mRNA Levels Of Na Research Articles

Autophagy activation, histopathological damage, and altered renal epithelial sodium channel and Na+,K+‐ATPase gene expression in offspring kidney after in utero exposure to allethrin

The goal of the current study was to determine the effects of the allethrin, as a pyrethroid-based insecticide, on kidney function in rat offspring. Sixty-day-old Wistar albino female rats were randomly allocated to three groups: control group and two allethrin-treated groups that were intragastrically administered with 34.2 or 68.5 mg/kg b.w. of allethrin starting from the day 6 of pregnancy until delivery. The results revealed that the high dose of allethrin (68.5 mg/kg) reduced the number of glomeruli. Results showed that the glomeruli diameter in animals treated with 34.2 mg/kg of allethrin significantly decreased when compared to the controls (p < 0.001) whereas at the dose of 68.5 mg/kg the glomeruli diameter was significantly higher than those of the control group (p < 0.05). Interestingly, the expression of the autophagic markers Beclin-1 and LC-3 was increased when compared to control indicating an induction of the autophagic mechanism. Furthermore, allethrin administration downregulated in a dose-dependent manner the expression of the epithelial sodium channel (ENaC) gene. The mRNA levels of Na+/K+- ATPase gene were significantly higher in the group treated with the low dose and significantly lower with the high dose, when compared to control. However, there was no effect of allethrin on the gene expression of NHE3 in both treated groups. Taken together, the findings of this study suggested that the exposure to allethrin might contribute to the nephrotoxicity by affecting the number and the diameter of glomeruli, causing histopathological alterations and inducing autophagic-related markers and affecting the mRNA levels of ENaC and Na+/ K + ATPase genes.

Developmental differences in myocardial transmembrane Na+ transport: implications for excitability and Na+ handling.

Little is currently known about possible developmental changes in myocardial Na+ handling, which may have impact on cell excitability and Ca2+ content. Resting intracellular Na+ concentration ([Na+ ]i ), measured in freshly isolated rat ventricular myocytes with CoroNa green, was not significantly different in neonates (3-5days old) and adults, but electrical stimulation caused marked [Na+ ]i rise only in neonates. Inhibition of L-type Ca2+ current by CdCl2 abolished not only systolic Ca2+ transients, but also activity-dependent intracellular Na+ accumulation in immature cells. This indicates that the main Na+ influx pathway during activity is the Na+ /Ca2+ exchanger, rather than voltage-dependent Na+ current (INa ), which was not affected by CdCl2 . In immature myocytes, INa density was two-fold greater, inactivation was faster, and the current peak occurred at less negative transmembrane potential (Em ) than in adults. Na+ channel steady-state activation and inactivation curves in neonates showed a rightward shift, which should increase channel availability at diastolic Em , but also require greater depolarization for excitation, which was observed experimentally and reproduced in computer simulations. Ventricular mRNA levels of Nav 1.1, Nav 1.4 and Nav 1.5 pore-forming isoforms were greater in neonate ventricles, while a decrease was seen for the β1subunit. Both molecular and biophysical changes in the channel profile may contribute to the differences in INa density and voltage-dependence, and also to the less negative threshold Em , in neonates compared to adults. The apparently lower excitability in immature ventricle may confer protection against the development of spontaneous activity in this tissue. KEY POINTS: Previous studies showed that myocardial preparations from immature rats are less sensitive to electrical field stimulation than adult preparations. Freshly isolated ventricular myocytes from neonatal rats showed lower excitability than adult cells, e.g. less negative threshold membrane potential and greater membrane depolarization required for action potential triggering. In addition to differences in mRNA levels for Na+ channel isoforms and greater Na+ current (INa ) density, Na+ channel voltage-dependence was shifted to the right in immature myocytes, which seems to be sufficient to decrease excitability, according to computer simulations. Only in neonatal myocytes did cyclic activity promote marked cytosolic Na+ accumulation, which was prevented by abolition of systolic Ca2+ transients by blockade of Ca2+ currents. Developmental changes in INa may account for the difference in action potential initiation parameters, but not for cytosolic Na+ accumulation, which seems to be due mainly to Na+ /Ca2+ exchanger-mediated Na+ influx.

