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Although vasodilatation evoked by acidosis at normal body temperature is well known, the reports regarding effect of acidosis on the reactivity of the isolated arteries at low temperatures are nonexistent. This study tested the hypothesis that the inhibitory effect of acidosis on the neurogenic vasoconstriction may be increased by cooling. Using wire myography, we recorded the neurogenic contraction of the rat tail artery segments to the electrical field stimulation in the absence and in the presence of 0.03-10.0 [micro]mol/L noradrenaline. The experiments were conducted at 37 [degrees]C or 25 [degrees]C and pH 7.4 or 6.6 which was decreased by means of C[O.sub.2]. Noradrenaline at concentration of 0.03-0.1 [micro]mol/L significantly potentiated the neurogenic vasoconstriction at 25 [degrees]C, and the potentiation was not inhibited by acidosis. Contrary to our hypothesis, acidosis at a low temperature did not affect the noradrenaline-induced tone and significantly increased the neurogenic contraction of the artery segments in the absence and presence of noradrenaline. These effects of acidosis were partly dependent on the endothelium and L- type [Ca.sup.2+] channels activation. The phenomenon described for the first time might be of importance for the reduction in the heat loss by virtue of decrease in the subcutaneous blood flow at low ambient temperatures. Key words: acidosis, low temperatures, noradrenaline, neurogenic vasoconstriction, rat tail artery. La vasodilatation evoquee par l'acidose a une temperature corporelle normale est un phenomene bien connu, mais il n'existe pas de publication quant a l'effet de l'acidose sur la reactivite des arteres isolees a des temperatures basses. Cette etude visait a confronter l'hypothese selon laquelle l'effet inhibiteur de l'acidose sur la vasoconstriction neurogenique pourrait augmenter avec le refroidissement. A l'aide de la myographie en fil, nous avons enregistre la contraction neurogenique de segments d'arteres caudales de rats en reaction a la stimulation par champ electrique en absence et en presence de noradrenaline a 0,03-10,0 [micro]mol/L. Nous avons mene les experiences a des temperatures de 37 ou de 25 [degrees]C et a un pH de 7,4 ou abaisse a 6,6 a l'aide de C[O.sub.2]. Aux concentrations de 0,03-0,1 [micro]mol/l la noradrenaline entrainait une potentialisation marquee de la vasoconstriction neurogenique a 25 [degrees]C, phenomene qui n'etait pas inhibe par l'acidose. Contrairement a ce que voulait notre hypothese, l'acidose a une temperature basse n'a pas eu d'effet sur le tonus obtenu avec la noradrenaline et entrainait une augmentation marquee de la contraction neurogenique des segments arteriels en absence et en presence de noradrenaline. Ces effets de l'acidose dependaient en partie de l'activation de l'endothelium et des canaux [Ca.sup.2+] de type L. Decrit pour la premiere fois, ce phenomene pourrait etre important dans la diminution de la deperdition de chaleur en vertu de la diminution du flux sanguin sous-cutane a des temperatures ambiantes basses. [Traduit par la Redaction] Mots-cles: acidose, temperatures basses, noradrenaline, vasoconstriction neurogenique, artere caudale de rat., Introduction Acidosis or decrease in pH of blood and extracellular fluid may occur under many different physiological and pathophysiological conditions including muscular work, respiratory and metabolic disorders, and diabetes (Austin [...]

Abstract: It has been proposed that, during metabolic acidosis, the liver downregulates mitochondrial ammonia detoxification via ureagenesis, a bicarbonate-consuming process. Since we previously demonstrated that hepatocyte mitochondrial aquaporin-8 channels (mtAQP8) [...]

The sodium-bicarbonate cotransporter NBCn1 (SLC4A7) is an acid-base transporter that normally moves [Na.sup.+] and HC[O.sup.-.sub.3] into the cell. This membrane protein is sensitive to cellular and systemic pH changes. We examined NBCn1 expression and localization in the brain and its response to chronic metabolic acidosis. Two new NBCn1 antibodies were generated by immunizing a rabbit and a guinea pig. The antibodies stained neurons in a variety of rat brain regions, including hippocampal pyramidal neurons, dentate gyrus granular neurons, posterior cortical neurons, and cerebellar Purkinje neurons. Choroid plexus epithelia were also stained. Double immunofluorescence labeling showed that NBCn1 and the postsynaptic density protein PSD-95 were found in the same hippocampal CA3 neurons and partially colocalized in dendrites. PSD-95 was pulled down from rat brain lysates with the GST/NBCn1 fusion protein and was also coimmunoprecipitated with NBCn1. Chronic metabolic acidosis was induced by feeding rats with normal chow or 0.4 M HCl-containing chow for 7 days. Real-time PCR and immunoblot showed upregulation of NBCn1 mRNA and protein in the hippocampus of acidotic rats. NBCn1 immunostaining was enhanced in CA3 neurons, posterior cortical neurons, and cerebellar granular cells. Intraperitoneal administration of N-methyl-D-aspartate caused neuronal death determined by caspase-3 activity, and this effect was more severe in acidotic rats. Administering N-methyl-D-aspartate also inhibited NBCn1 upregulation in acidotic rats. We conclude that NBCn1 in neurons is upregulated by chronic acid loads, and this upregulation is associated with glutamate excitotoxicity. acid/base; pH; ion transporter doi: 10.1152/ajpcell.00492.2009.

To determine the effect of metabolic acidosis on expression of L-Gln, L-Glu, and L-Asp metabolizing enzymes and transporters, the relative content of mRNA, protein, or mRNA and protein, of 6 enzymes and 5 transporters was determined by real-time reverse transcription-PCR and immunoblot analyses in homogenates of kidney, skeletal muscle, and liver of growing lambs fed a common diet supplemented with canola meal (control; n = 5) or HCl-treated canola meal (acidosis; n = 5). Acidotic sheep had a 790% greater (P = 0.050) expression of renal Na+-coupled neutral AA transporter 3 mRNA and a decreased expression of renal glutamine synthetase mRNA (47% reduction, P = 0.037) and protein (57% reduction, P = 0.015) than control sheep. No change in renal cytosolic phosphoenolpyruvate carboxykinase (protein and mRNA), glutaminase (mRNA), or L-Glu dehydrogenase (protein) was found. In skeletal muscle, acidotic sheep had 101% more (P = 0.026) aspartate transaminase protein than did control sheep, whereas no change in the content of 3 Na+-coupled neutral AA transporters (mRNA) or 2 high-affinity L-Glu transporter proteins was found. In liver, no change in the content of any assessed enzyme or transporter was found. Collectively, these findings suggest that tissue-level responses of sheep to metabolic acidosis are different than for nonruminants. More specifically, these results indicate the potential capacity for metabolism of L-Asp and L-Glu by skeletal muscle, and L-Gln absorption by kidneys, but no change in hepatic expression of L-Gln metabolism, elaborates previous metabolic studies by revealing molecular-level responses to metabolic acidosis in sheep. The reader is cautioned that the metabolic acidosis model employed in this study differs from the increased plasma lactate-induced metabolic acidosis commonly observed in ruminants fed a highly fermentable grain diet. Key words: acidosis, amino acid, aspartate transaminase, metabolism, sheep, [Na.sup.+]-coupled neutral amino acid transporter doi: 10.2527/jas.2009-2101

