On the functional interaction between nicotinic acetylcholine receptor and Na+,K+-ATPase

Previous studies have shown that nanomolar acetylcholine (ACh) produces a 2 to 4-mV hyperpolarization of skeletal muscle fibers putatively due to Na⁺,K⁺-ATPase activation. The present study elucidates the involvement of the nicotinic ACh receptor (nAChR) and of Na⁺,K⁺-ATPase isoform(s) in ACh-induced hyperpolarization of rat diaphragm muscle fibers. A variety of ligands of specific binding sites of nAChR and Na⁺,K⁺-ATPase were used. Dose–response curves for ouabain, a specific Na⁺,K⁺-ATPase inhibitor, were obtained to ascertain which Na⁺,K⁺-ATPase isoform(s) is involved. The ACh dose–response relationship for the hyperpolarization was also determined. The functional relationship between these two proteins was also studied in a less complex system, a membrane preparation from Torpedo electric organ. The possibility of a direct ACh effect on Na⁺,K⁺-ATPase was studied in purified lamb kidney Na⁺,K⁺-ATPase and in rat red blood cells, systems where no nAChR is present. The results indicate that binding of nAChR agonists to their specific sites results in modulation of ouabain-sensitive (most probably α2) isoform of Na⁺,K⁺-ATPase, leading to muscle membrane hyperpolarization. In the Torpedo preparation, ouabain modulates dansyl-C6-choline binding to nAChR, and vice versa. These results provide the first evidence of a functional interaction between nAChR and Na⁺,K⁺-ATPase. Possible interaction mechanisms are discussed.     

Na+/K+ pump and endothelial cell survival: [Na+]i/[K+]i-independent necrosis triggered by ouabain, and protection against apoptosis mediated by elevation of [Na+]i

Recent studies have demonstrated the tissue-specific effect of Na⁺/K⁺ pump inhibition by ouabain and other cardiac glycosides on cell viability. The vascular endothelium is an initial target of cardiac glycosides employed for the management of congestive heart failure as well as circulating endogenous ouabain-like substances (EOLS), the production of which is augmented in volume-expanded hypertension. This study examined the role of the Na⁺/K⁺ pump in the survival of cultured porcine aortic endothelial cells (PAEC). Complete Na⁺/K⁺ pump inhibition with ouabain led to PAEC death, indicated by cell detachment and decreased staining with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Based on cell swelling and resistance to benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (z-VAD.fmk) a pan-caspase inhibitor, this type of cell death was classified as necrosis. In contrast to ouabain, Na⁺/K⁺ pump inhibition in K⁺-free medium did not affect PAEC viability and sharply attenuated apoptosis triggered by ³H decay-induced DNA damage. Necrosis evoked by ouabain was preserved after dissipation of the transmembrane gradient of K⁺ and Na⁺, whereas dissipation of the Na⁺ gradient abolished the antiapoptotic action of K⁺-free medium. Comparative analysis of these results and modulation of intracellular Na⁺ and K⁺ content by the above-listed stimuli showed that interaction of ouabain with Na⁺/K⁺-ATPase triggered necrosis independently of inhibition of Na⁺/K⁺ pump-mediated ion fluxes and inversion of the [Na⁺]_i/[K⁺]_i ratio, whereas protection against apoptosis under Na⁺/K⁺ pump inhibition in K⁺-depleted medium was mediated by [Na⁺]_i elevation. The role of Na⁺/K⁺ pump-mediated regulation of endothelial cell survival and vascular remodelling seen in hypertension should be investigated further in context of EOLS and chronic treatment with digitalis.     

Calcium supplementation and thyroid hormone protect against gentamicin-induced inhibition of proximal tubular Na+, K+-ATPase activity and other renal functional changes