Swimming training promotes cardiac remodeling and alters the expression of mRNA and protein levels involved in calcium handling in hypertensive rats

AimThe aim of this study was to identify the effects of swimming training on the mRNA expression and protein levels of the calcium handling proteins in the hearts of renovascular hypertensive rats submitted to swimming protocol during 6weeks. Main methodsFischer rats with renovascular hypertension 2-kidney 1-clip (2K1C) and SHAM groups were divided among sedentary and exercised groups. The exercise protocol lasted for 6weeks (1h/day, 5×/week), and the mean arterial pressure, cardiomyocytes hypertrophy parameters, mRNA expression and protein levels of some calcium handling proteins in the left ventricle were evaluated. Key findingsSwimming training was able to reduce the levels of mean arterial pressure in the hypertensive group compared to 2K1C SED, and to promote cardiac hypertrophy in SHAM EX and 2K1C EX groups in comparison to the respective control groups. The mRNA levels of B-type natriuretic peptide were reduced in the 2K1C EX when compared to 2K1C SED. The mRNA and protein levels of the sarcoplasmic reticulum Ca2+-ATPase increased after the swimming training in SHAM and 2K1C groups. The mRNA and protein levels of phospholamban, displayed an increase in their levels in the exercised SHAM and in hypertensive rats in comparison to their respective controls; while mRNA levels of Na+/Ca2+ exchanger was reduced in the left ventricle comparing to the sedentary hypertensive rats. SignificanceTaken altogether, we provide evidence that the aerobic training may lead to cardiac remodeling, and modulate the calcium handling proteins expression in the heart of hypertensive rats.

OP0175 Monocarboxylate Transporter (MCT)-4, Associated with the Decrease of Synovial Fluid Ph, is A Novel Therapeutic Target of Rheumatoid Arthritis

BackgroundIt is well known that synovial fluid pH is decreased in rheumatoid arthritis (RA) patients (1), but the meaning and mechanism remain unclear.ObjectivesWe investigate the correlation between synovial fluid pH...

Supplementation with branched-chain amino acids to a low-protein diet regulates intestinal expression of amino acid and peptide transporters in weanling pigs

This study determined the effects of dietary branched-chain amino acids (AA) (BCAA) on growth performance, expression of jejunal AA and peptide transporters, and the colonic microflora of weanling piglets fed a low-protein (LP) diet. One hundred and eight Large White × Landrace × Duroc piglets (weaned at 28 days of age) were fed a normal protein diet (NP, 20.9 % crude protein), an LP diet (LP, 17.1 % crude protein), or an LP diet supplemented with BCAA (LP + BCAA, 17.9 % crude protein) for 14 days. Dietary protein restriction reduced piglet growth performance and small-intestinal villous height, which were restored by BCAA supplementation to the LP diet to values for the NP diet. Serum concentrations of BCAA were reduced in piglets fed the LP diet while those in piglets fed the LP + BCAA diet were similar to values for the NP group. mRNA levels for Na(+)-neutral AA exchanger-2, cationic AA transporter-1, b(0,+) AA transporter, and 4F2 heavy chain were more abundant in piglets fed the LP + BCAA diet than the LP diet. However, mRNA and protein levels for peptide transporter-1 were lower in piglets fed the LP + BCAA diet as compared to the LP diet. The colonic microflora did not differ among the three groups of pigs. In conclusion, growth performance, intestinal development, and intestinal expression of AA transporters in weanling piglets are enhanced by BCAA supplementation to LP diets. Our findings provide a new molecular basis for further understanding of BCAA as functional AA in animal nutrition.

