脑蛋白水解物对鼠缺血半暗带神经元Na+-K+-ATP酶活性影响

目的探讨脑蛋白水解物对大鼠急性脑缺血再灌注损伤后缺血半暗带神经元Na~+-K~+-ATP酶活性的影响。方法选取270只雄性Wistar大鼠随机分为假手术组、模型组、干预组,三组大鼠根据缺血再灌注不同时间又随机分为缺血2 h再灌注6 h、24 h、48 h、72 h、7 d五个亚组,每个亚组18只大鼠。采用线栓法制备大鼠缺血2 h再灌注模型,仅干预组采用药物进行干预。分别于再灌注6 h、24 h、48 h、72 h、7 d后观察大鼠神经症状评分,相应时间断头取脑,测定缺血半暗带脑组织Na~+-K~+-ATP酶活性、脑组织含水量、脑梗死范围的变化。结果模型组大鼠缺血半暗带神经元Na~+-K~+-ATP酶活性于6 h开始下降,48 h达最小值,72 h稍有回升,7 d趋于稳定;干预组大鼠缺血2 h后再灌注24 h、48 h、72 h各时间点Na~+-K~+-ATP酶活性与模型组大鼠相应时间点比较差异有统计学意义(P0.05)。结论脑蛋白水解物可保护大鼠急性脑缺血再灌注损伤后缺血半暗带神经元,该作用可能与提高缺血半暗带神经元Na~+-K~+-ATP酶活性有关。     

Some Properties of Erythrocyte Na+ -K+ -ATPase in Essenial Hypertension

The activity and some allosteric properties of Na⁺ -K⁺ -ATPase in erythrocytes and their membrane preparations (ghosts) from 57 patients with essential hypertension and 36 normotensive controls were studied. To reveal enzyme activity in whole erythrocytes the cells were pretreated with detergent Tween-20. It was found that in the patient erythrocytes the Na⁺ -K⁺ -ATPase activity was 33% less as compared to the control group. Moreover, in the patient erythmcytes the sensitivity of the enzyme to high concentrations of MgCI₂ was decreased. In contrast, no analogous changes of the enzyme were revealed in the patient ghosts. It is suggested that the erythrocytes of patients with essential hypertension contain an inhibitor of Na⁺ -K⁺ -ATPase.     

Metabolic basis for asthma and rhinitis: An integrated approach

Observations that suggest a metabolic basis for the association between asthma and rhinitis and their pathogenesis are reviewed. We have observed increased plasma lysophosphatidylcholine (LPC) levels and decreased (Na⁺-K⁺)-ATPase activity in the leukocytes of patients with asthma and rhinitis. In asthmatic patients leukocytes also showed significantly lower Ca⁺⁺-ATPase activity. LPC increases cell membrane permeability to Na⁺ and Ca⁺⁺, causes membrane depolarization, potentiates IgE response, promotes phagocytic activity, inhibits adenylate cyclase, and stimulates phosphodiesterase, thus decreasing cAMP levels in the cells. These changes, when accompanied by failure of Na/K and Ca pumps, would lead to increased cytosolic Ca⁺⁺ and enhanced release of mediators from the mast cells in the airway lumen, producing a state of airway inflammation. We have also observed abnormal fall in specific airway conductance at residual volume in these patients, thereby showing small airway obstruction in asthma as well as in rhinitis. This could be due to airway inflammation caused by mechanisms described above. Large airway obstruction seen in asthma could be due to failure of Ca pump, normal activity of which seems essential for proper efflux of Ca⁺⁺ from airway smooth muscle cells. We suspect that increased plasma LPC levels and failure of the homeostatic ionic pumps occur due to some defect in oxidative metabolism and suggest that asthma and rhinitis could be seen as metabolic disorders.     

