Arginine vasopressin enhances pHi regulation in the presence of HCO3- by stimulating three acid-base transport systems

Growth factors raise intracellular pH (pHi) by stimulating Na+/H+ exchange in the absence of HCO3-. In mutant cells that lack the Na+/H+ exchange activity, this alkalinization does not occur, and the cells do not proliferate without artificial elevation of pHi. It has therefore been widely suggested that an early pHi increase is a necessary signal for mitogenesis. In the presence of HCO3- however, growth factors fail to raise pHi in A431 cells, renal mesangial cells and 3T3 fibroblasts. In mesangial cells, arginine vasopressin (AVP) raises pHi in the absence of HCO3-, but lowers it when HCO3- is present; growth is stimulated under both conditions. We report here that, in the presence of HCO3-, AVP stimulates two potent HCO3- transporters, as well as the Na+/H+ exchanger. These are the Na+-dependent and Na+-independent Cl-/HCO3- exchangers. Our results indicate that AVP causes acidification in the presence of HCO3- because, at the resting pHi, it stimulates Na+-independent Cl-/HCO3- exchange (which lowers pHi) more than it stimulates the sum of Na+/H+ exchange and Na+-dependent Cl-/HCO3- exchange (both of which raise pHi). The stimulation of three acid-base transporters by the growth factor AVP greatly enhances the ability of the cell to regulate pHi.     

Basolateral KCl co-transport in a NaCl-absorbing epithelium

In NaCl-absorbing epithelia such as proximal renal tubule, small intestine and gallbladder, Na+-dependent Cl- entry across the luminal membrane is an electroneutral transport process that has been attributed to Na-Cl symport, Na-K-Cl symport, or a double (Na-H, Cl-HCO3) antiport. At the basolateral (antiluminal) membrane, Na+ extrusion can be attributed to the Na+-K+ pump, and Cl- transport could be explained in principle by electrodiffusion since the intracellular Cl- activity is higher than predicted for equilibrium distribution. However, in Necturus gallbladder, as in other epithelia, the electrodiffusional Cl- permeability of the membrane (PCl) is too low to account for the transepithelial Cl- transport rate. Because K+ is at a higher chemical potential in the cell than in the extracellular fluid, and because serosal Cl- substitutions have only small effects on membrane potential, the hypothesis of carrier-mediated electroneutral KCl co-transport was proposed. The experiments reported here were designed to test this hypothesis in Necturus gallbladder epithelium. Intracellular Cl- and K+ activities (aCli, aKi) were measured with ion-sensitive intracellular microelectrodes before, during and after ionic substitutions of the serosal (basolateral) bathing medium. The results demonstrate a Na+-independent basolateral membrane KCl symport.     

pH-sensitive activation of the intracellular-pH regulation system in squid axons by ATP-gamma-S

The regulation of intracellular pH (pHi) is essential for normal cell function, and controlled changes in pHi may play a central role in cell activation. Sodium-dependent Cl-HCO3 exchange is the dominant mechanism of pHi regulation in the invertebrate cells examined, and also occurs in mammalian cells. The transporter extrudes acid from the cell by exchanging extracellular Na+ and HCO3- (ref. 9) (or a related species) for intracellular Cl- (refs 3, 4). It is blocked by the stilbene derivatives DIDS (4,4'-diisothiocyano-stilbene-2,2'-disulphonate, ref. 10) and SITS (4-acetamido-4'-isothiocyano-stilbene-2,2'-disulphonate, ref. 3), and has a stoichiometry of two intracellular H+ neutralized for each Na+ taken up and each Cl- extruded by the axon. Because the inwardly-directed Na+ concentration gradient is sufficiently large to energize both the HCO3- influx and Cl- efflux, this electroneutral exchanger could be a classic secondary active transporter, thermodynamically independent of ATP hydrolysis. However, at least in the squid axon, the exchanger has an absolute requirement for ATP (ref. 3). Thus, a major unresolved issue is whether this Na-dependent Cl-HCO3 exchanger stoichiometrically hydrolyses ATP (the pump hypothesis), or whether ATP activates the transporter by a mechanism such as phosphorylation or simple binding (the activation hypothesis). We have now explored the role of ATP in pHi regulation by dialysing axons with the ATP analogue ATP-gamma-S. In many systems, ATP-gamma-S is an acceptable substrate for protein kinases, whereas the resulting thiophosphorylated proteins are not as readily hydrolysed by phosphatases as are phosphorylated proteins. Our results rule out the pump hypothesis, and show that the basis of the axon's ATP requirement is the pH-dependent activation (by, for instance, phosphorylation or ATP binding) of the exchanger itself, or of an essential activator.     

