Heat shock- and ethanol-induced ionic changes in C6 rat glioma cells determined by NMR and fluorescence spectroscopy

The effects of two different stressors, heat shock (HS; 44 degrees C, 20 min) and ethanol (1.2 M, 60 min), on ion content and membrane potential were investigated in C6 rat glioma cells. Both treatments were previously shown to induce the HS response [26]. Intracellular pH (pH(i)), sodium ion concentration ([NA+]i), potassium ion concentration ([K+]i) and membrane potential were determined by means of continuous 31P and 23Na nuclear magnetic resonance (NMR), continuous fluorescence spectroscopy and 86Rb uptake. Lactate extrusion was determined in addition with respect to pH(i) regulation. The aim of this study was a detailed picture of HS and ethanol-induced ion changes in a single cell type, because stress-induced changes in the intracellular ionic balance may be important factors for determining proliferation, stress response and apoptosis. HS lowered the pH(i) from 7.38 +/- 0.04 to about 7.05 +/- 0.04. [Na+]i decreased during HS to 50% of the control and recovered to normal level 95 min after HS treatment. During HS, [K+]i remained constant but increased after HS. The membrane potential hyperpolarized from -83 mV to -125 mV and returned to initial values during HS treatment. Lactate extrusion increased 3-fold after HS. Ethanol (1.2 M) lowered the pH(i) from pH 7.38 +/- 0.04 to pH 7.0 +/- 0.04, but in contrast to heat strongly increased [Na]i. It hyperpolarized the membrane potential from -83 to -125 mV. Ethanol also increased lactate extrusion similar to HS. Also in contrast to the effect of HS, the potassium concentration decreased during ethanol treatment. The Na(+)-H+ exchanger monensin was used to overcome the apparent inhibition of the cellular Na(+)-H+ exchanger by HS. At normal pH(e) (7.4) monensin increased [Na+]i and pH(i) considerably. A subsequent HS reduced [Na+]i only minimally. Acidification of the cells by low pH(e) (6.2) prior to HS did not abolish the HS-induced drop of pH(i), indicating that the Na(+)-H+ exchanger was also inhibited at low pH(i). At low pH(e), monensin transports H+ into the cell. A subsequent HS decreased pH(i) only little, showing the importance of inhibition of the Na(+)-H+ exchanger for the HS-induced pH(i) decrease. 100 microM amiloride reduced pH(i) and [Na+]i in a similar way as HS, but did not change pH(i) and [Na+]i much during a HS. These results indicate that some of the HS-induced ionic changes are mediated by inhibition of the Na(+)-H+ exchanger, activation of Na(+)-K(+)-ATPase and changes of membrane conductance for ions.     

Reduced Na(+)-K(+)-ATPase activity and plasma lysophosphatidylcholine concentrations in diabetic patients

A fraction from normal human plasma inhibiting Na(+)-K(+)-ATPase has been recently identified as lysophosphatidylcholine (LPC). The aim of this study was to investigate the existence of a relationship between the activity of the cellular membrane Na(+)-K(+)-ATPase and plasma LPC in human diabetes. We studied 10 patients with insulin-dependent-diabetes mellitus (IDDM), 14 patients with non-insulin-dependent diabetes mellitus (NIDDM), and 10 sex- and age-matched control subjects. Plasma LPC concentrations were increased in both IDDM and NIDDM patients compared with control subjects. Na(+)-K(+)-ATPase activity was reduced in both groups of patients in erythrocyte and platelet membranes. There was a significant correlation between the concentrations of plasma LPC and Na(+)-K(+)-ATPase activity in both erythrocyte and platelet membranes (P < 0.01). To investigate the effect of LPC on the enzyme, Na(+)-K(+)-ATPase activity was determined in erythrocyte membranes obtained from six healthy subjects after in vitro incubation with increasing concentrations of LPC (1-10 microM). Enzymatic activity was significantly reduced by in vitro LPC at a concentration of 2.5 microM, with a further decrease at 5 microM. These data suggest that the decrease in Na(+)-K(+)-ATPase activity in diabetes might be due to increased LPC concentrations.     

