Syndrome of apparent mineralocorticoid excess. A defect in the cortisol-cortisone shuttle.

The first adult case of 11 beta-hydroxysteroid dehydrogenase (11 beta-OHSD) deficiency is described. The impaired conversion of cortisol to cortisone (indicated by urinary cortisol and cortisone metabolites and failure to metabolize 11 alpha-[3H]cortisol to [3H]H2O), was associated with hypertension, hypokalemia, and suppression of the renin-angiotensin-aldosterone system. When established on a fixed Na+/K+ intake, dexamethasone, given orally, produced a natriuresis and potassium retention. Plasma renin activity became detectable. When hydrocortisone (10 mg daily s.c. for 4 d) was added, there was marked Na+ retention, a kaliuresis (urinary Na+/K+ falling from 1.2 to 0.15), with suppression of plasma renin activity and an increase in blood pressure. These changes were also seen with the subject on no treatment. Conversion of cortisone to cortisol was not affected. These results suggest that cortisol acts as a potent mineralocorticoid in 11 beta-OHSD deficiency. The major site for the oxidation of cortisol to cortisone is the kidney. In this patient congenital deficiency of 11 beta-OHSD results in high intrarenal cortisol levels which then act on renal type I mineralocorticoid receptors. This condition can be treated with dexamethasone, which suppresses cortisol secretion and binds to the type II glucocorticoid receptor. We suggest that 11 beta-OHSD exerts a critical paracrine role in determining the specificity of the type I receptor. In the normal state cortisol is converted by 11 beta-OHSD to cortisone which thus allows aldosterone to bind preferentially to the type I receptors in the kidney and gut. In this patient deficiency of 11 beta-OHSD results in high intrarenal cortisol concentrations that then bind to the type I receptor.

Importance of pH regulation and lactate/H+ transport capacity for work production during supramaximal exercise in humans.

We examine the influence of the cytosolic and membrane-bound contents of carbonic anhydrase (CA; CAII, CAIII, CAIV, and CAXIV) and the muscle content of proteins involved in lactate and proton transport [monocarboxylate transporter (MCT) 1, MCT4, and Na(+)/H(+) exchanger 1 (NHE1)] on work capacity during supramaximal exercise. Eight healthy, sedentary subjects performed exercises at 120% of the work rate corresponding to maximal oxygen uptake (W(max)) until exhaustion in placebo (Con) and metabolic alkalosis (Alk) conditions. The total (W(tot)) and supramaximal work performed (W(sup)) was measured. Muscle biopsies were obtained before and immediately after standardized exercises (se) at 120% W(max) in both conditions to determine the content of the targeted proteins, the decrease in muscle pH (DeltapH(m)), and the muscle lactate accumulation ([Lac](m)) per joule of W(sup) (DeltapH(m)/W(sup-se) and Delta[Lac](m)/W(sup-se), respectively) and the dynamic buffer capacity. In Con, W(sup) was positively [corrected] correlated with [corrected] MCT1, and tended to be positively correlated with MCT4 and NHE1. CAII + CAIII were correlated positively with DeltapH(m)/W(sup-se) and negatively with Delta[Lac](m)/W(sup-se), while CAIV was positively related to W(tot). The changes in W(sup) with Alk were correlated positively with those in dynamic buffer capacity and negatively with W(sup) in Con. Performance improvement with Alk was greater in subjects having a low content of proteins involved in pH regulation and lactate/proton transport. These results show the importance of pH regulating mechanisms and lactate/proton transport on work capacity and the role of the CA to delay decrease in pH(m) and accumulation in [Lac](m) during supramaximal exercise in humans.

Immunolocalization of vacuolar-type H+-ATPase in rat submandibular gland and adaptive changes induced by acid-base disturbances.