Gentamicin can cause proximal tubule necrosis. We have shown that inhibition of PT Na⁺, K⁺-ATPase activity is rapidly induced by gentamicin. We have now investigated whether manipulations known to attenuate the negative effects of gentamicin on renal excretory capacity, i.e. high calcium intake and L-thyroxine treatment, will also attenuate gentamicin-induced inhibition of Na⁺, K⁺-ATPase activity and ameliorated signs of proximal tubule damage. Rats were gentamicin- or vehicle-treated for 7 days. Sub-groups were given 4% calcium (Ca) supplements or L-thyroxine 20 μg 100 g^-1 body weight daily. Gentamicin significantly reduced the glomerular filtration rate and increased the urinary excretion of the proximal tubule lysosomal enzyme, N-acetyl-β-d -glucosaminidase. Gentamicin significantly reduced proximal tubule Na⁺, K⁺-ATPase activity, measured in single permeabilized proximal tubule segments. Sodium excretion was inversely correlated to proximal tubule Na⁺, K⁺-ATPase activity. Both calcium and L-thyroxine alleviated all gentamicin-induced side-effects on renal function as well as on proximal tubule Na⁺, K⁺-ATPase activity. Calcium and L-thyroxine had no significant effect on renal function. L-thyroxine, but not calcium, increased proximal tubule Na⁺, K⁺-ATPase activity in control rats. Renal cortical tissue gentamicin concentration was not influenced by calcium but was significantly lowered by L-thyroxine. Two procedures which, via different mechanisms, afford protection from gentamicin-induced changes in renal function also give protection from gentamicin-induced inhibition of Na⁺, K⁺-ATPase activity. This suggests that loss of integrity of the Na⁺, K⁺-ATPase enzyme contributes to gentamicin-induced nephrotoxicity.     

Unchanged sarcoplasmic reticulum Ca2+-ATPase activity, reduced Ca2+ sensitivity, and negative force-frequency relationship in transgenic rats overexpressing the mouse renin gene

Transgenic rats overexpressing the mouse Ren-2 gene [TG(mREN2)27 rats, TGR] were used to characterize alterations in force generation and relaxation following cardiac hypertrophy. Age-matched Sprague-Dawley rats were used as the control group. The β-adrenoceptor dependent increase in force of contraction was reduced in the transgenic animals but not the Ca²⁺-dependent increase in force generation. Additionally, force of contraction decreased after increasing stimulation frequencies (up to 7 Hz), but the frequency-dependent decrease in force of contraction was significantly more pronounced in the transgenic group. The Ca²⁺ sensitivity in chemically skinned fiber preparations of TGR was reduced than that in Sprague-Dawley rats while maximum effectiveness was the same. Unexpectedly, the sarcoplasmic reticulum Ca²⁺-ATPase activity measured in crude membrane preparations from TGR did not differ from that in Sprague-Dawley rats; however, the activity of the Na⁺/K⁺-ATPase was less while the Na⁺/Ca²⁺-exchanger activity was significantly greater. In the same preparations the protein expression of SERCA2 was reduced in TGR while expression of phospholamban and calsequestrin remained the same. Thus in the model of cardiac hypertrophy harboring the mouse Ren-2 gene the hypothesized correlation between SERCA2 function and force-frequency relationship was not observed. Possible reasons for the more negative force-frequency relationship in TGR included changes at the level of the myofilaments and altered intracellular Na⁺ homeostasis which may result from the reciprocal changes in the Na⁺/K⁺-ATPase and the Na⁺/Ca²⁺-exchanger activity. Received: 15 September 1997 / Accepted: 7 April 1998     

Sodium-dependent regulation of sodium,potassium-adenosine-tri-phosphatase (Na+, K+-ATPase) activity in medullary thick ascending limb of Henle segments. Effect of cyclic-adenosine-monophosphate guanosine-nucleotide-binding-protein activity and arginine vasopressin

This study examine the regulation Na⁺, K⁺-ATPase activity in the medullary thick ascending limb of Henle Na⁺, K⁺-ATPase activity was determined in medullary thick ascending limb of Henle (mtal) segments dissected from rat kidneys. The sodium concentration in the medium (Na,) was 20 or 70 mM. Since the segments were permeabilized, intracellular Na⁺ (Na,) was assumed to be the same as Na₂. Dibuturyl cyclic adenosine monophosphate (dbcAMP) and forskolin inhibited Na⁺, K⁺-ATPase activity independently of Na_m. Arginine vasopressin (AVP) receptors coupled to adenylate cyclase have been identified in the medullary thick ascending limb of Henle. At Na_m= 20 mMAVP caused a dose-dependent inhibition of Na⁺, K⁺-ATPase activity with a maximal effect (49%) at 10^-8 m. This inhibition was abolished in the presence of the adenylate cyclase inhibitor 2,5-dideoxyadenosine (2, 5-DDA). AVP had no effect on Na⁺, K⁺-ATPase activity in the mTAL at Na_m= 70 mM. The guanosine-diphosphate analogue GDPβS inhibited Na⁺, K⁺-ATPase activity at Na_m= 70 mM but not at Na_m= 20 mM. We conclude that increased cyclic adenosine monophosphate (CAMP) levels inhibit Na⁺, K⁺, ATPase activity in mTAL. AVP can, depending on Na₂ produce this effect by adenylate cyclase activation. The guanonine nucleotide binding protein G-protein might be the site of Na⁺-dependence.     