Endothelin 1 stimulates Na+,K(+)-ATPase and Na(+)-K(+)-Cl- cotransport through ETA receptors and protein kinase C-dependent pathway in cerebral capillary endothelium

The effect of endothelins (ET-1 and ET-3) on 86Rb+ uptake as a measure of K+ uptake was investigated in cultured rat brain capillary endothelium. ET-1 or ET-3 dose-dependently enhanced K+ uptake (EC50 = 0.60 +/- 0.15 and 21.5 +/- 4.1 nM, respectively), which was inhibited by the selective ETA receptor antagonist BQ 123 (cyclo-D-Trp-D-Asp-Pro-D-Val-Leu). Neither the selective ETB agonists IRL 1620 [N-succinyl-(Glu9,-Ala11,15)-ET-1] and sarafotoxin S6c, nor the ETB receptor antagonist IRL 1038 [(Cys11,Cys15)-ET-1] had any effect on K+ uptake. Ouabain (inhibitor of Na+,K(+)-ATPase) and bumetanide (inhibitor of Na(+)-K(+)-Cl- cotransport) reduced (up to 40% and up to 70%, respectively) the ET-1-stimulated K+ uptake. Complete inhibition was seen with both agents. Phorbol 12-myristate 13-acetate (PMA), activator of protein kinase C (PKC), stimulated Na+,K(+)-ATPase and Na(+)-K(+)-Cl- cotransport. ET-1- but not PMA-stimulated K+ uptake was inhibited by 5-(N-ethyl-N-isopropyl)amiloride (inhibitor of Na+/H+ exchange system), suggesting a linkage of Na+/H+ exchange with ET-1-stimulated Na+,K(+)-ATPase and Na(+)-K(+)-Cl- cotransport activity that is not mediated by PKC.     

Kaliuretic peptide: the most potent inhibitor of Na(+)-K+ ATPase of the atrial natriuretic peptides

The present investigation was designed to determine whether atrial natriuretic peptides consisting of amino acids 1-30 (i.e. long-acting natriuretic peptide), 31-67 (vessel dilator), 79-98 (kaliuretic peptide), and 99-126 [atrial natriuretic factor (ANF)] of the 126 amino acid ANF prohormone inhibit sodium-potassium-ATPase as part of their mechanism(s) of action for producing a natriuresis and/or kaliuresis. Kaliuretic peptide, long-acting natriuretic peptide, vessel dilator and ANF at their 10(-11) M concentrations inhibited Na(+)-K(+)-ATPase 39.5%, 27.8%, 19.2%, and 4% respectively, in bovine renal medulla, whereas their inhibition in renal cortical membranes was 37.5%, 27.5%, 20%, and 0%, respectively. Ouabain (0.5 mM) inhibited kidney medullary Na(+)-K(+)-ATPase 45% and in the cortex, 38%. There was no additive effect of any of these peptides with ouabain suggesting that they are interacting with the same site on the Na(+)-K(+)-ATPase as ouabain. To help elucidate the mechanism of these peptides' interaction with Na(+)-K(+)-ATPase, naproxen (0.5 mM), an inhibitor of prostaglandin synthesis, and direct measurement of prostaglandin E2 by RIA were used. Naproxen completely blocked the inhibition of Na(+)-K(+)-ATPase by kaliuretic peptide, long-acting natriuretic peptide, and vessel dilator suggesting that their inhibition of Na(+)-K(+)-ATPase in both the kidney medulla and cortex are mediated by prostaglandins. Direct measurement of prostaglandin E2 revealed that kaliuretic peptide > long-acting natriuretic peptide > vessel dilator increased prostaglandin E2 synthesis, whereas ANF did not have any effect. Of interest, angiotensin II and ouabain inhibition of Na(+)-K(+)-ATPase were also completely blocked by naproxen.     