Using antibodies against the 31-kD and 70-kD subunits of vacuolar type H+-ATPase (V-ATPase) and light microscopic immunocytochemistry, we have demonstrated the presence of this V-ATPase in rat submandibular gland. We have also investigated the adaptive changes of this transporter during acid-base disturbances such as acute and chronic metabolic acidosis or alkalosis. Our results show intracellularly distributed V-ATPase in striated, granular, and main excretory duct cells in controls, but no V-ATPase immunoreaction in acinar cells. Both acute and chronic metabolic acidosis caused a shift in V-ATPase away from diffuse distribution towards apical localization in striated and granular duct cells, suggesting that a V-ATPase could be involved in the regulation of acid-base homeostasis. In contrast, during acidosis the main excretory duct cells showed no changes in the V-ATPase distribution compared to controls. With acute and chronic metabolic alkalosis, no changes in the V-ATPase distribution occurred. (J Histochem Cytochem 46:91-100, 1998)

The effect of metabolic acidosis and alkalosis on the H+-ATPase of rat cerebral microvessels.

To determine the role of the proton translocating adenosine triphosphatase (H+-ATPase) of the blood-brain barrier, the density of the 31 Kd subunit of the vacuolar type H+-ATPase was quantitated in isolated rat cerebral microvessels with immunoblotting techniques. To establish the tissue specificity of the findings, synaptosomal membranes were also studied. Metabolic acidosis was induced with 1.5% ammonium chloride in drinking water for five days. Metabolic alkalosis was induced with 2.35% NaHCO3 in drinking water and daily injections of 10 mg/Kg furosemide intraperitoneally for 5 days. The quantity of the 31 Kd subunit (in arbitrary units) in cerebral microvessels was significantly increased in acidosis (3.98 +/- 0.45) (p<0.05) and was significantly decreased in metabolic alkalosis (0.49 +/- 0.16) (p<0.00) compared to controls (1.77 +/- 0.73). In synaptosomal membranes, metabolic alkalosis was associated with significant decrease in the quantity of the 31 Kd subunit-H+-ATPase (0.62 +/- 0.12 vs 0.92 +/- 0.01) p<0.05. The increase in the 31 Kd subunit in synaptosomal membranes with acidosis did not reach statistical significance. It is concluded that the quantity of vacuolar H+-ATPase in the blood-brain barrier is modulated by blood H+ or HCO3- content. These changes may be relevant to the physiology of the acid-base balance in the central nervous system.

Mechanism of alkalosis-induced constriction of rat cerebral penetrating arterioles.

Cerebral arterioles are in close contact with the supplied tissue and are strong regulators of cerebrovascular tone. Transient ischemia can cause brain intracellular alkalosis producing vasoconstriction. However, the mechanisms of alkalosis-induced cerebral arteriolar constriction are poorly understood. Here, we determined the vascular responses to alkalosis under different conditions by monitoring the internal diameter of pressurized penetrating arterioles isolated from the rat cerebrum with an operating microscope. The roles of Na+/H+ exchanger (NHE), Na+/Ca²+ exchanger (NCX), Na+/K+-adenosine triphosphatase (NKA), and potassium (K+) channels during alkalosis were examined using specific inhibitors. Our results indicated that the extent of constriction of the penetrating arterioles was dependent on alkaline pH. Moreover, the alkalosis-induced vasoconstriction was significantly attenuated by inhibitors of NHE, NCX, and NKA, but not K+ channel inhibitors. Therefore, we concluded that NHE, NKA, and NCX are important regulators involved in alkalosis-induced vasoconstriction of rat cerebral penetrating arterioles.

MAPK signaling pathways are needed for survival of H9c2 cardiac myoblasts under extracellular alkalosis.

