Influence of Cl- on pH(i) in oxynticopeptic cells of in vitro frog gastric mucosa

1990 ◽  
Vol 258 (5) ◽  
pp. G815-G824 ◽  
Author(s):  
A. Yanaka ◽  
K. J. Carter ◽  
H. H. Lee ◽  
W. Silen
Keyword(s):  
Gastric Mucosa ◽  
Intracellular Ph ◽  
Rana Catesbeiana ◽  
Basolateral Membrane ◽  
Fluorescent Dye ◽  
Disulfonic Acid ◽  
Fundic Mucosa ◽  
Ethanesulfonic Acid ◽  
Oxynticopeptic Cells

The effect of Cl- on intracellular pH (pH(i)) was studied using sheets of frog (Rana catesbeiana) fundic mucosa in which oxynticopeptic cells were selectively loaded with the acetomethoxy ester form of the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF/AM). Before the measurement of pH(i), tissues were exposed to either 10(-5) M forskolin in the serosal solution (stimulated tissues) or 3 x 10(-4) omeprazole in the serosal solution (inhibited tissues). In HCO3- and N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) buffers, pH(i) increased significantly after removal of Cl- from serosal and luminal solution, both in stimulated and inhibited tissues. The presence of Cl- in the luminal solution prevented this rise in pHi, an effect abolished by serosal 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 3 x 10(-4) M) but not by serosal amiloride (10(-3)M). In the presence of serosal Cl-, pH(i) increased after exposure to serosal DIDS, more prominently in the stimulated than in the inhibited tissues. These results confirm the presence of a Cl(-)-HCO3-exchanger in the basolateral membrane of oxynticopeptic cells in intact sheets of mucosa and suggest that luminal Cl- contributes to the regulation of pH(i) in oxynticopeptic cells.

Effect of PCO2 on intracellular pH in in vitro frog gastric mucosa

1989 ◽  
Vol 256 (1) ◽  
pp. G206-G213 ◽  
Author(s):  
K. J. Carter ◽  
I. Saario ◽  
U. Seidler ◽  
W. Silen
Keyword(s):  
Steady State ◽  
Gastric Mucosa ◽  
Intracellular Ph ◽  
Ringer Solution ◽  
Partial Recovery ◽  
Muscularis Mucosa ◽  
Fundic Mucosa ◽  
Ethanesulfonic Acid ◽  
Oxynticopeptic Cells

Steady-state intracellular pH (pHi) in 0, 5, and 10% CO2-buffered Ringer solution in sheets of in vitro frog gastric antral or fundic mucosa has been measured using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). In tissues perfused with N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-100% O2 buffer [extracellular pH (pHo) = 7.14], steady-state pHi in antral surface cells was 7.08 +/- 0.06 (n = 8), in fundic oxynticopeptic cells 6.91 +/- 0.03 (n = 13), in the muscularis mucosa 7.58 +/- 0.06 (n = 4). In mucosae perfused with 17.8 mM HCO3- -95% O2-5% CO2 buffer (pHo = 7.14), steady-state pHi in antral surface cells was 6.97 +/- 0.02 (n = 22), in fundic oxynticopeptic cells 7.00 +/- 0.04 (n = 18), and in fundic muscularis mucosa 7.39 +/- 0.05 (n = 8). In fundic oxynticopeptic cells perfused with 35.6 mM HCO3- -90% O2-10% CO2 (pHo = 7.14) steady-state pHi was 6.77 +/- 0.07 (n = 4). In tissues equilibrated initially with 100% O2 and changed to 5% CO2, antral surface cells acidified by 0.21 pH units and fundic oxynticopeptic cells by 0.10 pH units, with restoration of pHi to resting levels within 30 and 10 min, respectively. Exposure of tissues initially equilibrated with 5% CO2 to 100% O2 alkalinized antral surface cells by 0.22 pH units and fundic oxynticopeptic cells by 0.23 pH units, with only partial recovery of pHi by 30 min. These data suggest that steady-state pHi is equivalent in surface and oxynticopeptic cells and is lower than in the muscularis mucosa.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of ammonium ion and ammonia on function and morphology of in vitro frog gastric mucosa

