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Arterial pressure in most experimental and clinical hypertensions is exacerbated by salt. The effects of salt excess on right and left ventricular (RV and LV, respectively) functions and their respective coronary vasodilatory responses have been less explored. We therefore examined the effects of 8 wk of NaCl excess (8% in food) on arterial pressure, RV and LV functions (maximal rate of increase and decrease of ventricular pressure; dP/[dt.sub.max] and dP/[dt.sub.min]), coronary hemodynamics (microspheres), and collagen content (hydroxyproline assay and collagen volume fraction) in young adult normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR), aged 16 wk by the end of the study. Prolonged salt excess in WKY and SHR elevated pressure only modestly, but it markedly increased LV mass, especially in SHR. Moreover, salt excess significantly impaired RV and LV diastolic function in SHR but only LV diastolic function in WKY rats. However, salt loading affected neither RV nor LV contractile function in both strains. Interstitial and perivascular collagen deposition was increased, whereas coronary vasodilatory responses to dipyridamole diminished in both ventricles in the salt-loaded SHR but not in WKY rats. Therefore, accumulation of ventricular collagen as well as altered myocardial perfusion importantly contributed to the development of salt-related RV and LV dysfunctions in this model of naturally occurring hypertension. The unique effects of salt loading on both ventricles in SHR, but not WKY rats, strongly suggest that nonhemodynamic mechanisms in hypertensive disease participate pathophysiologically with salt-loading hypertension. These findings point to the conclusion that the concept of 'salt sensitivity' in hypertension is far more complex than simply its effects on arterial pressure or the LV. sodium excess: left and right ventricular function: spontaneously hypertensive rats; Wistar-Kyoto rats

Endothelial nitric oxide synthase-deficient mice exhibit increased susceptibility to endotoxin-induced acute renal failure. Am J Physiol Renal Physiol 287: F1044-F1048, 2004; doi:10.1152/ajprenal.00136.2004.--Acute renal failure (ARF) in septic patients drastically increases the mortality to 50-80%. Nitric oxide (NO) has been shown to be increased in sepsis. Endothelial nitric oxide synthase (eNOS) is one of the major regulators of arterial blood pressure and regional blood flow; however, its in vivo role in septic ARF is still unclear. We hypothesized that eNOS affords a protective effect against the renal vasoconstriction during endotoxemia. Because there are no specific inhibitors for eNOS, the study was therefore undertaken in eNOS knockout mice. There was no significant difference in baseline glomerular filtration rate (GFR) between the wild-type mice and the eNOS knockout mice (140 [+ or -] 10 vs. 157 [+ or -] 18 [micro]l/min, n = 9, P = not significant). However, renal blood flow (RBF) was significantly decreased in eNOS knockout mice compared with the wild-type controls (0.62 [+ or -] 0.05 ml/min, n = 6 vs. 0.98 [+ or -] 0.13 ml/min, n = 8, P < 0.05). Mean arterial pressure (MAP) was significantly higher in eNOS knockout mice than the wild-type controls (109 [+ or -] 5 vs. 80 [+ or -] 1 mmHg, n = 10, P < 0.01). Thus renal vascular resistance (RVR) was much higher in eNOS knockout mice than in the wild-type mice (176 [+ or -] 2, n = 6 vs. 82 [+ or -] 1 mmHg*[ml.sup.-1]*[min.sup.-1], n = 8, P < 0.01). When 1.0 mg/kg LPS was injected, there was no change in MAP in either the wild-type (84 [+ or -] 3 mmHg, n = 10) or the eNOS knockout mice (105 [+ or -] 5 mmHg, n = 10). Although GFR (154 [+ or -] 22 [micro]l/min, n = 8) and RBF (1.19 [+ or -] 0.05 ml/min, n = 9) remained unchanged with the 1.0-mg/kg dose of LPS in the wild-type mice, GFR (83 [+ or -] 18 vs. 140 [+ or -] 10 [micro]l/min, n = 6, P < 0.01) and RBF (0.36 [+ or -] 0.04 vs. 0.62 [+ or -] 0.05 ml/min, n = 6, P < 0.01 ) decreased significantly in the eNOS knockout mice. Fractional excretion of sodium increased significantly in eNOS knockout mice during endotoxemia (3.61 [+ or -] 0.78, n = 7 vs. 0.95 [+ or -] 0.14, n = 6, P < 0.01), whereas it remained unchanged in the wild-type mice (0.59 [+ or -] 0.16, n = 9 vs. 0.42 [+ or -] 0.05, n = 6, P = not significant). In summary, eNOS knockout mice have increased RVR and are more susceptible to endotoxemic ARF than wild-type mice despite higher MAP. tractional excretion of sodium; vasoconstriction; renal vascular resistance

Edelman et al. have empirically shown that plasma water sodium concentration ([[[Na.sup.+]].sub.pw]) is equal to 1.11([Na.sub.e] + [K.sub.e])/TBW--25.6 (Edelman IS, Leibman J, O'Meara MP, Birkenfeld LW. J Clin Invest 37: 1236-1256, 1958). However, the physiological significance of the slope and y-intercept in this equation has not been previously considered. Our analysis demonstrates that there are several clinically relevant parameters determining the magnitude of the y-intercept that independently alter [[[Na.sup.+]].sub.pw]: 1) osmotically inactive exchangeable [Na.sup.+] and [K.sup.+]; 2) plasma water [K.sup.+] concentration; and 3) osmotically active non-[Na.sup.+] and non-[K.sup.+] osmoles. In addition, we demonstrate quantitatively the physiological significance of the slope in the Edelman equation and its role in modulating [[[Na.sup.+]].sub.pw]. The slope of 1.11 in this equation which Edelman et al. determined empirically can be theoretically predicted by considering the combined effect of the osmotic coefficient of Na+ salts at physiological concentrations and Gibbs-Donnan equilibrium. In addition, our results demonstrate that the slope has an independent quantitative impact on the magnitude of the y-intercept in the Edelman equation. From a physiological standpoint, the components of both the slope and the y-intercept need to be addressed when considering the factors that modulate [[[Na.sup.+]].sub.pw]. osmotic equilibrium; dysnatremia; osmotic coefficient; potassium

Okamoto, Maristela Mitiko, Doris Hissako Sumida, Carla Roberta Oliveira Carvalho, Alessandra Martins Vargas, Joel Claudio Heimann, Beatriz D'Agord Schaan, and Ubiratan Fabres Machado. Changes in dietary sodium consumption modulate GLUT4 gene expression and early steps of insulin signaling. Am J Physiol Regul Integr Comp Physiol 286: R779-R785, 2004. First published December 11, 2003; 10.1152/ajpregu.00396.2003.--Previous studies have shown that chronic salt overload increases insulin sensitivity, while chronic salt restriction decreases it. In the present study we investigated the influence of dietary sodium on 1) GLUT4 gene expression, by Northern and Western blotting analysis; 2) in vivo GLUT4 protein translocation, by measuring the GLUT4 protein in plasma membrane and microsome, before and after insulin injection; and 3) insulin signaling, by analyzing basal and insulin-stimulated tyrosine phosphorylation of insulin receptor (IR)-[beta], insulin receptor substrate (IRS)-1, and IRS-2. Wistar rats were fed normal-sodium (NS-0.5%), low-sodium (LS-0.06%), or high-sodium diets (HS-3.12%) for 9 wk and were killed under pentobarbital anesthesia. Compared with NS rats, HS rats increased (P < 0.05) the GLUT4 protein in adipose tissue and skeletal muscle, whereas GLUT4 mRNA was increased only in adipose tissue. GLUT4 expression was unchanged in LS rats compared with NS rats. The GLUT4 translocation in HS rats was higher (P < 0.05) both in basal and insulin-stimulated conditions. On the other hand. LS rats did not increase the GLUT4 translocation after insulin stimulus. Compared with NS rats, LS rats showed reduced (P < 0.01) basal and insulin-stimulated tyrosine phosphorylatinn of IRS-1 in skeletal muscle and IRS-2 in liver, whereas HS rats showed enhanced basal tyrosine phosphorylation of IRS-1 in skeletal muscle (P < 0.05) and of IRS-2 in liver. In summary, increased insulin sensitivity in HS rats is related to increased GLUT4 gene expression, enhanced insulin signaling, and GLUT4 translocation, whereas decreased insulin sensitivity of LS rats does not involve changes in GLUT4 gene expression but is related to impaired insulin signaling. low-sodium diet; high-sodium diet; insulin resistance; GLUT4 gene expression; insulin signaling