pH is one of the most important physiological parameters, with its changes affecting the function of vital organs like the heart. However, the effects of alkalosis on the regulation of cardiac myocyte function have not been extensively investigated. Therefore, we decided to study whether the mitogen-activated protein kinase (MAPK) signaling pathways [c-Jun NH2-terminal kinases (JNKs), extracellular signal-regulated kinases (ERKs), and p38 MAPK] are activated by alkalosis induced with Tris-Tyrode buffer at two pH values, 8.5 and 9.5, in H9c2 rat cardiac myoblasts. These buffers also induced intracellular alkalinization comparable to that induced by 1 mM NH4Cl. The three MAPKs examined presented differential phosphorylation patterns that depended on the severity and the duration of the stimulus. Inhibition of Na+/H+ exchanger (NHE)1 by its inhibitor HOE-642 prevented alkalinization and partially attenuated the alkalosis (pH 8.5)-induced activation of these kinases. The same stimulus also promoted c-Jun phosphorylation and enhanced the binding at oligonucleotides bearing the activator protein-1 (AP-1) consensus sequence, all in a JNK-dependent manner. Additionally, mitogen- and stress-activated kinase 1 (MSK1) was transiently phosphorylated by alkalosis (pH 8.5), and this was abolished by the selective inhibitors of either p38 MAPK or ERK pathways. JNKs also mediated Bcl-2 phosphorylation in response to incubation with the alkaline medium (pH 8.5), while selective inhibitors of the three MAPKs diminished cell viability under these conditions. All these data suggest that alkalosis activates MAPKs in H9c2 cells and these kinases, in turn, modify proteins that regulate gene transcription and cell survival.

Teneurin carboxy (C)-terminal associated peptide-1 inhibits alkalosis-associated necrotic neuronal death by stimulating superoxide dismutase and catalase activity in immortalized mouse hypothalamic cells.

The teneurins and the teneurin C-terminal-associated peptides (TCAP) are implicated in the regulation of neuron growth and differentiation. However, current observations suggest that TCAP-1 may also have a neuroprotective action during times of pH-induced cellular stress in the brain such as during hypoxia-ischemia and brain alkalosis. To test this hypothesis, we cultured a TCAP-1-responsive mouse hypothalamic cell line, N38, using media buffered at pHs 6.8, 7.4, 8.0 and 8.4 subsequently treated with 100 nM TCAP-1. TCAP-1 significantly inhibited the decline in cell proliferation at pHs 8.0 and 8.4 as determined by direct cell viability assays and decreased the incidence of cells showing necrotic morphology. In addition, TCAP-1 decreased the number of cells undergoing necrosis by 4- to 5-fold as measured by uptake of ethidium homodimer III. Moreover, TCAP-1 significantly decreased the incidence of superoxide radicals and increased superoxide dismutase 1 (SOD1) expression. These results were accompanied by an increase in the SOD copper chaperone expression and increased catalase activity and expression. The results indicate that TCAP may play a neuroprotective role during periods of pH stress by upregulating oxygen radical scavenging systems. Thus, the TCAP-teneurin system may be part of a mechanism to protect neurons during trauma, such as hypoxia and ischemia.

Evidence of activation of the renal glutamate dehydrogenase pathway in intact acidotic dogs.

To determine if activity of the renal glutamate dehydrogenase (GD) pathway changes during chronic acidosis in intact dogs, we assessed the deamination of glutamate formed within renal cells during glutamine and alanine infusions. Infusing glutamine into chronically acidotic, normal and acutely alkalotic dogs enhanced renal ammonia production; more was formed as glutamine loading increased. In 4 acidotic dogs, the ratio of ammonia produced to glutamine extracted by the kidneys during exogenous glutamine loading was 1.93 compared with 0.99 for 5 alkalotic dogs and 1.23 for 2 control dogs. Little glutamate and alanine were released into the renal vein in acidotic dogs, whereas over 50% of the exogenous glutamine extracted in acutely alkalotic dogs could be accounted for as glutamate and alanine released into the renal vein. Renal glutamate concentrations were not elevated in acidosis compared with alkalosis despite greater deamidation. When glutamine infusions increased renal ammoniagenesis in acutely alkalotic and control dogs to levels seen in chronically acidotic dogs receiving no exogenous glutamine, approximately 4 to 6 times more glutamate was released from the kidneys. Infusing alanine into 7 chronically acidotic dogs enhanced ammoniagenesis significantly (p less than 0.01), but lesser augmentation was seen in 3 control dogs and no augmentation was seen in 6 acutely alkalotic dogs. The increases were secondary to enhanced glutamate deamination, not secondary to any changes in glutamine extraction and/or transaminase activity. We conclude that the glutamate dehydrogenase pathway is more active in intact acidotic dogs than it is in control and alkalotic dogs.