1993 ◽  
Vol 265 (2) ◽  
pp. G277-G288 ◽  
Author(s):  
A. Yanaka ◽  
H. Muto ◽  
S. Ito ◽  
W. Silen
Keyword(s):  
Gastric Mucosa ◽  
Electrical Resistance ◽  
Rana Catesbeiana ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Ammonium Ion ◽  
Gastric Epithelial Cells ◽  
Sudden Drop ◽  
Oxynticopeptic Cells

The effects of ammonium ion (NH+4) and ammonia (NH3) on function and morphology of gastric epithelial cells were studied in intact sheets of in vitro frog (Rana catesbeiana) gastric mucosa. Luminal 115 mM NH4Cl at luminal pH 8.0 (calculated [NH3] 2.7 mM), but not at 5.0 (calculated [NH3] 3 microM) induced 1) an increase in intracellular pH (pHi) in oxynticopeptic cells (OPC) and decreases in transmucosal potential difference (PD) and electrical resistance (R) in resting tissues, 2) a decrease in histamine-stimulated H+ secretion and an increase in H+ backdiffusion after removal of luminal NH4Cl, and 3) augmented acidification of OPC during luminal acidification. Serosal 30 mM NH4Cl at serosal pH 7.2 (calculated [NH3] 0.47 mM) induced 1) an increase in pHi in OPC and inhibition of the alkalinization of OPC after removal of ambient Cl-, 2) a decrease in PD associated with the increase in R and decrease in short-circuit current, effects attenuated by serosal 15 mM K+, accentuated by 0.2 mM Ba2+, and abolished by removal of ambient Cl-, 3) a sudden drop of PD in resting, but not in stimulated tissues, effects prevented by high serosal pH (7.8), serosal HCO3-, or removal of luminal Cl-, 4) a decrease in histamine-stimulated H+ secretion and an increase in H+ backdiffusion after removal of NH4Cl, and 5) augmented acidification of OPC during luminal acidification. These results suggest that 1) luminal NH3, but not NH+4, increases backdiffusion of H+ from the lumen to the mucosa, 2) serosal NH3 and/or NH+4 induces depolarization of OPC and decreases electrogenic Cl- transport, thereby attenuating the activity of the basolateral Cl(-)-HCO3- exchanger in OPC, and 3) both of these effects contribute to the augmented acidification of OPC during exposure to high luminal [H+].

Regulation of intracellular pH in proximal tubules of avian loopless reptilian-type nephrons

1997 ◽  
Vol 273 (6) ◽  
pp. R1845-R1854 ◽  
Author(s):  
Christina L. Martinez ◽  
Olga H. Brokl ◽  
Apichai Shuprisha ◽  
Diane E. Abbott ◽  
William H. Dantzler
Keyword(s):  
Intracellular Ph ◽  
Acid Level ◽  
Ph Sensitive ◽  
Fluorescent Dye ◽  
Proximal Tubules ◽  
Disulfonic Acid ◽  
Control Value ◽  
High K ◽  
Rate Of Recovery ◽  
Ethanesulfonic Acid

In proximal tubules isolated from chicken superficial loopless reptilian-type nephrons, intracellular pH (pHi), measured with pH-sensitive fluorescent dye 2′,7′-bis(carboxyethyl)-5(6)-carboxyfluorescein, was ∼7.1–7.2 under control conditions ( N-2-hydroxyethylpiperazine- N′-2-ethanesulfonic acid-buffered medium with pH 7.4 at 37°C), and was reduced to ∼6.9 in response to NH4Cl pulse. The rate of recovery of pHi(control value ≅ 5 × 10−3 pH U/s) from this acid level was 1) significantly decreased by removal of Na+ or both Na+ and Cl− from the bath or addition of 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (0.25 mM) to the bath, 2) significantly increased by high bath K+ (75 mM), and 3) unchanged by removal of Cl− alone from the bath or addition of ethylisopropylamiloride (1 mM) or Ba2+ (5 mM) to the bath. Resting pHi was 1) significantly decreased by Na+ or simultaneous Na+ and Cl− removal, 2) significantly increased by high K+, and 3) unchanged by Cl− removal alone or addition of Ba2+. The data do not fit the concept of pHi regulation by the most commonly suggested basolateral transporters (Na+/H+exchanger, Na+-dependent and Na+-independent Cl−/[Formula: see text]exchangers, or Na+-[Formula: see text]-[Formula: see text]cotransporter).