Membrane polarity of the Na(+)-K+ pump in primary cultures of Xenopus retinal pigment epithelium

The retinal pigment epithelium is a transporting epithelium that helps regulate the volume and composition of the subretinal space surrounding photoreceptor outer segments. The capacity of the RPE to actively transport Na+ and K+ between the retina and the blood supply depends on the localization of the Na+, K(+)-ATPase to the apical membrane, but in culture this polar distribution can be lost. Using primary cultures of Xenopus RPE, we examined the anatomical and functional polarity of this electrogenic pump. Confluent monolayers were established on Matrigel-coated microporous filters and cultured for 2-4 weeks in serum-free defined medium. Electrogenic pump activity at the apical and basolateral membranes was assayed by mounting the monolayer and filter in an Ussing chamber and exposing one or the other surface to ouabain while recording the apical (Vap) and basolateral (Vba) membrane potentials with an intracellular microelectrode. The addition of 0.2 mM ouabain to the apical bath caused Vap to rapidly depolarize by about 4 mV, consistent with the inhibition of a hyperpolarizing pump current at that membrane. When ouabain was added to the basal bath, however, it had no effect on Vba, suggesting the absence of a functional Na(+)-K+ pump on the basolateral membrane. To confirm these electrophysiological results, we examined the distribution of the Na+, K(+)-ATPase catalytic component using an antiserum specific for the bovine kidney alpha subunit. Antibody labeling of cultures was highly polarized, with strong reaction present on the apical microvilli, but not the basolateral cell surfaces. The findings of this study indicate that the Na(+)-K+ pump in monolayers of Xenopus RPE, as in native RPE, is located mainly in the apical membrane, providing evidence of a functionally intact transport pathway in these primary cultures.     

Human brain activity related to speed discrimination tasks

This study examined the ionic mechanism of ibutilide, a class III antiarrhythmic in clinical use, on freshly isolated human atrial cells. Cells had resting potentials of -71.4 +/- 2.4 mV, action potentials with overshoot of 36.8 +/- 1.8 mV, duration of 265 +/- 89 msec at 90% repolarization and slow repolarization (n = 16). Ibutilide, at 10(-7) M, markedly increased action potential duration. Four types of outward currents were detected: Ito, Iso, a delayed rectifier and IK1. Ibutilide had no inhibitory effect on these outward currents at 10(-7) M (n = 28). In K(+)-free solutions and -40 mV holding potential, mean peak inward current at 20 mV was -1478 +/- 103 pA (n = 12). Ibutilide increased this current to -2347 +/- 75 pA at 10(-7) M, with half maximal effect (Kd) of 0.1 to 0.9 nM between -10 and +40 mV (n = 21). At similar concentrations, the drug increased APD, with Kd of 0.7 and 0.23 nM at 70 and 90% repolarization, respectively (n = 8). Ibutilide shifted the mid-point of the steady-state inactivation curve from -21 to -12.2 mV (n = 6), and reduced current decline during repetitive depolarization (n = 5). The drug induced inward current was carried by Na+o through a nifedipine inhibited inward channel because Na+o removal eliminated the effect, and nifedipine abolished the inward current and the drug induced APD prolongation. We propose that a Na+ current through the L-type Ca++ channel mediates ibutilide's potent clinical class III antiarrhythmic action.     

Na/H exchange and H-K ATPase increase distal tubule acidification in chronic alkalosis

We examined whether H(+)-ATPase, H(+)-K(+)-ATPase, and or Na+/H+ exchange mediates increased distal tubule acidification in animals with chronic metabolic alkalosis using pharmacological inhibitors of these H+ transporters in in vivo-perfused tubules of anesthetized rats. Chronic metabolic alkalosis was induced with furosemide followed by minimum electrolyte diet and HCO3 drinking water. The reduction in net HCO3 reabsorption was greater in distal tubules of alkalotic compared to control animals perfused with Schering 28080 to inhibit H(+)-K(+)-ATPase (-6.4 +/- 0.9 vs. -1.4 +/- 0.5 pmol/mm.min-1, P < 0.02) and with EIPA to inhibit Na+/H+ exchange (-11.1 +/- 1.7 vs. -6.6 +/- 0.9 pmol/mm.min-1, P < 0.01) but was similar in distal tubules of alkalotic and control animals perfused with bafilomycin to inhibit H(+)-ATPase. The greater reduction of distal tubule net HCO3 reabsorption in alkalotic compared to control animals induced by EIPA was eliminated by systemic infusion of the endothelin receptor antagonist bosentan (-4.6 +/- 0.7 vs. -4.4 +/- 0.7 pmol/mm.min-1, P = NS) but the greater reduction induced by Schering 28080 persisted. Urine endothelin-1 (ET-1) excretion was higher in animals with maintained alkalosis (164.5 +/- 23.7 vs. 76.6 +/- 10.8 fmol/day, P < 0.03), but decreased following KCl repletion to a value (86.7 +/- 10.0 fmol/day, P < 0.02 vs. respective before-KCl value) that was not different from that for KCl-repleted control animals (79.9 +/- 8.7 fmol/day, P = NS vs. KCl-repleted alkalotic animals). The data support that augmented distal tubule acidification in alkalotic animals is due to increased H(+)-K(+)-ATPase and Na+/H+ exchange activity, the latter stimulated by endogenous endothelins.     