During the brain's innate immune response microglia, astroglia and ependymal cells resolve/repair damaged tissue and control infection. Released interleukin-1[beta] (IL-1[beta]) reaching cerebroventricles stimulates circumventricular organs (CVOs; subfornical organ, SFO; organum vasculosum lamina terminalis, OVLT), the median preoptic nucleus (MePO), and magnocellular and parvocellular neurons in the supraoptic (SON) and paraventricular (PVN) nuclei. Hypertonic saline (HS) also activates these osmosensory CVOs and neuroendocrine systems, but, in contrast to IL-1[beta], inhibits the peripheral immune response. To examine whether the brain's innate immune response is attenuated by osmotic stimulation, sterile acidic perfusion fluid was microdialyzed (2 [micro]l/ min) in the SON area of conscious rats for 6 h with sterile HS (1.5 M NaCl) injected subcutaneously (15 ml/kg) at 5 h. Immunohistochemistry identified cytokine sources (IL-1[[beta].sup.+]; OX-[42.sup.+] microglia) and targets (IL-[1R.sup.+]; inducible cyclooxygenase, COX-[2.sup.+]; c-[Fos.sup.+]) near the probe, in CVOs, MePO, ependymal cells, periventricular hypothalamus, SON, and PVN. Inserting the probe stimulated magnocellular neurons (c-[Fos.sup.+]; SON; PVN) via the MePO (c-[Fos.sup.+]), a response enhanced by HS. Microdialysis activated microglia (OX-[42.sup.+]; amoeboid/hypertrophied; IL-1[[beta].sup.+]) in the adjacent SON and bilaterally in perivascular areas of the PVN, periventricular hypothalamus and ependyma, coincident with c-Fos expression in ependymal cells and COX-2 in the vasculature. These microglial responses were attenuated by HS, coincident with activating parvocellular and magnocellular neuroendocrine systems and elevating circulating IL-1[beta], oxytocin, and vasopressin. Acidosis-induced cellular injury from microdialysis activated the brain's innate immune response by a mechanism inhibited by peripheral osmotic stimulation. hypertonic saline; interleukin-1[beta]; acidosis; magnocellular neurons; c-Fos doi: 10.1152/ajpregu.00340.2009.

The mammalian [Na.sup.+]/[H.sup.+] exchanger isoform 1 (NHE1) is a ubiquitously expressed membrane protein that regulates intracellular pH in the myocardium. NHE1 is also important in mediating myocardial hypertrophy, and the blockage of NHE1 activity prevents hypertrophy and reduces ischemia-reperfusion injury in animal models. We recently demonstrated that extracellular-regulated kinase (ERK)-mediated activation of NHE1 occurs during ischemia-reperfusion of the myocardium. To understand the regulation of NHE1 in the myocardium by phosphorylation, we expressed a series of adenoviruses that express wild-type and mutant cDNA for NHE1. All exogenous cDNA for NHE1 had additional mutations [[Leut.sup.163]Phe/[Gly.sup.174]Ser], which increases NHE1 resistance to EMD-87580 (a specific blocker of NHE 1) 100-fold, and allowed the measurement of exogenous NHE1 while inhibiting endogenous NHE1. By examining the effects of a series of mutations of the NHE1 cytosolic region, we determined that the amino acids [Ser.sup.770] and [Ser.sup.771] were essential for the acute activation of NHE1 activity in rat cardiomyocytes. The specific mutation of either residue prevented the rapid activation of exchanger activity by a sustained intracellular acidosis through ERK-dependent pathways. The same amino acids were critical to phenylephrine-mediated, ERK-dependent activation of NHE1 activity and increased the phosphorylation in intact rat cardiomyocytes. The results demonstrate that both sustained intracellular acidosis and phenylephrine rapidly activate the NHE1 protein in intact cardiac cells through ERK-dependent pathways that act on a common pathway mediated by amino acids [Ser.sup.770] and [Ser.sup.771] of the cytosolic tail of the protein. extracellular-regulated kinases-1/2; ischemia-reperfusion; sodium-hydrogen exchanger

Byline: Hiroshi Morisaki (1), Satoshi Yajima (1), Yoko Watanabe (1), Takeshi Suzuki (1), Michiko Yamamoto (1), Nobuyuki Katori (1), Saori Hashiguchi (1), Junzo Takeda (1) Keywords: Hypercapnia; Bacterial translocation; Intramucosal pH; Myeoloperoxidase Abstract: Objective Recent evidence demonstrated that hypercapnic acidosis due to lung protective strategy was not only permissive but also even therapeutic for injured lung. Since the effects of hypercapnic acidosis on extra-pulmonary organs remain to be clarified, we tested the hypothesis that hypercapnic acidosis protects gut mucosal barrier function by modulating inflammation in a rabbit model of endotoxemia. Design Prospective randomized animal study. Setting University research laboratory. Subjects Male New Zealand white rabbits. Interventions Thirty-two animals were randomly allocated into two groups: normocapnia (n = 17) and hypercapnia (n = 15). The latter group received [F.sub.I]CO.sub.2 5% under mechanical ventilation to achieve hypercapnia throughout the study periods, whereas the former with [F.sub.I]CO.sub.2 0%. Measurements and results Arterial blood gas, intramucosal pH (pHi) and portal blood flow were assessed at baseline, 2-h and 4-h infusion of lipopolysaccharide. At 4 h, ileal myeloperoxidase (MPO) activity and intestinal permeability were measured. The animals in the hypercapnia group showed apparent hypercapnic acidosis and progressive intramucosal acidosis at 4 h, accompanied by significantly lower intestinal permeability versus normocapnia group. Ileal MPO activity was comparable between the study groups. Conclusions Hypercapnic acidosis attenuates endotoxin-induced gut barrier dysfunction possibly through neutrophil-independent mechanisms. Author Affiliation: (1) Department of Anesthesiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan Article History: Registration Date: 03/07/2008 Received Date: 16/11/2007 Accepted Date: 19/06/2008 Online Date: 15/07/2008 Article note: Electronic supplementary material The online version of this article (doi: 10.1007/s00134-008-1212-7) contains supplementary material, which is available to authorized users.

Sensory neurons can detect ischemia and transmit pain from various organs. Whereas the primary stimulus in ischemia is assumed to be acidosis, little is known about how the inevitable metabolic challenge influences neuron function. In this study we have investigated the effects of anoxia, aglycemia, and acidosis upon intracellular [Mg.sup.2+] concentration [[[Mg.sup.2+]].sub.i] and intracellular pH (p[H.sub.i]) in isolated sensory neurons. Anoxia, anoxic aglycemia, and acidosis all caused a rise in [[[Mg.sup.2+]].sub.i] and a fall in p[H.sub.i]. The rise in [[[Mg.sup.2+]].sub.i] in response to acidosis appears to be due to [H.sup.+] competing for intracellular [Mg.sup.2+] binding sites. The effects of anoxia and aglycemia were mimicked by metabolic inhibition and, in a dorsal root ganglia (DRG)-derived cell line, the rise in [[[Mg.sup.2+]].sub.i] during metabolic blockade was closely correlated with fall in intracellular ATP concentration ([[ATP].sub.i]). Increase in [[[Mg.sup.2+]].sub.i] during anoxia and aglycemia were therefore assumed to be due to MgATP hydrolysis. Even brief periods of anoxia ( magnesium; ischemia

During systemic acidosis, renal proximal tubular cells exhibit enhanced rates of bicarbonate and ammonium ion synthesis and undergo extensive hypertrophy. The former adaptations are accomplished, in part, by increased expression of glutaminase (GA). LLC-[PK.sub.1-FB[Pase.sup.+] cells, a gluconeogenie line of porcine kidney ceils, exhibit a rapid activation of the ERK1/2 and p38 MAPK pathways and a two- to threefold increase in GA mRNA when transferred to acidic medium (pH 6.9). Transforming growth factor-[beta] (TGF-[beta]), a potent activator of MAPK and Smad signaling cascades, also causes extensive renal hypertrophy. Thus the potential role of TGF-[beta] in the renal response to metabolic acidosis was investigated. Western blot analyses established that in LLC-[PK.sub.1]-FB[Pase.sup.+] cells, TGF-[beta] activated the ERK1/2, p38 MAPK, and Smadl/5/8 pathways, but not the JNK and Smad2/3 pathways. Addition of TGF-[beta] to cells cultured in normal medium (pH 7.4) produced a steady increase in GA mRNA, resulting in a twofold induction after 18 h. Western blot analysis indicated that treatment with either TGF-[beta] or acidic medium resulted in an increased level of fibronectin. However, the effects of the two treatments on both GA mRNA and fibronectin levels occurred with different time courses and were additive. In addition, the rates of ammonia production were decreased slightly by addition of TGF-[beta]. Finally, a GA-luciferase reporter construct, which is activated 3.5-fold by treatment with acidic medium, is not affected by TGF-[beta]. Therefore, TGF-[beta] and metabolic acidosis activate some of the same signaling pathways in LLC-[PK.sub.1]FB[Pase.sup.+] cells, but produce separate effects on GA expression. metabolic acidosis; renal hypertrophy; ERK1/2; p38 MAPK; Smad