青海黄鼠湾—何家庄河谷区地下水水化学特征

随着黄鼠湾-何家庄的工业园区发展,周边生态环境受到了影响。地下水作为生态重要组成部分,研究地下水水化学特征对该地区地下水保护和生态建设具有重要意义。因此,本文2019年在黄鼠湾-何家庄河谷区采集29组水样进行检测分析,运用经典统计学、Piper三线图、Gibbs图、主要离子浓度关系图等方法,对地下水特征及成因进行分析。结果表明：(1)优势阳离子为钙(Ca2+)、镁(Mg2+)、钠(Na+)。优势阴离子为碳酸氢(HCO3-)、硫酸根(SO42-)、氯(Cl-)。(2)研究区水化学类型分为三类。HCO3?Cl-Ca?Na?Mg类、HCO3-Mg?Na?Ca类均分布在研究区上游。HCO3?SO4-Mg?Na?Ca类分布于研究区中游与下游。(3)阳离子受蒸发浓缩、岩石风化与离子交换作用影响。阴离子受岩石风化作用影响。研究区Na+、K+、Cl-主要来源于岩盐与硅酸盐的溶解, Ca2+、Mg2+、HCO-3、SO2-4主要来源于碳酸盐、硅酸盐和蒸发岩的溶解。Ca2+、Na+受到阳离子交换作用影响。过高的K+、Na+、SO42-与人类工业活动有关。     

Glucose/galactose malabsorption caused by a defect in the Na+/glucose cotransporter

Glucose/galactose malabsorption (GGM) is an autosomal recessive disease manifesting within the first weeks of life and characterized by a selective failure to absorb dietary glucose and galactose from the intestine. The consequent severe diarrhoea and dehydration are usually fatal unless these sugars are eliminated from the diet. Intestinal biopsies of GGM patients have revealed a specific defect in Na(+)-dependent absorption of glucose in the brush border. Normal glucose absorption is mediated by the Na+/glucose cotransporter in the brush border membrane of the intestinal epithelium. Cellular influx is driven by the transmembrane Na+ electrochemical potential gradient; thereafter the sugar moves to the blood across the basolateral membrane via the facilitated glucose carrier. We have previously cloned and sequenced a Na+/glucose cotransporter from normal human ileum and shown that this gene, SGLT1, resides on the distal q arm of chromosome 22. We have now amplified SGLT1 complementary DNA and genomic DNA from members of a family affected with GGM by the polymerase chain reaction. Sequence analysis of the amplified products has revealed a single missense mutation in SGLT1 which cosegregates with the GGM phenotype and results in a complete loss of Na(+)-dependent glucose transport in Xenopus oocytes injected with this complementary RNA.     

Activation of chloride channels in normal and cystic fibrosis airway epithelial cells by multifunctional calcium/calmodulin-dependent protein kinase.