Dissimilar alterations of sodium-coupled uptake by platinum-coordination complexes in renal proximal tubular cells in primary culture

The potent anticancer drug cis-diamminedichloroplatinum (II) (CDDP) interferes early with electrolyte transport by the renal proximal tubule. To study the early effects of platinum coordination complexes on apical Na(+)-coupled transport systems, we examined the effect of increasing concentrations of CDDP, trans-diamminedichloroplatinum (II) (TDDP) and cis-diammine-1,1-cyclobutane-dicarboxylate platinum (II) (CBDCA) on Na(+)-coupled uptake of P(i), methyl-alpha-D-glucopyranoside (MGP) and L-alanine by rabbit proximal tubule cells in primary culture. At 17 microM CDDP and 540 microM CBDCA, 1) cell viability (lactate dehydrogenase release) and ATP content were unaffected, 2) Na(+)-K(+)-ATPase activity was reduced by 40%, 3) Na(+)-coupled uptake of MGP and P(i) was reduced, whereas 4) Na(+)-coupled uptake of alanine rose to twice the control value. Alterations of Na(+)-coupled uptake of P(i), MGP and alanine were due to changes in Km, with no significant change in Vmax. At 333 microM TDDP, Na(+)-dependent P(i) and MGP uptake decreased, whereas Na(+)-independent uptake increased markedly and was associated with a decline in cell viability and ATP content. We conclude that 1) the TDDP-induced decrease in Na+/P(i) and Na+/glucose cotransport was associated with reduced cell viability, 2) both CDDP and CBDCA had different effects on Na+/P(i), Na+/glucose and Na+/alanine cotransport, arguing against an alteration of the Na+ gradient due to reduced Na(+)-K(+)-ATPase activity and 3) CBDCA induced alterations of Na(+)-coupled uptake similar to those of CDDP at concentrations 20 to 30 times higher.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of time in culture on modulation of Na(+)-K(+)-ATPase activity in rat alveolar type II cells

To determine the effect of time in culture on epithelial cell function, we evaluated the modulation of Na(+)-K(+)-ATPase activity in rat alveolar type II cells in culture. Ouabain sensitivity testing revealed that the alpha-1 predominance in the enzyme's isoforms was maintained over the 120 hours in culture. Basal Na(+)-K(+)-ATPase activity in the whole cell homogenate did not differ significantly between cells cultured for 48 hours and those cultured for 120 hours. Terbutaline (10 mM) did not activate Na(+)-K(+)-ATPase in the cells cultured for 48 hours, but, it significantly increased the activity of this enzyme in the cells cultured for 120 hours cells cultured for 48 hours, produced intracellular cyclic AMP after exposure to 10 mM of terbutaline. These results indicate that the coupling between Na(+)-K(+)-ATPase and the beta-adrenergic pathway in alveolar type II cells can be influenced by the time in cell culture.     

