pH regulation in ileum: Na(+)-H+ and Cl(-)-HCO3- exchange in isolated crypt and villus cells

1991 ◽  
Vol 260 (3) ◽  
pp. G440-G449 ◽  
Author(s):  
U. Sundaram ◽  
R. G. Knickelbein ◽  
J. W. Dobbins
Keyword(s):  
Intracellular Ph ◽  
Ph Regulation ◽  
Second Messengers ◽  
Cell Types ◽  
Current Evidence ◽  
Leucine Incorporation ◽  
Trypan Blue Exclusion ◽  
Morphological Criteria ◽  
Acid Load ◽  
Separation Cell

Current evidence suggests that intestinal crypt and villus cells have different functions in electrolyte transport. To study the regulation of transporters, we isolated and separated these two cell types. This was accomplished by sequential collection of enterocytes from rabbit ileal loops incubated with buffered solutions of calcium chelators. Alkaline phosphatase and thymidine kinase activity, sodium-glucose cotransport, and morphological criteria were used to determine cell separation. Cell viability was evaluated with trypan blue exclusion, leucine incorporation into protein, and morphological features. The role of Na(+)-H+ and Cl(-)-HCO3- exchange in the regulation of intracellular pH was analyzed using an intracellular pH sensitive dye, BCECF. Removal of external Na+ or the addition of amiloride resulted in acidification of both crypt and villus cells. Removal of Cl- or the addition of DIDS resulted in alkalinization of both cell types. The cells could be acidified with NH4Cl, and recovery from this acid load was dependent on Na+ and inhibited by amiloride. Similarly, the cells could be alkalinized with propionate and recovery was Cl- dependent and DIDS sensitive. These data are consistent with the presence of Na(+)-H+ and Cl(-)-HCO3- exchange in both crypt and villus cells. Both exchanges appear to be involved in the regulation of basal pH as well as in recovery from alterations in intracellular pH. Having demonstrated the presence of Na(+)-H+ and Cl(-)-HCO3- exchange activity in both crypt and villus cells, we can now use these cells to determine the regulation of these exchangers by intracellular second messengers.

Intracellular pH regulation in the rabbit cortical collecting duct A-type intercalated cell

1997 ◽  
Vol 273 (3) ◽  
pp. F340-F347 ◽  
Author(s):  
A. E. Milton ◽  
I. D. Weiner
Keyword(s):  
Intracellular Ph ◽  
Ph Regulation ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
Bafilomycin A1 ◽  
Proton Secretion ◽  
Acid Load ◽  
A Cell ◽  
Phi Regulation ◽  
Intracellular Acid

The A cell may possess multiple H+ transporters, including H(+)-adenosinetriphosphatase (H(+)-ATPase) and H(+)-K(+)-ATPase. The current study examines the relative roles of proton transporters in the A cell by observing their contribution to both basal intracellular pH (pHi) regulation and pHi recovery from an intracellular acid load. CCD were studied using in vitro microperfusion, and pHi was measured in the individual A cell using the fluorescent, pH-sensitive dye, 2',7'-bis(carboxyethyl)-5(6)-carboxy-fluorescein (BCECF). Inhibiting H(+)-ATPase with luminal bafilomycin A1 decreased basal pHi, whereas inhibiting apical H(+)-K(+)-ATPase with either luminal Sch-28080 or luminal potassium removal did not. The predominant mechanism of pHi, recovery from an intracellular acid load was peritubular sodium dependent and peritubular ethylisopropylamiloride (EIPA) sensitive, identifying basolateral Na+/H+ exchange activity. In the absence of peritubular sodium, pHi recovery was inhibited by luminal bafilomycin A1 but not by luminal Sch-28080 addition or by luminal potassium removal. However, when Na+/H+ exchange was inhibited with EIPA, both bafilomycin A1 sensitive and potassium dependent, Sch-28080-sensitive components of pHi recovery were present. Quantitatively, the rate of H(+)-ATPase proton secretion was greater than the rate of H(+)-K(+)-ATPase proton secretion. We conclude that basolateral Na+/H+ exchange is the predominant mechanism of A cell pHi recovery from an intracellular acid load. An apical H(+)-ATPase is the primary apical transporter contributing to A cell pHi regulation. An apical H(+)-K(+)-ATPase, while present, plays a more limited role under the conditions tested.