Effect of metabolic acidosis and alkalosis on NEM-sensitive ATPase in rat nephron segments.

An N-ethylmaleimide (NEM)-sensitive adenosinetriphosphatase (ATPase) displaying the kinetic and pharmacological properties of an electrogenic proton pump has been described in the different segments of rat nephron, where it mediates part of the active tubular proton secretion. This study was therefore designed to evaluate whether changes in urinary acidification observed during metabolic acidosis or alkalosis were associated with alterations of the activity of tubular NEM-sensitive ATPase, and if so, to localize the nephron segments responsible for these changes. Within 1 wk after the onset of ammonium chloride treatment, rats developed a metabolic acidosis, and NEM-sensitive ATPase activity was markedly increased in the medullary thick ascending limb of Henle's loop and outer medullary collecting tubule, and slightly increased in the cortical collecting tubule. Conversely, treatment with sodium bicarbonate induced a metabolic alkalosis that was accompanied by decreased NEM-sensitive ATPase activity in medullary thick ascending limb and outer medullary collecting tubule. NEM-sensitive ATPase activity was not altered in any other nephron segment tested in alkalotic and acidotic rats, i.e., the proximal tubule and the cortical thick ascending limb of Henle's loop. Changes qualitatively similar were observed as soon as 3 h after the onset of NaHCO3 or NH4Cl-loading. In the medullary collecting tubule, alterations of NEM-sensitive ATPase activity are in part due to hyperaldosteronism observed in both acidotic and alkalotic rats.(ABSTRACT TRUNCATED AT 250 WORDS)

[The influence of arterial partial pressure of CO2 and of the arterial value of pH on the oxygen consumption of the whole body during extracorporal circulation (author's transl)]. / Der Einfluss der arteriellen Kohlensäurespannung und des arteriellen pH-Wertes auf den Sauerstoffverbrauch des Gesamtorganismus während der extrakorporalen Zirkulation.

134 patients with coronary heart disease, with defects of the cardiac values or with inborn heart diseases were studied referring to the influences of the arterial partial pressure of CO2 and the arterial value of pH on oxygen consumption of the whole body during total hypothermic extracorporal circulation. Oxygen consumption of the whole body increases during respiratory or metabolic alkalosis whereas respiratory or metabolic acidosis decreases oxygen consumption significantly. These typical changes of oxygen consumption of the whole body during extraorporal circulation due to alkalosis or acidosis may be explained by different facts. Within the cellular area glycolysis is in relation of value for pH, and phosphofructokinase-reaction slows up due to ATP-level also in relationship of value for pH. In addition to cellular parameters hemodynamic parameters may explain the changes of oxygen consumption due to alkalosis and acidosis. Alkalosis effects a decrease of the total peripheral resistane whereas acidosis effects an increase of the total peripheral resistance. Peripheral oxygen consumption vaires in addition to other parameters in inverse proportion of the level of totalperipheral resistance.

H+-K+-ATPases: regulation and role in pathophysiological states.

Molecular cloning experiments have identified the existence of two H+-K+-ATPases (HKAs), colonic and gastric. Recent functional and molecular studies indicate the presence of both transporters in the kidney, which are presumed to mediate the exchange of intracellular H+ for extracellular K+. On the basis of these studies, a picture is evolving that indicates differential regulation of HKAs at the molecular level in acid-base and electrolyte disorders. Of the two transporters, gastric HKA is expressed constitutively along the length of the collecting duct and is responsible for H+ secretion and K+ reabsorption under normal conditions and may be stimulated with acid-base perturbations and/or K+ depletion. This regulation may be species specific. To date there are no data to indicate that the colonic HKA (HKAc) plays a role in H+ secretion or K+ reabsorption under normal conditions. However, HKAc shows adaptive regulation in pathophysiological conditions such as K+ depletion, NaCl deficiency, and proximal renal tubular acidosis, suggesting an important role for this exchanger in potassium, HCO-3, and sodium (or chloride) reabsorption in disease states. The purpose of this review is to summarize recent functional and molecular studies on the regulation of HKAs in physiological and pathophysiological states. Possible signals responsible for regulation of HKAs in these conditions will be discussed. Furthermore, the role of these transporters in acid-base and electrolyte homeostasis will be evaluated in the context of genetically altered animals deficient in HKAc.