Voltage-gated sodium channel expression and action potential generation in differentiated NG108-15 cells

BackgroundThe generation of action potential is required for stimulus-evoked neurotransmitter release in most neurons. Although various voltage-gated ion channels are involved in action potential production, the initiation of the action potential is mainly mediated by voltage-gated Na+ channels. In the present study, differentiation-induced changes of mRNA and protein expression of Na+ channels, Na+ currents, and cell membrane excitability were investigated in NG108-15 cells.ResultsWhole-cell patch-clamp results showed that differentiation (9 days) didn’t change cell membrane excitability, compared to undifferentiated state. But differentiation (21 days) induced the action potential generation in 45.5% of NG108-15 cells (25/55 cells). In 9-day-differentiated cells, Na+ currents were mildly increased, which was also found in 21-day differentiated cells without action potential. In 21-day differentiated cells with action potential, Na+ currents were significantly enhanced. Western blot data showed that the expression of Na+ channels was increased with differentiated-time dependent manner. Single-cell real-time PCR data demonstrated that the expression of Na+ channel mRNA was increased by 21 days of differentiation in NG108-15 cells. More importantly, the mRNA level of Na+ channels in cells with action potential was higher than that in cells without action potential.ConclusionDifferentiation induces expression of voltage-gated Na+ channels and action potential generation in NG108-15 cells. A high level of the Na+ channel density is required for differentiation-triggered action potential generation.

EFFECTS OF HIGH SALT DIET ON ARTERIAL REMODELLING AND THE INTERVENTION OF TELMISARTAN IN WISTAR RATS

ObjectivesTo investigate the effects of dietary salt on aorta and mesenteric artery remodelling in Wistar Rats and explore the possible mechanism of salt-induced arterial remodelling and AngiotensinII receptor blockers telmisartan...

Overexpression of NaV1.6 channels is associated with the invasion capacity of human cervical cancer

Functional activity of voltage-gated sodium channels (VGSC) has been associated to the invasion and metastasis behaviors of prostate, breast and some other types of cancer. We previously reported the functional expression of VGSC in primary cultures and biopsies derived from cervical cancer (CaC). Here, we investigate the relative expression levels of VGSC subunits and its possible role in CaC. Quantitative real-time PCR revealed that mRNA levels of Na(V) 1.6 α-subunit in CaC samples were ∼40-fold higher than in noncancerous cervical (NCC) biopsies. A Na(V) 1.7 α-subunit variant also showed increased mRNA levels in CaC (∼20-fold). All four Na(V) β subunits were also detected in CaC samples, being Na(V) β1 the most abundant. Proteins of Na(V) 1.6 and Na(V) 1.7 α-subunits were immunolocalized in both NCC and CaC biopsies and in CaC primary cultures as well; however, although in NCC sections proteins were mainly relegated to the plasma membrane, in CaC biopsies and primary cultures the respective signal was stronger and widely distributed in both cytoplasm and plasma membrane. Functional activity of Na(V) 1.6 channels in the plasma membrane of CaC cells was confirmed by whole-cell patch-clamp experiments using Cn2, a Na(V) 1.6-specific toxin, which blocked ∼30% of the total sodium current. Blocking of sodium channels VGSC with tetrodotoxin and Cn2 did not affect proliferation neither migration, but reduced by ∼20% the invasiveness of CaC primary culture cells in vitro assays. We conclude that Na(V) 1.6 is upregulated in CaC and could serve as a novel molecular marker for the metastatic behavior of this carcinoma.

Changes in gill H+-ATPase and Na+/K+-ATPase expression and activity during freshwater acclimation of Atlantic salmon (Salmo salar)

Few studies have examined changes in salmon gill ion transporter expression during the transition from seawater to freshwater, a pivotal moment in the salmonid life cycle. Seawater-acclimated Atlantic salmon were transferred to freshwater and blood and gill tissue were sampled over 30 days of acclimation. Salmon held in seawater had stable plasma osmolality and sodium and chloride levels throughout the experiment. Following freshwater exposure, plasma sodium and chloride levels and total osmolality decreased significantly before returning towards control levels over time. Gill H(+)-ATPase activity increased by more than 45% 14 days after exposure to freshwater, whereas H(+)-ATPase mRNA levels were not affected by the salinity change. Within 4 days of freshwater exposure, gill Na(+)/K(+)-ATPase activity increased ∼43% over control levels, remaining significantly higher until the 30 day sampling group when it declined back to control levels. This increase in activity was associated with a more than 7-fold increase in Na(+)/K(+)-ATPase isoform α1a mRNA level and a ∼60% decrease in Na(+)/K(+)-ATPase isoform β1b mRNA level. The mRNA levels of Na(+)/K(+)-ATPase isoforms α1c and α3 did not change as a result of freshwater exposure. The time courses for mRNA expression of the small membrane protein FXYD 11 and the β1-subunit were very similar, with levels increasing significantly 7 days following freshwater exposure before subsiding back to control levels at 30 days. Taken together, these data suggest an important role for Na(+)/K(+)-ATPase in freshwater acclimation in Atlantic salmon.