Frassetto LA, Morris RC, Jr., Sebastian A. Dietary sodium chloride intake independently predicts the degree of hyperchloremic metabolic acidosis in healthy humans consuming a net acid-producing diet. Am J Physiol Renal Physiol 293: F521-F525, 2007. First published May 23, 2007; doi:10.1152/ajprenal.00048.2007.--We previously demonstrated that typical American net acid-producing diets predict a low-grade metabolic acidosis of severity proportional to the diet net acid load as indexed by the steady-state renal net acid excretion rate (NAE). We now investigate whether a sodium (Na) chloride (Cl) containing diet likewise associates with a low-grade metabolic acidosis of severity proportional to the sodium chloride content of the diet as indexed by the steady-state Na and Cl excretion rates. In the steady-state preintervention periods of our previously reported studies comprising 77 healthy subjects, we averaged in each subject three to six values of blood hydrogen ion concentration ([H]b), plasma bicarbonate concentration ([HC[O.sup.-.sub.3]]p), the partial pressure of carbon dioxide (Pco2), the urinary excretion rates of Na, Cl, NAE, and renal function as measured by creatinine clearance (CrCl), and performed multivariate analyses. Dietary Cl strongly correlated positively with dietary Na (P < 0.001) and was an independent negative predictor of [HC[O.sup.-.sub.3]]P after adjustment for diet net acid load, Pco2 and CrC1, and positive and negative predictors, respectively, of [H]b and [HC[O.sup.-.sub.3]]p after adjustment for diet acid load and [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII]. These data provide the first evidence that, in healthy humans, the diet loads of NaCl and net acid independently predict systemic acid-base status, with increasing degrees of low-grade hyperchloremic metabolic acidosis as the loads increase. Assuming a causal relationship, over their respective ranges of variation, NaCl has ~50-100% of the acidosis-producing effect of the diet net acid load. salt; acid-base; pathophysiology

Blood pH controls the activity of important regulatory enzymes in the metabolism. Serine dehydratase (SerDH) transforms L-serine into pyruvate and ammonium and is involved in the regulation of gluconeogenesis from serine in the rat liver. In this work, we investigate the effect of chronic metabolic acidosis on the kinetics, specific protein level, tissue location, and mRNA levels of rat liver SerDH. Experimental acidosis was induced in rats by ingestion of 0.28 M ammonium chloride solution for 10 days. Acidosis significantly (P < 0.05) decreased SerDH activity at all substrate concentrations assayed. Moreover, the [V.sub.max] value was 38.50 [+ o r-] 3.51 mU/mg (n = 7)of mitochondrial protein in the acidotic rats and 92.49 [+ or -] 6.79 mU/mg (n = 7) in the control rats. Western blot analysis revealed a significant reduction (14%) in the level of SerDH protein content in the rat liver during acidosis. Immunohistochemical analysis showed that SerDH location did not change in response to chronic metabolic acidosis and confirmed previous results on SerDH protein levels. Moreover, the SerDH mRNA level, estimated by RT-PCR, was also significantly 33.8% lower than in control. These results suggest that during experimental acidosis a specific repression of rat-liver SerDH gene transcription could result, lowering the amount and activity of this enzyme. The changes found in SerDH expression are part of an overall metabolic response of liver to maintain acid-base homeostasis during acidosis. N[H.sub.4]Cl; reverse transcriptase-polymerase chain reaction; serine catabolism

The human electrogenic renal Na-HC[O.sub.3] cotransporter (NBCel-A; SLC4A4) is localized to the basolateral membrane of proximal tubule cells. Mutations in the SLC4A4 gene cause an autosomal recessive proximal renal tubular acidosis (pRTA), a disease characterized by impaired ability of the proximal tubule to reabsorb HC[O.sub.3] from the glomerular filtrate. Other symptoms can include mental retardation and ocular abnormalities. Recently, a novel homozygous missense mutant (R881C) of NBCel-A was reported from a patient with a severe pRTA phenotype. The mutant protein was described as having a lower than normal activity when expressed in Xenopus oocytes, despite having normal [Na.sup.+] affinity. However, without trafficking data, it is impossible to determine the molecular basis for the phenotype. In the present study, we expressed wild-type NBCe1-A (WT) and mutant NBCe1-A (R881C), tagged at the COOH terminus with enhanced green fluorescent protein (EGFP). This approach permitted semi-quantification of surface expression in individual Xenopus oocytes before assay by two-electrode voltage clamp or measurements of intracellular pH. These data show that the mutation reduces the surface expression rather than the activity of the individual protein molecules. Confocal microscopy on polarized mammalian epithelial kidney cells [Madin-Darby canine kidney (MDCK)I] expressing nontagged WT or R881C demonstrates that WT is expressed at the basolateral membrane of these cells, whereas R881C is retained in the endoplasmic reticulum. In summary, the pathophysiology of pRTA caused by the R881C mutation is likely due to a deficit of NBCe1-A at the proximal tubule basolateral membrane, rather than a defect in the transport activity of individual molecules. bicarbonate; intracellular pH; acidbase; SLC4A4; [Na.sup.+]-HC[O.sub.3] cotransporter 1

We studied the signal pathway through which troglitazone (TRO) acts in inducing cellular acidosis in LLC-P[K.sub.1]-[F.sup.+] cells in relation to ammoniagenesis and DNA synthesis. Cells were grown to confluent monolayers in 30-mm chambers and monitored for intracellular pH ([pH.sub.i] by the BCECF assay and activated ERK by phospo-ERK1/2 antibodies. TRO induces a severe cellular acidosis ([pH.sub.i] 6.68 [+ or -] 0.10 vs. 7.28 [+ or -] 0.07 time control at 4 min, P < 0.01), whereas phospho-ERK1/2 to total ERK1/2 ratio increases 3.4-fold (P < 0.01). To determine whether ERK1/2 was activated by cellular acidosis or TRO was acting via MEK1/2 to activate ERK1/2, cells were pretreated with specific inhibitors of MEK1/2 activity, PD-098059 and U-0126, followed by the addition of TRO or vehicle. With MEK1/2 activity inhibited, TRO treatment tailed to activate ERK1/2. Preventing ERK1/2 activation abrogated the TRO-induced cellular acidosis and maintained the [pH.sub.i] within the low normal range (7.06 [+ or -] 0.11). To determine whether blocking ERK activation prevents TRO's inhibitory effect on NHE activity, cells were acid-loaded and the recovery response was monitored as [DELTA][pH.sub.i]/t over a 4-min recovery period. TRO inhibited NHE activity by 85% (P < 0.01), whereas blocking ERK activation restored the response. We measured activated ERK levels and [pH.sub.i] after 3- and 18-h exposure to TRO or extracellular acidosis (pHe = 6.95) to determine whether ERK activation was sustained. Whereas both TRO and extracellular acidosis increased activated ERK and decreased [pH.sub.i] after 3 h, only TRO sustained this response at 18 h. Furthermore, both enhanced ammoniagenesis and decreased DNA synthesis reflected the effect of TRO to induce and sustain a cellular acidosis. peroxisome proliferator-activated receptor-[gamma] independent; intracellular pH; P-ERK; sodium hydrogen exchanger; ammoniagenesis; nonvolatile acid production; DNA synthesis