Myocyte cytochrome oxidase activity and neuromuscular conduction associated with metabolic alkalemia.

Many studies have shown increased affinity of hemoglobin for oxygen during metabolic alkalemia and dependence of intramitochondrial cytochrome oxidase activity on arterial oxyhemoglobin saturation. The present studies tested the hypothesis that metabolic alkalemia produces tissue hypoxia independent of arterial oxygen desaturation. Neuromuscular conduction latency was used as an indicator of functional impairment, and was measured following electrostimulation of the sciatic nerve and recording of the electromyogram from the gastrocnemius muscle of rats anesthetized with pentobarbital sodium. To increase the affinity of hemoglobin for oxygen, sodium bicarbonate was administered in graded doses every 15 min. Statistical significance of changes was estimated by the paired Student's t test. Arterial bicarbonate ion concentration increased from 25 +/- 1.3 to 39.0 +/- 3.0 mM while arterial pH increased from 7.30 +/- 0.02 to 7.50 +/- 0.03 (P less than 0.01). Neuromuscular conduction latency increased from 1.9 +/- 0.13 to 2.7 +/- 0.18 ms (P less than 0.01). Tissue hypoxia was suggested by a greater decrease in mass spectrometric determinations of gastrocnemius muscle oxygen tension (PO2) in separate groups of control (arterial pH 7.38 +/- 0.04) and experimental (arterial pH 7.48 +/- 0.03) rats. These changes were accompanied by markedly decreased uptake of 3.3'-diaminobenzidine (DAB) by gastrocnemius muscle mitochondria, suggesting decreased intracellular activity of cytochrome oxidase and intracellular oxygen availability to myocytes in the absence of arterial oxygen desaturation.

NEM-sensitive ATPase activity in rat nephron: effect of metabolic acidosis and alkalosis.

The present study was designed to quantitate the amount and to map the localization of N-ethylmaleimide (NEM)-sensitive adenosinetriphosphatase (ATPase) activity in microdissected segments of the rat nephron. After complete nephron mapping the effect of chronic metabolic acidosis and alkalosis on enzyme activity was determined. In control animals the highest enzyme activity was found in the early proximal convoluted tubule of juxtamedullary nephrons; superficial early proximal tubule as well as medullary and cortical thick ascending limbs and collecting ducts also contained substantial activity. Enzyme activity in the papillary collecting duct before entry into the ducts of Bellini was 329 +/- 93 pmol.mm-1.h-1 (n = 8); after entry, however, enzyme activity was approximately one-fourth that value (60 +/- 9 pmol.mm-1.h-1, n = 8, P less than 0.01). No NEM-sensitive ATPase activity was found in the thin limbs of the loop of Henle. Enzyme activity increased in both the medullary and cortical thick ascending limbs as well as in the cortical collecting tubule in response to NH4Cl-induced chronic metabolic acidosis; in the cortical collecting duct, metabolic acidosis increased maximum activity (Vmax) but did not change Michaelis-Menten constant (Km). In the proximal convoluted tubule, enzyme activity decreased with metabolic acidosis. Bicarbonate loading had no effect on enzyme activity except in the most distal portion of the collecting duct where it was stimulated. These results show that NEM-sensitive ATPase activity exists throughout much of the rat nephron. These data suggest that both the cortical collecting tubule and thick ascending limb are regulatory sites of distal urinary acidification during acid loading.