K+ fluxes mediated by Na(+)-K(+)-Cl- cotransport and Na(+)-K(+)-ATPase pumps in renal tubule cell lines transformed by wild-type and temperature-sensitive strains of Simian virus 40

The relative contributions of Na(+)-K(+)-ATPase pumps and Na(+)-K(+)-Cl- cotransport to total rubidium (Rb+) influx into primary cultures of renal tubule cells (PC.RC) and cells transformed either with the wild-type or a temperature-sensitive mutant of the simian virus 40 (SV40), were measured under various growth conditions. The Na(+)-K(+)-ATPase-mediated component represented 74% and 44-48% of total Rb+ influx into PC.RC and SV40-transformed cells, respectively. Proliferating transformed cells showed substantial ouabain-resistant bumetanide-sensitive (Or-Bs) Rb+ influx (41-45% of total) which indicated the presence of a Na(+)-K(+)-Cl- cotransport. The Or-Bs component of Rb+ influx was greatly reduced when temperature-sensitive transformed renal cells (RC.SVtsA58) grown in Petri dishes or on permeable filters were shifted from the permissive (33 degrees C) to the restrictive temperature (39.5 degrees C) to arrest cell growth. The ouabain-sensitive Rb+ influx mediated by the Na(+)-K(+)-ATPase, the total and amiloride-sensitive Na+ uptakes were not modified following inhibition of cell proliferation. A similar fall in the Or-Bs influx was obtained when renal tubule cells transformed by the wild-type SV40 (RC.SV) were incubated with the K+ channel blocker, tetraethylammonium (TEA) ion, which we had previously shown to arrest cell growth without affecting cell viability (Teulon et al.: J. Cell. Physiol., 151:113-125, 1992). Reinitiation of cell growth by removal of TEA or return to 33 degrees C of the temperature-sensitive cells restored the Or-Bs component of Rb influx. Taken together, these results indicate that the Na(+)-K(+)-Cl- cotransport activity is critically dependent on cell growth conditions.     

Coordinate interaction between ATP-sensitive K+ channel and Na+, K+-ATPase modulates ischemic preconditioning

BACKGROUND: We reported that digoxin abolishes the infarct size (IS)-limiting effect of ischemic preconditioning (IPC). Because ATP-sensitive K+ (KATP) channels are involved in IPC, we studied whether Na+,K+-ATPase and KATP channels functionally interact, thereby modulating IPC. METHODS AND RESULTS: Rabbits received 30 minutes of coronary artery occlusion followed by 3 hours of reperfusion. IPC was elicited by 5 minutes of occlusion followed by 10 minutes of reperfusion. The IS, expressed as a percentage of the area at risk, was 40.2+/-2.8% in control and 39.8+/-5.0% in digoxin pretreatment rabbits. Both IPC and pretreatment with cromakalim, a KATP channel opener, reduced IS to 11.8+/-1.8% and 13.4+/-2.6% (P<0. 05 versus control). Digoxin abolished the reduction in IS induced by IPC (33.5+/-3.3%), whereas it did not change that induced by cromakalim (18.8+/-3.0%). In patch-clamp experiments, digoxin was found to inhibit the opening of KATP channels in single ventricular myocytes in which ATP depletion had been induced by metabolic stress. In contrast, digoxin had little effect on the channel opening induced by cromakalim. Moreover, the inhibitory action of digoxin on channel activities was dependent on subsarcolemmal ATP concentration. CONCLUSIONS: The IS-limiting effect of IPC is modulated by an interaction between KATP channels and Na+,K+-ATPase through subsarcolemmal ATP.     

Short-term effect of epidermal growth factor (EGF) on sodium and glucose cotransport of isolated jejunal epithelial cells

This study was undertaken to assess the short-term effects of EGF on sodium and glucose uptake, glucose metabolism and Na+/K(+)-ATPase activity in isolated enterocytes of rats. Jejunal cells exposed to EGF had a significantly greater total uptake of sodium compared to controls after 6 min. Kinetic analysis of glucose transport across BBMV's demonstrated similar Km values but a significant increase of the Vmax in vesicles prepared from cells first exposed to EGF as compared to controls. EGF was also associated with a significant increase in glucose metabolism of jejunal enterocytes after 15 min. The activity of Na+/K(+)-ATPase increased in jejunal enterocytes exposed to EGF. The increase in Na+/K(+)-ATPase activity of the cells following EGF exposure was not accompanied by an increase in immunodetectable total or assembled Na+/K(+)-ATPase protein. EGF's effect on enzyme activity was abolished by removing NaCl from the incubation solution, and by preincubating the enterocytes with phlorizin prior to addition of EGF. Preincubation with amiloride did not inhibit the effect of EGF on Na+/K(+)-ATPase. The results confirm that EGF promotes uptake of both sodium and glucose by the jejunal mucosal cells, and suggest the effect of EGF on glucose and sodium is mediated through the brush-border membrane glucose-sodium transporter. The increase in Na+/K(+)-ATPase activity that occurs with EGF appears to be secondary to a rise in intracellular Na+ concentration. The short-term effects of EGF on glucose and sodium transport by the small intestine may have potential therapeutic implications.     