Forskolin–induced down–regulation of Na+,K+–ATPase activity is not associated with internalization of the enzyme

Activation by protein kinase A by forskolin phosphorylates and inactivates Na⁺,K⁺-ATPase in COS-7 cells (Cheng et al. 1997b). In this study we show, using [³H]ouabain binding, that forskolin-induced inhibition of Na⁺,K⁺-ATPase activity is not because of internalization of the enzyme. The effect of forskolin on Na⁺,K⁺-ATPase activity was examined by two independent methods, ouabain-sensitive ⁸⁶Rb⁺ uptake in intact cells and ATP hydrolysis in microsomal preparations from cells. The change in number of functional pumps on cell surface before and after protein kinase A activation was assessed by [³H]ouabain binding measured under equilibrium conditions. Cells, which had been ATP-depleted by antimycin A and 2-deoxyglucose treatment, served as a positive control for the internalization of Na⁺,K⁺-ATPase. Activation of protein kinase A with forskolin in combination with the phosphodiesterase inhibitor 3-isobutyl-1-methyl xanthine, inhibited Na⁺,K⁺-ATPase activity, but this treatment had no effect on specific ouabain binding. No change in ouabain binding was found following activation of protein kinase C by phorbol ester or diacyl glycerol analogue treatment in cells. These data suggest that protein kinase A phosphorylation and inhibition of Na⁺,K⁺-ATPase activity does not lead to any internalization of the enzyme in COS-7 cells.     

Additive positive inotropic effects of milrinone,ouabain and calcium in diseased human ventricular myocardium

Summary The interactions of milrinone, ouabain and calcium on force of contraction in isolated, contracting human papillary muscle strips were measured. Milrinone (EC₅₀, 8 × 10^–5 M) increased force of contraction maximally by 2.8±0.8 mN at 5 × 10^–4M; significantly less than either ouabain (1 × 10^–7M; 4.8±0.5 mN increase) or calcium (15 mM; 6.2±0.6 mN increase). A submaximal, but not a maximal, inotropic effect of ouabain could be increased by the addition of milrinone; in contrast, both ouabain and calcium increased the maximal inotropic effect of milrinone by 1.7±0.2 mN and 2.7±0.3 mN, respectively. The combined inotropic effect of milrinone with either ouabain of 4.2±0.3 mN or calcium of 5.6±0.4 mN was not different from that with calcium or ouabain alone. We conclude that further positive inotropic effects should be expected when digitalis is given to patients with congestive heart failure who are already optimally treated with milrinone.Abbreviations K _D dissociation constant - (Na⁺ + K⁺)-ATPase sodium and potassium activated adenosine triphosphatase (EC 3.6.1.3) This study was carried out using human heart samples provided by: Prof. E. Kreuzer, Prof. B. Kemkes, Dr. C. Weinhold and their colleagues, Herzchirurgische Klinik der Universität, Klinikum Großhadern, München     

A proposed mechanism of action of the inotropic agent digitalis

A theory based on hydrogen bonded water structure is put forward to explain the action of digitalis and other cardiac glycosides on cardiac tissue. The activity of free Ca⁺⁺ is altered when the Na⁺ + K⁺ ATPase is inhibited, and there results a redistribution between sarcoplasm and other calcium pools.     