The electroneutral sodium bicarbonate cotransporter 3 (NBC3) coimmunoprecipitates from renal lysates with the vacuolar [H.sup.+]-ATPase. In renal type A and B intercalated cells, NBC3 colocalizes with the vacuolar [H.sup.+]ATPase. The involvement of the COOH termini of NBC3 and the 56-kDa subunit of the proton pump in the interaction of these proteins was investigated. The intact and modified COOH termini of NBC3 and the 56-kDa subunit of the proton pump were synthesized, coupled to Sepharose beads, and used to pull down kidney membrane proteins. Both the 56-and the 70-kDa subunits of the proton pump, as well as a PDZ domain containing protein [Na.sup.+]/[H.sup.+] exchanger regulatory factor 1 (NHERF-1), were bound to the intact 18 amino acid NBC3 COOH terminus. A peptide truncated by five COOH-terminal amino acids did not bind these proteins. Replacement of the COOH-terminal leucine with glycine blocked binding of both the proton pump subunits but did not affect binding of NHERF-1. The 18 amino acid COOH terminus of the 56-kDa subunit of the proton pump bound NHERF-1 and NBC3, but the truncated and modified peptide did not. A complex of NBC3, the 56-kDa subunit of the proton pump, and NHERF-1 was identified in rat kidney. The data indicate that the COOH termini of NBC3 and the 56-kDa subunit of the vacuolar proton pump are PDZ-interacting motifs that are necessary for the interaction of these proteins. NHERF-1 is involved in the interaction of NBC3 and the vacuolar proton pump. sodium; bicarbonate; cotransporters; membrane; binding

Research was conducted to identify the effect produced by anoxia on Na+-K+ pump activity and to determine how trout erythrocytes maintain energy equilibrium. Microcalorimetric measurements of the heat production of cell suspensions quantified the changes in metabolic activity that were linked to normoxia-to-anoxia transitions. Results demonstrate that the cells maintained their ATP contents stable and were viable for hours in the absence of oxygen although the metabolic activity was reduced during anoxia to one-third of the normoxic value.

The effects of cortisol treatment on renal cortical Na+-K+-ATPase were studied in relation to proximal tubule salt transport. The results showed that physiologically relevant maturation of Na+-K+-ATPase with physiological doses of glucocorticoids in 132 day fetal sheep could be induced. Increased protein abundance was pointed to as a possible source of increased activity but the remainder appeared to have resulted from posttranscriptional and posttranslational influences.

A study was conducted to analyze whether dopamine transporter activity could be minimized by a treatment that induces the drive underlying the motivated behavior of sodium appetite in rats. The animals were treated with diuretic furosemide before euthanasia. Results indicated that a direct action of also is one possible mechanism by which sodium depletion promotes a reduction in dopamine transporter activity in the nucleus accumbens.

The role of electroneutral Na+/H+ exchange (NHE) in extracellular acidification and hormone secretion was investigated. Results indicated that stimulation of cholecystokinin (CCK) secretion is functionally linked with activation of NHE through a mechanism that may involve pH-sensitive changes in membrane potential. It was proposed that inhibition of NHE lowers intracellular pH and causes membrane depolarization through the effects of H+ on K+ permeability.

Research was conducted to show that the chimeric AS-oligos selectively knock down (Na+/Ca2+ Exchanger) NCX protein expression and inhibit NCX function in primary cultured rat cardiac myocytes. Protein t(super 1/2) was determined by (35S)methionine with a pulse-chase protocol while the 35S signal in NCX was determined by immunoprecipitation and Western blotting. Results are in good agreement with the physiological evidence of NCX knockdown in ~60% of cells.

A topological analysis of the ubiquitous Na+/H+ exchanger (NHE1) using chymotryptic cleavage. Proteases, glycosidases and impermeant biotin derivatives were utilized in combination of antibodies in the analysis. Results indicate that at least one lysine residue in human NHE1 is readily exposed extracellularly. These sites were found to be located near the NH2 terminus since they were cleaved by short incubation with chymotrypsin.

Research was conducted to examine the differential inhibition of Na+/Ca2+ exchanger isoforms by divalent cations and isothiourea derivative. The properties of three mammalian Na+/Ca2+ exchanger isoforms, NCX1, NCX2 and NCX3, were compared through an analysis of the effects of Ni2+ and other cations and the inhibitor isothiourea derivatives on intracellular Na+-dependent 45Ca2+ uptake into CCL-39 fibroblasts stably expressing each isoform. It was observed that there were distinct differences in the kinetic and pharmacological characteristics between the three isoforms.

A study was conducted on potassium and sodium balance during dietary sodium loading and restriction in enalapril-treated adult Wistar rats. The rats exhibited sodium retention during loading and normal adaptation and transient renal potassium wastage during restriction. Enalapril-treated rats also exhibited reduced renal sodium and potassium handling as well as weak urinary concentrating ability.

A study was conducted on the influence of gustatory neural responses in the nucleus of the solitary tract in rats. Furosemide sodium depletion was found to change NaCl best neuron responsiveness. Results indicate that gustatory system intensity coding is significantly affected by deprivation conditions as well as by the manner in which deprivation is achieved. Results were found opposite to previous studies for nucleus of the solitary neurons involving dietary sodium restriction.

An analysis of the mechanisms compensating Na and water retention induced by long term reduction of renal perfusion pressure was conducted among chronically instrumented and freely moving beagle dogs. Results of the analysis indicated that endogenous downregulation of components of the renin-angiotensin-aldosterone system is needed to reduce sodium and water retention. Natriuretic and diuretic factors is of least importance.

NACA1, NACA2 and NACA3 are three unique isoforms recognized in the Na/Ca exchanger in rabbit. NACA1 obtained from rabbit heart appears the same as that obtained from other species. The kidney yields an NACA2 isoform which is sequenced to reveal that it is a minor kidney isoform. NACA3 is the important isoform in the kidney. It is novel and varies substantially from the minor isoform. Mechanical tissue-specific variation in behavior is provided by these isoforms.

The function of acidity and extracellular Na+ in the production of oxygen by the rat alveolar macrophage was examined. Baseline intracellular acidity (pHi) and transient changes in the pHi were determined by a cytoplasmic pH- sensitive fluorochrome. Changes in intracellular Ca2+ under conditions of extracellular sodium were also measured. The interaction of extracellular Na+ and pH in the modulation of the respiratory burst of rat alveolar macrophage was demonstrated.