β-Site APP-cleaving enzyme 1 (BACE1) cleaves cerebellar Na+ channel β4-subunit and promotes Purkinje cell firing by slowing the decay of resurgent Na+ current

In cerebellar Purkinje cells, the β4-subunit of voltage-dependent Na(+) channels has been proposed to serve as an open-channel blocker giving rise to a "resurgent" Na(+) current (I (NaR)) upon membrane repolarization. Notably, the β4-subunit was recently identified as a novel substrate of the β-secretase, BACE1, a key enzyme of the amyloidogenic pathway in Alzheimer's disease. Here, we asked whether BACE1-mediated cleavage of β4-subunit has an impact on I (NaR) and, consequently, on the firing properties of Purkinje cells. In cerebellar tissue of BACE1-/- mice, mRNA levels of Na(+) channel α-subunits 1.1, 1.2, and 1.6 and of β-subunits 1-4 remained unchanged, but processing of β4 peptide was profoundly altered. Patch-clamp recordings from acutely isolated Purkinje cells of BACE1-/- and WT mice did not reveal any differences in steady-state properties and in current densities of transient, persistent, and resurgent Na(+) currents. However, I (NaR) was found to decay significantly faster in BACE1-deficient Purkinje cells than in WT cells. In modeling studies, the altered time course of I (NaR) decay could be replicated when we decreased the efficiency of open-channel block. In current-clamp recordings, BACE1-/- Purkinje cells displayed lower spontaneous firing rate than normal cells. Computer simulations supported the hypothesis that the accelerated decay kinetics of I (NaR) are responsible for the slower firing rate. Our study elucidates a novel function of BACE1 in the regulation of neuronal excitability that serves to tune the firing pattern of Purkinje cells and presumably other neurons endowed with I (NaR).

E0037 Effects of enalapril and irbesartan on aorta remodelling and ion pumps in renovascular hypertensive rats

ObjectiveTo investigate the effects of single-drug or combination therapy of enalapril and irbesartan on aorta remodelling and its mechanisms.MethodsRenovascular hypertensive rats (RHD) induced by two-kidney one-clip method were treated with...

E0121 The effects of endothein-1 and BQ-123 on ATPase activity and mRNA expression in aortic smooth muscle cells from spontanously hypertensive rats

AimTo study the effects of endothelin-1 (ET-1) and BQ-123 (ETA receptor antagonist) on activities and mRNA expression of ATPase in aortic smooth muscle cells (ASMCs) from spontaneously hypertensive rats (SHR)...

Bepridil up‐regulates cardiac Na+ channels as a long‐term effect by blunting proteasome signals through inhibition of calmodulin activity

Bepridil is an anti-arrhythmic agent with anti-electrical remodelling effects that target many cardiac ion channels, including the voltage-gated Na+ channel. However, long-term effects of bepridil on the Na+ channel remain unclear. We explored the long-term effect of bepridil on the Na+ channel in isolated neonatal rat cardiomyocytes and in a heterologous expression system of human Na(v)1.5 channel. Na+ currents were recorded by whole-cell voltage-clamp technique. Na+ channel message and protein were evaluated by real-time RT-PCR and Western blot analysis. Treatment of cardiomyocytes with 10 micromol.L(-1) bepridil for 24 h augmented Na+ channel current (I(Na)) in a dose- and time-dependent manner. This long-term effect of bepridil was mimicked or masked by application of W-7, a calmodulin inhibitor, but not KN93 [2-[N-(2-hydroxyethyl)-N-(4-methoxy benzenesulphonyl)]-amino-N-(4-chlorocinnamyl)-N-methylbenzylamine], a Ca2+/calmodulin-dependent kinase inhibitor. During inhibition of protein synthesis by cycloheximide, the I(Na) increase due to bepridil was larger than the increase without cycloheximide. Bepridil and W-7 significantly slowed the time course of Na(v)1.5 protein degradation in neonatal cardiomyocytes, although the mRNA levels of Na(v)1.5 were not modified. Bepridil and W-7 did not increase I(Na) any further in the presence of the proteasome inhibitor MG132 [N-[(phenylmethoxy)carbonyl]-L-leucyl-N-[(1S)-1-formyl-3-methylbutyl]-L-leucinamide]. Bepridil, W-7 and MG132 but not KN93 significantly decreased 20S proteasome activity in a concentration-dependent manner. We conclude that long-term exposure of cardiomyocytes to bepridil at therapeutic concentrations inhibits calmodulin action, which decreased degradation of the Na(v)1.5 alpha-subunit, which in turn increased Na+ current.