Contribution of Elevated Peripheral Resistance to Acute Volume Loaded Hypertension in Dogs

ABSTRACT

The role of peripheral resistance in volume loaded hypertension was examined by measuring changes in hemodynamic and humoral factors during dextran infusion for one hour, and for three hours after stopping the infusion in 30 anesthetized dogs. The mean blood pressure was elevated significantly (118%, p<0. 01), accompanied by increased cardiac output (170%, p<0. 01) during volume loading. In the recovery period, cardiac output returned to the basal level while the total peripheral resistance gradually increased (118%, p<0. 01)contributing to the maintenance of high blood pressure (108%, p<0. 01). There was no significant correlation between the hematocrit change and the increase in peripheral resistance in the recovery period. The levels of plasma norepinephrine, epinephrine, renin activity and vasopressin were suppressed by volume loading. These results demonstrated the importance of increase in peripheral resistance for maintaining high blood pressure in this acute volume excess model. The participation of the renin-angiotensin system, the sympathetic nervous system and vasopressin were all excluded and vascular structural changes were most unlikely to be involved in this acute experiment. It remains unknown whether this conversion is attributable to the summation of local autoregulation or to some other factors.     

Relative abundance of alpha2 Na(+) pump isoform influences Na(+)-Ca(2+) exchanger currents and Ca(2+) transients in mouse ventricular myocytes

In the mouse, genetic reduction in the Na(+), K(+)-ATPase alpha1 or alpha2 isoforms results in different functional phenotypes: heterozygous alpha2 isolated hearts are hypercontractile, whereas heterozygous alpha1 hearts are hypocontractile. We examined Na(+)/Ca(2+) exchange (NCX) currents in voltage clamped myocytes (pipette [Na(+)]=15 mM) induced by abrupt removal of extracellular Na(+). In wild-type (WT) myocytes, peak exchanger currents were 0.59+/-0.04 pA/pF (mean+/-S.E.M., n=10). In alpha1(+/-) myocytes (alpha2 isoform increased by 54%), NCX current was reduced to 0.33+/-0.05 (n=9, P<0.001) indicating a lower subsarcolemmal [Na(+)]. In alpha2(+/-) myocytes (alpha2 isoform reduced by 54%), the NCX current was increased to 0.89+/-0.11 (n=8, P=0.03). The peak sarcolemmal Na(+) pump currents activated by abrupt increase in [K(+)](o) to 4 mM in voltage clamped myocytes in which the Na(+) pump had been completely inhibited for 5 min by exposure to 0 [K(+)](o) were similar in alpha1(+/-) (0.86+/-0.12, n=10) and alpha2(+/-) myocytes (0.94+/-0.08 pA/pF, n=16), and were slightly but insignificantly reduced relative to WT (1.03+/-0.05, n=24). The fluo-3 [Ca(2+)](i) transient (F/F(o)) in WT myocytes paced at 0.5 Hz was 2.18+/-0.09, n=34, was increased in alpha2(+/-) myocytes (F/F(o)=2.56+/-0.14, n=24, P=0.02), and was decreased in alpha1(+/-) myocytes (F/F(o)=1.93+/-0.08, n=28, P<0.05). Thus the alpha2 isoform rather than the alpha1 appears to influence Na(+)/Ca(2+) exchanger currents [Ca(2+)](i) transients, and contractility. This finding is consistent with the proposal that alpha2 isoform of the Na pump preferentially alters [Na(+)] in a subsarcolemmal micro-domain adjacent to Na(+)/Ca(2+) exchanger molecules and SR Ca(2+) release sites.     

Insulin does not regulate vascular smooth muscle Na+, K+-ATPase activity in rabbit aorta

Summary To determine whether insulin regulates vascular smooth muscle Na⁺, K⁺-ATPase activity and if impaired insulin stimulation of vascular smooth muscle Na⁺, K⁺-ATPase activity could be a cause of increased vascular reactivity to norepinephrine and angiotensin II in diabetic states, the effects of insulin on Na⁺, K⁺-ATPase activity were examined in normal rabbit aortic intima-media incubated with normal plasma glucose and myo-inositol levels for 30 min. Insulin at 100 U/ml (600 pmol/l) had no effect on Na⁺, K⁺-ATPase activity. At 250 U/ml it caused a 4.2±0.8% increase, and at 500 U/ml insulin caused a 17.7±1.4% increase in Na⁺, K⁺-ATPase activity that was completely inhibited by amiloride (1 mmol/l). Human insulin-like growth factor I (600 pmol/l) caused an 18.0±1.0% increase in Na⁺, K⁺-ATPase activity that was inhibited by amiloride. Insulin does not regulate (stimulate) aortic vascular smooth muscle Na⁺, K⁺-ATPase activity. Supraphysiological insulin concentrations, probably acting through an insulin-like growth factor I receptor, stimulate Na⁺/H⁺ exchange in aortic vascular smooth muscle and cause small secondary increases in Na⁺, K⁺-ATPase activity. In aortic intima-media incubated with normal plasma glucose and myo-inositol levels, endogenously released adenosine stimulates and maintains a component of resting Na⁺, K⁺-ATPase activity and stimulates acute increases in activity when norepinephrine (1 mol/l) or angiotensin II (100 nmol/l) is added. These adenosine-stimulated components of Na⁺, K⁺-ATPase activity are selectively inhibited when the medium glucose is raised to 30 mmol/l during a 30-min equilibration and 30-min incubation. Insulin (100 U/ml) added during the incubation had no effect on the alterations in Na⁺, K⁺-ATPase activity induced by glucose at an elevated plasma level. Impaired insulin stimulation of vascular smooth muscle Na⁺, K⁺-ATPase activity is not a possible cause for alterations in vascular reactivity in diabetes.     

