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.

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.

scholarly journals Effect of electroneutral luminal and basolateral lactate transport on intracellular pH in salamander proximal tubules.

1987 ◽  
Vol 90 (6) ◽  
pp. 799-831 ◽  
Author(s):  
A W Siebens ◽  
W F Boron
Keyword(s):  
Intracellular Ph ◽  
Initial Rate ◽  
Basolateral Membrane ◽  
Proximal Tubules ◽  
Tiger Salamander ◽  
Organic Substrates ◽  
Luminal Membrane ◽  
Acid Extrusion ◽  
Intracellular Alkalinization ◽  
Basolateral Membrane Potential

We used microelectrodes to examine the effects of organic substrates, particularly lactate (Lac-), on the intracellular pH (pHi) and basolateral membrane potential (Vbl) in isolated, perfused proximal tubules of the tiger salamander. Exposure of the luminal and basolateral membranes to 3.6 mM Lac- caused pHi to increase by approximately 0.2, opposite to the decrease expected from nonionic diffusion of lactic acid (HLac) into the cell. Addition of Lac- to only the lumen also caused alkalinization, but only if Na+ was present. This alkalinization was not accompanied by immediate Vbl changes, which suggests that it involves luminal, electroneutral Na/Lac cotransport. Addition of Lac- to only the basolateral solution caused pHi to decrease by approximately 0.08. The initial rate of this acidification was a saturable function of [Lac-], was not affected by removal of Na+, and was reversibly reduced by alpha-cyano-4-hydroxycinnamate (CHC). Thus, the pHi decrease induced by basolateral Lac- appears to be due to the basolateral entry of H+ and Lac-, mediated by an H/Lac cotransporter (or a Lac-base exchanger). Our data suggest that this transporter is electroneutral and is not present at the luminal membrane. A key question is how the addition of Lac- to the lumen increases pHi. We found that inhibition of basolateral H/Lac cotransport by basolateral CHC reduced the initial rate of pHi increase caused by luminal Lac-. On the other hand, luminal CHC had no effect on the luminal Lac(-)-induced alkalinization. These data suggest that when Lac- is present in the lumen, it enters the cell from the lumen via electroneutral Na/Lac cotransport and then exists with H+ across the basolateral membrane via electroneutral H/Lac cotransport. The net effect is transepithelial Lac- reabsorption, basolateral acid extrusion, and intracellular alkalinization.

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.

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.

Regulation of intracellular pH in proximal tubules of avian long-looped mammalian-type nephrons

1998 ◽  
Vol 274 (6) ◽  
pp. R1526-R1535 ◽  
Author(s):  
Olga H. Brokl ◽  
Christina L. Martinez ◽  
Apichai Shuprisha ◽  
Diane E. Abbott ◽  
William H. Dantzler
Keyword(s):  
Intracellular Ph ◽  
Ph Sensitive ◽  
Fluorescent Dye ◽  
Proximal Tubules ◽  
Acid Base ◽  
Rate Of Recovery

In nonperfused proximal tubules isolated from chicken long-looped mammalian-type nephrons, intracellular pH (pHi), measured with the pH-sensitive fluorescent dye 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, was ∼7.3 under control conditions (HEPES-buffered medium with pH 7.4 at 37°C) and was reduced to ∼7.0 in response to NH4Cl pulse. The rate of recovery of pHi from this level to the resting level was 1) significantly reduced by the removal of Na+ from the bath, 2) significantly increased by the removal of Cl− from the bath, 3) unchanged by the removal of both Na+ and Cl− from the bath, 4) significantly reduced by the addition of either ethylisopropylamiloride or DIDS to the bath, 5) significantly increased by a high bath K+ concentration, and 6) unchanged by the addition of Ba2+ to the bath. These data suggest that both Na+-coupled and Cl−-coupled basolateral acid-base fluxes are involved in determining the rate of recovery of pHi after acidification. The most likely ones to be important in regulating pHi are a Na+/H+exchanger and a Na+-coupled Cl−/[Formula: see text]exchanger. In birds, long-looped mammalian-type nephrons resemble short-looped transitional nephrons but differ markedly from superficial loopless reptilian-type nephrons.