Extracellular acidosis (EA) has profound effects on vascular homeostasis, including vascular bed-specific alterations in vascular tone. Regulation of gene expression by EA has been observed in a variety of cells including vascular endothelial cells. Whether EA regulates gene expression in vascular smooth muscle cells (VSMCs) is not known. Heine oxygenase (HO)-1 is expressed in vascular cells, and its expression is regulated by cellular stressors such as heat, radiation, and hypoxia. Increased HO-1 expression in VSMCs leads to increased production of CO and its second messenger cGMP, which are important regulators of vascular tone and paracrine interactions in the vasculature. We examined whether EA regulates the expression of HO-1 in VSMCs. Exposure of VSMCs to acidic medium (pH 6.8) significantly increased HO-1 mRNA and protein compared with exposure to medium of physiological pH (pH 7.4). The acidic induction of HO-1 expression was time dependent and involved both transcriptional activation of the HO-1 gene and enhanced stability of HO-1 mRNA. Nitric oxide did not appear to mediate this response. We conclude that HO-1 is transcriptionally and posttranscriptionally upregulated by EA in VSMCs. This induction is time dependent and reversible. We speculate that EA, as an important tissue and cellular stressor for VSMCs, may elicit changes in gene expression patterns that contribute to the maintenance or disruption of vascular homeostasis. transcriptional activation; ribonucleic acid stabilization

Respiratory acidosis, a decrease in blood pH caused by a rise in [C[O.sub.2]], rapidly triggers a compensatory response in which the kidney markedly increases its secretion of [H.sup.+] from blood to urine. However, in this and other acid-base disturbances, the equilibrium C[O.sub.2] + [H.sub.2]O [??] HC[O.sup.-.sub.3] + [H.sup.+] makes it impossible to determine whether the critical parameter is [C[O.sub.2]], [HC[O.sup.-.sub.3]], and/or pH. Here, we used out-of-equilibrium C[O.sub.2]/HC[O.sup.-.sub.3] solutions to alter basolateral (BL) [HC[O.sup.-.sub.3]], [C[O.sub.2]], or pH, systematically and one at a time, on isolated perfused S2 rabbit proximal tubules. We found that increasing [[HC[O.sup.-.sub.3]]BL] from 0 to 44 mM, at a fixed [[C[O.sub.2]]BL] of 5% and a fixed [pH.sub.BL] of 7.40, caused HC[O.sup.-.sub.3] reabsorption ([MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.]) to fall by half but did not significantly affect volume reabsorption ([J.sub.V]). Increasing [[C[O.sub.2]]BL] from 0% to 20%, at a fixed [[HC[O.sup.-.sub.3]]BL] of 22 mM and [pH.sub.BL] of 7.40, caused [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.] to rise 2.5-fold but did not significantly affect [J.sub.V]. Finally, increasing [pH.sub.BL] from 6.80 to 8.00, at a fixed [[HC[O.sup.-.sub.3]]BL] of 22 mM and [[C[O.sub.2]]BL] of 5%, did not affect either [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.] or [J.sub.V]. Analysis of the [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.] and [J.sub.V] data implies that, as the tubule alters [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.], it compensates the reabsorption of other solutes to keep [J.sub.V] approximately constant. Because the cells cannot respond acutely to pH changes, we propose that the responses of [MATHEMATICAL EXPRESSION NOT REPRODUCIBLE IN ASCII.] and the reabsorption of other solutes to changes in [[HC[O.sup.-.sub.3]]BL] or [[C[O.sub.2]]BL] involve sensors for basolateral HC[O.sup.-.sub.3] and C[O.sub.2]. kidney | out-of-equilibrium solution | acid-base disturbances | volume reabsorption

Systemic acidosis has detrimental effects on the skeleton, and local acidosis coincides with bone destruction in inflammatory and metastatic diseases. Acidification dramatically enhances osteoclastic resorption, although the underlying mechanism has remained elusive. We investigated the effect of acidosis on the osteoclastogenic transcription factor NFATc1, which upon dephosphorylation translocates from the cytoplasm to nuclei. Lowering extracellular pH dramatically increased accumulation of NFATc1 in nuclei of rat and rabbit osteoclasts to levels comparable with those induced by the proresorptive cytokine receptor activator of NF-[kappa]B ligand (RANKL). Activation of NFATc1 by RANKL was mediated by means of prolonged stimulation of the [Ca.sup.2+]/calmodulin-dependent protein phosphatase, calcineurin. In contrast, NFATc1 activation by acidosis involved stimulation of calcineurin and suppression of NFATc1 inactivation. Acidosis, like RANKL, induced transient elevation of cytosolic free [Ca.sup.2+] concentration ([[[Ca.sup.2+]].sub.i]), which persisted in [Ca.sup.2+]-free media and was abolished by inhibition of phospholipase C or depletion of intracellular [Ca.sup.2+] stores. Real-time-PCR of osteoclast-like ceils generated from RAW 264.7 cells revealed high levels of expression of ovarian cancer G protein-coupled receptor 1, which links extracellular acidification to elevation of [[[Ca.sup.2+]].sub.i]. In addition, the calcineurin inhibitor cyclosporin A suppressed the stimulatory effect of acidification on resorption, implicating NFAT in mediating the actions of acidosis on osteoclast activity. In summary, acidification and RANKL induce signals in osteoclasts that converge on the [Ca.sup.2+]/calcineurin/NFAT pathway. Acidosis acts directly on osteoclasts to activate NFATc1 and stimulate resorption. bone resorption | cyclosporin A | ovarian cancer G protein-coupled receptor 1 | phospholipase C | protons

The development of acidosis during intense exercise has traditionally been explained by the increased production of lactic acid, causing the release of a proton and the formation of the acid salt sodium lactate. On the basis of this explanation, if the rate of lactate production is high enough, the cellular proton buffering capacity can be exceeded, resulting in a decrease in cellular pH. These biochemical events have been termed lactic acidosis. The lactic acidosis of exercise has been a classic explanation of the biochemistry of acidosis for more than 80 years. This belief has led to the interpretation that lactate production causes acidosis and, in turn, that increased lactate production is one of the several causes of muscle fatigue during intense exercise. This review presents clear evidence that there is no biochemical support for lactate production causing acidosis. Lactate production retards, not causes, acidosis. Similarly, there is a wealth of research evidence to show that acidosis is caused by reactions other than lactate production. Every time ATP is broken down to ADP and [P.sub.i], a proton is released. When the ATP demand of muscle contraction is met by mitochondrial respiration, there is no proton accumulation in the cell, as protons are used by the mitochondria for oxidative phosphorylation and to maintain the proton gradient in the intermembranous space. It is only when the exercise intensity increases beyond steady state that there is a need for greater reliance on ATP regeneration from glycolysis and the phosphagen system. The ATP that is supplied from these nonmitochondrial sources and is eventually used to fuel muscle contraction increases proton release and causes the acidosis of intense exercise. Lactate production increases under these cellular conditions to prevent pyruvate accumulation and supply the NA[D.sup.+] needed for phase 2 of glycolysis. Thus increased lactate production coincides with cellular acidosis and remains a good indirect marker for cell metabolic conditions that induce metabolic acidosis. If muscle did not produce lactate, acidosis and muscle fatigue would occur more quickly and exercise performance would be severely impaired. metabolism; skeletal muscle; lactate; acid-base; lactic acidosis