Ouabain-Like and Non-Ouabain-Like Factors in Plasma of Patients with Essential Hypertension

Circulating inhibitor of Na⁺, K⁺-ATPase and ouabain-like immuno-reactivity were studied in patients with essential hypertension. In the plasma of patients, two types of Na⁺, K⁺-ATPase inhibitors (ouabain-like and non-ouabain-like inhibitors) and ouabain-like immunoreactivity were detected. Ouabain-like inhibitor was clearly detected at a low KC1 concentration (0.1 mM) in the assay buffer, and non-ouabain-like inhibitor was detected at a high KC1 concentration (10 mM). The plasma level of ouabain-like inhibitor correlated significantly with that of ouabain-like immunoreactivity (p<0.001) and with a mean blood pressure (p<0.01). The plasma level of non-ouabain-like inhibitor was not correlated with the levels of either ouabain-like immunoreactivity or mean blood pressure. The level of plasma ouabain-like inhibitor did not correlate with that of plasma non-ouabain like inhibitor. Both ouabain-like inhibitor and ouabain-like immunoreactivity in the plasma of patients with essential hypertension were significantly higher than those in normotensive subjects, but the plasma level of non-ouabain-like inhibitor in patients with essential hypertension was not higher than that in normotensive subjects. These results suggest that the plasma from patients with essential hypertension contains ouabain-like factor(s) which is important to mantain the high blood pressure.     

SIK1 is part of a cell sodium-sensing network that regulates active sodium transport through a calcium-dependent process

In mammalian cells, active sodium transport and its derived functions (e.g., plasma membrane potential) are dictated by the activity of the Na(+),K(+)-ATPase (NK), whose regulation is essential for maintaining cell volume and composition, as well as other vital cell functions. Here we report the existence of a salt-inducible kinase-1 (SIK1) that associates constitutively with the NK regulatory complex and is responsible for increases in its catalytic activity following small elevations in intracellular sodium concentrations. Increases in intracellular sodium are paralleled by elevations in intracellular calcium through the reversible Na(+)/Ca(2+) exchanger, leading to the activation of SIK1 (Thr-322 phosphorylation) by a calcium calmodulin-dependent kinase. Activation of SIK1 results in the dephosphorylation of the NK alpha-subunit and an increase in its catalytic activity. A protein phosphatase 2A/phosphatase methylesterase-1 (PME-1) complex, which constitutively associates with the NK alpha-subunit, is activated by SIK1 through phosphorylation of PME-1 and its dissociation from the complex. These observations illustrate the existence of a distinct intracellular signaling network, with SIK1 at its core, which is triggered by a monovalent cation (Na(+)) and links sodium permeability to its active transport.     