Both enalapril and losartan attenuate sarcolemmal Na+-K+-ATPase remodeling in failing rat heart due to myocardial infarctionThis article is one of a selection of papers published in the special issue Bridging the Gap: Where Progress in Cardiovascular and Neurophysiologic Research Meet.

To investigate the mechanisms underlying the depressed sarcolemmal (SL) Na(+)-K(+)-ATPase activity in congestive heart failure (CHF), different isoforms and gene expression of Na(+)-K(+)-ATPase were examined in the failing left ventricle (LV) at 8 weeks after myocardial infarction (MI). In view of the increased activity of renin-angiotensin system (RAS) in CHF, these parameters were also studied after 5 weeks of treatment with enalapril (10 mg x kg-1 x day-1), an angiotensin-converting enzyme inhibitor, and losartan (20 mg.kg-1.day-1), an angiotensin II type 1 receptor antagonist, starting at 3 weeks after the coronary ligation in rats. The infarcted animals showed LV dysfunction and depressed SL Na(+)-K(+)-ATPase activity. Protein content and mRNA levels for Na(+)-K(+)-ATPase alpha2 isoform were decreased whereas those for Na(+)-K(+)-ATPase alpha3 isoform were increased in the failing LV. On the other hand, no significant changes were observed in protein content or mRNA levels for Na(+)-K(+)-ATPase alpha1 and beta1 isoforms. The treatment of infarcted animals with enalapril or losartan improved LV function and attenuated the depression in Na(+)-K(+)-ATPase alpha2 isoform as well as the increase in alpha3 isoform, at both the protein and mRNA levels; however, combination therapy with enalapril and losartan did not produce any additive effects. These results provide further evidence that CHF due to MI is associated with remodeling of SL membrane and suggest that the blockade of RAS plays an important role in preventing these alterations in the failing heart.

Hypoxia-induced changes in the expression of rat hepatobiliary transporter genes

Cholestatic disorders may arise from liver ischemia (e.g., in liver transplantation) through various mechanisms. We have examined the potential of hypoxia to induce changes in the expression of hepatobiliary transporter genes. In a model of arterial liver ischemia subsequent to complete arterial deprivation of the rat liver, the mRNA levels of VEGF, a hypoxia-inducible gene, were increased fivefold after 24 h. The pattern of VEGF-induced expression and ultrastructural changes, including swelling of the endoplasmic reticulum, indicated that hypoxia affected primarily cholangiocytes, but also hepatocytes, predominantly in the periportal area. Serum and bile analyses demonstrated liver dysfunction of cholestatic type with reduced bile acid biliary excretion. Fluorescence-labeled ursodeoxycholic acid used as a tracer displayed no regurgitation, eliminating bile leakage as a significant mechanism of cholestasis in this model. In liver tissue, a marked reduction in the mRNA levels of Na(+)-taurocholate-cotransporting polypeptide (Ntcp), bile salt export protein (Bsep), and multidrug resistance-associated protein 2 (Mrp2) and an increase in those of Cftr were detected before bile duct proliferation occurred. In cultured hepatocytes, a nontoxic hypoxic treatment caused a decrease in the mRNA and protein expression of Ntcp, Bsep, and Mrp2 and in the mRNA levels of nuclear factors involved in the transactivation of these genes, i.e., HNF4alpha, RXRalpha, and FXR. In bile duct preparations, hypoxic treatment elicited an increase in Cftr transcripts, along with a rise in cAMP, a major regulator of Cftr expression and function. In conclusion, hypoxia triggers a downregulation of hepatocellular transporters, which may contribute to cholestasis, whereas Cftr, which drives secretion in cholangiocytes, is upregulated.