scholarly journals Regulation of intracellular pH during oocyte growth and maturation in mammals

Reproduction ◽  
2009 ◽  
Vol 138 (4) ◽  
pp. 619-627 ◽  
Author(s):  
Greg FitzHarris ◽  
Jay M Baltz
Keyword(s):  
Intracellular Ph ◽  
Ph Regulation ◽  
Cell Types ◽  
Mammalian Development ◽  
Ph Homeostasis ◽  
Oocyte Growth ◽  
Homeostatic Process ◽  
Mapk Signalling ◽  
Recent Developments ◽  
Early Mammalian Development

Regulation of intracellular pH (pHi) is a fundamental homeostatic process essential for the survival and proliferation of virtually all cell types. The mammalian preimplantation embryo, for example, possesses Na+/H+and HCO3−/Cl−exchangers that robustly regulate against acidosis and alkalosis respectively. Inhibition of these transporters prevents pH corrections and, perhaps unsurprisingly, leads to impaired embryogenesis. However, recent studies have revealed that the role and regulation of pHiis somewhat more complex in the case of the developing and maturing oocyte. Small meiotically incompetent growing oocytes are apparently incapable of regulating their own pHi, and instead rely upon the surrounding granulosa cells to correct ooplasmic pH, until such a time that the oocyte has developed the capacity to regulate its own pHi. Later, during meiotic maturation, pHi-regulating activities that were developed during growth are inactivated, apparently under the control of MAPK signalling, until the oocyte is successfully fertilized. Here, we will discuss pH homeostasis in early mammalian development, focussing on recent developments highlighting the unusual and unexpected scenario of pH regulation during oocyte growth and maturation.

Potential-induced changes in intracellular pH

1994 ◽  
Vol 266 (5) ◽  
pp. F685-F696 ◽  
Author(s):  
V. Lyall ◽  
T. U. Biber
Keyword(s):  
Metal Ions ◽  
Intracellular Ph ◽  
Second Messengers ◽  
Strong Dependence ◽  
Cell Types ◽  
Divalent Metal ◽  
Divalent Metal Ions ◽  
Low Concentrations ◽  
Intracellular Second Messengers ◽  
Induced Changes

In a variety of cell types and tissues there is a strong dependence of intracellular pH (pHi) on membrane potential (Vm). Since cell Vm values can be altered by hormones, ion concentrations, and changes in membrane conductances, the potential-dependent changes in pHi may serve as an important mechanism by which cells can alter their pHi to an environmental stimulus. The H+ flux across the cell membranes is thought to take place via putative H+ channels that are blocked by low concentrations of divalent metal ions. However, in Na(+)-transporting epithelia, a major part of the H+ flux seems to be via the amiloride-sensitive apical Na+ channels, which are not sensitive to divalent metal ions. The H+ flux via the Na+ channels can be modulated by natriferic hormones and intracellular second messengers. The H(+)-conductive pathways may play an important role in signal transduction in some cells.

Intracellular pH regulation in trout urinary bladder epithelium: Na(+)-H+(NH4+) exchange

1991 ◽  
Vol 261 (3) ◽  
pp. R652-R658
Author(s):  
W. S. Marshall ◽  
S. E. Bryson
Keyword(s):  
Urinary Bladder ◽  
Intracellular Ph ◽  
Ph Regulation ◽  
Electrochemical Gradient ◽  
Bladder Epithelium ◽  
Basolateral Side ◽  
Acid Load ◽  
Mucosal Side ◽  
Phi Regulation

We measured intracellular pH (pHi) of single epithelial cells in situ in the urinary bladder epithelium using microspectrofluorometry and the cytoplasmically trapped pH-sensitive fluorophore, 2',7'-bis(2-carboxyethyl)-5(6)- carboxyfluorescein (BCECF). The resting pHi was 7.21 +/- 0.03 (n = 40 bladders, 489 cells) in pH 7.8 bathing solutions, indicating that H+ is not passively distributed across the plasma membrane and is extruded against its electrochemical gradient. Whereas exposure to hypercapnia (5% CO2 saturation) reversibly decreased pHi, mucosally added 20 mM NH4+ reversibly increased pHi. Recovery from the NH4+ effect was slow and lacked an acid-load pHi undershoot; this is interpreted as suggesting significant NH4+ permeability. Recovery from hypercapnic acidosis was blocked by mucosally added amiloride, indicating that apical Na(+)-H+ exchange is involved in pHi regulation. Addition of 0.5 mM NH4+ to the basolateral side when the mucosal side was bathed in mock urine (2 mM NaCl) significantly increased undirectional mucosal-to-serosal Na+ flux, and the increase was blocked by mucosally added amiloride. We conclude that an apically located Na(+)-H+ exchange is important in pHi regulation and may also accept NH4+ as the counterion for Na+.