Program

Dietary potassium supplementation lowers blood pressure (BP) and attenuates complications in hypertensive subjects, particularly those with the low renin volume expanded (LRVE) variety. We and others have shown that the plasma level of a digitalis like substance (DLS) is elevated in this type of hypertension. We therefore, examined the effect of increases in dietary potassium on the plasma level of endogenous DLS, myocardial and renal Na⁺, K⁺-ATPase (NKA) activities, BP, and renal excretory function in reduced renal mass (RRM)-salt hypertension in the rat, a classical model of LRVE hypertension. 70% RRM rats were divided in 4 groups, namely those consuming: 1) a sodium free and normal potassium (1.3% as KCl) diet (RRM-0 Na), 2) a normal sodium and normal potassium diet (RRM-NaK), 3) a normal sodium and high potassium (2 X normal) diet (RRM-Na2K), and 4) a normal sodium and 4 times normal potassium diet (RRM-Na4K). At the end of 4 weeks of dietary treatment, direct BP was recorded, plasma level of DLS determined by bioassay and with a radioimmunoassay for digoxin (DIF) and myocardial and renal NKA activities were measured. As expected, compared to RRM-0Na rats, RRM-NaK rats developed hypertension. BP increased significantly less in RRM-Na2K, whereas BP did not increase in RRM-Na4K rats. Hypertension in RRM-NaK rats was associated with an increase in plasma DLS and DIF and decrease in renal and myocardial NKA activities. DLS was increased (DIF was not changed) and myocardial NKA also decreased in rats consuming double potassium. However, quadrupling potassium in the diet (RRM-Na4K) normalized DLS and DIF and increased myocardial and renal NKA activities, compared to RRM-0Na rats. Also compared to RRM-0Na, water consumption, urinary volume excretion, sodium, and potassium increased in the other 3 groups, more so in RRM-Na4K rats. These data show that quadrupling the potassium in the diet prevents the BP increase in RRM rats and this is associated with diuresis/natriuresis and normalization of DLS, perhaps because the diuresis/natriuresis normalizes blood volume.     

Preparation and characterization of plasma membrane of cardiac tissue

A method of preparing highly purified plasma membrane from cardiac muscle is described using continuous ficoll-sucrose density gradient centrifugation in zonal rotors. 5′-Nucleotidase and ATPase (Mg²⁺ dependent, Na⁺ + K⁺ stimulated and ouabain inhibited) were found to be concentrated in the plasma membrane fraction. At 11% ficollsucrose Ca²⁺ and (Ca²⁺ + K⁺) stimulation of the ATPase was very small (13%) and very low level of succinic dehydrogenase activity was present. Electron microscopic studies of the fraction revealed the presence of membrane vesicles and sheets. Composition of these membranes was analyzed by 7% polyacrylamide gel electrophoresis in the presence of 1% SDS. One protein band of 68 000 Daltons accounted for 18% of protein on the gel. Lipid and carbohydrate stains revealed six and two bands respectively. Studies on the conformation of proteins within membranes were achieved using circular dichroism (CD) and optical rotatory dispersion (ORD) techniques. Corrections were applied using the pseudo reference state approach.     

Alterations of Na/K-ATPase function in caveolin-1 knockout cardiac fibroblasts

Recent studies have demonstrated that the Na⁺/K⁺-ATPase is not only an ion pump, but also a membrane receptor that confers the ligand-like effects of cardiotonic steroids (CTS) such as ouabain on protein kinases and cell growth. Because CTS have been implicated in cardiac fibrosis, this study examined the role of caveolae in the regulation of Na⁺/K⁺-ATPase function and CTS signaling in cardiac fibroblasts. In cardiac fibroblasts prepared from wild-type and caveolin-1 knockout [Cav-1(−/−)] mice, we found that the absence of caveolin-1 did not affect total cellular amount or surface expression of Na⁺/K⁺-ATPase α1 subunit. However, it did increase ouabain-sensitive ⁸⁶Rb⁺ uptake. While knockout of caveolin-1 increased basal activities of Src and ERK1/2, it abolished the activation of these kinases induced by ouabain but not angiotensin II. Finally, ouabain stimulated collagen synthesis and cell proliferation in wild type but not Cav-1(−/−) cardiac fibroblasts. Thus, we conclude that caveolae are important for regulating both pumping and signal transducing functions of Na⁺/K⁺-ATPase. While depletion of caveolae increases the pumping function of Na⁺/K⁺-ATPase, it suppresses CTS-induced signal transduction, growth, and collagen production in cardiac fibroblasts.     