Na(+)-H+ antiporter and Na(+)-(HCO3-)n symporter regulate intracellular pH in mouse medullary thick limbs of Henle

1990 ◽  
Vol 258 (3) ◽  
pp. F445-F456 ◽  
Author(s):  
D. Kikeri ◽  
S. Azar ◽  
A. Sun ◽  
M. L. Zeidel ◽  
S. C. Hebert
Keyword(s):  
Steady State ◽  
Intracellular Ph ◽  
Dominant Role ◽  
Disulfonic Acid ◽  
Outer Medulla ◽  
Ethanesulfonic Acid ◽  
Antibody Labeling ◽  
Intracellular Alkalinization ◽  
Phi Regulation

To determine mechanisms of intracellular pH (pHi) regulation in mouse medullary thick limbs (MTAL), pHi was measured in MTAL suspensions and in the isolated perfused MTAL by use of 2',7'-bis(carboxyethyl)-5(6)carboxyfluorescein (BCECF). A method to obtain MTAL suspensions from the mouse outer medulla is reported. Characterization of suspensions with microscopy, anti-Tamm-Horsfall antibody labeling, measurement of O2 consumption, and adenosine 3',5'-cyclic monophosphate responses to antidiuretic hormone indicated that these suspensions were highly purified for viable MTAL tubules. The resting pHi was 7.41 +/- 0.02 (means +/- SE) in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered media and 7.23 +/- 0.02 in CO2- HCO3(-)-buffered media, both at extracellular pH 7.4. MTAL tubules exhibited rapid pHi recovery from intracellular acidification. Recovery of pHi was dependent on luminal Na+ (apparent Km = 13.2 +/- 3.2 mM) and was inhibited by amiloride (apparent Ki = 10.6 microM), consistent with the activity of an apical Na(+)-H+ antiporter. Antiporter activity was enhanced by acidification and was diminished at the resting pHi. Recovery from intracellular alkalinization (rapid withdrawal of CO2- HCO3-) was sensitive to the stilbene anion transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, Cl(-)-insensitive, and Na(+)-sensitive, consistent with the activity of a Na(+)-(HCO3-)n symporter. Both transporters were significantly involved in steady-state pHi regulation in the presence of CO2- HCO3-. In contrast, the Na(+)-H+ antiporter played the dominant role in steady-state pHi regulation in the absence of CO2- HCO3-.

Na(+)-HCO3- symport modulates intracellular pH in alveolar epithelial cells

1991 ◽  
Vol 260 (6) ◽  
pp. L555-L561 ◽  
Author(s):  
R. L. Lubman ◽  
E. D. Crandall
Keyword(s):  
Intracellular Ph ◽  
Initial Rate ◽  
Alveolar Epithelium ◽  
Alveolar Epithelial Cells ◽  
Transepithelial Transport ◽  
Disulfonic Acid ◽  
Experimental Conditions ◽  
Alveolar Epithelial ◽  
Ethanesulfonic Acid ◽  
Acid Extrusion