Modification of sarcolemmal Na+-K+-ATPase and Na+/Ca2+exchanger expression in heart failure by blockade of renin-angiotensin system

The activities of both sarcolemmal (SL) Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchanger, which maintain the intracellular cation homeostasis, have been shown to be depressed in heart failure due to myocardial infarction (MI). Because the renin-angiotensin system (RAS) is activated in heart failure, this study tested the hypothesis that attenuation of cardiac SL changes in congestive heart failure (CHF) by angiotensin-converting enzyme (ACE) inhibitors is associated with prevention of alterations in gene expression for SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchanger. CHF in rats due to MI was induced by occluding the coronary artery, and 3 wk later the animals were treated with an ACE inhibitor, imidapril (1 mg.kg(-1).day(-1)), for 4 wk. Heart dysfunction and cardiac hypertrophy in the infarcted animals were associated with depressed SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchange activities. Protein content and mRNA levels for Na(+)/Ca(2+) exchanger as well as Na(+)-K(+)-ATPase alpha(1)-, alpha(2)- and beta(1)-isoforms were depressed, whereas those for alpha(3)-isoform were increased in the failing heart. These changes in SL activities, protein content, and gene expression were attenuated by treating the infarcted animals with imidapril. The beneficial effects of imidapril treatment on heart function and cardiac hypertrophy as well as SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchange activities in the infarcted animals were simulated by enalapril, an ACE inhibitor, and losartan, an angiotensin receptor antagonist. These results suggest that blockade of RAS in CHF improves SL Na(+)-K(+)-ATPase and Na(+)/Ca(2+) exchange activities in the failing heart by preventing changes in gene expression for SL proteins.

Preconditioning attenuates ischemia-reperfusion-induced remodeling of Na+-K+-ATPase in hearts.

The aim of this study was to determine whether changes in protein content and/or gene expression of Na+-K+-ATPase subunits underlie its decreased enzyme activity during ischemia and reperfusion. We measured protein and mRNA subunit levels in isolated rat hearts subjected to 30 min of ischemia and 30 min of reperfusion (I/R). The effect of ischemic preconditioning (IP), induced by three cycles of ischemia and reperfusion (10 min each), was also assessed on the molecular changes in Na+-K+-ATPase subunit composition due to I/R. I/R reduced the protein levels of the alpha2-, alpha3-, beta1-, and beta2-isoforms by 71%, 85%, 27%, and 65%, respectively, whereas the alpha1-isoform was decreased by <15%. A similar reduction in mRNA levels also occurred for the isoforms of Na+-K+-ATPase. IP attenuated the reduction in protein levels of Na+-K+-ATPase alpha2-, alpha3-, and beta2-isoforms induced by I/R, without affecting the alpha1- and beta1-isoforms. Furthermore, IP prevented the reduction in mRNA levels of Na+-K+-ATPase alpha2-, alpha3-, and beta1-isoforms following I/R. Similar alterations in protein contents and mRNA levels for the Na+/Ca2+ exchanger were seen due to I/R as well as IP. These findings indicate that remodeling of Na+-K+-ATPase may occur because of I/R injury, and this may partly explain the reduction in enzyme activity in ischemic heart disease. Furthermore, IP may produce beneficial effects by attenuating the remodeling of Na+-K+-ATPase and changes in Na+/Ca2+ exchanger in hearts after I/R.

Differential effects of short and prolonged exposure to carvedilol on voltage-dependent Na(+) channels in cultured bovine adrenal medullary cells.