Kinetics of Red Cell Na+ and K+ Transport in Prague Hypertensive Rats

Kinetics of ouabain-sensitive, furosemide-sensitive (FS), bumetanide-sensitive (BS) and -resistant Na⁺ and K⁺ transport were studied in erythrocytes of Prague hypertensive rats (PHR) and Prague normotensive rats (PNR). Maximal transport rates (V_max) and apparent affinities for either intracellular Na⁺ or extracellular K⁺ (replaced by Rb⁺) were determined in red cells in which Na⁺ content varied around the physiological range and that were incubated in Na⁺ media. No major differences between PHR and PNR were disclosed in the kinetics of ion transport mediated by the Na⁺-K⁺ pump or BS inward Na⁺-K⁺ cotransport. FS Rb⁺ uptake was higher (due to a greater V_max) in red cells of PHR as compared to PNR. In cells with a lowered Na⁺ content this elevation of FS Rb⁺ uptake was largely due to an augmented K⁺-Cl^? cotransport which exhibits a low affinity for Rb⁺_o and is blocked by 1 mM furosemide but not by 10 μM bumetanide. Red cells of PHR and PNR strains did not differ in either Na⁺ or Rb⁺ leaks. A slight increase of red cell Na⁺ content in PHR was evaluated in terms of the pump-leak concept. The present study did not reveal any obvious kinetic abnormalities of red cell cation transport the presence of which in tissues involved in blood pressure regulation would favor the development or the maintenance of genetic hypertension in PHR.     

Na+-K+ pump inhibition caused by chronic amiodarone in guinea pig myocardium

Summary Although it is known that amiodarone inhibits myocardial Na⁺-K⁺ pump activity, the potency and the time course of this inhibition are unknown. The aim of this study was to investigate these aspects with reference to digoxin, using guinea pigs treated with either intraperitoneal amiodarone (20mg/kg per week, up to 12 weeks,n = 26) or the same amount of vehicle as a control (n = 24). ECG recording and microelectrode experiments were conducted every 2 weeks. QT interval corrected by heart rate and action potential duration were prolonged as a function of the time of exposure to amiodarone. Hyperpolarization observed immediately after the overdrive (1.0Hz) termination or K⁺-replenishment following K⁺-depletion in the presence of 0.1mM Ba²⁺ was compared in the amiodarone-treated and untreated groups, as an index of the Na⁺-K⁺ pump activity. The resting membrane potential recovery from overdrive-induced depolarization was slower and the amplitude of K⁺-induced hyperpolarization was smaller in the amiodarone-treated group than in the untreated group. These changes were evident as the chronic amiodarone treatment progressed, although the changes in these parameters were greater in the case of acute application of 50µM digoxin. In conclusion, this study indicates that treatment with amiodarone for longer than several weeks moderately inhibits the myocardial Na⁺-K⁺ pump.This work was partly supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan (11877125).     

Assay of a Circulating Sodium Pump Inhibitor in Patients with Essential Hypertension and Normotensive Subjects

The plasma levels of a sodium pump inhibitor (Na⁺PI) were measured by a modified method of Hamlyn et al, using dog kidney Na⁺, K⁺-ATPase. When the level of Na⁺PI was expressed as the % inhibition of the enzyme and compared with that of a control solution, it was found to be 9.0 ± 0.7% in 43 untreated patients with essential hypertension. This was significantly higher than 5.0 ± 0.4% for 56 normotensive subjects (p < 0.01). Male patients with essential hypertension showed the highest mean value of 10.5 ± 1.1%, disclosing an apparent sex difference in the patient group (p< 0.01). Only in female patients was there a significant positive correlation between the inhibitor's level and the mean blood pressure (r = 0.649, p < 0.01). These results provided additional evidence for increased Na⁺ PI in the plasma of patients with essential hypertension, which might bear an important role in the pathogenesis of the disease.     