Changes in extracellular pH occur during both physiological neuronal activity and pathological conditions such as epilepsy and stroke. Such pH changes are known to exert profound effects on neuronal activity and survival. Heteromeric KCNQ2/3 potassium channels constitute a potential target for modulation by [H.sup.+] ions as they are expressed widely within the CNS and have been proposed to underlie the M-current, an important determinant of excitability in neuronal cells. Whole-cell and single-channel recordings demonstrated a modulation of heterologously expressed KCNQ2/3 channels by extracellular [H.sup.+] ions. KCNQ2/3 current was inhibited by [H.sup.+] ions with an I[C.sub.50] of 52 nM (pH 7.3) at -60 mV, rising to 2 [micro]M (pH 5.7) at -10 mV. Neuronal M-current exhibited a similar sensitivity. Extracellular [H.sub.+] ions affected two distinct properties of KCNQ2/3 current: the maximum current attainable upon depolarization ([I.sub.max]) and the voltage dependence of steady-state activation. Reduction of [I.sub.max] was antagonized by extracellular [K.sup.+] ions and affected by mutations within the outer-pore turret, indicating an outer-pore based process. This reduction of [I.sub.max] was shown to be due primarily to a decrease in the maximum open-probability of single KCNQ2/3 channels. Single-channel open times were shortened by acidosis (pH 5.9), while closed times were increased. Acidosis also recruited a longer-lasting closed state, and caused a switch of single-channel activity from the full-conductance state (~8 pS) to a subconductance state (~5 pS). A depolarizing shift in the activation curve of macroscopic KCNQ2/3 currents and single KCNQ2/3 channels was caused by acidosis, while alkalosis caused a hyperpolarizing shift. Activation and deactivation kinetics were slowed by acidosis, indicating specific effects of [H.sup.+] ions on elements involved in gating. Contrasting modulation of homomeric KCNQ2 and KCNQ3 currents revealed that high sensitivity to [H.sup.+] ions was conferred by the KCNQ3 subunit. KEY WORDS: acidosis * conductance * [K.sup.+] channel * M-current * pH

Chronic metabolic acidosis increases urinary calcium excretion without altering intestinal calcium absorption, suggesting that bone mineral is the source of the additional urinary calcium. In vivo and in vitro studies have shown that metabolic acidosis causes a loss of mineral calcium while buffering the additional hydrogen ions. Previously, we studied changes in femoral, midcortical ion concentrations after 7 days of in vivo metabolic acidosis induced by oral ammonium chloride. We found that, compared with mice drinking only distilled water, ammonium chloride induced a loss of bone sodium and potassium and a depletion of mineral HC[O.sup.-.sub.3] and phosphate. There is more phosphate than carbonate in neonatal mouse bone. In the present in vitro study, we utilized a high-resolution scanning ion microprobe with secondary ion mass spectroscopy to test the hypothesis that chronic acidosis would decrease bulk (cross-sectional) bone phosphate to a greater extent than HC[O.sup.-.sub.3] by localizing and comparing changes in bone HC[O.sup.-.sub.3] and phosphate after chronic incubation of neonatal mouse calvariae in acidic medium. Calvariae were cultured for a total of 51 h in medium acidified by a reduction in HC[O.sup.-.sub.3] concentration ([HC[O.sup.-.sub.3]]; pH ~7.14, [HC[O.sup.-.sub.3] ~13) or in control medium (pH ~7.45, HC[O.sup.-.sub.3] ~26). Compared with incubation in control medium, incubation in acidic medium caused no change in surface total phosphate but a significant fall in cross-sectional phosphate, with respect to the carbon-carbon bond (C.sub.2) and the carbon-nitrogen bond (CN). Compared with incubation in control medium, incubation in acidic medium caused no change in surface HC[O.sup.-.sub.3] but a significant fall in cross-sectional HC[0.sup.-.sub.3] with respect to [C.sub.2] and CN. The fall in cross-sectional phosphate was significantly greater than the fall in cross-sectional HC[O.sup.-.sub.3]. The fall in phosphate indicates release of mineral phosphates, and the fall in HC[O.sup.-.sub.3] indicates release of mineral HC[O.sup.-.sub.3], both of which would be expected to buffer the additional protons and help restore the pH toward normal. Thus a model of chronic acidosis depletes bulk bone proton buffers, with phosphate depletion exceeding that of HC[O.sup.-.sub.3]. ion microprobe; calcium; proton; metabolic acidosis

Byline: Antje Pittner (1), Marek Nalos (1), Pierre Asfar (1), Yan Yang (1), Can Ince (2), Michael Georgieff (1), Uwe Bruckner (3), Peter Radermacher (1), Gebhard Froba (1) Keywords: Endotoxin Ileal mucosal-arterial PCO2 gap Villous Microcirculation Laser Doppler flowmetry Remission spectrophotometry Orthogonal polarization spectral imaging Abstract: Abstract Objective. To determine the mechanisms of improved gut mucosal acidosis associated with selective inducible nitric oxide synthase (iNOS) inhibition. Design. Prospective, controlled experimental study. Setting. Animal research laboratory. Animals. Fourteen domestic pigs. Interventions. Anesthetized and mechanically ventilated pigs received continuous i.v. endotoxin for 24 h. A selective iNOS-inhibitor (1400W, n=8) or vehicle (control, n=6) was started at 12 h of endotoxin and infused until the end of the experiment. Measurements and results. Before as well as at 12 and 24 h of endotoxin, portal venous flow (ultrasound probe), intestinal oxygen (O.sub.2) extraction, portal venous-arterial carbon dioxide (CO.sub.2) content difference and ileal mucosal-arterial PCO.sub.2 gap (fiberoptic sensor) were assessed together with video recordings of the villous microcirculation (number of perfused/unperfused villi) using orthogonal polarization spectral imaging via an ileostomy. The gut wall microvascular blood flow (units) and hemoglobin O.sub.2 saturation (AuHb-O.sub.2) were assessed with a combined laser Doppler flow and remission spectrophotometry probe. 1400W blunted the otherwise progressive rise in the PCO.sub.2 gap without affecting portal venous flow, regional O.sub.2 and CO.sub.2 exchange or the number of unperfused villi. While endotoxin markedly aggravated the heterogeneity of the microvascular blood flow and oxygenation, 1400W had no further effect. Conclusions. Given the uninfluenced parameters of the ileal mucosal microcirculation in our model of long-term porcine endotoxemia, selective iNOS inhibition probably improved the PCO.sub.2 gap due to a redistribution of the microvascular perfusion within the gut wall and/or an amelioration of the cellular respiration. Author Affiliation: (1) Sektion Anasthesiologische Pathophysiologie und Verfahrensentwicklung, Universitatsklinikum, Parkstrasse 11, 89073 Ulm, Germany (2) Afdeling Experimentele Anaesthesiologie, Academisch Medisch Centrum, Amsterdam, The Netherlands (3) Sektion Chirurgische Forschung, Universitatsklinikum, Parkstrasse 11, 89073 Ulm, Germany (4) Present address: Anesteziologicko resuscitacni klinika, Karlova Universita, PlzeA, Czech Republic (5) Present address: Service de Reanimation Medicale, CHU, Angers, France Article History: Received Date: 24/04/2002 Accepted Date: 18/10/2002 Article note: Electronic Publication

The effects of chronic dietary acid loads on shifts in bone mineral reserves and physiological concentrations of cations and anions in extracellular fluids were assessed in growing swine. Four trials were conducted with a total of 38 (8.16 [+ or -] 0.30 kg, mean [+ or -] SEM) Large White x Landrace x Duroc pigs randomly assigned to one of three dietary treatments. Semipurifled diets, fed for 13 to 17 d, provided an analyzed dietary electrolyte balance (dEB, meq/kg diet = [Na.sup.+] + [K.sup.+] - [Cl.sup.-]) of -35, 112, and 212 for the acidogenic, control, and alkalinogenic diets, respectively. Growth performance, arterial blood gas, serum chemistry, urine pH, mineral balance, bone mineral content gain, bone-breaking strength, bone ash, and percentage of bone ash were determined. Dietary treatments created a range of metabolic acid loads without affecting (P > 0.10) growth or feed intake. Urine pH was 5.71, 6.02, and 7.65 [+ or -] 0.48 (mean + SEM) and arterial blood pH was 7.478, 7.485, and 7.526 [+ or -] 0.006 for pigs fed acidogenic, control, and alkalinogenic treatments, respectively. A lower dEB resulted in an increased (P < 0.001) apparent [Cl.sup.-] retention (106.6, 55.4, and 41.2 [+ or -] 6.3 meq/d), of which only 1.6% was accounted for by expansion of the extracellular fluid [Cl.sup.-] pool as calculated from serum [Cl.sup.-] (105.5, 103.4, 101.6 [+ or -] 0.94 meq/L (mean [+ or -] SEM) for pigs fed acidogenic, control, and alkalinogenic treatments, respectively. A lower dEB did not decrease (P > 0.10) bone mineral content gain, bone-breaking strength, bone ash, percentage of bone ash, or calcium and phosphate balance. In conclusion, bone mineral (phosphate) was not depleted to buffer the dietary acid load in growing pigs over a 3-wk period. Key Words: Bones, Calcium, Chlorides, Electrolytes, Phosphates, Pigs