Husted, Russell F., Rita D. Sigmund, and John B. Stokes. Mechanisms of inactivation of the action of aldosterone on collecting duct by TGF-[Beta]. Am. J. Physiol. Renal Physiol. 278: F425-F433, 2000.--The purpose of these experiments was to investigate the mechanisms whereby transforming growth factor-[Beta] (TGF-[Beta]) antagonizes the action of adrenocorticoid hormones on [Na.sup.+] transport by the rat inner medullary collecting duct in primary culture. Steroid hormones 1) increased [Na.sup.+] transport by three- to fourfold, 2) increased the maximum capacity of the [Na.sup.+]-[K.sup.+] pump by 30-50%, 3) increased the steady-state levels of the [[Alpha].sub.1]-subunit of the [Na.sup.+]-[K.sup.+]-ATPase by ~30%, and 4) increased the steady-state levels of the [Alpha]-subunit of the rat epithelial [Na.sup.+] channel ([Alpha]-rENaC) by nearly fourfold. TGF-[Beta] blocked the effects of steroids on the increase in [Na.sup.+] transport and the stimulation of the [Na.sup.+]-[K.sup.+]-ATPase and pump capacity. However, there was no effect of TGF-[Beta] on the steroid-induced increase in mRNA levels of [Alpha]-rENaC. The effects of TGF-[Beta] were not secondary to the decrease in [Na.sup.+] transport per se, inasmuch as benzamil inhibited the increase in [Na.sup.+] transport but did not block the increase in pump capacity or [Na.sup.+]-[K.sup.+]-ATPase mRNA. The results indicate that TGF-[Beta] does not inactivate the steroid receptor or its translocation to the nucleus. Rather, they indicate complex pathways involving interruption of the enhancement of pump activity and activation/ inactivation of pathways distal to the steroid-induced increase in the transcription of [Alpha]-rENaC. sodium transport; inner medullary collecting duct; epithelial sodium channel; sodium-potassium-adenosinetriphosphatase; benzamil; glucocorticoid; mineralocorticoid; Northern blot; ribonuclease protection assay; electrophysiology

Nawano, Masao, Akira Oku, Kiichiro Ueta, Itsuro Umebayashi, Tomomi Ishirahara, Kenji Arakawa, Akira Saito, Motonobu Anai, Masatoshi Kikuchi, and Tomoichiro Asano. Hyperglycemia contributes insulin resistance in hepatic and adipose tissue but not skeletal muscle of ZDF rats. Am. J. Physiol. Endocrinol. Metab. 278: E535-E543, 2000.--To determine the contribution of hyperglycemia to the insulin resistance in various insulin-sensitive tissues of Zucker diabetic fatty (ZDF) rats, T-1095, an oral sodium-dependent glucose transporter (SGLT) inhibitor, was administered by being mixed into food. Long-term treatment with T-1095 lowered both fed and fasting blood glucose levels to near normal ranges. A hyperinsulinemic euglycemic clamp study that was performed after 4 wk of T-1095 treatment demonstrated partial recovery of the reduced glucose infusion rate (GIR) in the T-1095-treated group. In the livers of T-1095-treated ZDF rats, hepatic glucose production rate (HGP) and glucose utilization rate (GUR) showed marked recovery, with almost complete normalization of reduced glucokinase/glucose-6-phosphatase (G-6-Pase) activities ratio. In adipose tissues, decreased GUR was also shown to be significantly improved with a normalization of insulin-induced GLUT-4 translocation. In contrast, in skeletal muscles, the reduced GUR was not significantly improved in response to amelioration of hyperglycemia by T-1095 treatment. These results suggest that the contribution of hyperglycemia to insulin resistance in ZDF rats is very high in the liver and considerably elevated in adipose tissues, although it is very low in skeletal muscle. glucose toxicity; Zucker diabetic fatty rat; sodium-dependent glucose transporter

Praetorius, J., D. Andreasen, B. L. Jensen, M. A. Ainsworth, U. G. Friis, and T. Johansen. NHE1, NHE2, and NHE3 contribute to regulation of intracellular pH in murine duodenal epithelial cells. Am. J. Physiol. Gastrointest. Liver Physiol. 278: G197--G206, 2000.--[Na.sup.+]/[H.sup.+]-exchangers (NHE) mediate acid extrusion from duodenal epithelial cells, but the isoforms involved have not previously been determined. Thus we investigated 1) the contribution of [Na.sup.+]-dependent processes to acid extrusion, 2) sensitivity to [Na.sup.+]/[H.sup.+] exchange inhibitors, and 3) molecular expression of NHE isoforms. By fluorescence spectroscopy the recovery of intracellular pH ([pH.sub.i]) was measured on suspensions of isolated acidified murine duodenal epithelial cells loaded with 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Expression of NHE isoforms was studied by RT-PCR and Western blot analysis. Reduction of extracellular [Na.sup.+] concentration ([[[Na.sup.+]].sub.o]) during phi recovery decreased [H.sup.+] efflux to minimally 12.5% of control with a relatively high apparent Michaelis constant for extracellular [Na.sup.+]. The [Na.sup.+]/[H.sup.+] exchange inhibitors ethylisopropylamiloride and amiloride inhibited [H.sup.+] efflux maximally by 57 and 80%, respectively. NHE1, NHE2, and NHE3 were expressed at the mRNA level (RT-PCR) as well as at the protein level (Western blot analysis). On the basis of the effects of low [[[Na.sup.+]].sub.o] and inhibitors we propose that acid extrusion in duodenal epithelial cells involves [Na.sup.+]/[H.sup.+] exchange by isoforms NHE 1, NHE2, and NHE3. intracellular pH; 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein; ethylisopropylamiloride; amiloride; fluorescence spectroscopy; reverse transcriptase-polymerase chain reaction; Western blot; duodenum; sodium/hydrogen exchanger isoforms

D'Cotta, Helena, Claudiane Valotaire, Florence Le Gac, and Patrick Prunet. Synthesis of gill [Na.sup.+]-[K.sup.+]-ATPase in Atlantic salmon smolts: differences in [Alpha]-mRNA and [Alpha]-protein levels. Am. J. Physiol. Regulatory Integrative Comp. Physiol. 278: R101-R110, 2000.--Several parameters were analyzed to determine the mechanisms responsible for the enhancement of the gill [Na.sup.+]-[K.sup.+]-ATPase activity of Atlantic salmon smolts. A major [Alpha]-subunit transcript of 3.7 kb was revealed by Northern blot in both parr and smolt gills when hybridized with two distinct cDNA probes. The [Alpha]-mRNA abundance demonstrated an increase to maximal levels in smolts at an early stage of the parr-smolt transformation. This was followed by a gradual rise in [Alpha]-protein levels, revealed by Western blots with specific antibodies and by an increase in gill [Na.sup.+]-[K.sup.+]-ATPase hydrolytic activity, both only reaching maximum levels a month later, at the peak of the transformation process. Parr fish experienced a decrease in [Alpha]-mRNA abundance and had basal levels of [Alpha]-protein and enzyme activity. Measurement of the binding of [[sup.3]H]ouabain to [Na.sup.+]-[K.sup.+]-ATPase was characterized in smolts and parr gill membranes showing more than a twofold elevation in smolts and was of high affinity in both groups (dissociation constant = 20-23 nM). Modulation of the enzyme due to increased salinity was also observed in seawater-transferred smolts, as demonstrated by an increase in [Alpha]-mRNA levels after 24 h with a rise in [Na.sup.+]-[K.sup.+]-ATPase activity occurring only after 11 days. No qualitative change in [Alpha]-expression was revealed at either the mRNA or protein level. Immunological identification of the [Alpha]-protein was performed with polyclonal antibodies directed against the rat [Alpha]-specific isoforms, revealing that parr, freshwater, and seawater smolts have an [[Alpha].sub.3]-like isoform. This study shows that the increase in [Na.sup.+]-[K.sup.+]-ATPase activity in smolt gills depends first on an increase in the [Alpha]-mRNA expression and is followed by a slower rise in [Alpha]-protein abundance that eventually leads to a higher synthesis of [Na.sup.+]-[K.sup.+] pumps. sodium pump; salmonids; smoltification; [[sup.3]H]ouabain binding; transcript

Andersen, Jens Lundbaek, Lars Juel Andersen, Niels C. F. Sandgaard, and Peter Bie. Volume expansion natriuresis during servo control of systemic blood pressure in conscious dogs. Am. J. Physiol. Regulatory Integrative Comp. Physiol. 278: R19-R27, 2000.--The importance of arterial blood pressure (BP) and ANG II for the renal natriuretic response (NaEx) to volume expansion (3.5% body wt) was investigated during converting enzyme blockade (enalaprilate, 2 mg/kg). In separate experiments, BP was clamped either 30 mmHg above or a few millimeters mercury below baseline by servo-controlled infusion of ANG II or sodium nitroprusside, respectively, so that volume expansion did not change BP. Enalapril decreased BP by 8 mmHg. Without clamping, volume expansion returned BP to that of preenalapril control and increased NaEx 10-fold (40 [+ or -] 10 to 377 [+ or -] 69 [micro]mol/min). During high pressure clamping (133 [+ or -] 2 mmHg), peak NaEx after volume expansion was 6% of control experiments. During low pressure clamping, NaEx was 68% of control experiments (45 [+ or -] 15 to 256 [+ or -] 64 [micro]mol/min). The results show that 1) in absence of ANG II, volume expansion elicited pronounced natriuresis without increases in BP beyond baseline, 2) in the presence of hypertensive amounts of ANG II, the volume expansion-induced natriuresis was almost eliminated, and 3) nitroprusside prevented the increase in BP but not sodium excretion during volume expansion. ANG II appears to dominate the control of NaEx; however, when absent, volume expansion may still induce marked natriuresis even at constant BP, possibly via nitric oxidemediated mechanisms. sodium excretion; pressure natriuresis; angiotensin II; converting enzyme inhibition; nitric oxide