Regulation of intracellular pH in avian renal proximal tubules

1997 ◽  
Vol 272 (1) ◽  
pp. R341-R349 ◽  
Author(s):  
Y. K. Kim ◽  
O. H. Brokl ◽  
W. H. Dantzler
Keyword(s):  
Intracellular Ph ◽  
Acid Level ◽  
Basolateral Membrane ◽  
Ph Sensitive ◽  
Fluorescent Dye ◽  
Proximal Tubules ◽  
Acid Base ◽  
Renal Proximal Tubules ◽  
Rate Of Recovery ◽  
Ethanesulfonic Acid

In proximal tubules isolated from chicken transitional nephrons, intracellular pH (pHi), measured with the pH-sensitive fluorescent dye 2'.7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF), was approximately 7.3-7.4 under control conditions [N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered medium with pH 7.4 at 39 degrees C] and was reduced to approximately 6.8 in response to NH4Cl pulse. The rate of recovery of pHi (dpHi/dt) from this acid level to the resting level and the resting pHi were 1) significantly reduced by the removal of Na+ from the bath, 2) significantly increased by the removal of Cl from the bath, and 3) unchanged by the removal of both Na+ and Cl from the bath. The addition of either amiloride or 4,4'-diisothiocyanostilbene-2,2'-disulfonate to the bath reduced dpHi/dt to about the same extent as the removal of Na+. These data suggest that both Na(+)-coupled and Cl-coupled acid-base fluxes at the basolateral membrane are involved in determining the resting pHi and the rate of recovery of pHi after acidification. The most likely possibilities appear to be a basolateral Na+/Hi exchanger, a basolateral Na(+)-coupled Cl/HCO3 exchanger, a basolateral Na(+)-HCO3(-)CO(3)2 cotransporter, and a basolateral Na(+)-independent Cl-/HCO3 exchanger.

Intracellular pH in snake renal proximal tubules

1995 ◽  
Vol 269 (4) ◽  
pp. R822-R829 ◽  
Author(s):  
Y. K. Kim ◽  
W. H. Dantzler
Keyword(s):  
Intracellular Ph ◽  
Basolateral Membrane ◽  
Distal Portion ◽  
Proximal Tubules ◽  
Disulfonic Acid ◽  
Renal Tubules ◽  
Garter Snakes ◽  
Luminal Membrane ◽  
Rate Of Recovery ◽  
Ethanesulfonic Acid

Intracellular pH (pHi) was studied in isolated proximal renal tubules of garter snakes (Thamnophis spp.) with oil-filled lumens under control conditions [N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered medium with pH 7.4 at 25 degrees C] and in response to NH4Cl pulse. pHi was measured with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). Control resting pHi (7.1) and acidification in response to NH4Cl pulse (minimum pHi, 6.6) were essentially the same in snake tubules with oil-filled lumens or perfused lumens and in rabbit S2 proximal tubules with oil-filled lumens. Rate of recovery of pHi (dpHi/dt) from acid to resting level in snake tubules (2.5 x 10(-3) pH U/s was about one-third of that in rabbit tubules. Resting pHi and dpHi/dt from acid to resting level were Na+ dependent in the distal portion but not the proximal portion of snake proximal tubules. However, dpHi/dt was not influenced by amiloride or 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid in snake proximal tubules, suggesting that the effect of Na+ on dpHi/dt and resting pHi may involve membrane potential. This study also indicates that oil-filled lumens do not interfere with measurements of resting pHi and do permit evaluation of pHi regulation at the basolateral membrane without complications from transport at the luminal membrane.