Cystic fibrosis is associated with defective regulation of apical membrane chloride channels in airway epithelial cells. These channels in normal cells are activated by cyclic AMP-dependent protein kinase and protein kinase C. In cystic fibrosis these kinases fail to activate otherwise normal Cl- channels. But Cl- flux in cystic fibrosis cells, as in normal cells, can be activated by raising intracellular Ca2+ (refs 5-10). We report here whole-cell patch clamp studies of normal and cystic fibrosis-derived airway epithelial cells showing that Cl- channel activation by Ca2+ is mediated by multifunctional Ca2+/calmodulin-dependent protein kinase. We find that intracellular application of activated kinase and ATP activates a Cl- current similar to that activated by a Ca2+ ionophore, that peptide inhibitors of either the kinase or calmodulin block Ca2(+)-dependent activation of Cl- channels, and that a peptide inhibitor of protein kinase C does not block Ca2(+)-dependent activation. Ca2+/calmodulin activation of Cl- channels presents a pathway with therapeutic potential for circumventing defective regulation of Cl- channels in cystic fibrosis.     

Expression cloning and cDNA sequencing of the Na+/glucose co-transporter

Organic substrates (sugars, amino acids, carboxylic acids and neutrotransmitters) are actively transported into eukaryotic cells by Na+ co-transport. Some of the transport proteins have been identified--for example, intestinal brush border Na+/glucose and Na+/proline transporters and the brain Na+/CI-/GABA transporter--and progress has been made in locating their active sites and probing their conformational states. The archetypical Na+-driven transporter is the intestinal brush border Na+/glucose co-transporter (see ref. 8), and a defect in the co-transporter is the origin of the congenital glucose-galactose malabsorption syndrome. Here we describe cloning of this co-transporter by a method new to membrane proteins. We have sequenced the cloned DNA and have found no homology between the Na+/glucose co-transporter and either the mammalian facilitated glucose carrier or the bacterial sugar transport proteins. This suggests that the mammalian Na+-driven transporter has no evolutionary relationship to the other sugar transporters.     

Effects of ATP and vanadate on calcium efflux from barnacle muscle fibres

Calcium ions carry the inward current during depolarization of barnacle muscle fibres and are involved in the contraction process. Intracellular ionized calcium ([Ca2+]i) in barnacle muscle, as in other cells, is kept at a very low concentration, against a large electrochemical gradient. This large gradient is maintained by Ca2+ extrusion mechanisms. When [Ca2+]i is below the contraction threshold, Ca2+ efflux from giant barnacle muscle fibres is, largely, both ATP dependent and external Na+ (Na+0) dependent (see also refs 5,6). When [Ca2+]i is raised to the level expected during muscle contraction (2-5 muM), most of the Ca2+ efflux from perfused fibres is Na0 dependent; as in squid axons, this Na+0-dependent Ca2+ efflux is ATP independent. Orthovanadate is an inhibitor of (Na+ + K+) ATPase and the red cell Ca2+-ATpase. We report here that vanadate inhibits ATP-promoted, Na+0-dependent Ca2+ efflux from barnacle muscle fibres perfused with low [Ca2+]i (0.2-0.5 microM), but has little effect on the Na+0-dependent, ATP-independent Ca2+ efflux from fibres with a high [Ca]i (2-5 microM). Nevertheless, ATP depletion or vanadate treatment of high [Ca2+]i fibres causes an approximately 50-fold increase of Ca2+ efflux into Ca2+-containing lithium seawater. These results demonstrate that both vanadate and ATP affect Ca2+ extrusion, including the Na+0-dependent Ca2+ efflux (Na-Ca exchange), in barnacle muscle.     

Aberrant CFTR-dependent HCO3- transport in mutations associated with cystic fibrosis

Cystic fibrosis (CF) is a disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR). Initially, Cl- conductance in the sweat duct was discovered to be impaired in CF, a finding that has been extended to all CFTR-expressing cells. Subsequent cloning of the gene showed that CFTR functions as a cyclic-AMP-regulated Cl- channel; and some CF-causing mutations inhibit CFTR Cl- channel activity. The identification of additional CF-causing mutants with normal Cl- channel activity indicates, however, that other CFTR-dependent processes contribute to the disease. Indeed, CFTR regulates other transporters, including Cl(-)-coupled HCO3- transport. Alkaline fluids are secreted by normal tissues, whereas acidic fluids are secreted by mutant CFTR-expressing tissues, indicating the importance of this activity. HCO3- and pH affect mucin viscosity and bacterial binding. We have examined Cl(-)-coupled HCO3- transport by CFTR mutants that retain substantial or normal Cl- channel activity. Here we show that mutants reported to be associated with CF with pancreatic insufficiency do not support HCO3- transport, and those associated with pancreatic sufficiency show reduced HCO3- transport. Our findings demonstrate the importance of HCO3- transport in the function of secretory epithelia and in CF.     