Sandgaard, Niels C. F., Jens Lundbaek Andersen, and Peter Bie. Hormonal regulation of renal sodium and water excretion during normotensive sodium loading in conscious dogs. Am. J. Physiol. Regulatory Integrative Comp. Physiol. 278: R11-R18, 2000.--Saline was infused intravenously for 90 min to normal, sodium-replete conscious dogs at three different rates (6, 20, and 30 [micro]mol [multiplied by] [kg.sup.-1] [multiplied by] [min.sup.-1]) as hypertonic solutions (HyperLoad-6, HyperLoad-20, and HyperLoad-30, respectively) or as isotonic solutions (IsoLoad-6, IsoLoad-20, and IsoLoad-30, respectively). Mean arterial blood pressure did not change with any infusion of 6 or 20 [micro]mol [multiplied by] [kg.sup.-1] [multiplied by] [min.sup.-1]. During HyperLoad-6, plasma vasopressin increased by 30%, although the increase in plasma osmolality (1.0 mosmol/kg) was insignificant. During HyperLoad-20, plasma ANG II decreased from 14 [+ or -] 2 to 7 [+ or -] 2 pg/ml and sodium excretion increased markedly (2.3 [+ or -] 0.8 to 19 [+ or -] 8 [micro]mol/min), whereas glomerular filtration rate (GFR) remained constant. IsoLoad-20 decreased plasma ANG II similarly (13 [+ or -] 3 to 7 [+ or -] 1 pg/ml) concomitant with an increase in GFR and a smaller increase in sodium excretion (1.9 [+ or -] 1.0 to 11 [+ or -] 6 [micro]mol/min). HyperLoad-30 and IsoLoad-30 increased mean arterial blood pressure by 6-7 mmHg and decreased plasma ANG II to ~6 pg/ml, whereas sodium excretion increased to ~60 [micro]mol/min. The data demonstrate that, during slow sodium loading, the rate of excretion of sodium may increase 10-fold without changes in mean arterial blood pressure and GFR and suggest that the increase may be mediated by a decrease in plasma ANG II. Furthermore, the vasopressin system may respond to changes in plasma osmolality undetectable by conventional osmometry. angiotensin II; volume expansion; vasopressin; blood pressure; natriuresis.

Wei, Chiming, and John C. Burnett, Jr. Inhibition by calcium antagonism of circulating and renal endothelin in experimental congestive heart failure. Am. J. Physiol. Heart Circ. Physiol. 278: H263-H268, 2000.--Endothelin (ET) is a potent vasoconstrictor and sodium-regulating peptide whose tissue and plasma concentrations are increased in congestive heart failure (CHF). ET may mediate its vasoconstrictor and sodium-regulatory actions secondary to an increase in intracellular calcium. Calcium influx may augment ET synthesis. Although felodipine, a dihydropyridine calcium-channel antagonist, is effective in reducing vascular resistance in generalized vasoconstriction, its actions in CHF on circulating and local tissue ET remain undefined. The current studies were designed to determine the modulating actions of felodipine (oral, 40 mg/day for 7 days; n = 6) in an experimental canine model of CHF produced by chronic thoracic inferior vena caval constriction (TIVCC) compared with normal (n = 7) and TIVCC-alone (n = 7) dogs. We hypothesized that felodipine would decrease circulating and renal ET. Plasma ET was significantly increased in TIVCC compared with normal dogs (26 [+ or -] 0.5 vs. 12 [+ or -] 0.7 pg/ml, P [is less than] 0.05) and was markedly decreased by felodipine compared with TIVCC alone (14 [+ or -] 3 vs. 26 [+ or -] 0.5 pg/ml, P [is less than] 0.05). Renal ET immunohistochemical staining demonstrated the presence of ET in normal kidney, which was markedly increased in renal cortex and medulla in TIVCC dogs. Renal cortical and medullary ET staining densities were markedly decreased with felodipine compared with those with TIVCC alone. In the TIVCC + felodipine group, cardiovascular hemodynamics also was markedly improved compared with the TIVCC-alone group [systemic vascular resistance: 27 [+ or -] 2 vs. 44 [+ or -] 3 resistance units (RU), P [is less than] 0.05; pulmonary vascular resistance: 3.3 [+ or -] 0.1 vs. 5.7 [+ or -] 0.4 RU, P [is less than] 0.05; cardiac output: 2.9 [+ or -] 0.2 vs. 1.7 [+ or -] 0.11/min, P [is less than] 0.05]. This study demonstrates important modulating inhibitory actions of felodipine on renal and plasma ET in an experimental model of CHF. vasoconstrictor; vasodilator; calcium-channel antagonist

Kuo, Terry B. J., and Cheryl C. H. Yang. Altered frequency characteristic of central vasomotor control in SHR. Am. J. Physiol. Heart Circ. Physiol. 278: H201-H207, 2000. --Previous work from our laboratory has demonstrated that the very low-frequency (VLF: 0-0.25 Hz) and low-frequency (LF: 0.25-0.8 Hz) power of arterial pressure variability (APV) are related to vasomotor reactivity in response to control signals from the rostral ventrolateral medulla (RVLM) via the sympathetic system in the rat. The present study evaluated the differences in the dynamic property of central vasomotor control between spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). Experiments were carried out in 10- to 12-wk-old rats that were anesthetized with continuous infusion of pentobarbital sodium, paralyzed with pancuronium, and maintained on mechanical ventilation. We found that SHR exhibited significantly higher arterial pressure (AP), heart rate (HR), and VLF, LF, and high-frequency (0.8-2.4 Hz) power of APV than WKY under resting state. Broad-band electrical stimulation of the RVLM elicited parallel APV in the VLF and LF ranges in both rat strains. The evoked APV and transfer magnitude of the APV to stimulus spike rate variability (RVLM-AP magnitude) were significantly higher in SHR, especially in the LF range. The response frequency of central vasomotor control, represented by the high-cut frequency of RVLM-AP magnitude, was also extended in SHR. The disparity in RVLM-AP transfer magnitude between SHR and WKY became virtually absent after combined [Alpha]- and [Beta]-adrenoceptor blockade by phentolamine and propranolol. These results suggest that the dynamic control of RVLM on AP reactivity is enhanced in SHR, in which the adrenergic system may play a major role. arterial pressure variability; rostral ventrolateral medulla; broad-band stimulation; coherence; transfer function