Effects of digoxin on muscle reflexes in normal humans

Blockade of the skeletal muscle Na⁺–K⁺-ATPase pump by digoxin could result in a more marked hyperkaliema during a forearm exercise, which in turn could stimulate the mechano- and metaboreceptors. In a randomized, double-blinded, placebo-controlled, and cross-over-design study, we measured mean blood pressure (MBP), heart rate (HR), ventilation (V _E), oxygen saturation (SpO₂), muscle sympathetic nerve activity (MSNA), venous plasma potassium and lactic acid during dynamic handgrip exercises, and local circulatory arrest in 11 healthy subjects. Digoxin enhanced MBP during exercise but not during the post-handgrip ischemia and had no effect on HR, V _E, SpO₂, and MSNA. Venous plasma potassium and lactic acid were also not affected by digoxin-induced skeletal muscle Na⁺–K⁺-ATPase blockade. We conclude that digoxin increased MBP during dynamic exercise in healthy humans, independently of changes in potassium and lactic acid. A modest direct sensitization of the muscle mechanoreceptors is unlikely and other mechanisms, independent of muscle reflexes and related to the inotropic effects of digoxin, might be implicated.     

Isoform of Na+, K+-ATPase from rumen epithelium identified and quantified by immunochemical methods

Using biopsies of rumen epithelium papillae a net influx of [⁸⁶Rb⁺] was measured corresponding to a high concentration of Na⁺, K⁺-pumps found in [³H]ouabain-binding studies ( 18 ). In the present study the Na⁺, K⁺-ATPase in papillae homogenates is compared with purified (Na⁺, K⁺)-ATPase from different sources, immunochemically characterized with respect to the isoform of the hydrolytic α subunit and the concentration of pumps substantiated by a novel immunochemical method. Na⁺, K⁺-ATPase purified from bovine kidney was shown to contain one homogeneous high-affinity population of [³H]ouabain-binding sites (K_d 1.37 n M ). The ouabain-binding capacity was 0.82 nmol (mg protein)^?1. Site-directed polyclonal antibodies raised to isoform-specific sequences of the three known α-subunit isoforms and monoclonal α₁-specific antibodies were used for isoform characterization on western blots of peptides separated by SDS-polyacrylamide gel electrophoresis. All three isoforms were present in Na⁺, K⁺-ATPase prepared from bovine brain. The α isoform of bovine kidney Na⁺, K⁺-ATPase and of rumen epithelium homogenate appeared to be α₁ whereas α₂ and α₃ were undetectable. Using an α₁-specific antibody and ¹²⁵I-labelled antimouse IgG the content of (Na⁺, K⁺)-ATPase in rumen epithelium was determined by comparison of the signal from known amount of bovine kidney Na⁺, K⁺-ATPase on western blots. By this method rumen epithelium was found to contain 2.6 nmol Na⁺, K⁺-ATPase (g wet wt)^?1, i.e. a similarly high or even higher concentration than previously seen in ouabain-binding studies on biopsies.     

Inotropic effect of ouabain in hypertrophied rat heart

The contribution of the heterogeneous digitalis receptors to the inotropic effect of ouabain was studied in hypertrophied rat hearts (aortic stenosis) by using isolated Langendorff heart preparations. Development and washout of the biological effects as well as the dose/ response curves revealed two inotropic components of high and low drug sensitivity. Maximal inotropy was observed with 100 M ouabain in both control and hypertrophied rat hearts. The high-sensitivity component accounted for only one-third of the response in control hearts but for two-thirds in hypertrophied hearts. The respective apparent affinities (10–20 nM and 10–20 M) of the two inotropic components found in isolated hearts were similar to those of the high- and low-affinity Na⁺,K⁺-ATPase activities detected in isolated cardiac sarcolemma. Onset and reversion of the pharmacological effects of ouabain occurred at respective rates that were similar to those of the association and dissociation of ouabain to the Na⁺,K⁺-ATPase level. In hypertrophied heart, the high- and low-sensitivity components (or receptors) reacted seven- and threefold, respectively, more slowly than the corresponding sites in normal hearts. These alterations in inotropic responsiveness and propertries of the digitalis receptors in cardiac hypertrophy suggest that new regulations should be taken into account in the adaptation to pressure overload.     