H(+)-K(+)-ATPase contributes to regulation of pHi in frog oxynticopeptic cells

1991 ◽  
Vol 261 (5) ◽  
pp. G781-G789 ◽  
Author(s):  
A. Yanaka ◽  
K. J. Carter ◽  
P. J. Goddard ◽  
M. C. Heissenberg ◽  
W. Silen
Keyword(s):  
Steady State ◽  
Gastric Mucosa ◽  
Histamine Release ◽  
Rana Catesbeiana ◽  
Intracellular Acidosis ◽  
Endogenous Histamine ◽  
Decreasing Ph ◽  
Oxynticopeptic Cells

The effect of intracellular acidosis on luminal H+ secretion and the role of H(+)-K(+)-ATPase in regulation of intracellular pH (pHi) in oxynticopeptic cells (OPC) (measured with a pH-sensitive fluorescent dye) were examined in intact sheets of in vitro frog (Rana catesbeiana) gastric mucosa. Intracellular acidosis of OPC induced by decreasing pH in the serosal solution (pHs) from 7.2 to 6.0 reversibly increased forskolin-stimulated H+ secretion without increasing endogenous histamine release. The observed increase in H+ secretion was unaffected by either 1 mM cimetidine or 1 mM histamine, but was accentuated by 1 mM amiloride, an effect abolished by 0.3 mM omeprazole. Steady-state pHi values in stimulated or resting OPC at pHs 7.2 were not significantly different. However, pHi in OPC was significantly higher in stimulated than in resting tissues at pHs 6.9, a difference accentuated by decreasing pHs to 6.4 or by 1 mM amiloride. Amiloride completely prevented recovery from intracellular acidosis induced by pHs 6.4 or 6.9 in omeprazole-treated tissues, but only partially mitigated recovery in cimetidine- or forskolin-treated tissues. At pHs 6.4, high luminal [K+] (100 mM) increased H+ secretion and hastened recovery of pHi in cimetidine-treated tissues in the presence of amiloride. These results suggest that, in intact sheets of in vitro frog gastric mucosa, 1) intracellular acidosis stimulates luminal H+ secretion via histamine-independent mechanisms and 2) H(+)-K(+)-ATPase contributes to the recovery of OPC from intracellular acidosis.

scholarly journals Neonatal rabbit proximal tubule basolateral membrane Na+/H+antiporter and Cl−/base exchange

1999 ◽  
Vol 276 (6) ◽  
pp. R1792-R1797 ◽  
Author(s):  
Mehul Shah ◽  
Raymond Quigley ◽  
Michel Baum
Keyword(s):  
Proximal Tubule ◽  
Intracellular Ph ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Fluorescent Dye ◽  
Adult Rabbit ◽  
Base Exchange ◽  
Straight Tubules ◽  
Exchange Activity