Arnon, Assaf, John M. Hamlyn, and Mordecai P. Blaustein. [Na.sup.+] entry via store-operated channels modulates [Ca.sup.2+] signaling in arterial myocytes. Am J. Physiol. Cell Physiol. 278: C163-C173, 2000.--In many nonexcitable cells, hormones and neutransmitters activate [Na.sup.+] influx and mobilize [Ca.sup.2+] from intracellular stores. The stores are replenished by [Ca.sup.2+] influx via 'store-operated' [Ca.sup.2+] channels (SOC). The main routes of [Na.sup.+] entry in these cells are unresolved, and no role for [Na.sup.+] in signaling has been recognized. We demonstrate that the SOC are a major [Na.sup.+] entry route in arterial myocytes. Unloading of the [Ca.sup.2+] stores with cyclopiazonic acid (a sarcoplasmic reticulum [Ca.sup.2+] pump inhibitor) and caffeine induces a large external [Na.sup.+]-dependent rise in the cytosolic [Na.sup.+] concentration. One component of this rise in cytosolic [Na.sup.+] concentration is likely due to [Na.sup.+]/[Ca.sup.2+] exchange; it depends on elevation of cytosolic [Ca.sup.2+] and is insensitive to 10 mM [Mg.sup.2+] and 10 [micro]M [La.sup.3+]. Another component is inhibited by [Mg.sup.2+] and [La.sup.3+], blockers of SOC; this component persists in cells preloaded with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N',-tetraacetic acid to buffer [Ca.sup.2+] transients and prevent [Na.sup.+]/[Ca.sup.2+] exchange-mediated [Na.sup.+] entry. This [Na.sup.+] entry apparently is mediated by SOC. The [Na.sup.+] entry influences [Na.sup.+] pump activity and [Na.sup.+]/ [Ca.sup.2+] exchange and has unexpectedly large effects on cell-wide [Ca.sup.2+] signaling. The SOC pathway may be a general mechanism by which [Na.sup.+] participates in signaling in many types of cells. sodium-calcium exchange; magnesium; sodium pump; cytosolic sodium concentration

Su, Xin, Ruth E. Wachtel, and G. F. Gebhart. Capsaicin sensitivity and voltage-gated sodium currents in colon sensory neurons from rat dorsal root ganglia. Am. J. Physiol. 277 (Gastrointest. Liver Physiol. 40): G1180-G1188, 1999.--DiI-labeled colon sensory neurons were acutely dissociated from S1 rat dorsal root ganglia (DRG) and studied using perforated whole cell patch-clamp techniques. Forty-six percent (54/116) of labeled sensory neurons responded to capsaicin ([10.sup.-8]-[10.sup.-5] M) with an increase in inward current, which was a nonspecific cation conductance. Responses to capsaicin applied by puffer ejection were dependent on dose, with a half-maximal response at 4.9 x [10.sup.-7] M; bath application was characterized by marked desensitization. Voltage-gated [Na.sup.+] currents in 23 of 30 DRG cells exhibited both TTX-sensitive and TTX-resistant components. In these cells, capsaicin induced an inward current in 11 of 17 cells tested. Of the cells containing only a TTX-sensitive component, none of six cells tested was sensitive to capsaicin. In all cells that responded to capsaicin with an increase in inward current, capsaicin abolished voltage-gated [Na.sup.+] currents (n = 21). Capsazepine ([10.sup.-6] M) significantly attenuated both the increase in inward current and the reduction in [Na.sup.+] currents. [Na.sup.+] currents were not significantly altered by adenosine, bradykinin, histamine, [PGE.sub.2], or serotonin at [10.sup.-6] M and [10.sup.-5] M. These findings may have important implications for understanding both the irritant and analgesic properties of capsaicin. visceral pain; bradykinin; prostaglandin [E.sub.2]; serotonin; tetrodotoxin

Haworth, Robert S., James Sinnett-Smith, Enrique Rozengurt, and Metin Avkiran. Protein kinase D inhibits plasma membrane [Na.sup.+]/[H.sup.+] exchanger activity. Am. J. Physiol. 277 (Cell Physiol. 46): C1202-C1209, 1999.--The regulation of plasma membrane [Na.sup.+]/[H.sup.+] exchanger (NHE) activity by protein kinase D (PKD), a novel protein kinase C- and phorbol ester-regulated kinase, was investigated. To determine the effect of PKD on NHE activity in vivo, intracellular pH ([pH.sub.i]) measurements were made in COS-7 cells by microepifluorescence using the pH indicator cSNARF-1. Cells were transfected with empty vector (control), wild-type PKD, or its kinase-deficient mutant PKD-K618M, together with green fluorescent protein (GFP). NHE activity, as reflected by the rate of acid efflux ([J.sub.H]), was determined in single GFP-positive cells following intracellular acidification. Overexpression of wild-type PKD had no significant effect on [J.sub.H] (3.48 [+ or -] 0.25 vs. 3.78 [+ or -] 0.24 mM/min in control at [pH.sub.i] 7.0). In contrast, overexpression of PKD-K618M increased [J.sub.H] (5.31 [+ or -] 0.57 mM/min at [pH.sub.i] 7.0; P [is less than] 0.05 vs. control). Transfection with these constructs produced similar effects also in A-10 cells, indicating that native PKD may have an inhibitory effect on NHE in both cell types, which is relieved by a dominant-negative action of PKD-K618M. Exposure of COS-7 cells to phorbol ester significantly increased [J.sub.H] in control cells but failed to do so in cells overexpressing either wild-type PKD (due to inhibition by the overexpressed PKD) or PKD-K618M (because basal [J.sub.H] was already near maximal). A fusion protein containing the cytosolic regulatory domain (amino acids 637-815) of NHE1 (the ubiquitous NHE isoform) was phosphorylated in vitro by wild-type PKD, but with low stoichiometry. These data suggest that PKD inhibits NHE activity, probably through an indirect mechanism, and represents a novel pathway in the regulation of the exchanger. pH regulation; COS-7; A-10; green fluorescent protein; sodium/ hydrogen exchanger type 1; protein kinase C[micro]

Tomlinson, L. A., T. C. Carpenter, E. H. Baker, J. B. Bridges, and J. V. Weil. Hypoxia reduces airway epithelial sodium transport in rats. Am. J. Physiol. 277 (Lung Cell. Mol. Physiol. 21): L881-L886, 1999.--Ascent to high altitude leads to pulmonary edema formation in some individuals. Recent laboratory evidence supports the hypothesis that hypoxia may impair the function of the alveolar epithelium and thus augment edema accumulation via reduced clearance of lung liquid. We investigated the effect of hypobaric hypoxia on epithelial sodium transport in adult Sprague-Dawley rats by measuring the nasal transepithelial potential difference (PD) as an index of airway sodium transport. Baseline PDs were similar to those previously reported in other species. Administration of amiloride resulted in a significant fall in nasal PD, as did ouabain administration for 24 h (-27.8 vs. -18.8 mV; P = 0.001; n = 5 rats). Exposure to hypobaric hypoxia (0.5 atm) for 24 h caused a significant fall in nasal PD (-23.7 vs. -18.8 mV; P = 0.002; n = 15 rats), which was not additive to the changes in nasal PD produced by amiloride or ouabain. We conclude that subacute exposure to moderate hypobaric hypoxia can inhibit sodium transport by the airway epithelium in rats. transepithelial potential difference; ouabain

Traebert, Martin, Olaf Hattenhauer, Heini Murer, Brigitte Kaissling, and Jurg Biber. Expression of type II Na-[P.sub.i] cotransporter in alveolar type II cells. Am. J. Physiol. 277 (Lung Cell. Mol. Physiol. 21): L868-L873, 1999.--Type II Na-[P.sub.i] cotransporters (type IIa and type IIb) represent apically located Na-[P.sub.i] cotransporters in epithelia of proximal tubules (type IIa) and small intestine (type IIb). Here we provide evidence that the type IIb (but not the type IIa) Na-[P.sub.i] cotransporter is also expressed in the lung. With the use of immunohistochemistry, location of the type IIb protein was found exclusively in the apical membrane of type II cells of the alveolar epithelium. Such a location of the type IIb cotransporter suggests an involvement in the reuptake of phosphate necessary for the synthesis of surfactant. A possible regulation of the abundance of the type IIb cotransporter in the lung was studied after adaptation of mice to a low-[P.sub.i] diet. After a chronic adaptation to a low-[P.sub.i] diet, no changes in the type IIb protein and the type IIb transcript were observed. These results exclude dietary intake of phosphate as a regulatory factor of the type IIb Na-[P.sub.i] cotransporter in alveolar type II cells. sodium-inorganic phosphate cotransporter; immunohistochemistry