We examined the effects of short and prolonged exposure to carvedilol, an antihypertensive and beta-adrenoceptor blocking drug, on voltage-dependent Na(+) channels in cultured bovine adrenal medullary cells. Carvedilol (1-100 microM) reduced (22)Na(+) influx induced by veratridine, an activator of voltage-dependent Na(+) channels. Carvedilol also suppressed veratridine-induced (45)Ca(2+) influx and catecholamine secretion in a concentration-dependent manner similar to that of (22)Na(+) influx. Prolonged exposure of the cells to 10 microM carvedilol increased [(3)H]saxitoxin ([(3)H]STX) binding, which reached a plateau at 12 h and was still observed at 48 to 72 h. Scatchard analysis of [(3)H]STX binding revealed that carvedilol increased the B(max) value (control, 14.9 +/- 0.9 fmol/10(6) cells; carvedilol, 23.8 +/- 1.2 fmol/10(6) cells) (n = 3, P < 0.05) without altering the K(d) value, suggesting a rise in the number of cell surface Na(+) channels. The increase in [(3)H]STX binding by carvedilol was prevented by cycloheximide, an inhibitor of protein synthesis, whereas carvedilol changed neither alpha- nor beta(1)-subunit mRNA levels of Na(+) channels. The carvedilol-induced increase of [(3)H]STX binding was abolished by brefeldin A and H-89, inhibitors of intracellular vesicular trafficking of proteins from the trans-Golgi network and of cyclic AMP-dependent protein kinase (protein kinase A), respectively. The present findings suggest that short-term treatment with carvedilol reduces the activity of Na(+) channels, whereas prolonged exposure to carvedilol up-regulates cell surface Na(+) channels. This may add new pharmacological effects of carvedilol to our understanding in the treatment of heart failure and hypertension.

Seawater tolerance and gene expression in two strains of Atlantic salmon smolts

The seawater tolerance of Atlantic salmon (Salmo salar) smolts reared under identical hatchery conditions was assessed in two Norwegian strains: AquaGen and Imsa. Plasma ion levels were disrupted in both strains following seawater exposure, but these disruptions were more profound in the AquaGen fish. To investigate the mechanisms underlying these differences, we measured gill Na+,K+-adenosine triphosphatase (ATPase) activity and mRNA levels of Na+,K+-ATPase α-subunit and two isoforms of the cystic fibrosis transmembrane conductance regulator (CFTR). Gill Na+,K+-ATPase activity rose significantly in both strains following seawater exposure. Both Na+,K+-ATPase α-subunit and CFTR I mRNA levels were significantly elevated for the entire 2-week period following seawater exposure, whereas CFTR II levels were transiently elevated during the first 24 h only. There were no differences in enzyme activity or gene expression between strains, with the exception of CFTR II, which was significantly lower in the Imsa strain 2 weeks following seawater exposure. This suggests that although changes in mRNA and protein expression for these genes are associated with seawater transfer, they are not the basis of observed physiological differences between strains.

Sepsis increases the plasma membrane content of alpha1 and alpha2 isoforms of Na+-K+ adenosine triphosphatase in rat skeletal muscle.

Increased Na(+)-K(+) adenosine triphosphatase (ATPase) activity in skeletal muscle during sepsis is caused by transient increases in enzyme content within the plasma membrane. Randomized controlled study. University laboratory. Eighty-eight adult male Wistar rats were randomly assigned to undergo cecal ligation and puncture (CLP) or sham operation. Gastrocnemius muscles were harvested 6, 12, 24, and 48 hours after operation and Na(+)-K(+) ATPase activities were measured spectrofluorimetrically. Messenger RNA (mRNA) levels for the alpha1 and alpha2 isoforms of Na(+)-K(+) ATPase were determined by Northern blot analysis. Crude membranes, internal membranes, and purified plasma membranes were isolated from gastrocnemius muscles and protein levels of alpha1 and alpha2 isoforms were determined by Western blot analysis. Na(+)-K(+) ATPase activity in the CLP group was significantly higher compared with the sham group 24 hours after operation (P<.05). However, there were no differences between the sham and CLP groups 6, 12, or 48 hours after operation. No significant differences between the CLP and sham groups were noted in mRNA levels for Na(+)-K(+) ATPase alpha1 and alpha2 isoforms. Western blot analysis revealed that the plasma membrane (but not internal membrane or crude membrane) content of alpha2 and alpha1 isoforms from the CLP group was significantly increased compared with the sham group 24 hours after operation (P<.05). Na(+)-K(+) ATPase activity increases 24 hours after CLP in gastrocnemius muscle and then declines. This increase is caused by increased Na(+)-K(+) ATPase protein levels in the plasma membrane.