Pendrin is an apical [Cl.sup.-]/[OH.sup.-]/HC[O.sup.-.sub.3] exchanger in [beta]-intercalated cells ([beta]-ICs) of rat and mouse cortical collecting duct (CCD). However, little is known about its regulation in acid-base disorders. Here, we examined the regulation of pendrin in metabolic acidosis, a condition known to decrease HC[0.sup.-.sub.3] secretion in CCD. Rats were subjected to [NH.sub.4]Cl loading for 4 days, which resulted in metabolic acidosis. Apical [Cl.sup.-]/HC[O.sup.-.sub.3] exchanger activity in [beta]-ICs was determined as amplitude and rate of intracellular pH change when Cl was removed in isolated, microperfused CCDs. Intracellular pH was measured by single-cell digital ratiometric imaging using fluorescent pH-sensitive dye 2',7'-bis-(3-carboxypropyl)-5-(and-6)-carboxyfluorescein-AM. Pendrin mRNA expression in kidney cortex was examined by Northern blot hybridizations. Expression of pendrin protein was assessed by indirect immunofluorescence. Microperfused CCDs isolated from acidotic rats demonstrated ~60% reduction in apical [Cl.sup.-]/HC[O.sup.-.sub.3] exchanger activity in [beta]-ICs (P < 0.001 vs. control). Northern blot hybridizations indicated that the mRNA expression of pendrin in kidney cortex decreased by 68% in acidotic animals (P < 0.02 vs. control). Immunofluorescence labeling demonstrated significant reduction in pendrin expression in CCDs of acidotic rats. We conclude that metabolic acidosis decreases the activity of the apical [Cl.sup.-]/HC[O.sup.-.sub.3] exchanger in [beta]-ICs of the rat CCD by reducing the expression of pendrin. Adaptive downregulation of pendrin in metabolic acidosis indicates the important role of this exchanger in acid-base regulation in the CCD. kidney; intercalated cells

Acidosis inhibits current through the Ky1.4 [K.sup.+] channel, perhaps as a result of enhancement of C-type inactivation. The mechanism of action of acidosis on C-type inactivation has been studied. A mutant Kv1.4 channel that lacks N-type inactivation (fKv1.4 [DELTA]2-146) was expressed in Xenopus oocytes, and currents were recorded using two-microelectrode voltage clamp. Acidosis increased fKv1.4 [DELTA]2-146 C-type inactivation. Replacement of a pore histidine with cysteine (H508C) abolished the increase. Application of positively charged thiol-specific methanethiosulfonate to fKv1.4 [DELTA]2-146 H508C increased C-type inactivation, mimicking the effect of acidosis. Replacement of a pore lysine with cysteine (K532C) abolished the acidosis-induced increase of C-type inactivation. A model of the Kv1.4 pore, based on the crystal structure of KcsA, shows that H508 and K532 lie close together. It is suggested that the acidosis-induced increase of C-type inactivation involves the charge on H508 and K532. acidosis; C-type inactivation; Kv1.4

To characterize the effects of ischemia on cGMP synthesis in microvascular endothelium, cultured endothelial cells from adult rat hearts were exposed to hypoxia or normoxia at pH 6.4 or 7.4. Cellular cGMP and soluble (sGC) and membrane guanylyl cyclase (mGC) activities were measured after stimulation of sGC (S-nitroso-N-acetyl-penicillamine) or mGC (urodilatin) or after no stimulation. Cell death (lactate dehydrogenase release) was negligible in all experiments. Hypoxia at pH 6.4 induced a rapid ~90% decrease in cellular cGMP after sGC and mGC stimulation. This effect was reproduced by acidosis. Hypoxia at pH 7.4 elicited a less pronounced (~50%) and slower reduction in cGMP synthesis. Reoxygenation after 2 h of hypoxia at either pH 6.4 or 7.4 normalized the response to mGC stimulation but further deteriorated the sGC response; normalization of pH rapidly reversed the effects of acidosis. At pH 7.4, the response to GC stimulation correlated well with cellular ATP. We conclude that simulated ischemia severely depresses cGMP synthesis in microvascular coronary endothelial cells through ATP depletion and acidosis without intrinsic protein alteration. guanylyl cyclases; ATP depletion; pH; ischemia-reperfusion; nitric oxide; natriuretic factors

The effect of acidosis on the electrical activity of isolated rat atrial myocytes was investigated using the patch-clamp technique. Reducing the pH of the bathing solution from 7.4 to 6.5 shortened the action potential. Acidosis had no significant effect on transient outward or inward rectifier currents but increased steady-state outward current. This increase was still present, although reduced, when intracellular [Ca.sup.2+] was buffered by 1,2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid (BAPTA); BAPTA also inhibited acidosis-induced shortening of the action potential. [Ni.sup.2+] (5 mM) had no significant effect on the acidosis-induced shortening of the action potential. Acidosis also increased inward current at -80 mV and depolarized the resting membrane potential Acidosis activated an inwardly rectifying [Cl.sup.-] current that was blocked by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), which also inhibited the acidosis-induced depolarization of the resting membrane potential. It is concluded that an acidosis-induced increase in steady-state outward [K.sup.+] current underlies the shortening of the action potential and that an acidosis-induced increase in inwardly rectifying [Cl.sup.-] current underlies the depolarization of the resting membrane potential during acidosis. action potential; potassium current; chloride current; perforated patch

Electrophysiological response of rat ventricular myocytes to acidosis. Am J Physiol Heart Circ Physiol 283: H412-H422, 2002. First published March 28, 2002; 10.1152/ajpheart.01042.2001.--The effects of acidosis on the action potential, resting potential, L-type C[a.sub.2+]([I.sub.Ca]), inward rectifier potassium ([I.sup.K1]), delayed rectifier potassium ([I.sub.K]), steady-state ([I.sub.SS]), and inwardly rectifying chloride ([I.sup.Cl,ir]) currents of rat subepicardial (Epi) and subendocardial (Endo) ventricular myocytes were investigated uslng the patch-clamp technique. Action potential duration was shorter in Epi than in Endo cells. Acidosis (extracellular pH decreased from 7.4 to 6.5) depolarized the resting membrane potential and prolonged the time for 50% repolarization of the action potential in Epi and Endo cells, although the prolongation was larger in Endo cells. At control pH, [I.sub.Ca], [I.sub.K1], and [I.sub.SS] were not significantly different in Epi and Endo cells, but [I.sub.K] was larger in Epi cells. Acidosis did not alter [I.sub.Ca], [I.sub.K1], or [I.sub.K] but decreased/ss; this decrease was larger in Endo cells. It is suggested that the acidosis-induced decrease in/ss underlies the prolongation of the action potential. [I.sub.Cl,ir] at control pH was [Cd.sup.2+] sensitive but 4,4'-disothiocyanato-stilbene-2,2'-disulfonic acid resistant. Acidosis increased [I.sub.Cl,ir]; it is suggested that the acidosis-induced increase in [I.sub.Cl,ir] underlies the depolarization of the resting membrane potential. action potential; chloride current; potassium current; perforated patch