scholarly journals Intracellular pH Regulation in Cultured Astrocytes from Rat Hippocampus

1997 ◽  
Vol 110 (4) ◽  
pp. 453-465 ◽  
Author(s):  
Mark O. Bevensee ◽  
Regina A. Weed ◽  
Walter F. Boron
Keyword(s):  
Intracellular Ph ◽  
Ph Regulation ◽  
Disulfonic Acid ◽  
Bath Solution ◽  
Major Cation ◽  
Cultured Astrocytes ◽  
Buffering Power ◽  
Acid Load ◽  
Acid Extrusion ◽  
Best Fit

We studied the regulation of intracellular pH (pHi) in single cultured astrocytes passaged once from the hippocampus of the rat, using the dye 2′,7′-biscarboxyethyl-5,6-carboxyfluorescein (BCECF) to monitor pHi. Intrinsic buffering power (βI) was 10.5 mM (pH unit)−1 at pHi 7.0, and decreased linearly with pHi; the best-fit line to the data had a slope of −10.0 mM (pH unit)−2. In the absence of HCO3−, pHi recovery from an acid load was mediated predominantly by a Na-H exchanger because the recovery was inhibited 88% by amiloride and 79% by ethylisopropylamiloride (EIPA) at pHi 6.05. The ethylisopropylamiloride-sensitive component of acid extrusion fell linearly with pHi. Acid extrusion was inhibited 68% (pHi 6.23) by substituting Li+ for Na+ in the bath solution. Switching from a CO2/HCO3−-free to a CO2/HCO3−-containing bath solution caused mean steady state pHi to increase from 6.82 to 6.90, due to a Na+-driven HCO3− transporter. The HCO3−-induced pHi increase was unaffected by amiloride, but was inhibited 75% (pHi 6.85) by 400 μM 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS), and 65% (pHi 6.55–6.75) by pretreating astrocytes for up to ∼6.3 h with 400 μM 4-acetamide-4′-isothiocyanatostilbene-2,2′-disulfonic acid (SITS). The CO2/HCO3−-induced pHi increase was blocked when external Na+ was replaced with N-methyl-d-glucammonium (NMDG+). In the presence of HCO3−, the Na+-driven HCO3− transporter contributed to the pHi recovery from an acid load. For example, HCO3− shifted the plot of acid-extrusion rate vs. pHi by 0.15–0.3 pH units in the alkaline direction. Also, with Na-H exchange inhibited by amiloride, HCO3− increased acid extrusion 3.8-fold (pHi 6.20). When astrocytes were acid loaded in amiloride, with Li+ as the major cation, HCO3− failed to elicit a substantial increase in pHi. Thus, Li+ does not appear to substitute well for Na+ on the HCO3− transporter. We conclude that an amiloride-sensitive Na-H exchanger and a Na+-driven HCO3− transporter are the predominant acid extruders in astrocytes.

Influence of surface pH on intracellular pH regulation in cardiac and skeletal muscle

1986 ◽  
Vol 250 (5) ◽  
pp. C748-C760 ◽  
Author(s):  
B. Vanheel ◽  
A. de Hemptinne ◽  
I. Leusen
Keyword(s):  
Intracellular Ph ◽  
Soleus Muscle ◽  
Ph Regulation ◽  
Purkinje Fiber ◽  
Intracellular Acidification ◽  
Surface Ph ◽  
Acid Load ◽  
Rat Soleus Muscle ◽  
Rate Of Recovery ◽  
Intracellular Acid