Effect of furosemide-induced hypokalemic metabolic alkalosis on renal transport enzymes.

Hypokalemic metabolic alkalosis is one of the most common complications of chronic furosemide administration. In this study we examined acid-base composition and ATPase enzyme activities in medullary thick ascending limb of Henle's loop (MTAL) and collecting tubule (CCT and MCT) after seven days of chronic furosemide therapy. All of the studies were conducted in adrenal intact (AI) rats or in adrenalectomized (ADX) glucocorticoid replete rats replaced with a physiological dose of aldosterone (Aldo). Furosemide (F) was administered to each rat by mini-osmotic pump. In the AI+F group, plasma Aldo was high and obvious metabolic alkalosis occurred (HCO3- = 37 +/- 2 mEq/liter vs. 22 +/- 2 mEq/liter in controls, P < 0.005); activities of H-K-ATPase, H-ATPase, and Na-K-ATPase were increased approximately twofold in both CCT and MCT. In the ADX+F group (HCO3- = 28 +/- 2 mEq/liter, P < 0.05 from control), H-ATPase activity was normal in CCT and it was slightly increased in MCT. CCT and MCT H-K-ATPase activities were markedly increased (approximately twofold). Na-K-ATPase activity was the same as control in CCT but it was increased in MCT. In ADX+F+Vanadate (V) group which also had normal Aldo levels, acid-base changes were modest (20 +/- 2 mEq/liter, NS from control); in CCT and MCT H-K-ATPase and Na-K-ATPase activities were markedly reduced, but H-ATPase activity in MCT was increased. In all three experimental groups Na-K-ATPase activity in MTAL was reduced fivefold. Hypokalemia developed in both intact and ADX animals receiving furosemide.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of pH and bicarbonate on phosphoenolpyruvate carboxykinase and glutaminase mRNA levels in cultured renal epithelial cells.

A gluconeogenic strain of renal epithelial cells (LLC-PK1-F+) was used to characterize the effect of pH and bicarbonate concentration on the levels of phosphoenolpyruvate carboxykinase (PCK) and glutaminase (GA) mRNAs. The levels of both mRNAs are markedly dependent upon medium glucose concentration. The level of PCK mRNA is increased with increasing glucose concentration from 0 to 40 mM, whereas the level of GA mRNA is maximal between 3 and 5 mM glucose. When LLC-PK1-F+ cells are grown with 5 mM glucose and then subjected to an acute decrease in pH (from 7.4 to 6.9) and bicarbonate concentration (from 25 to 10 mM), the level of PCK mRNA exhibits a biphasic response. The PCK mRNA is initially increased 4-fold within 3 h, then decreases slightly and subsequently increases between 10 and 20 h to a level that is 17-fold greater than normal. Only the initial increase parallels the changes observed in vivo. In contrast, after onset of acidosis, the level of GA mRNA initially remains unchanged, is then increased 8-fold between 10 and 16 h, and then decreases slightly. This response closely mimics the results obtained in vivo. A decrease in media pH at constant bicarbonate causes a marked increase in both mRNAs. However, the levels of the two mRNAs are also elevated by decreasing bicarbonate at a constant pH. Thus, both parameters independently affect the level of the two mRNAs. The use of actinomycin D to measure the half-lives of PCK and GA mRNAs at pH 7.4 and 6.9 indicates that stabilization may fully account for the induction of GA mRNA and contributes to the inductive effects of decreased pH and/or bicarbonate on PCK mRNA. Following recovery from acidic conditions, the two mRNAs exhibit a rapid and coordinate decrease (t1/2 approximately 20 min). Dexamethasone had no effect on the level of either mRNA, whereas cAMP increased only PCK mRNA. The latter effect was additive with the increase caused by decreased pH and/or bicarbonate and was reversed by incubating in alkalotic media. Thus, the induction of PCK and GA mRNAs during acidosis is initiated in direct response to a decrease in extracellular pH and/or bicarbonate.