The present in vitro microperfusion study examined the maturation of Na+/H+antiporter and Cl−/base exchanger on the basolateral membrane of rabbit superficial proximal straight tubules (PST). Intracellular pH (pHi) was measured with the pH-sensitive fluorescent dye 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein in neonatal and adult superficial PST. Na+/H+antiporter activity was examined after basolateral Na+ addition in tubules initially perfused and bathed without Na+. Neonatal Na+/H+antiporter activity was ∼40% that of adult segment (9.7 ± 1.5 vs. 23.7 ± 3.2 pmol ⋅ mm−1 ⋅ min−1; P < 0.001). The effect of bath Cl− removal on pHi was used to assess the rates of basolateral Cl−/base exchange. In both neonatal and adult PST, the Cl−/base exchange activity was significantly higher in the presence of 25 mM[Formula: see text] than in the absence of[Formula: see text] and was inhibited by cyanide and acetazolamide, consistent with Cl−/[Formula: see text]exchange. The proton flux rates in the presence of bicarbonate in neonatal and adult tubules were 14.1 ± 3.6 and 19.5 ± 3.5 pmol ⋅ mm−1min−1, respectively ( P = NS), consistent with a mature rate of Cl−/[Formula: see text]exchanger activity in neonatal tubules. Basolateral Cl−/base exchange activity in the absence of CO2 and[Formula: see text], with luminal and bath cyanide and acetazolamide, was greater in adult than in neonatal PST and inhibited by bath DIDS consistent with a maturational increase in Cl−/OH−exchange. We have previously shown that the rates of the apical membrane Na+/H+antiporter and Cl−/base exchanger were approximately fivefold lower in neonatal compared with adult rabbit superficial PST. These data demonstrate that neonatal PST basolateral membrane Na+/H+antiporter and Cl−/base exchanger activities are relatively more mature than the Na+/H+antiporter and Cl−/base exchangers on the apical membrane.

Evidence for Cl(-)-independent HCO3- transport in basolateral membranes of Necturus oxyntopeptic cells

1996 ◽  
Vol 271 (6) ◽  
pp. G1096-G1103
Author(s):  
M. E. Klingensmith ◽  
R. R. Cima ◽  
A. E. Gadacz ◽  
D. I. Soybel
Keyword(s):  
Gastric Mucosa ◽  
Recent Work ◽  
Intracellular Ph ◽  
Disulfonic Acid ◽  
Substantial Part ◽  
Ph Indicator ◽  
Basolateral Membranes ◽  
Hcl Secretion ◽  
Ethanesulfonic Acid

Luminal H+ secretion by gastric mucosa is accompanied by basolateral HCO3- release. A basolateral Cl-/HCO3- exchanger is known to mediate HCO3- extrusion from oxyntopeptic cells during resting and secretagogue-induced apical HCl secretion. From recent work, we hypothesized that there might be a Cl(-)-independent pathway for basolateral HCO3- exit in Necturus oxyntopeptic cells. In this study, we used a fluorescent pH indicator [2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein] to evaluate Cl(-)-independent HCO3- transport across the basolateral membranes of intact oxyntopeptic cells. Removal of serosal Cl- increased intracellular pH (pHi) (7.05 to 7.25), consistent with Cl(-)-dependent HCO3- extrusion. Removal of serosal Na+ in the absence of Cl- resulted in significant acidification of pHi (7.10 to 6.89), but studies involving amiloride, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), and 0 HCO3(-)-N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered solutions suggest that Na(+)-dependent changes in pHi are due to Na+/H+ exchange. Our studies demonstrate a marked concentration-dependent alkalinization when tissues are exposed to increases in serosal K+. A substantial part of this alkalinization in response to increases in serosal K+ (pHi 7.00 to 7.46) appears to be a HCO3- exit pathway that is independent of both Na+ and Cl-, unaffected by bumetanide or amiloride, but sensitive to DIDS. We propose the presence of a Cl(-)- and Na(+)-independent K(+)-dependent HCO3- cotransporter in Necturus oxyntopeptic cell basolateral membranes.

Electrogenic Na/HCO3 cotransport across basolateral membrane of isolated perfused Necturus proximal tubule

1987 ◽  
Vol 253 (2) ◽  
pp. F340-F350 ◽  
Author(s):  
A. G. Lopes ◽  
A. W. Siebens ◽  
G. Giebisch ◽  
W. F. Boron
Keyword(s):  
Proximal Tubule ◽  
Liquid Membrane ◽  
Initial Rate ◽  
Basolateral Membrane ◽  
Proximal Tubules ◽  
Disulfonic Acid ◽  
Necturus Maculosus ◽  
Ethanesulfonic Acid ◽  
Ph Microelectrodes

