biological activity of some new steroidal guanylhydrazones.

The synthesis, proof of structure, and biological activity of some new steroidal 17beta-formyl guanylhydrazones are described. The guanylhydrazones of nondigitalis-like steroids inhibited myocardial Na+,K+-ATPase but had only a depressant effect on myocardial contractility. By comparison, the corresponding guanylhydrazone of a digitalis-like steroid gave a positive inotropic effect in concentrations that also inhibited Na+,K+-ATPase. The nondigitalis-like guanylhydrazones also inhibited membrane Mg2+-ATPase and this may infer that the compounds act nonspecifically by membrane stabilization rather than by interaction with stereoselective receptors. Biological activity was determined in the guinea pig.     

The effect of ouabain on tension in isolated respiratory tract smooth muscle of humans and other species.

1. The Na+,K+-pump has been implicated in animal models of airway hyperreactivity. We examined the effects of inhibiting the Na+,K+-pump and Na+,Ca2+-exchange on isometric tone of isolated trachealis from humans and other species. 2. In preparations from 5 out of 9 humans, strong spontaneous contractions (36-48 h-1; up to 1.8 g) developed within 25 min. 3. Ouabain (10(-7)-10(-5) M) caused an immediate and sustained contraction. This response was not blocked by atropine, diphenhydramine, or cimetidine. 4. Contractions were also elicited when the normal physiological solution was changed to a K+-free solution, a procedure which inhibits the Na+,K+-pump, and in reduced (15 mM) Na+ solution, which inhibits Na+,Ca2+ exchange. 5. In preparations of dog and guinea-pig isolated trachea, ouabain (10(-5) M) caused a multiphasic response; in the rabbit, ouabain was without effect. K+-free solution was without effect in the dog preparations and produced relaxation of the guinea-pig trachea. Guinea-pig tracheae responded to a low Na+ solution with a strong contraction. 6. Our findings indicate that: (a) human airway smooth muscle may be a spontaneously contracting muscle, at least in vitro, (b) a prolonged contraction to ouabain is unique for the human airway smooth muscle among the animals tested, as is the contraction in a K+-free medium, and (c) the contractile response does not involve acetylcholine or histamine release, but may involve a Na+,Ca2+-exchange mechanism. These results suggest that the level of Na+,K+-pump activity could play a role in determining the degree of bronchomotor tone in humans.     

Relationship of cardiac muscle tension to Na+,K+-adenosine triphosphatase activity after chronic digitoxin administration in cats

To obtain a better understanding of the mechanism of action of the cardiac glycosides, we examined inotropic and biochemical effects of digitoxin in myocardium from cats chronically exposed to the drug. The mechanical function of papillary muscles was tested isometrically and left ventricular tissue was analyzed for Na+,K+-dependent adenosine triphosphatase ATPase activity. Muscles from control cat hearts developed tension at 2.5 +/- 0.7 g/mm2; muscles from cats that received subcutaneous digitoxin--100 micrograms/kg on day 1, followed by 40 micrograms/kg/day for 4 days (group A), and 75 micrograms/kg on day 1, followed by 25 micrograms/kg/day for 9 days (group B)--developed significantly greater (p less than 0.05) tension of 4.8 +/- 0.3 and 3.6 +/- 0.6 g/mm2, respectively. Further, in vitro maximal responsiveness to digitoxin was greater in the muscles from digitalized groups than in controls (p less than 0.05): Muscles from control cats had a maximal response to in vitro addition of digitoxin of 3.5 +/- 0.1 g/mm2; muscles from cats in group A reached 4.9 +/- 0.3 g/mm2, and those from group B, 4.5 +/- 0.7 g/mm2. Specific activity of microsomal Na+,K+-ATPase from hearts of digitalized groups A and B was inhibited by 50-70% (p less than 0.01). Developed tension, specific Na+,K+-ATPase activity, and in vitro maximal responsiveness to digitoxin in a third group (C) of cats receiving the least daily digitoxin (75 micrograms/kg on day 1, followed by 15 micrograms/kg/day for 29 days) were not different from controls. Mean plasma digitoxin concentrations were 33, 16, and 3 ng/ml in groups A, B, and C, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of human erythrocyte and leukocyte Na+, K(+)-pump activity by lysophosphatidylcholines

Synthetic lysophosphatidylcholines (LPCs) were examined for their effects on erythrocyte and leukocyte Na+,K(+)-pump activity, on erythrocyte Na+,K(+)-cotransport activity and on the passive permeability of the red blood cell membrane. Erythrocyte and leukocyte Na+,K(+)-pump activity was estimated by ouabain-sensitive 86Rb-uptake and erythrocyte Na+,K(+)-cotransport activity by bumetanide-sensitive 86Rb-uptake. Bumetanide, ouabain-resistant 86Rb-uptake was considered as a measure of the passive permeability of the red blood cell membrane. LPCs containing long chain fatty acids such as myristoyl, palmitoyl, lauroyl, stearoyl and oleoyl inhibited erythrocyte and leukocyte Na+,K(+)-pump activity and erythrocyte Na+,K(+)-cotransport activity, while they stimulated the passive membrane permeability of the red blood cells. LPCs containing lauroyl, the shortest fatty acid in this group, had the lowest inhibitory activity, while LPCs with intermediate chain length fatty acids such as caproyl and decanoyl had no effect. The order of inhibitory action of these LPCs on erythrocyte and leukocyte Na+,K(+)-pump activity was: palmitoyl greater than stearoyl greater than myristoyl greater than oleoyl greater than lauroyl.     