Multiple transport modes of the cardiac Na+/Ca2+ exchanger

The cardiac Na+/Ca2+ exchanger (NCX1; ref. 2) is a bi-directional Ca2+ transporter that contributes to the electrical activity of the heart. When, and if, Ca2+ is exported or imported depends on the Na+/Ca2+ exchange ratio. Whereas a ratio of 3:1 (Na+:Ca2+) has been indicated by Ca2+ flux equilibrium studies, a ratio closer to 4:1 has been indicated by exchange current reversal potentials. Here we show, using an ion-selective electrode technique to quantify ion fluxes in giant patches, that ion flux ratios are approximately 3.2 for maximal transport in either direction. With Na+ and Ca2+ on both sides of the membrane, net current and Ca2+ flux can reverse at different membrane potentials, and inward current can be generated in the absence of cytoplasmic Ca2+, but not Na+. We propose that NCX1 can transport not only 1 Ca2+ or 3 Na+ ions, but also 1 Ca2+ with 1 Na+ ion at a low rate. Therefore, in addition to the major 3:1 transport mode, import of 1 Na+ with 1 Ca2+ defines a Na+-conducting mode that exports 1 Ca2+, and an electroneutral Ca2+ influx mode that exports 3 Na+. The two minor transport modes can potentially determine resting free Ca2+ and background inward current in heart.     

新疆博斯腾湖周围白刺植物下土壤盐分组成及分布特征研究

文章在野外考察、GPS定点和土壤采样分析的基础上,借助Excel、SPSS13等统计软件对新疆博斯腾湖周围白刺植物下土壤总碱度、盐离子含量以及盐离子间相关性作了探讨。结果表明,土壤pH值为8．47（平均值）,呈碱性。在0～15cm土层盐分含量2．775％,15～45cm土层盐分含量4．506％,45～70cm土层盐分含量3．096％;不同深度的盐分含量依次为15～45cm＞45～70cm＞0～15cm,各阴离子含量为 SO42-＞HCO3-＞Cl-,而CO32-在实验中未检测到,阳离子含量为K＋＋Na＋＞Ca2＋＞Ma2＋。在0～15cm土层Ca2＋与K＋＋Na＋达极显著正相关,Ca2＋与HCO3-呈显著正相关,Mg2＋与Ca2＋、K＋＋Na＋呈显著正相关,pH值与HCO3-、Mg2＋呈显著反相关;在15～45cm土层Ca2＋与K＋＋Na＋达极显著正相关,HCO3-与Cl-呈显著正相关,pH值与SO42-、HCO3-、K＋＋Na＋呈显著正相关;而在45～70cm土层Ca2＋与K＋＋Na＋呈显著正相关,SO42-与Ca2＋、K＋＋Na＋、Mg2＋呈显著正相关。     

张家口柴宣盆地浅层地下水水化学特征及水质评价

地下水作为张家口地区的主要供水水源,近年来受人类活动影响水质有恶化的趋势,影响了当地的用水安全,确定地下水化学特征与水质状况对水资源合理利用具有重要意义。基于2020年水质数据,选取Ca²⁺、Mg²⁺、K⁺、Na⁺、HCO^-₃、TH、pH、TDS、SO₄^2-、Cl^-、Al³⁺、NO^-₃、F^-、Cr⁶⁺等水质因子,通过数理统计法、Piper三线图、Gibbs图和岩石风化端元图对张家口柴宣盆地地区浅层地下水水化学特征进行分析,并采用模糊综合评价和改进内梅罗指数法对水质进行评价。结果表明：研究区地下水是Ca²⁺和HCO^-₃为主的弱碱性淡水,微硬水、硬水和极硬水分别占39.51%、34.57%和25.93%;沿地下水流向,地下...     