Energy transduction in the thermophilic anaerobic bacterium Clostridium fervidus is exclusively coupled to sodium ions

The thermophilic, peptidolytic, anaerobic bacterium Clostridium fervidus is unable to generate a pH gradient in the range of 5.5-8.0, which limits growth of the organism to a narrow pH range (6.3-7.7). A significant membrane potential (delta psi approximately -60 mV) and chemical gradient of Na+ (-Z delta pNa approximately -60 mV) are formed in the presence of metabolizable substrates. Energy-dependent Na+ efflux is inhibited by the Na+/H+ ionophore monensin but is stimulated by uncouplers, suggesting that the Na+ gradient is formed by a primary pumping mechanism rather than by secondary Na+/H+ antiport. This primary sodium pump was found to be an ATPase that has been characterized in inside-out membrane vesicles and in proteoliposomes in which solubilized ATPase was reconstituted. The enzyme is stimulated by Na+, resistant to vanadate, and sensitive to nitrate, which is indicative of an F/V-type Na(+)-ATPase. In the proteoliposomes Na+ accumulation depends on the presence of ATP, is inhibited by the ATPase inhibitor nitrate, and is completely prevented by the ionophore monensin but is stimulated by protonophores and valinomycin. These and previous observations, which indicated that secondary amino acid transport uses solely Na+ as coupling ion, demonstrate that energy transduction at the membrane in C. fervidus is exclusively dependent on a Na+ cycle.     

5-HPETE is a potent inhibitor of neuronal Na+, K(+)-ATPase activity

The effects of 1 microM concentrations of arachidonic acid hydroperoxide (HPETES) products of 5-, 12- and 15-lipoxygenase on Na+, K(+)-ATPase activity were investigated in synaptosomal membrane preparations from rat cerebral cortex. 5-HPETE inhibited Na+, K(+)-ATPase activity by up to 67 %. In contrast, 12-HPETE and 15-HPETE did not inhibit Na+, K(+)-ATPase activity. In addition, neither 5-HETE or LTA4 inhibited Na+, K(+)-ATPase activity. Dose-response studies indicated that 5-HPETE was a potent (IC25 = 10(-8) M) inhibitor of Na+, K(+)-ATPase activity. These findings indicate that 5-HPETE inhibits Na+, K(+)-ATPase activity by a mechanism that is dependent on the hydroperoxide position and independent of further metabolism by 5-lipoxygenase. It is proposed that 5-HPETE production by 5-lipoxygenase and subsequent inhibition of neuronal Na+, K(+)-ATPase activity may be a mechansim for modulating synaptic transmission.     

The Na(+)-K(+)-ATPase beta 1 subunit is associated with the HK alpha 2 protein in the rat kidney

The Na-K-ATPase beta 1 subunit acts as the beta subunit for the HK alpha 2 protein in the rat kidney. The colonic H(+)-K(+)-ATPase is a member of the P-type ATPases, and has been shown to contribute to potassium transport by the mammalian kidney and colon. The P-type ATPases often consist of an alpha subunit that contains the catalytic site and a beta subunit that participates in regulation of enzyme activity and targeting of the enzyme to the plasma membrane. The cDNA of the alpha subunit (HK alpha 2) has been cloned and the HK alpha 2 protein has been isolated from the rat kidney and colon. However, a unique beta subunit for the colonic H(+)-K(+)-ATPase has not been described. To determine if one of the known beta subunits present in the kidney might act as the beta subunit for the colonic H(+)-K(+)-ATPase, microsomes enriched in the colonic H(+)-K(+)-ATPase were isolated using an HK alpha 2-specific antibody (AS 31.7) and the Minimac magnetic separation system. Immunoblots of rat kidney microsomal protein isolated with antibody AS 31.7 were probed with antibodies directed against the gastric HK beta subunit, Na(+)-K(+)-ATPase alpha 1, and Na(+)-K(+)-ATPase beta 1 subunits. A band of the appropriate size was detected with Na(+)-K(+)-ATPase beta 1-specific antibodies, but not those directed against HK beta 1. These data suggest that Na(+)-K(+)-ATPase beta 1 could be the beta subunit for the colonic H(+)-K(+)-ATPase in the kidney.     