Heterogeneity of Na+/K+-ATPase from rectal gland of Squalus acanthias is not due to α isoform diversity

 Purified Na⁺/K⁺-ATPase (EC 3.6.1.37) isolated from the rectal gland of Squalus acanthias was characterized in ouabain-binding studies and with respect to isoform(s) of the α peptide. To avoid enzyme inactivation [³H]ouabain equilibrium binding was carried out at 20°C. The heterogeneity of Na⁺/K⁺-ATPase isolated from shark rectal gland was similar in [³H]ouabain binding as previously seen in hydrolytic studies. The binding isotherms were compatible with the existence of a high-affinity (K _dis 0.69 nM) and a low-affinity (K _dis 42 nM) component of 1.46 and 0.79 nmol^.(mg protein)^–1, respectively. In Western blots the α peptide of the enzyme hybridized with an isoform-specific polyclonal antibody raised to an α₃-specific region of the large intracellular domain of rat Na⁺/K⁺-ATPase, but not with the supposed α₃-specific monoclonal antibody MA3-915 with its epitope near the N-terminus. Semi-quantitative analysis of the reaction of the α₃-specific polyclonal antibody with the α peptide from the shark enzyme compared to the reaction with α peptide from rat brain enzyme indicated that this region is not exactly the same in the two species. The α peptide of shark enzyme was not recognized by α₁- or α₂-specific polyclonal antibodies, or by the α₁-specific monoclonal antibodies 3B and F6. The large intracellular domain of Na⁺/K⁺-ATPase from shark rectal gland thus seems to be α₃-like and no α isoform heterogeneity seems able to account for the heterogeneity seen in ouabain binding. Received: 7 August 1998 / Accepted: 6 November 1998     

Treatment of life-threatening digitalis intoxication with digoxin-specific Fab antibody fragments: experience in 26 cases

Purified Fab fragments of digoxin-specific antibodies obtained from sheep were used to treat 26 patients with advanced, life-threatening digoxin (23 cases) or digitoxin (3 cases) toxicity. These patients had advanced cardiac arrhythmias, and in some cases hyperkalemia, which were resistant to conventional treatment. All patients had an initial favorable response to doses of Fab fragments calculated (in most cases) to be equivalent, on a molar basis, to the amount of cardiac glycoside in the patient's body. In four patients treated after prolonged hypotension and low cardiac output, death ensued from cerebral or myocardial hypoperfusion. In one case the available Fab fragment supply was inadequate to reverse a massive suicidal ingestion of digoxin, and the patient died after recurrent ventricular arrhythmias. In the remaining 21 patients, cardiac rhythm disturbances and hyperkalemia were rapidly reversed, and full recovery ensued. There were no adverse reactions to the treatment. We conclude that the use of purified digoxin-specific Fab fragments is a safe and effective means to reverse advanced, life-threatening digitalis intoxication.     

Occurrence and properties of a (Na+−K+)-activated ATPase in the mucosa of the rat intestine

Summary The existence of an ouabain-sensitive (Na⁺–K⁺)-activated ATPase system has been demonstrated in the total intestine of the rat. The (Na⁺–K⁺)-ATPase activity was about 10–15% of the total ATPase in 4 equal parts of the small intestine; in the colon about 35% of the total ATPase was (Na⁺–K⁺)-activated ATPase. The highest (Na⁺–K⁺)-ATPase activity has been observed in the first and second part of the small intestine, while in the colon the activity was 2–4 times higher than in the ileum.The (Na⁺–K⁺)-ATPase of rat colon required both Na⁺ (K _m=8.3 mmoles/l) and K⁺ (K _m=0.6 mmoles/l). Maximal activation of the (Na⁺–K⁺)-ATPase system required 2 mM Mg²⁺ at an ATP concentration of 2 mM. The pH optimum for (Na⁺–K⁺)-ATPase of rat colon was 7.5, while the Mg²⁺-activated ATPase activity had a pH optimum of 8.6. The (Na⁺–K⁺)-ATPase was inhibited by ouabain (pI ₅₀=3.6).The relation between the differences in (Na⁺–K⁺)-ATPase activity and Na⁺-absorption on different parts of the intestine is discussed.     