Therien, Alex G., and Rhoda Blostein. [K.sup.+]/[Na.sup.+] antagonism at cytoplasmic sites of [Na.sup.+]-[K.sup.+]-ATPase: a tissue-specific mechanism of sodium pump regulation. Am. J. Physiol. 277 (Cell Physiol. 46): C891-C898, 1999.--Tissue-distinct interactions of the [Na.sup.+]-[K.sup.+]-ATPase with [Na.sup.+] and [K.sup.+], independent of isoform-specific properties, were reported previously (A. G. Therien, N. B. Nestor, W. J. Ball, and R. Blostein. J. Biol. Chem. 271: 7104-7112, 1996). In this paper, we describe a detailed analysis of tissue-specific kinetics particularly relevant to regulation of pump activity by intracellular [K.sup.+], namely [K.sup.+] inhibition at cytoplasmic [Na.sup.+] sites. Our results show that the order of susceptibilities of [[Alpha].sub.1] pumps of various rat tissues to [K.sup.+]/[Na.sup.+] antagonism, represented by the ratio of the apparent affinity for [Na.sup.+] binding at cytoplasmic activation sites in the absence of [K.sup.+] to the affinity constant for [K.sup.+] as a competitive inhibitor of [Na.sup.+] binding at cytoplasmic sites, is red blood cell [is less than] axolemma [approximately equals] rat [[Alpha].sub.1]-transfected HeLa cells [is less than] small intestine [is less than] kidney [is less than] heart. In addition, we have carried out an extensive analysis of the kinetics of [K.sup.+] binding and occlusion to the cytoplasmic cation binding site and find that, for most tissues, there is a relationship between the rate of [K.sup.+] binding/occlusion and the apparent affinity for [K.sup.+] as a competitive inhibitor of [Na.sup.+] activation, the order for both parameters being heart [is greater than or equal to] kidney [is greater than] small intestine [approximately equals] rat [[Alpha].sub.1]-transfected HeLa cells. The notion that modulations in cytoplasmic [K.sup.+]/[Na.sup.+] antagonism are a potential mode of pump regulation is underscored by evidence of its reversibility. Thus the relatively high [K.sup.+]/[Na.sup.+] antagonism characteristic of kidney pumps was reduced when rat kidney microsomal membranes were fused into the dog red blood cell. [[Alpha].sub.1]-isoform; heart; kidney

Guner, Yigit S., Pawel R. Kiela, Hua Xu, James F. Collins, and Fayez K. Ghishan. Differential regulation of renal sodium-phosphate transporter by glucocorticoids during rat ontogeny. Am. J. Physiol. 277 (Cell Physiol. 46): C884-C890, 1999.--The effects of chronic administration of methylprednisolone (MP) were studied on the ontogeny of the renal type II Na-[P.sub.i] transporter (NaPi-2). Immunoblot analysis showed that MP did not alter the expression of NaPi-2 protein levels in suckling and weanling rats; however, there was an ~50% decrease in adolescent and adult rats. There was no change in Na-dependent [P.sub.i] uptake in brush-border membrane vesicles in suckling rats, but there was an almost twofold decrease in adolescent rats induced by MP treatment. MP administration did not alter mRNA levels in suckling or adolescent rats. Dual injections with the glucocorticoid receptor blocker RU-486 (mifepristone) and MP did not reverse the downregulation of NaPi-2 immunoreactive protein levels in adolescent rats. To control for RU-486 antagonism efficiency, Na/H exchanger isoform 3 (NHE3) protein levels were also assayed after injection with RU-486 and MP. As expected, NHE3 protein levels increased after MP injection; however, the increase was blocked in adolescent rats by RU-486. We conclude that there is an age-dependent responsiveness to glucocorticoids and that the marked decrease in NaPi-2 immunoreactive protein levels and activity in adolescent rats is due to posttranscriptional mechanisms. sodium-hydrogen exchanger isoform 3; RU-486; methylprednisolone; rat development; kidney

Xue, Xiao-Hua, Larry V. Hryshko, Debora A. Nicoll, Kenneth D. Philipson, and Glen F. Tibbits. Cloning, expression, and characterization of the trout cardiac [Na.sup.+]/ [Ca.sup.2+] exchanger. Am. J. Physiol. 277 (Cell Physiol. 46): C693-C700, 1999.--Isoform 1 of the cardiac [Na.sup.+]/[Ca.sup.2+] exchanger (NCX1) is an important regulator of cytosolic [Ca.sup.2+] concentration in contraction and relaxation. Studies with trout heart sarcolemmal vesicles have shown NCX to have a high level of activity at 7 [degrees] C, and this unique property is likely due to differences in protein structure. In this study, we describe the cloning of an NCX (NCX-TR1) from a Lambda ZAP II cDNA library constructed from rainbow trout (Oncorhynchus mykiss) heart RNA. The NCX-TR1 cDNA has an open reading frame that codes for a protein of 968 amino acids with a deduced molecular mass of 108 kDa. A hydropathy plot indicates the protein contains 12 hydrophobic segments (of which the first is predicted to be a cleaved leader peptide) and a large cytoplasmic loop. By analogy to NCX1, NCX-TR1 is predicted to have nine transmembrane segments. The sequences demonstrated to be the exchanger inhibitory peptide site and the regulatory [Ca.sup.2+] binding site in the cytoplasmic loop of mammalian NCX1 are almost completely conserved in NCX-TR1. NCX-TR1 cRNA was injected into Xenopus oocytes, and after 3-4 days currents were measured by the giant excised patch technique. NCX-TR1 currents measured at ~23[degrees]C demonstrated [Na.sup.+]-dependent inactivation and [Ca.sup.2+]-dependent activation in a manner qualitatively similar to that for NCX1 currents. teleosts; myocardium; contractility; sodium/calcium exchange

Sato, Yoshihiko, Hiroyuki Hanai, Atsuhiro Nogaki, Koki Hirasawa, Eizo Kaneko, Hisayoshi Hayashi, and Yuichi Suzuki. Role of the vasopressin [V.sub.1] receptor in regulating the epithelial functions of the guinea pig distal colon. Am. J. Physiol. 277 (Gastrointest. Liver Physiol. 40): G819-G828, 1999.--Vasopressin has a wide spectrum of biological action. In this study, the role of vasopressin in regulating electrolyte transport in the colon was elucidated by measuring the short-circuit current ([I.sub.sc]) as well as the [Na.sup.+], [K.sup.+], and [Cl.sup.-] flux in a chamber-mounted mucosal sheet. The cytosolic [Ca.sup.2+] concentration ([[Ca.sup.2+]].sub.i]) was also measured in fura 2-loaded cells by fluorescence imaging. Serosal vasopressin decreased [I.sub.sc] at [10.sup.-9] M and increased [I.sub.sc] at [10.sup.-7]-[10.sup.-6] M. The decrease in [I.sub.sc] was accompanied by two effects: one was a decrease in the amiloride-sensitive [Na.sup.+] absorption, whereas the other was an increase in the bumetanidesensitive [K.sup.+] secretion. The increase in [I.sub.sc] was accompanied by an increase in the [CL.sup.-] secretion that can be inhibited by serosal bumetanide or mucosal diphenylamine-2-carboxylate. Vasopressin caused an increase in [[Ca.sup.2+]].sub.i] in crypt cells. These responses of [I.sub.sc] and the [[Ca.sup.2+]].sub.i] increase in crypt cells were all more potently inhibited by the vasopressin [V.sub.1] receptor antagonist than by the [V.sub.2] receptor antagonist. These results suggest that vasopressin inhibits electrogenic [Na.sup.+] absorption and stimulates electrogenic [K.sup.+] and [Cl.sup.-] secretion. In all of these responses, the [V.sub.1] receptor is involved, and the [[Ca.sup.2+]].sub.i] increase may play an important role. sodium absorption; potassium secretion; chloride secretion; intracellular calcium; intestinal transport