A protocol to improve analgesia use in the accident and emergency department

In this experiment, intracellular K+ concentration ([K+]i) and ATPase activity of myocardiocytes were measured in early stage of burn injury. Comparing with control group, it was found that, 1. [K+]i were decreased after burn injury, [K+]i of 1st, 3rd, 8th and 24th hours were decreased to 96.2 +/- 1.3%, 85.8 +/- 1.3%, 65.9 +/- 1.0% and 73.7 +/- 1.1% of normal, respectively. 2. Cardiac sarcolemma total ATPase, Mg(2+)-ATPase and Na(+)-K(+)-ATPase activities were all reduced significantly at 8th hour after injury. These results suggest that, burn injury accelerates K+ efflux current, but inhibits K+ influx current, and the reduction of Na(+)-K(+)-ATPase activity is one reason of decrease of [K+]i after injury.     

Peripheral nerve blocks improve analgesia after total knee replacement surgery

Using an in vitro cell system and Cs+ NMR techniques we were able to show that porcine aortic endothelial cells (PAEC) reduce their Na(+)-K(+)-ATPase activity upon an increase in intracellular cAMP. Reduction in the pump rate was due to phosphorylation of the alpha-subunit of the ATPase as shown by immunoprecipitation. Apart from a pump inhibiton using 8-Br-cAMP and IBMX, we were also able to show that changes in the Na(+)-K(+)-ATPase activity could be mediated by the adenosine-A2 and prostaglandin receptor agonists 5'-N-Ethylcarboxamidoadenosine and Iloprost, respectively. Parallel to a decrease in pump activity we also observed a decrease in intracellular Cs+, indicating opening of K+ channels.     

Electrogenic property of Na+, K(+)-ATPase through computer simulation

A computer simulation of the electrogenic nature of the membrane-bound Na+, K(+)-ATPase is presented. The model involves coupling two simulation systems for passive and active transports, using a minimum of empirical parameters, and studies the contribution of the pump to the membrane potential. The simulation results indicate that electrogenic active transport accelerates the restoration of the resting electrochemical gradients and contributes approximately 0.44-1.1 mV to the resting potential of the membrane, depending on the Na:K coupling ratio. The effect of membrane potential and the physical positioning of the enzyme from the passive transporting channel on the enzyme function is also presented. The validity of the model is checked by comparing our results with reported literature values.     

Antinociceptive effects of SC-39566, an opioid dipeptide arylalkylamide, in the Rhesus monkey

We stably expressed the rat D1A dopamine receptor in mouse fibroblast LTK- cells and obtained specific ligand binding and functional activity characteristic of the D1A dopamine receptor coupled to stimulation of adenylyl cyclase. In the transfected cells, the selective D1 agonist fenoldopam caused a concentration-dependent inhibition of Na+/K(+)-ATPase activity, achieving maximum inhibition of approximately 30%. The latter was abolished by the selective D1 antagonist (+)-SCH 23390 and by the specific protein kinase A inhibitor protein kinase inhibitor-(6-22) amide. In the nontransfected cells, fenoldopam did not affect Na+/K(+)-ATPase activity. 8-Chlorophenylthio-cAMP inhibited Na+/K(+)-ATPase activity in both transfected and nontransfected cells; this effect was blocked by protein kinase inhibitor-(6-22). These results indicate that the inhibition of Na+/K(+)-ATPase activity induced by agonist occupancy of D1A receptors is mediated by protein kinase A.