The interaction between the deacetylated (amine) metabolite of diamphenethide (DAMD) and cytochemically demonstrable Na+/K+-ATPase activity in the tegument of Fasciola hepatica

The relative effects on tegumental Na⁺/ K⁺ -ATPase activity in Fasciola hepatica of the deacetylated (amine) metabolite of diamphenethide (DAMD) (10 g/ml, 18 h) and the Na⁺/K⁺-ATPase inhibitor ouabain (0.1 mM, 0.5 h) have been assessed cytochemically. In the normal tegument, Na⁺/K⁺-ATPase activity is particularly concentrated along the invaginations of the apical plasma membrane and the infoldings of the basal plasma membrane. Ouabain pretreatment significantly reduces the overall level of Na⁺/K⁺-ATPase activity, but does not induce swelling of the basal infolds. In contrast, DAMD does not significantly reduce either the level or distribution of Na⁺/K⁺-ATPase activity, but does cause a pronounced swelling of the basal infolds. The results are discussed in relation to the postulated action of diamphenethide as an inhibitor of Na⁺/K⁺-ATPase activity.     

Distribution of sodium-potassium ATPase in the rat testis and epididymis

Sodium-potassium ATPase (Na⁺ K⁺-ATPase) is a ubiquitous plasma membrane enzyme which uses the hydrolysis of ATP to regulate cellular Na⁺ and K⁺ levels and fluid volume. This ion pumping action is also thought to be involved in fluid movement across certain epithelia. There are several different genes for this enzyme, some of which are tissue specific. Using an antibody specific for the catalytic subunit of canine kidney Na⁺ K⁺-ATPase, we have localized immunoreactivity in the seminiferous and epididymal epithelium of rats of various ages. There was no specific staining of 10-day-old rat testis. Faint staining was detected at 13 days and appeared to be associated with the borders of Sertoli cells. At 16 days prominent apical and lateral staining but no basal staining of Sertoli cell membranes was observed. This type of distribution continued until spermatids were present in the epithelium. In the adult rat testis, specific staining was detected in Sertoli cell crypts associated with elongating spermatids, and on the apical and lateral Sertoli cell membrane. In some instances immunoreactivity was concentrated at presumed sites of junctional specializations. In the excurrent ducts of immature and mature rats, Na⁺ K⁺-ATPase staining was heavy in the efferent ducts and somewhat lighter in the epididymis. In all regions, the staining was basolateral although there were variations in intensity among the different parts of the epididymis. These results show (1) that rat testis and epididymal Na⁺ K⁺-ATPase share some immunological determinants with the canine enzyme; (2) that the epididymal enzyme is located in the conventional basolateral position; and (3) that the distribution of Sertoli cell Na⁺ K⁺-ATPase is probably apical and lateral rather than basal.     

Renal Na+, K+-ATPase in Dahl salt-sensitive rats: K+ dependence,effect of cell environment and protein kinases

Na⁺, K⁺-ATPase in renal epithelial cells plays an important role in the regulation of Na⁺ balance, extracellular volume and blood pressure. The function of renal Na⁺, K⁺-ATPase in Dahl salt-sensitive (DS) rats, an animal model for salt-sensitive hypertension, and Dahl salt-resistant (DR) rats has been studied. In Na⁺, K⁺-ATPase partially purified from renal cortex, affinities and the Hill coefficients for Na⁺ and K⁺ activation were similar in DS and DR rats. Only one component of low ouabain affinity site was found in both strains, indicating the presence of the al isoform. Protein kinase C and cAMP-dependent protein kinase phosphorylated Na⁺, K⁺-ATPase α subunit in DS and DR rats, and the phosphorylation by protein kinase C was associated with an inhibition of enzyme activity. The kinetic parameters for K⁺ activation were also studied in a preparation of basolateral membranes and were found to be similar in DS and DR rats. In a preparation of cortical tubule cells, Na⁺, K⁺-ATPase activity was determined as ouabain-sensitive oxygen consumption (OS Qo₂). Maximal OS Qo₂, measured in Na⁺ loaded cells, was the same in DS and DR rats. The K₀₆ for K⁺ was significantly lower in DS than DR rats (0.163 ±0.042 vs. 0.447 + 0.061 mM, P < 0.05), indicating that factors regulating Na⁺, K⁺-ATPase activity in intact cells are altered in DS rats. Kinetic parameters for Na⁺ activation in cells were the same in both strains. In summary, the function of renal Na⁺, K⁺-ATPase molecule is not altered in DS rats. The intracellular systems that regulate renal Na⁺, K⁺-ATPase activity might be different in DS and DR rats.     