A study was conducted to determine whether bacterial direct-fed microbials (DFM) could be used to minimize the risk of acidosis in feedlot cattle receiving high concentrate diets. Six ruminally cannulated steers, previously adapted to a high concentrate diet, were used in a double 3 x 3 Latin square to study the effects of DFM on feed intake, ruminal pH, and ruminal and blood characteristics. Steers were provided ad libitum access to a diet containing steam-rolled barley, barley silage, and a protein-mineral supplement at 87, 9, and 4% (DM basis), respectively. Treatments were as follows: control, Propionibacterium P15 (P15), and Propionibacterium P15 and Enterococcus faecium EF212 (PE). The bacterial treatments ([10.sup.9] cfu/g) plus whey powder carrier, or whey powder alone for control, were top-dressed once daily at the time of feeding (10 g/[steer/d]). Periods consisted of 2 wk of adaptation and 1 wk of measurements. Ruminal pH was continuously measured for 6 d using indwelling electrodes. Dry matter intake and ruminal pH (mean, minimum, hours, and area pH < 5.8 or < 5.5) were not affected by treatment (P > 0.05). However, supplementation with P15 increased protozoal numbers (P < 0.05) with a concomitant increase in ruminal N[H.sub.3] concentration (P < 0.01) and a decrease in the number of amylolytic bacteria (P < 0.05) compared with the control. Streptococcus bovis, enumerated using a selective medium, was numerically reduced with supplementation of PE. Although blood pH and blood glucose were not affected by DFM supplementation, steers fed PE had numerically lower concentrations of blood [CO.sub.2] than control steers, which is consistent with a reduced risk of metabolic acidosis. Although the bacterial DFM used in this study did not induce changes in DMI or ruminal and blood pH, some rumen and blood variables indicated that the bacterial DFM used in this study may decrease the risk of acidosis in feedlot cattle. Key Words: Acidosis, Beef Cattle, Enterococcus, Microbial Flora, Probiotics, Propionibacterium

We studied the effect of troglitazone on cellular acid-base balance and alanine formation in isolated rat mesangial cells. Mesangial cells were grown to confluency in RPMI 1640 media on 30-mm chambers used to monitor both cellular pH using the pH-sensitive dye 2'7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein and metabolic acid production as well as glutamine metabolism. Troglitazone (10 [micro]M) induced a spontaneous cellular acidosis (6.95 [+ or -] 0.02 vs. 7.47 [+ or -] 0.04, respectively; P < 0.0001) but without an increase in lactic acid production. Alanine production was reduced 64% (P < 0.01) consistent with inhibition of the glutamate transamination. These findings pointed to a decrease in acid extrusion rather than an increase in acid production as the underlying mechanism leading to the cellular acidosis. To test their acid extrusion capabilities, mesangial cells were acid loaded with N[H.sup.+.sub.4] and then allowed to recover in Krebs-Henseleit media or in Krebs-Henseleit media minus bicarbonate (HEPES substituted), and the recovery response ([DELTA]p[H.sub.i]/min) was monitored. In the presence of 10 [micro]M troglitazone, the recovery response to the N[H.sup.+.sub.4] acid load was virtually eliminated in the bicarbonate-buffered media (0.00 [+ or -] 0.001 vs. 0.06 [+ or -] 0.02 p[H.sub.i]/min, P < 0.0001 vs. control) and reduced 75% in HEPES-buffered media (0.01 [+ or -] 0.01 vs. 0.04 [+ or -] 0.02 p[H.sub.i]min, P < 0.002 vs. control). These results show that troglitazone induces a spontaneous cellular acidosis resulting from a reduction in cellular acid extrusion. 2'7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein; lactic acid; sodium/hydrogen exchange; bicarbonate-activated acid extrusion; glutamine metabolism

Acidosis and angiotensin II (ANG II) stimulate ammonia production and transport by the proximal tubule. We examined the effect of short-term (18 h) in vivo acid loading with N[H.sub.4]Cl on ammonia production and secretion rates by mouse S2 proximal tubule segments microperfused in vitro with or without ANG II in the luminal microperfusion solution. S2 tubules from N[H.sub.4]Cl-treated mice displayed higher rates of luminal ammonia secretion compared with those from control mice. The adaptive increase in ammonia secretion in N[H.sub.4]Cl-treated mice was eliminated when losartan was coadministered in vivo with N[H.sub.4]Cl. Ammonia secretion rates from both N[H.sub.4]Cl-treated and control mice were largely inhibited by amiloride. Addition of ANG II to the microperfusion solution enhanced ammonia secretion and production rates to a greater extent in tubules from N[H.sub.4]Cl-treated mice compared with those from controls, and the stimulatory effects of ANG II were blocked by losartan. These results demonstrate that a short-term acid challenge induces an adaptive increase in ammonia secretion by the proximal tubule and suggest that ANG II plays an important role in the adaptive enhancement of ammonia secretion that is observed with short-term acid challenges. transport; ammoniagenesis; acid-base physiology; losartan; ammonium chloride

Chronic metabolic acidosis upregulates rat kidney Na-HC[O.sup.-.sub.3] cotransporters NBCn1 and NBC3 but not NBC1. Am J Physiol Renal Physiol 282: F341-F351, 2002. First published September 21, 2001; 10.1152/ajprenal. 00104.2001.--Several members of the Na-HC[O.sup.-.sub.3] cotransporter (NBC) family have recently been identified functionally and partly characterized, including rkNBC1, NBCn1, and NBC3. Regulation of these NBCs may play a role in the maintenance of intracellular pH and in the regulation of renal acid-base balance. However, it is unknown whether the expressions of these NBCs are regulated in response to changes in acid-base status. We therefore tested whether chronic metabolic acidosis (CMA) affects the abundance of these NBCs in kidneys using two conventional protocols. In protocol 1, rats were treated with N[H.sub.4]Cl in their drinking water (12 [+ or -] 1 mmol * [rat.sup.-1] * [day.sup.-1]) for 2 wk with free access to water (n = 8). Semiquantitative immunoblotting demonstrated that whole kidney abundance of NBCn1 and NBC3 in rats with CMA was dramatically increased to 995 [+ or -] 87 and 224 [+ or -] 35%, respectively, of control levels (P < 0.05), whereas whole kidney rkNBC1 was unchanged (88 [+ or -] 14%). In protocol 2, rats were given N[H.sub.4]Cl in their food (10 [+ or -] 1 mmol * [rat.sup.-1] * [day.sup.-1]) for 7 days, with a fixed daily water intake (n = 6). Consistent with protocol 1, whole kidney abundances of NBCnl (262 [+ or -] 42%) and NBC3 (160 [+ or -] 31%) were significantly increased compared with controls (n = 6), whereas whole kidney rkNBC1 was unchanged (84 [+ or -] 17%). In both protocols, immunocytochemistry confirmed upregulation of NBCn1 and NBC3 with no change in the segmental distribution along the nephron. Consistent with the increase in NBCn1, measurements of pH transients in medullary thick ascending limb (mTAL) cells in kidney slices revealed two- to threefold increases in DIDS- sensitive, [Na.sup.+]-dependent HC[O.sup.-.sub.3] uptake in rats with CMA. In conclusion, CMA is associated with a marked increase in the abundance of NBCn1 in the mTAL and NBC3 in intercalated cells, whereas the abundance of NBC1 in the proximal tubule was not altered. The increased abundance of NBCn1 may play a role in the reabsorption of N[H.sup.-.sub.4] in the mTAL and increased NBC3 in reabsorbing HC[O.sup.-.sub.3]. acid-base balance; bicarbonate transport; intracellular pH; intercalated cell; thick ascending limb

Study of mutations associated with acid-base transport disease will likely improve understanding of protein structure-function relationships and their effects on physiology. The related diseases involve plasmalemmal and organellar transporters of hydrogen ions, chloride ions, and HCO(sub.3)(super.-) ions, and autosomal-dominant and -recessive forms of distal renal tubular acidosis (dRTA) caused by mutations in ion transporters of the acid-secreting Type A intercalated cell of the renal collecting duct. Causes of recessive proximal RTA with ocular disease, recessive mixed proximal-distal RTA accompanied by osteopetrosis and mental retardation, the metabolic alkalosis of congenital chloride-losing diarrhea, and recessive osteopetrosis are covered.