The influence of the surface pH (pHs) on the intracellular pH (pHi) and the recovery of pHi after an imposed intracellular acid load was investigated in isolated sheep cardiac Purkinje fiber, rabbit papillary muscle, and mouse and rat soleus muscle. pHs and pHi, respectively, were continuously measured by use of single- and double-barreled pH-sensitive glass microelectrodes. Surface acidosis, usually obtained by superfusion with solutions of acid pH, was also produced with low buffered (5 mM N-2-hydroxyethylpiperazine-N'-2-ethane-sulfonic acid) solutions at control pH. The pHs decrease (delta pHs) induced by low buffering was smallest (-0.08 pH unit) in Purkinje fiber and largest (-0.31 pH unit) in rat soleus muscle, which already had a more acid surface in control conditions. delta pHs was somewhat dependent on the superfusion rate. Higher superfusion rates decreased but did not abolish delta pHs. Surface acidosis was associated with a small intracellular acidification. Intracellular acid loads were produced by adding and subsequently withdrawing 20 meq/l NH4+ from the superfusate. In all preparations, the rate of recovery of pHi after NH4+ withdrawal was notably decreased at acidified pHs. This effect was amiloride sensitive. It is concluded that, in superfused multi-cellular preparations, pHs and therefore the buffer concentration of a superfusate can considerably influence steady-state pHi and pHi recovery from an imposed intracellular acid load.

Intracellular pH regulation in isolated myocytes from adult rat heart in HCO-3-containing and HCO-3-free media

1993 ◽  
Vol 84 (2) ◽  
pp. 133-139 ◽  
Author(s):  
L. L. Ng ◽  
J. E. Davies ◽  
P. Quinn
Keyword(s):  
Intracellular Ph ◽  
Ventricular Myocytes ◽  
Ph Regulation ◽  
Specific Inhibitor ◽  
Buffering Capacity ◽  
Intracellular Acidosis ◽  
Rat Ventricular Myocytes ◽  
Adult Rat ◽  
Acid Load ◽  
Free Media

1. Using microfluorimetry, intracellular pH, buffering capacity and intracellular pH recovery from intracellular acidosis were determined in isolated adult rat ventricular myocytes, in buffers with and without HCO-3. 2. In nominally HCO-3-free media, the intracellular pH was higher than in HCO-3-containing media. Buffering capacity at resting intracellular pH and at a pH of about 6.3 was also lower in HCO-3-free media. 3. In HCO-3-free media, recovery from an acid load after an NH4C1 prepulse was almost completely inhibited by the Na+/H+ antiport activity specific inhibitor ethylisopropyl amiloride. However, in the presence of HCO-3, H+ efflux rate was enhanced, and ethylisopropyl amiloride led to only partial inhibition of H+ efflux. Complete inhibition was achieved only with further addition of the anion-transport inhibitor 4,4′-di-isothiocyanatostilbene-2,2′-disulphonate. 4. Thus, in adult rat ventricular myocytes, recovery from intracellular acidosis in the absence of HCO-3 was almost wholly due to Na+/H+ antiport activity. In the more physiological situation with HCO-3 present, a third of the recovery from an intracellular acid load was attributed to an additional external Na+-dependent di-isothiocyanatostilbene-disulphonate-sensitive H+ efflux.

Regulation of intracellular pH in two cell populations of inner stripe of rabbit outer medullary collecting duct

1993 ◽  
Vol 265 (3) ◽  
pp. F406-F415 ◽  
Author(s):  
I. D. Weiner ◽  
C. S. Wingo ◽  
L. L. Hamm
Keyword(s):  
Intracellular Ph ◽  
Cell Population ◽  
Collecting Duct ◽  
Organic Anion ◽  
Cell Types ◽  
Disulfonic Acid ◽  
Cell Populations ◽  
Acetoxymethyl Ester ◽  
Acid Load ◽  
Medullary Collecting Duct

The inner stripe of the outer medullary collecting duct (OMCDis) is a major site of HCO3- reabsorption and urinary acidification. Whether this nephron segment consists of a single or multiple cell types remains unclear. Apical incubation of rabbit OMCDis via luminal perfusion with 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester resulted in heterogeneous fluorescence, suggesting two cell types. This heterogeneity was not prevented by inhibition of either carbonic anhydrase or organic anion transport. Subsequent studies were directed at characterizing the major intracellular pH (pHi) regulatory transporters in these two cell populations. Both cell populations demonstrated similar rates of Na+/H+ exchange, as assessed by peritubular Na(+)-dependent, amiloride-sensitive pHi recovery from an intracellular acid load. In contrast, Na(+)-independent, HCO3(-)-independent pHi recovery from an acid load was present in both cell populations but had two to three times greater activity in a minority cell population. In vivo deoxycorticosterone acetate administration increases this rate in both populations but to a greater extent in the minority cell population. In CO2/HCO3(-)-containing solutions, Cl- removal from the peritubular solution caused 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid-sensitive alkalinization of all cells. Again, the magnitude and rate of alkalinization were significantly greater in the minority cell population. These studies demonstrate that the OMCDis consists of qualitatively similar cells in different states of functional activity. Although they are similar in most characteristics, a minority of cells more actively secrete H+ (independent of Na+) and reabsorb HCO3-.