Stimulation of Na+, K+-pump activity in skeletal muscle by methylxanthines: evidence and proposed mechanisms

Evidence is presented to support the hypothesis that submillimolar concentrations of methylxanthines stimulate Na⁺, K⁺-ATPase activity in skeletal muscle. Administration of methylxanthines to skeletal muscle results in plasma membrane hyperpolarization and increased rates of K⁺ uptake and Na⁺ efflux. These effects are both dose- and time-dependent and inhibited by blockers of the Na⁺, K⁺ ATPase. The mechanisms for stimulation of Na⁺, K⁺-ATPase activity and the signal transduction pathways are not known. The methylxanthine concentrations required for stimulation of Na⁺, K⁺-ATPase activity are less than those required to cause a 50% inhibition of phosphodiesterase activity, and therefore increases in cyclic AMP due to inhibition of the enzyme are not involved. Possible mechanisms by which methylxanthines may increase Na⁺, K⁺-ATPase activity include: (1) a role for increased intracellular [Ca²⁺]; (2) Ca²⁺ or adenosine-receptor-mediated increases in intracellular cyclic AMP; and (3) a direct action of methylxanthines on the Na⁺, K⁺ ATPase.     

Influence of bile acids on the (Na+−K+)-activated- and Mg2+-activated ATPase of rat colon

Summary The influence of various bile acids on the (Na⁺−K⁺)-ATPase and Mg²⁺-ATPase activity of rat colon is described. At a concentration of 0.6 mmol/l C and TC did not inhibit the (Na⁺−K⁺)-ATPase activity in contrast to GC. The taurine derivates TC, TCDC and TDC did not influence or even enhanced the (Na⁺−K⁺)-ATPase activity. All bile acids except C, TC and CDC depressed the Mg²⁺-ATPase activity. At higher concentrations only C and TC did not influence the (Na⁺−K⁺)-ATPase activity. C, GC and TC at 2.5 mmol/l decreased the (Na⁺−K⁺)-activated phosphatase with ATP as substrate. All other substrates tested did not influence the enzymic activity significantly. The results indicate that bile acids can inhibit the Na⁺-absorbing system in rat colon. Hence this inhibition can cause diarrhea.     

High salt diet down-regulates proximal tubule Na+, K+-ATPase activity in Dahl salt-resistant but not in Dahl salt-sensitive rats: evidence of defective dopamine regulation

Nishi , A., Bertorello , A. M. & Aperia , A. 1992. High salt diet down-regulates proximal tubule Na⁺, K⁺-ATPase activity in Dahl salt-resistant but not in Dahl salt-sensitive rats; evidence of defective dopamine regulation. Acta Ptiysiol Scand 144 , 263–267. Received 26 July 1991, accepted 25 October 1991. ISSN 0001–6772. Department of Paediatrics, Karolinska Institute, Sweden We examined the regulation of Na⁺, K⁺-ATPase activity in proximal tubule segments during a high salt diet in prehypertensive Dahl salt-sensitive and salt-resistant rats. Rats were placed on normal salt or high salt diets (0.9% saline as drinking water). During the normal salt diet, Na⁺, K⁺-ATPase activity was not different between Dahl salt-sensitive and salt-resistant rats. After 2 days and 10 days on a high salt diet, Na⁺, K⁺-ATPase activity in Dahl salt-resistant rats significantly decreased when compared to Dahl salt-resistant rats on a normal salt diet (P < 0.01). The decreased Na⁺, K⁺-ATPase activity in Dahl salt-resistant rats during a high salt diet was reversed by treatment with an inhibitor of aromatic l -amino acid decarboxylase (dopamine synthesizing enzyme), benserazide. In contrast, Na⁺, K⁺-ATPase activity did not decrease during the high salt diet and benserazide had no effect on Na⁺, K⁺-ATPase activity in Dahl salt-sensitive rats. These results indicate that Dahl salt-sensitive rats do not have the capacity to down-regulate the proximal tubule Na⁺, K⁺-ATPase activity during a high salt diet. Indirect evidence suggests that the regulation of Na⁺, K⁺-ATPase activity by locally produced dopamine is absent in Dahl salt-sensitive rats.