Effects of Na+,K+-pump inhibitors and membrane depolarizing agents on acetylcholine-induced endothelium-dependent relaxation and cyclic GMP accumulation in rat aorta

The purpose of this study was to investigate the effects of inhibitors of the Na+,K+-pump and membrane depolarizing agents on endothelium-dependent relaxation and elevated cyclic GMP levels induced by acetylcholine in rat thoracic aorta. Ouabain or exposure to K+-free or Mg2+-free Krebs-Ringer bicarbonate solution, agents and procedures known to inhibit the Na+,K+-pump, inhibited acetylcholine-induced relaxation and the associated increased levels of cyclic GMP. However, the inhibitory effect of ouabain on cyclic GMP levels was abolished in the absence of norepinephrine or in the presence of norepinephrine and the alpha-adrenergic receptor antagonist phentolamine. The membrane depolarizing agents KCl and tetraethylammonium also inhibited the acetylcholine-induced relaxation and the elevated cyclic GMP levels. Exposure to norepinephrine reduced the increased levels of cyclic GMP due to acetylcholine as compared to rested controls. This effect was inhibited by prior exposure to phentolamine, but not by the beta-adrenergic receptor antagonist, propranolol. These results suggest that increased activity of the Na+,K+-pump may mediate, in part, endothelium-dependent relaxation; inhibition of relaxation may be due to membrane depolarization; the endothelium-dependent increased levels of cyclic GMP may increase Na+,K+-pump activity; a complex interaction exists between membrane polarization, the Na+,K+-pump and alpha-adrenergic stimulation in regulation of cyclic GMP accumulation and relaxation.     

Suggestive evidence for inhibitory action of amiodarone on Na+, K+-pump activity in guinea pig heart

1. Effects of amiodarone, a powerful antiarrhythmic agent, on Na+, K+-pump activity were examined on ventricular papillary muscle of guinea pig, by means of conventional microelectrode technique. 2. The activity of Na+, K+-pump was measured by two methods. One of them was to measure the amplitudes of depolarization observed during overdrive stimulation (3.3 Hz) and of hyperpolarization observed after the overdrive stimulation (post-overdrive hyperpolarization), and the other, to measure the hyperpolarization observed following introduction of 10 mM K+, after exposure to K+ free solution for a certain duration. 3. Amiodarone significantly decreased the amplitude of depolarization during overdrive stimulation and the amplitude of post-overdrive hyperpolarization. 4. In the latter method, the deactivation process of hyperpolarization recorded by the introduction of 10 mM K+ following K+ depletion slowed down by amiodarone. 5. These findings suggest that amiodarone may inhibit, at least in part, the Na+, K+-pump activity in the ventricular muscle of guinea pig.     

The action of amlodipine on voltage-operated calcium channels in vascular smooth muscle.

1. The activity of the Na+/K(+)-pump in rat peritoneal mast cells was measured at various time intervals after induction of cellular histamine release by compound 48/80 or by the antigen-antibody reaction. The Na+/K(+)-pump activity was assessed as the ouabain-sensitive potassium uptake of the cells using 86Rb+ as a tracer for potassium (K+(86Rb+)-uptake). 2. Stimulation of the cells with compound 48/80 induced a time and concentration dependent increase of the Na+/K(+)-pump activity. The pump activity was maximal 2 min after stimulation of the cells. Then, the activity gradually decreased and reached a level not significantly different from the controls after 2 h of incubation. 3. When the cells were stimulated by the antigen-antibody reaction, there was also a rapid (within 5 min) stimulation of the Na+/K(+)-pump. In contrast to the stimulation with compound 48/80, the pump activity returned to the control level after 60 min of incubation with antigen. 4. The ouabain-resistant potassium uptake of the cells was increased after stimulation of the cells, regardless of the secretagogue used. This probably reflects the increased surface area of the cells present after the histamine release. 5. On the basis of the present results, we suggest a role for the Na+/K(+)-pump in the recovery process of the mast cell following histamine release.     

In vitro effect of xipamide on sodium-potassium transport systems in human erythrocytes

The in vitro effects of xipamide in a concentration range of 10(-8) to 10(-2) M were investigated on various Na+ and K+ transport systems in human red blood cells. Xipamide inhibited the anion carrier or DIDS-sensitive LiCO3- -influx starting from a concentration of 10(-5) M. However, a decrease in the Na+, K+-pump and the Na+, K+-cotransport activity and a rise in the passive permeability of the cell membrane was only observed starting from a concentration of 10(-4) M xipamide.     