Yanagawa, Norimoto, Chi Pham, Remi N. J. Shih, Stephen Miao, and Oak Don Jo. Chloride dependency of renal brush-border membrane phosphate transport. Am. J. Physiol. 277 (Renal Physiol. 46): F506-F512, 1999.--In our present study, we examined the effect of [Cl.sup.-] on rabbit renal brush-border membrane (BBM) phosphate ([P.sub.i]) uptake. It was found that the [Na.sup.+]-dependent BBM [sup.32]P uptake was significantly inhibited by [Cl.sup.-] replacement in the uptake solution with other anions, or by [Cl.sup.-] transport inhibitors, including DIDS, SITS, diphenylamine-2-carboxylate (DPC), niflumic acid (NF), and 5-nitro-2-(3-phenylpropylamino)benzoate (NPPB). Intravesicular formate or [Cl.sup.-] increased BBM [sup.36][Cl.sup.-] uptake but did not affect BBM [sup.32]P uptake. BBM [sup.22][Na.sup.+] uptake was lowered by [Cl.sup.-] replacement in the uptake solution but not by [Cl.sup.-] transport inhibitors. Changes in transmembrane electrical potential altered BBM [sup.36][Cl.sup.-] and [sup.32]P uptake in directions consistent with a net inward movement of negative and positive charges, respectively. However, the [Cl.sup.-] dependent BBM Pi uptake was not affected by changes in transmembrane electrical potential. Finally, a similar [Cl.sup.-] dependency of Pi uptake was also found with BBM derived from rat and mouse kidneys. In summary, our study showed that a component of [Na.sup.+] dependent [P.sub.i] uptake was also [Cl.sup.-] dependent in rabbit, rat, and mouse renal BBM. The mechanism underlying this [Cl.sup.-] dependency remains to be identified. proximal tubule; chloride channel; chloride channel inhibitor; sodium cotransport; hereditary nephrolithiasis

Denton, D. A., J. R. Blair-West, M. I. McBurnie, J. A. P. Miller, R. S. Weisinger, and R. M. Williams. Effect of adrenocorticotrophic hormone on sodium appetite in mice. Am. J. Physiol. 277 (Regulatory Integrative Comp. Physiol. 46): R1033-R1040, 1999.--A main vector of the effects of stress is secretion of corticotrophin releasing factor (CRF), adrenocorticotrophin (ACTH), and adrenal steroids. Systemic administration of ACTH (2.8 [micro]g/day sc) for 7 days in BALB/c mice caused a very large increase of voluntary intake of 0.3 M NaCl equivalent to turnover of total body sodium content each day. Intracerebroventricular infusion of ACTH (20 ng/ day) had no effect. Intracerebroventricular infusion of ovine CRF (10 ng/h for 7 days) caused an increase of sodium intake. The large sodium appetite-stimulating effect of systemic ACTH was not influenced by concurrent systemic infusion of captopril (2 mg/day). Induction of stress by immobilization of mice on a running wheel caused an increase in Na appetite associated with a 50% decrease of thymus weight, indicative of corticosteroid effects. The present data suggest that stress and the hormone cascade initiated by stress evoke a large sodium appetite in mice, which may be an important survival mechanism in environmental conditions causing stress. corticotrophin releasing factor; captopril; immobilization; stress; sodium intake; water intake

Sciortino, Christopher M., and Michael F. Romero. Cation and voltage dependence of rat kidney electrogenic [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] cotransporter, rkNBC, expressed in oocytes. Am. J. Physiol. 277 (Renal Physiol. 46): F611-F623, 1999.--Recently, we reported the cloning and expression of the rat renal electrogenic [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] cotransporter (rkNBC) in Xenopus oocytes [M. F. Romero, P. Fong, U. V. Berger, M. A. Hediger, and W. F. Boron. Am. J. Physiol. 274 (Renal Physiol. 43): F425-F432, 1998]. Thus far, all NBC cDNAs are at least 95% homologous. Additionally, when expressed in oocytes the NBCs are 1) electrogenic, 2) [Na.sup.+] dependent, 3) [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] dependent, and 4) inhibited by stilbenes such as DIDS. The apparent [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] coupling ratio ranges from 3:1 in kidney to 2:1 in pancreas and brain to 1:1 in the heart. This study investigates the cation and voltage dependence of rkNBC expressed in Xenopus oocytes to better understand NBC's apparent tissue-specific physiology. Using two-electrode voltage clamp, we studied the cation specificity, [Na.sup.+] dependence, and the current-voltage (I-V) profile of rkNBC. These experiments indicate that [K.sup.+] and choline do not stimulate [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] -sensitive currents via rkNBC, and [Li.sup.+] elicits only 3 [+ or -] 2% of the total [Na.sup.+] current. The [Na.sup.+] dose response studies show that the apparent affinity of rkNBC for extracellular [Na.sup.+] (~30 mM [[[Na.sup.+]].sub.o]) is voltage and [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] independent, whereas the rkNBC I-V relationship is [Na.sup.+] dependent. At [[[Na.sup.+]].sub.o] [V.sub.max] (96 mM), the I-V response is approximately linear; both inward and outward [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] cotransport are observed. In contrast, only outward cotransport occurs at low [[[Na.sup.+]].sub.o] ([is less than] 1 mM [[[Na.sup.+]].sub.o]). All rkNBC currents are inhibited by extracellular application of DIDS, independent of voltage and [[[Na.sup.+]].sub.o]. Using ion-selective microelectrodes, we monitored intracellular pH and [Na.sup.+] activity. We then calculated intracellular [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] and, with the observed reversal potentials, calculated the stoichiometry of rkNBC over a range of [[[Na.sup.+]].sub.o] values from 10 to 96 mM at 10 and 33 mM [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII]. rkNBC stoichiometry is 2 [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] over this entire [Na.sup.+] range at both [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] concentrations. Our results indicate that rkNBC is highly selective for [Na.sup.+], with transport direction and magnitude sensitive to [[[Na.sup.+]].sub.o] as well as membrane potential. Since the rkNBC protein alone in oocytes exhibits a stoichiometry of less than the 3 [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] thought necessary for [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII] reabsorption by the renal proximal tubule, a control mechanism or signal that alters its in vivo function is hypothesized. sodium/bicarbonate cotransport; NBC; Xenopus oocyte expression; intracellular pH; sodium transport; bicarbonate transport; kinetics; voltage clamp