Bushinsky, David A., Walter R. Parker, Kristen M. Alexander, and Nancy S. Krieger. Metabolic, but not respiratory, acidosis increases bone PG[E.sub.2] levels and calcium release. Am J Physiol Renal Physiol 281: F1058-F1066, 2001. First published July 12, 2001; 10.1152/ajprenal.00355.2001.-- A decrease in blood pH may be due to either a reduction in bicarbonate concentration ([HC[O.sup.-.sub.3]; metabolic acidosis) or to an increase in P[CO.sub.2] (respiratory acidosis). In mammals, metabolic, but not respiratory, acidosis increases urine calcium excretion without altering intestinal calcium absorption, indicating that the additional urinary calcium is derived from bone. In cultured bone, chronic metabolic, but not respiratory, acidosis increases net calcium efflux ([J.sub.Ca]), decreases osteoblastic collagen synthesis, and increases osteoclastic bone resorption. Metabolic acidosis increases bone PG[E.sub.2] production, which is correlated with [J.sub.Ca], and inhibition of PG[E.sub.2] production inhibits this acid-induced [J.sub.Ca]. Given the marked differences in the osseous response to metabolic and respiratory acidosis, we hypothesized that incubation of neonatal mouse calvariae in medium simulating respiratory acidosis would not increase medium PG[E.sub.2] levels, as observed during metabolic acidosis. To test this hypothesis, we determined medium PG[E.sub.2] levels and [J.sub.Ca] from calvariae incubated at pH ~7.1 to model either metabolic (Met; [HC[O.sup.-.sub.3]] ~11 mM) or respiratory (Resp; P[CO.sub.2] ~83 Torr) acidosis, or at pH ~7.5 as a control (Ntl). We found that after 24-48 and 48-51 h in culture, periods when cell-mediated [J.sub.Ca] predominates, medium PG[E.sub.2] levels and [J.sub.Ca] were increased with Met, but not Resp, compared with Ntl, and there was a direct correlation between medium PG[E.sub.2] levels and [J.sub.Ca]. Thus metabolic, but not respiratory, acidosis induces the release of bone PG[E.sub.2], which mediates [J.sub.Ca] from bone. calvariae; osteoblasts; protons; pH Received 1 December 2000; accepted in final form 10 July 2001

The effect of metabolic acidosis (MA) on Na-S(sub i) cotransport activity and NaS(sub i)-1 protein and mRNA expression was investigated. In rats with MA for 24 hrs, there was substantial increase in the fractional excretion of S(sub i), which was related with a 2.4-fold reduction in brush-border membrane (BBM) Na-S(sub i) cotransport activity. Results showed that MA decreases S(sub i) reabsorption by inducing reductions in BBM Na-S(sub i) cotransport activity and that reductions in the expression of NaS(sub i)-1 protein and mRNA abundance, at least partially, have a significant role in the inhibition of Na-S(sub i) cotransport activity during MA.

The physiologic adaptation of the outer medullary collecting duct (OMCD) to metabolic acidosis in vivo and vitro was investigated in rabbits through the measurement of the net HCO(super -)(sub 3) absorption mediated by changes in the concentrations of H+ and adenosine triphosphatase. The H+ secretion rate of the collecting duct segments was believed to be highest at the OMCD under baseline conditions. It was revealed that OMCD adaptation to low-pH incubation was similar to the in vivo metabolic acidosis adaptation. The shuttling and exocytosis of H+ pumps mediated metabolic acidosis adaptation.

The rat rostral ventrolateral medulla (RVLM) catechol signal was studied in rats after mild systemic acidosis to test the idea that systemic acidosis causes activates RVLM adrenergic neurons. Findings showed that a catechol activation in the RVLM results when arterial pressure is maintained during isocapnic systemic metabolic acidosis. Adrenergic RVLM neurons may relay inputs from the central respiratory generator to the sympathetic system and/or assume the role of chemosensors for H+ next to the surface of the ventrolateral medulla.

Carbonic anhydrase (CAD) II-deficient mice were used as models of moderate chronic metabolic acidosis in the investigation of the levels of growth hormone (GH) and insulin-like growth factors, and the response to recombinant growth hormone (rGH) supplementation of animals with chronic acidosis. Results reveal that males have depressed levels of GH compared to controls and female mice with CAD deficiency. Further, the response to rGH is more pronounced among CAD-deficient male mice.

Research was undertaken to assess the physiological effect of metabolic acidosis on the cellular reabsorption of sodium chloride (NaCl) in the murine proximal convoluted tubule. It was established that NaCl reabsorption decreased the capacity of proximal tubule during metabolic acidosis. Results revealed possible processes involved in metabolic acidosis. Metabolic acidosis downregulated NaCl transport resulting from organic anion. The process involved in the NaCl transport was the passive paracellular mechanism.

The expression of carbonic anhydrase (CA) from the cytosol (CA II) and from the membrane (CA IV) in various segments of the nephron is investigated using reverse-transcriptase polymerase chain reaction. CA IV mRNA is expressed in various segments of the proximal tubule and the colleting ducts while CA II is expressed throughout the length of the nephron. Acid loading stimulates CA mRNA transcription.

Isolated small cerebral arteries from rats are used in the investigation of the role of calcium in vasorelaxation in acidotic conditions. Acidosis was induced by increasing carbon dioxide or decreasing bicarbonate concentrations in the media resulting in hypercapnic acidosis and normocapnic acidosis respectively. Results indicate that intracellular calcium plays a role in acidosis-related vasorelaxation but the mechanism of intracellular calcium regulation differs between hypercapnic acidosis and normocapnic acidosis as there are different membrane potential responses.

Acidosis is associated with myocardial ischemia and several reports indicate the greater vulnerability of the aged heart to ischemic dysfunction. We investigated the effects of hypercapnic acidosis on isolated heart (n = 14) and papillary muscle (n = 10) from adult and senescent rats. Acidosis (pH from 7.36 to 6.91) induced a decrease in left ventricular developed pressure together with an increase in left ventricular end-diastolic pressure, but was significantly more evident in senescent than in adult hearts (p < .01). The return to normal pH induced a further increase in the end-diastolic pressure parallel to the development of arrhythmias that were greater in senescent than in adult hearts. In isolated papillary muscle, acidosis confirmed its greater negative inotropic effect on senescent than adult muscles (p < .01), while intracellular sodium activity (??) increased to a similar extent in both adult and senescent papillary muscles (p = NS). 5-(N,N-dimethyl)-amiloride hydrochloride (DMA), a specific inhibitor of [Na.sup.+]/[H.sup.+] exchanger, produced similar modification of tension and [Mathematical Expression Omitted] in both adult and senescent muscles. When DMA was superfused in acidotic solution, the contractility was markedly compromised in senescent than in adult muscles (p < .01), but the [Mathematical Expression Omitted] modifications were similar in adult and senescent muscles (p = NS). Our results show that acidosis induced a greater reduction of contractility in senescent than in adult hearts. The similarity of contractility during DMA administration between adult and senescent muscle and of modifications of [Mathematical Expression Omitted] suggests that depression of contractility with acidosis may be related to pathophysiologic mechanisms other than the [Na.sup.+]/[H.sup.+] exchanger.

Research was conducted to study the influence of acidosis on the intracellular Ca2+ concentration and contractile properties of intact mammalian cardiac muscle during tetanic and twitch contractions. Results indicated that the decrease in Ca2+ binding to troponin C is directly correlated with the decline in tetanic and twitch contraction applied to the determination of induced less intracellular Ca2+ concentration change in acidosis.

Tubule microperfusion studies were conducted on acute and chronic respiratory acid-base manipulations to determine the response of the rat loop on Henle (LOH) in vivo. The results revealed no evidence on the effect of acute respiratory alkalosis and acidosis on LOH bicarbonate transport. On the other hand, bicarbonate reabsorption was inhibited but increases during partial recovery, where the urinary bicarbonate excretion increases with decreasig blood carbonate concentration.

The role of thiazide-type diuretic (TZR) receptors in the regulation of normal acid-base balance was analyzed in rats that exhibited ammonium chloride-induced metabolic acidosis and sodium bicarbonate-induced metabolic alkalosis. Induction of metabolic acidosis via ammonium chloride administration in rats increased renal bicarbonate reabsorption in the Henle's loop and hydrogen radical secretion from the collecting duct cells. Furthermore, metabolic acidosis reduced TZR density while metabolic alkalosis reduced TZR density.