This study was undertaken to determine whether the proximal tubule of the mud puppy Necturus maculosus possesses a basolateral Na/HCO3 cotransporter. We examined the effects on basolateral membrane potential (Vbl) and intracellular pH (pHi) of 1) lowering basolateral [HCO3-] at constant PCO2, and 2) replacing Na+ with N-methyl-D-glucamine. Vbl and pHi were measured with Ling-Gerard and liquid-membrane pH microelectrodes, respectively, in isolated tubules perfused in vitro. We found that decreasing basolateral [HCO3-] from 10 mM (pH 7.5) to 2 mM (pH 6.8) resulted in an immediate depolarization of 14.9 mV, and a pHi decrease of 0.35. SITS (4-acetamido-4'-isothiocyanostibene-2,2'-disulfonic acid, 0.5 mM) inhibited the HCO3-induced depolarization by 87% and inhibited the initial rate of the pHi decrease by 79%. Replacement of basolateral Na+ with N-methyl-D-glucamine resulted in an immediate depolarization of 11.3 mV, and a pHi decrease of 0.36. SITS inhibited the zero Na-induced depolarization by 86% and the initial rate of the pHi decrease by 81%. Nominal removal of basolateral HCO3- (replaced with N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) inhibited the zero Na-induced depolarization by 64%, whereas nominal removal of Na+ inhibited the 2 mM HCO3-induced depolarization by 67%. Replacement of all basolateral Cl- with glucuronate did not inhibit the changes in Vbl induced by changing [HCO3-] or [Na+]. Observations similar to those described above have been made previously on Ambystoma proximal tubules, and attributed to an electrogenic Na/HCO3 cotransport mechanism that carries HCO3-, Na+, and net negative charge in the same direction. We conclude that Necturus proximal tubules possess a similar, if not identical, electrogenic Na/HCO3 cotransport mechanism.

Prostaglandin stimulates Cl(-)-HCO3- exchange in amphibian oxynticopeptic cells

1992 ◽  
Vol 262 (1) ◽  
pp. G44-G49
Author(s):  
A. Yanaka ◽  
K. J. Carter ◽  
P. J. Goddard ◽  
W. Silen
Keyword(s):  
Gastric Mucosa ◽  
Prostaglandin E2 ◽  
Disulfonic Acid ◽  
Muscularis Mucosae ◽  
Cl Transport ◽  
The Relationship ◽  
Different Tissues ◽  
Oxynticopeptic Cells ◽  
Stimulation Of

Prostaglandins, shown to stimulate Cl- transport in epithelial cells of several different tissues, protect gastric mucosa against physiological injury induced by luminal acid. To clarify the relationship between the stimulation of Cl(-)-transport and the protection of gastric mucosa, the effect of prostaglandin on Cl(-)-HCO3- exchange in oxynticopeptic cells (OPC) was examined in intact sheets of in vitro frog gastric mucosa, in which OPC were selectively loaded with the pH-sensitive fluorescent dye 2',7'-bis(carboxyethyl)-5(6')-carboxyfluorescein (BCECF). In omeprazole (0.3 mM)-pretreated frog fundic mucosae, in which H+ secretion was totally inhibited, 16,16-dimethyl prostaglandin E2 (dmPGE2) induced a significant decrease in intracellular pH (pHi) in OPC simultaneously with a significant increase in pHi in adjacent muscularis mucosae, an effect abolished by removal of ambient Cl- or addition of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) (0.5 mM). dmPGE2 accentuated the rates of alkalinization of OPC after either removal of ambient Cl- or addition of serosal H2DIDS. During exposure to luminal or serosal acid, dmPGE2 significantly attenuated acidification of OPC induced by the exogenous H+, effects abolished either by removal of ambient Cl- or by addition of H2DIDS (0.5 mM). These results suggest that 1) dmPGE2 stimulates extrusion of HCO3- through the basolateral Cl(-)-HCO3- exchanger in resting OPC (H+ secretion inhibited) and that 2) relatively high extracellular [HCO3-] on the basolateral surface afforded by dmPGE2 protects OPC from acidification during exposure to luminal or serosal acid.

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