Control of dynamic CFTR selectivity by glutamate and ATP in epithelial cells

Cystic fibrosis is caused by mutations in cystic fibrosis transmembrane conductance regulator (CFTR), an anion channel. Phosphorylation and ATP hydrolysis are generally believed to be indispensable for activating CFTR. Here we report phosphorylation- and ATP-independent activation of CFTR by cytoplasmic glutamate that exclusively elicits Cl-, but not HCO3-, conductance in the human sweat duct. We also report that the anion selectivity of glutamate-activated CFTR is not intrinsically fixed, but can undergo a dynamic shift to conduct HCO3- by a process involving ATP hydrolysis. Duct cells from patients with DeltaF508 mutant CFTR showed no glutamate/ATP activated Cl- or HCO3- conductance. In contrast, duct cells from heterozygous patients with R117H/DeltaF508 mutant CFTR also lost most of the Cl- conductance, yet retained significant HCO3- conductance. Hence, not only does glutamate control neuronal ion channels, as is well known, but it can also regulate anion conductance and selectivity of CFTR in native epithelial cells. The loss of this uniquely regulated HCO3- conductance is most probably responsible for the more severe forms of cystic fibrosis pathology.     

Extrusion of calcium from rod outer segments is driven by both sodium and potassium gradients

Calcium is transported across the surface membrane of both nerve and muscle by a Na+-dependent mechanism, usually termed the Na:Ca exchange. It is well established from experiments on rod outer segments that one net positive charge enters the cell for every Ca2+ ion extruded by the exchange, which is generally interpreted to imply an exchange stoichiometry of 3 Na+:1 Ca2+. We have measured the currents associated with the operation of the exchange in both forward and reversed modes in isolated rod outer segments and we find that the reversed mode, in which Ca2+ enters the cell in exchange for Na+, depends strongly on the presence of external K+. The ability of changes in external K+ concentration ([K+]o) to perturb the equilibrium level of [Ca2+]i indicates that K+ is co-transported with calcium. From an examination of the relative changes of [Ca2+]o, [Na+]o, [K+]o and membrane potential required to maintain the exchange at equilibrium, we conclude that the exchange stoichiometry is 4 Na+:1 Ca2+, 1 K+ and we propose that the exchange should be renamed the Na:Ca, K exchange. Harnessing the outward K+ gradient should allow the exchange to maintain a Ca2+ efflux down to levels of internal [Ca2+] that are considerably lower than would be possible with a 3 Na+:1 Ca2+ exchange.     

正常麻醉犬动脉血和脑脊液酸碱电解质关系的初步分析

３０只健康麻醉犬动脉血和脑脊液（ＣＳＦ）酸碱变量及主要电解质检测结果分析发现：ＣＳＦｐＨ、［Ｋ~＋］＜ｐＨａ和［Ｋ~＋］_ａ，相互之间相关显著，其回归方程分别为ＣＳＦｐＨ＝０．４７７×ｐＨａ＋３．７８，ＣＳＦ［Ｋ~＋］＝０．４１８×［Ｋ~＋］_ａ＋１．１２。ＣＳＦＰＣＯ２、［Ｃｌ~－］、［乳酸］［Ｌａｃｔ］＞ＰａＣＯ２、［Ｃｌ－］_ａ和［Ｌａｃｔ］_ａ，相互之间相关显著。其回归方程分别为ＣＳＦＰＣＯ２＝０．５６４×ＰａＣＯ２＋３．５３２５，ＣＳＦ［Ｃｌ~－］＝０．５×［Ｃｌ~－］_ａ＋７６．７，ＣＳＦ［Ｌａｃｔ］＋０．９５×［Ｌａｃｔ］_ａ＋１．０。ＣＳＦ［ＨＣＯ_３~－］与ＣＳＦＰＣＯ２相关显著，回归方程为ＣＳＦ［ＨＣＯ_３~－］＝１．８９８×ＣＳＦＰＣＯ２＋１０．６。     