Endoxin Antagonist Lessens Myocardial Ischemia Reperfusion Injury

OBJECTIVE: To elucidate whether endoxin is one of important factors involved in myocardial ischemia reperfusion (MIR) injury, the change of myocardial endoxin level was determined in rats with MIR injury model and the effects of anti-digoxin antiserum (ADA), an endoxin specific antagonist, on MIR injury were studied. METHODS: MIR injury model was obtained by ligating left anterior descending coronary artery 30 min followed by 45 min reperfusion. Sprague-Dawley rats were randomly divided into six groups of 10 rats, each. Sham group, MIR group, normal saline group, ADA 9, 18 and 36 mg.kg(-1). ECG was continuously recorded. After reperfusion left ventricular myocardium samples of ischemic area were processed immediately. Myocardial endoxin level, Na(+)-K(+)-ATPase, Ca(2+)-ATPase, Mg(2+)-ATPase activities, and intramitochondrial Ca(2+) content were measured. RESULTS: Myocardial endoxin level was significantly increased; Na(+)-K(+)-ATPase, Ca(2+)-ATPase, and Mg(2+)-ATPase activities were remarkably decreased; intramitochondrial Ca(2+) content was remarkably raised; ST segments of ECG were significantly elevated and occurrence and scores of ventricular arrhythmias were significantly increased in early stage of reperfusion in rats with MIR. In all groups with ADA, myocardial endoxin level was remarkably decreased; Na(+)-K(+)-ATPase, Ca(2+)-ATPase and Mg(2+)-ATPase activities were drastically increased; intramitochondrial Ca(2+) content was declined; ST segments and ventricular arrhythmias were improved. CONCLUSION: Myocardial endoxin level was increased in MIR, which implies that the elevated endoxin may be one of major factors inducing MIR injury. This postulate is supported by the observation that ADA has protective and therapeutic effects against MIR injury probably by antagonizing the action of endoxin. The underlying mechanism may be ascribed to restoration of energy metabolism, and attenuation of intracellular Ca(2+) overload.     

The Effect of High Salt Intake on Na+, K+-Atpase Activity of Tissues in the Rat

1. The effect of chronic feeding of high salt diet on Na⁺, K⁺-ATPase activity of heart, liver, skeletal muscle, kidney and aorta was studied in the rat.

2. Groups of rats were either given tap water or 18 g/L saline to drink. After 7 days, 3 months or 12 months, the control group and salt loaded     

Inhibition of Na+, K+-ATPase Activity by the Metabolites Accumulating in Homocystinuria

Homocystinuria is an inborn error of sulfur amino acid metabolism characterized predominantly by vascular and nervous system dysfunction. In this study we determined the in vitro effects of homocysteine and methionine, metabolites which accumulate in homocystinuria, on Na⁺, K⁺-ATPase, and Mg²⁺-ATPase activities in synaptic membranes from the hippocampus of rats. The results showed that both metabolites significantly inhibit Na⁺, K⁺-ATPase but not Mg²⁺-ATPase activity at concentrations usually observed in plasma of homocystinuric patients. Furthermore, incubation of hippocampal homogenates with homocysteine also elicited an inhibition of the enzyme activity which was however prevented by the simultaneous addition of cysteine to the medium. In addition, cysteine or methionine per se did not modify the two enzymatic activities. These findings indicate that oxidation of critical groups in the enzyme may possibly be involved in homocysteine inhibitory effect. Moreover, kinetic studies performed to investigate the interaction between homocysteine and methionine on Na⁺, K⁺-ATPase inhibition suggested a common site for the two amino acids in the enzyme. Considering the critical role exerted by Na⁺, K⁺-ATPase in brain, it is proposed that the inhibition provoked by homocysteine and methionine on the enzyme activity may be possibly related to the brain dysfunction characteristic of homocystinuria.     

Cytosolic pH regulation in density-defined subpopulations of bronchoalveolar macrophages

Bronchoalveolar macrophages (m) represent a heterogeneous population of morphologically and functionally distinct cells. In mixed populations of bronchoalveolar m, cytosolic pH (pH _i ) regulation has been shown to involve both Na⁺-dependent and -independent mechanisms for H⁺ extrusion, i.e., passive H⁺ extrusion in exchange for extracellular Na⁺ (Na⁺-H⁺ exchange or NHE) and active H⁺ extrusion by plasmalemmal vacuolar-type H⁺-ATPase (V-ATPase), respectively. The present studies explored the possibility that individual subpopulations of bronchoalveolar m possess distinct ensembles of H⁺ extrusion mechanisms. Rabbit bronchoalveolar m were separated into five density-defined subpopulations using a discontinuous density gradient. Scanning and transmission electron microscopy revealed morphological differences between the subpopulations. The number of plasmalemmal projections and electron-dense inclusions increased with increments in cell density. The subpopulations were also functionally distinct. Fe receptor-mediated phagocytosis increased in the increasing density subpopulations. Despite these differences, all subpopulations displayed Na⁺-dependent and -independent mechanisms for pH _i recovery from intracellular acid loads (ammonia prepulse technique). We conclude that NHE and V-ATPase activities were present in each subpopulation. These findings support the use of mixed populations to study pH _i homeostasis in bronchoalveolar m. Offprint requests to: Akhil Bidani