Effectiveness of cardiac glycosides in human myocardium with and without "downregulated" beta-adrenoceptors.

We investigated the "receptor-effector-coupling" in the beta-adrenoceptor- and the Na+, K(+)-ATPase-mediated systems in nonfailing hearts and terminally failing human myocardium from patients with cardiomyopathy. The density of beta-adrenoceptors in the failing human myocardium was significantly (p less than 0.01) lower as compared with nonfailing hearts, whereas the receptor density and affinity measured by [3H]ouabain binding (cardiac glycoside receptor) was not different in either group. The maximal inotropic response to isoprenaline was significantly reduced in papillary muscle strips from failing human hearts (2.1 +/- 0.5 mN) as compared with control hearts (8.0 +/- 1.0 mN; p less than 0.05). Ouabain remained effective in both groups (6.8 +/- 1.0 vs. 5.5 +/- 0.6 mN; NS). The positive inotropic response due to extracellular Ca2+ elevation (1.8-15 mM) was studied for comparison. Maximal Ca2+ effects were reduced by 30% in failing human myocardium (7.2 +/- 0.5 mN vs. 5.1 +/- 0.8 mN, p less than 0.05). Ouabain had effectiveness (95%) similar to that of Ca2+ in nonfailing and failing human cardiac muscle. It is concluded that treatment with cardiac glycosides may still be effective in end-stage heart failure with "downregulated" beta-adrenoceptors, as judged from these in vitro studies.     

Endogenous digitalis-like factors: in vitro comparison of biological and immunological activities of peptide and steroid candidates

Endogenous substances that modulate the activity of (Na+ + K+)-ATPase through interaction at the cardiac glycoside site have been postulated. Reports of digitalis-like biological and immunological activity exhibited by certain ACTH/MSH peptides and 14-OH steroids make these compounds potential candidates as endogenous digitalis-like factors. We tested several ACTH/MSH peptides and 14 alpha-OH steroids in four in vitro assays and detected no significant cardiac glycoside-like activity. On the other hand, chlormadinone acetate, a progesterone derivative shown to bind with high affinity to the digitalis receptor, was nearly equipotent to digoxigenin in a [3H]ouabain radioreceptor assay. In a [3H]digoxin radioimmunoassay, however, digoxigenin and digoxin were equipotent but chlormadinone acetate was inactive. A clear dissociation between radioreceptor assay and radioimmunoassay activity was also observed using 15 beta-OH-progesterone. Our findings indicate that (a) ACTH/MSH peptides and 14 alpha-OH steroids are not viable candidates as endogenous digitalis-like factors, (b) digoxin antibodies are not necessarily directed at molecular determinants critical for biological activity, and (c) among the compounds reported to exhibit digitalis-like activity and postulated to share structural features with an endogenous steroidal digitalis-like factor, only chlormadinone acetate and its congeners appear to constitute tenable models.     

Metabolic effects of high dose amiloride and spironolactone: a comparative study in normal subjects.

Amiloride (75 mg daily) and spironolactone (300 mg daily) were given to five normal subjects for 7 days in order to compare metabolic effects at maximal doses. Blood pressure, body weight, Na+ and K+ balance, and plasma concentrations of Na+, K+, active and total renin, angiotensin II, aldosterone, 11-deoxycorticosterone (DOC), 18-hydroxydeoxycorticosterone (18-OH DOC), corticosterone (B), 18-hydroxycorticosterone (18-OH B) and cortisol were measured before and on each day of treatment. Natriuresis and K+ retention were significantly greater with amiloride. Plasma K+ increased from 4.1 +/- 0.2 to 4.9 +/- 0.2 mmol/l (mean +/- s.d.) on amiloride and from 4.0 +/- 0.2 to 4.4 +/- 0.2 mmol/l with spironolactone. Stimulation of renin, angiotensin II, aldosterone and 18-OH B occurred with both drugs but was greater with amiloride in each case. A transient decrease in systolic and diastolic blood pressure was observed after 2 days of spironolactone treatment but not with amiloride. The slope of the regression of aldosterone on angiotensin II during spironolactone treatment was less than that with amiloride, consistent with partial blockade of aldosterone synthesis by spironolactone. These data suggest that the maximum metabolic effects of amiloride exceed those of spironolactone.     

In vitro effect of cilazaprilat on sodium-potassium transport systems in human erythrocytes

The in vitro effects of cilazaprilat in a concentration range of 10 nM to 1 mM were investigated on various Na+ and K+ transport systems in human red blood cells. Cilazaprilat inhibited the anion carrier or DIDS-sensitive LiCO3(-)-influx and the bumetanide-sensitive K(+)-efflux. However, no effect of cilazaprilat on the Na+, K(+)-pump activity, the number of active Na+ pump units and on the Na+, Li(+)-countertransport activity could be demonstrated. It is suggested that through a decreased anion carrier indicating a decreased Na(+)-influx, the lower intracellular Na+ concentration observed in vivo during angiotensin-converting enzyme inhibition, can be at least partly explained.