Phorbol ester and diacylglycerol mimic growth factors in raising cytoplasmic pH

There is now good evidence that cytoplasmic pH (pHi) may have an important role in the metabolic activation of quiescent cells. In particular, growth stimulation of mammalian fibroblasts leads to a rapid increase in pHi (refs 3-6), due to activation of a Na+/H+ exchanger in the plasma membrane, and this alkalinization is necessary for the initiation of DNA synthesis. However, the mechanism by which mitogens activate the Na+/H+ exchanger to raise pHi is not known, although an increase in cytoplasmic free Ca2+ ([Ca2+]i) has been postulated as the primary trigger. We now present data suggesting that the Na+/H+ exchanger is set in motion through protein kinase C, a phospholipid- and Ca2+-dependent enzyme normally activated by diacylglycerol produced from inositol phospholipids in response to external stimuli. Using newly developed pH microelectrodes and fluorimetric techniques, we show that a tumour promoting phorbol ester and synthetic diacylglycerol, both potent activators of kinase C (refs 12-15), mimic the action of mitogens in rapidly elevating pHi in different cell types. Furthermore, we demonstrate that, contrary to previous views, an early rise in [Ca2+]i is not essential for the activation of Na+/H+ exchange and the resultant increase in pHi. Finally, we suggest that an alkaline pHi shift, mediated by Na+/H+ exchange, may be a common signal in the action of those hormones which elicit the breakdown of inositol phospholipids.     

云南高原湖泊—程海湖水和底泥干雨季各元素含量的变迁及其放线菌群

程海湖水的pH为9,总碱度40左右,属碱性湖泊。干季水中各水域的K~+、Na~+、Ca~(++)、Mg~(++)、Cl~-、HCO_3~-、PO_4~(3+)等离子含量均高于雨季,而SiO_2和F~-、CO_3~(2-)、SO_4~(2-)离子低于雨季。各水域底泥除K含量雨季高于干季外,其Na、Ca、Mg、S、P、Fe、Mn、Zn、Ni含量以及C/N比值,雨季均低于干季。从底泥中分离到的放线菌有18个属之多,其中对细菌有抑制作用的占40％,对真菌有抑制作用的占20％。20％左右的菌有分解脂肪、纤维活性。1～2％的菌还产生蛋白酶抑制剂。     

Mechanism of beta-adrenergic relaxation of smooth muscle.

The mechanism of beta-adrenergic relaxation was investigated in isolated smooth muscle cells. Beta-adrenergic agents stimulate cyclic AMP-dependent phosphorylation, enhance Na+/K+ transport and induce relaxation. The stimulation of Na+/K+ transport is obligatory for relaxation, and we suggest that this stimulation induces relaxation through enhanced Na+/Ca2+ exchange.     

Detection of K+ and Cl-channels from calf cardiac sarcolemma in planar lipid bilayer membranes

The ionic currents underlying the cardiac action potential are believed to be much more complex than those in nerve. During the cardiac action potential, various membrane channels control the flow of K+, Na+, Ca2+ and Cl- across the sarcolemma of cardiac muscle cells. Thus, it has become increasingly clear that a detailed knowledge of the mechanisms that activate (or inactivate) heart channels is required to understand cardiac excitability. We report here the use of planar lipid bilayer techniques to detect and characterize K+ and Cl- channels in purified heart sarcolemma membrane vesicles. We have identified four different types of channel on the basis of their selectivity, conductance and gating kinetics. We present in some detail the properties of a K+ channel and a Cl- channel. We have tentatively identified the K+ channel with the ix type of current found in Purkinje, myocardial ventricular and atrial fibres. The chloride channel might be related to the transient chloride current found in Purkinje fibres.