Ionic requirements for intracellular pH regulation in rainbow trout hepatocytes

1986 ◽  
Vol 250 (1) ◽  
pp. R24-R29 ◽  
Author(s):  
P. J. Walsh
Keyword(s):  
Rainbow Trout ◽  
Intracellular Ph ◽  
Ph Regulation ◽  
Disulfonic Acid ◽  
Salmo Gairdneri ◽  
Exchange System ◽  
Exchange Processes ◽  
Acid Load ◽  
Rate Of Recovery ◽  
22Na Uptake

The ionic requirements for pH regulation in isolated rainbow trout (Salmo gairdneri) hepatocytes were determined by manipulation of intracellular pH (pHi; measured by the dimethyloxazolidinedione distribution technique) by NH4Cl prepulse and changes in external [CO2] in the presence and absence of various drugs and external ions. The presence of a Na+/H+(NH+4) exchange system is supported by the following results: 1) the rate of recovery from an acid load is decreased by amiloride (0.5 mM) or reduction of external [Na+]; 2) the rate of 22Na uptake is increased during recovery from an acid load, and this increase in amiloride sensitive. The presence of a Cl-/HCO3- exchange system is supported by the observations that 1) pHi is increased, and 2) rates of recovery of pHi from acid loading are enhanced, by exposure to 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (0.5 mM) and reductions in external [Cl-]. Further studies are required to determine the role of these exchange processes during physiological pHi perturbations.

Effect of ursodeoxycholic acid on intracellular pH regulation in isolated rat bile duct epithelial cells

1993 ◽  
Vol 265 (4) ◽  
pp. G783-G791 ◽  
Author(s):  
D. Alvaro ◽  
A. Mennone ◽  
J. L. Boyer
Keyword(s):  
Bile Duct ◽  
Ursodeoxycholic Acid ◽  
Cholic Acid ◽  
Intracellular Ph ◽  
Ph Regulation ◽  
Weak Acid ◽  
Normal Rat ◽  
Acid Load ◽  
Ethanesulfonic Acid ◽  
Isolated Rat

To determine if ursodeoxycholic acid (UDCA) induces a HCO3(-)-rich hypercholeresis by stimulating HCO3- secretion from bile duct epithelial (BDE) cells, we studied the effect of UDCA, sodium tauroursodeoxycholate (TUDCA), and cholic acid on intracellular pH (pHi) regulation and HCO3- excretion in BDE cells isolated from normal rat liver. Exposure of BDE cells to UDCA (0.5-1.5 mM) produced a dose-dependent initial acidification [from -0.05 to -0.16 pH units (pHu)], which was lower in Krebs-Ringer bicarbonate than in N-2-hydroxyethylpiperazine-N'-2- ethanesulfonic acid (HEPES), because of the higher cell-buffering power in the presence of HCO3-. In contrast, TUDCA (1 mM) had no effect on pHi in either media. BDE acidification induced by UDCA (1.5 mM) in KRB was not inhibited by Cl- depletion excluding activation of Cl(-)-HCO3- exchange. Most BDE cells spontaneously recovered their basal pHi during the UDCA infusion (0.5-1 mM) by a secondary activation of the Na(+)-H+ exchanger (amiloride inhibition of pHi recovery; n = 4), and pHi overshot basal levels by 0.1-0.2 pHu after UDCA withdrawal. The activity of Cl(-)-HCO3- exchange (Cl- removal/readmission maneuver) as well as the activities of Na(+)-H+ exchange and Na(+)-HCO3- symport (NH4Cl acid load in HEPES and KRB, respectively) were unaffected by UDCA (0.5 mM) compared with controls. Cholic acid (1.5 mM), which does not produce a hypercholeresis, also acidified BDE cells in KRB media. These studies indicate that UDCA does not stimulate HCO3- excretion from isolated rat BDE cells but modifies pHi in BDE cells as a weak acid.

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