Pei, Jian-Ming, Xiao-Chun Yu, Jin-Song Bian, and Tak-Ming Wong. Acidosis antagonizes intracellular calcium response to K-opioid receptor stimulation in the rat heart. Am. J. Physiol. 277 (Cell Physiol. 46): C492-C500, 1999.--To study the effects of [Kappa]-opioid receptor stimulation on intracellular [Ca.sub.2+] concentration ([[[Ca.sup.2+]].sub.i]) homeostasis during extracellular acidosis, we determined the effects of [Kappa]-opioid receptor stimulation on [[[Ca.sup.2+]].sub.i] responses during extracellular acidosis in isolated single rat ventricular myocytes, by a spectrofiuorometric method. U-50488H (10-30 [micro]M), a selective [Kappa]-opioid receptor agonist, dose dependently decreased the electrically induced [[[Ca.sub.2+]].sub.i] transient, which results from the influx of [Ca.sub.2+] and the subsequent mobilization of [Ca.sub.2+] from the sarcoplasmic reticulum (SR). U-50488H (30 [micro]M) also increased the resting [[[Ca.sub.2+]].sub.i] and inhibited the [[[Ca.sup.2+]].sub.i] transient induced by caffeine, which mobilizes [Ca.sup.2+] from the SR, indicating that the effects of the K-opioid receptor agonist involved mobilization of [Ca.sub.2+] from its intracellular pool into the cytoplasm. The [Ca.sub.2+] responses to 30 [micro]M U-50488H were abolished by 5 [micro]M nor-binaltorphimine, a selective [Kappa]-opioid receptor antagonist, indicating that the event was mediated by the [Kappa]-opioid receptor. The effects of the agonist on [[[Ca.sub.2+]].sub.i] and the electrically induced [[[Ca.sub.2+]].sub.i] transient were significantly attenuated when the extracellular pH ([pH.sub.e]) was lowered to 6.8, which itself reduced intracellular pH ([pH.sub.i]) and increased [[[Ca.sub.2+]].sub.i]. The inhibitory effects of U-50488H were restored during extracellular acidosis in the presence of 10 pM ethylisopropyl amiloride, a potent [Na.sup.+]/[H.sup.+] exchange blocker, or 0.2 mM [Ni.sup.2+], a putative [Na.sup.+]/[Ca.sup.2+] exchange blocker. The observations indicate that acidosis may antagonize the effects of [Kappa]-opioid receptor stimulation via [Na.sup.+]/[H.sup.+] and [Na.sup.+]/[Ca.sub.2+] exchanges. When glucose at 50 mM, known to activate the [Na.sup.+]/[H.sup.+] exchange, was added, both the resting [[[Ca.sub.2+]].sub.i] and [pH.sub.i] increased. Interestingly, the effects of U-50488H on [[Ca.sub.2+].sub.i] and the electrically induced [[[Ca.sup.2+]].sub.i] transient during superfusion with glucose were significantly attenuated; this mimicked the responses during extracellular acidosis. When a high-[Ca.sub.2+] (3 mM) solution was superfused, the resting [[[Ca.sub.2+]].sub.i] increased; the increase was abolished by 0.2 mM [Ni.sup.2+], but the [pH.sub.i] remained unchanged. Like the responses to superfusion with high-concentration glucose and extracellular acidosis, the responses of the [[[Ca.sub.2+]].sub.i] and electrically induced [[[Ca.sub.2+]].sub.i] transients to 30 [micro]M U-50488H were also significantly attenuated. Results from the present study demonstrated for the first time that extracellular acidosis antagonizes the effects of [Kappa]-opioid receptor stimulation on the mobilization of [Ca.sub.2+] from SR. Activation of both [Na.sup.+]/[H.sup.+] and [Na.sup.+]/[Ca.sup.2+] exchanges, leading to an elevation of [[[Ca.sub.2+]].sub.i], may be responsible for the antagonistic action of extracellular acidosis against [Kappa]-opioid receptor stimulation. sodium/hydrogen exchange; sodium/calcium exchange; sarcoplasmic reticulum

Blazer-Yost, Bonnie L., Teodor G. Paunescu, Sandy I. Helman, Kimberly D. Lee, and Chris J. Vlahos. Phosphoinositide 3-kinase is required for aldosterone-regulated sodium reabsorption. Am. J. Physiol. 277 (Cell Physiol. 46): C531-C536, 1999.--Aldosterone, a steroid hormone, regulates renal [Na.sup.+] reabsorption and, therefore, plays an important role in the maintenance of salt and water balance. In a model renal epithelial cell line (A6) we have found that phosphoinositide 3-kinase (PI 3-kinase) activity is required for aldosterone-stimulated [Na.sup.+] reabsorption. Inhibition of PI 3-kinase by the specific inhibitor LY-294002 markedly reduces both basal and aldosterone-stimulated [Na.sup.+] transport. Further, one of the products of PI 3-kinase, phosphatidylinositol 3,4,5-trisphosphate, is increased in response to aldosterone in intact A6 monolayers. This increase occurs just before the manifestation of the functional effect of the hormone and is also inhibited by LY-294002. With the use of blocker-induced noise analysis, it has been demonstrated that inhibition of phosphoinositide formation causes an inhibition of [Na.sup.+] entry in both control and aldosterone-pretreated cultures by reducing the number of open functional epithelial [Na.sup.+] channels (ENaCs)in the apical membrane of the A6 cells. These novel observations indicate that phosphoinositides are required for ENaC expression and suggest a mechanism for aldosterone regulation of channel function. epithelial sodium channels; noise analysis; electrophysiology; kidney; cortical collecting ducts; A6 cell line

Hecht, Gail, and Athanasia Koutsouris. Myosin regulation of NKCC1: effects on cAMP-mediated [Cl.sup.-] secretion in intestinal epithelia. Am. J. Physiol. 277 (Cell Physiol. 46): C441-C447, 1999.--The basally located actin cytoskeleton has been demonstrated previously to regulate [Cl.sup.-] secretion from intestinal epithelia via its effects on the [Na.sup.+]-[K.sup.+]-[2Cl.sup.-] cotransporter (NKCC1). In nontransporting epithelia, inhibition of myosin light chain kinase (MLCK) prevents cell-shrinkage-induced activation of NKCC1. The aim of this study was to investigate the role of myosin in the regulation of secretagogue-stimulated [Cl.sup.-] secretion in intestinal epithelia. The human intestinal epithelial cell line T84 was used for these studies. Prevention of myosin light chain phosphorylation with the MLCK inhibitor ML-9 or ML-7 and inhibition of myosin ATPase with butanedione monoxime (BDM) attenuated cAMP but not [Ca.sup.2+]-mediated [Cl.sup.-] secretion. Both ML-9 and BDM diminished cAMP activation of NKCC1. Neither apical [Cl.sup.-] channel activity, basolateral [K.sup.+] channel activity, nor [Na.sup.+]-[K.sup.+]-ATPase were affected by these agents. Cytochalasin D prevented such attenuation, cAMP-induced rearrangement of basal actin microfilaments was prevented by both ML-9 and BDM. The phosphorylation of mosin light chain and subsequent contraction of basal actin-myosin bundles are crucial to the cAMP-driven activation of NKCC1 and subsequent apical [Cl.sup.-] efflux. sodium-potassium-chloride cotransporter; myosin light chain kinase; chloride secretion; myosin phosphorylation

Buhagiar, Kerrie A., Peter S. Hansen, David F. Gray, Anastasia S. Mihailidou, and Helge H. Rasmussen. Angiotensin regulates the selectivity of the [Na.sup.+]-[K.sup.+] pump for intracellular [Na.sup.+] Am. J. Physiol. 277 (Cell Physiol. 46): C461-C468, 1999.--Treatment of rabbits with angiotensin converting enzyme (ACE) inhibitors increases the apparent affinity of the [Na.sup.+]-[K.sup.+] pump for [Na.sup.+] To explore the mechanism, we voltage clamped myocytes from control rabbits and rabbits treated with captopril with patch pipettes containing 10 mM [Na.sup.+]. When pipette solutions were [K.sup.+] free, pump current ([I.sub.p]) for myocytes from captopril-treated rabbits was nearly identical to that for myocytes from controls. However, treatment caused a significant increase in [p measured with pipettes containing [K.sup.+]. A similar difference was observed when myocytes from rabbits treated with the ANG II receptor antagonist losartan and myocytes from controls were compared. Treatment-induced differences in [I.sub.p] were eliminated by in vitro exposure to ANG II or phorbol 12-myristate 13-acetate or inclusion of the protein kinase C fragment composed of amino acids 530-558 in pipette solutions. Treatment with captopril had no effect on the voltage dependence of [I.sub.p]. We conclude that ANG II regulates the pump's selectivity for intracellular [Na.sup.+] at sites near the cytoplasmic surface. Protein kinase C is implicated in the messenger cascade. cardiac myocytes; sodium; protein kinase C

A study was conducted to analyze the cloning and functional expression of the Na+/H+ exchanger (NHE) from Amphiuma red blood cells. The first-strand cDNA templates were amplified in th presence of primers while the rapid amplification of cDNA ends was carried out by ligating adapters. Experimental results indicated a high similarity between the sequences of human hNHE1 and Amphiuma tridactylum NHE1.

Volume 78, July 1998 Pages 583-686: J. T. Fitzsimons. 'Angiotensin, Thirst, and Sodium Appetite.' Page 591: In the first paragraph of the right column, the sentence beginning on line 20 [...]

Research was conducted to examine the effects of human cytomegalovirus (HCMV) infection on the Na+-K+-Cl- cotransporter (NKCC) in a human fibroblast cell line. It was observed through the use of the Cl-sensitive dye MQAE, that the mock-infected MRC-5 cells express a functional NKCC. Results indicate that HCMV infection leads to the abolition of NKCC function coincident with the severe reduction in the amount of NKCC protein expressed.