We investigated Na(+)-HCO3- cotransport as a mechanism for regulation of intracellular pH (pHi) in rat alveolar pneumocytes grown in primary culture. pHi was monitored using the fluorescent pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Cells incubated in 6 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) medium at pH 7.4 were subjected to rapid acidification by CO2 pulse. pHi recovered in the presence of Na+ with an initial rate (dpHi/dt) of 0.15 min-1, which was reduced by 67% when Na+ was replaced by choline, unaffected by substitution of gluconate for Cl-, reduced 40% in the presence of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 500 microM), and unchanged by amiloride (1 mM). In parallel experiments, cells were incubated at pH 7.4 with 20 mM HCO3- and pHi acutely lowered by NH3 prepulse. dpHi/dt in these experiments was 0.14 min-1 in the presence of Na+ and HCO3-, and reduced 79% under Na(+)-free conditions. These data indicate the presence of a Na(+)-dependent, Cl(-)-independent, DIDS-sensitive and amiloride-insensitive mechanism of recovery from acute intracellular acidification in alveolar pneumocytes, most consistent with Na(+)-HCO3- cotransport (symport) effecting acid extrusion under these experimental conditions. This ion transport mechanism may contribute to regulation of pHi in alveolar pneumocytes, transepithelial transport of acid-base equivalents across the alveolar epithelium, and modulation of pH of alveolar fluid in adult mammalian lungs.

Proximal duodenal enterocyte transport: evidence for Na(+)-H+ and Cl(-)-HCO3- exchange and NaHCO3 cotransport

1993 ◽  
Vol 265 (4) ◽  
pp. G677-G685 ◽  
Author(s):  
J. I. Isenberg ◽  
M. Ljungstrom ◽  
B. Safsten ◽  
G. Flemstrom
Keyword(s):  
Intracellular Ph ◽  
Collagen Matrix ◽  
Ph Sensitive ◽  
Transport Processes ◽  
High Rate ◽  
Disulfonic Acid ◽  
Acid Base ◽  
Acetoxymethyl Ester ◽  
Addition And Subtraction ◽  
Ethanesulfonic Acid

The duodenum, in contrast to the jejunum, actively secretes HCO3- at a high rate, a process that protects the mucosa from acid/peptic injury. Our purpose was to define the mechanisms involved in HCO3- transport by studying the acid-base transport processes in isolated duodenal enterocytes. Individual rat duodenocytes, isolated by a combination of Ca2+ chelation and collagenase, attached to a collagen matrix were loaded with the pH-sensitive fluoroprobe 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester (BCECF-AM), and intracellular pH was monitored by microfluorospectrophotometry. To identify Na(+)-H+ transport, cells in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid 1) were pulsed with NH4Cl (40 mM) in the absence and presence of amiloride and 2) were removed of Na+. To examine Cl(-)-HCO3- exchange, Cl- was removed from Ringer-HCO3- superfusate in the presence and absence of dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (H2DIDS). The NaHCO3 cotransporter was studied by addition and subtraction of Na+ to amiloride-treated and Cl(-)-depleted enterocytes perfused with Na(+)- and Cl(-)-free Ringer-HCO3- buffer with and without H2DIDS. Mammalian duodenocytes contain at least three acid-base transporters: an amiloride-sensitive Na(+)-H+ exchanger that extrudes acid, a DIDS-sensitive Cl(-)-HCO3- exchanger that extrudes base, and a NaHCO3 cotransporter, also DIDS sensitive, that functions as a base loader. These acid-base transporters likely play a key role in duodenal mucosal HCO3- secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of norepinephrine on intracellular pH in kidney proximal tubule: role of Na+-( HCO 3 − ) n cotransport

1998 ◽  
Vol 275 (1) ◽  
pp. F33-F45 ◽  
Author(s):  
Solange Abdulnour-Nakhoul ◽  
Raja N. Khuri ◽  
Nazih L. Nakhoul
Keyword(s):  
Membrane Potential ◽  
Proximal Tubule ◽  
Intracellular Ph ◽  
Basolateral Membrane ◽  
Inhibitory Effect ◽  
Rate Of Change ◽  
Kidney Proximal Tubule ◽  
Luminal Membrane ◽  
Basolateral Membrane Potential

We examined the effect of norepinephrine (NE) on intracellular pH (pHi) and activity of Na+([Formula: see text]) in the isolated perfused kidney proximal tubule of Ambystoma, using single-barreled voltage and ion-selective microelectrodes. In control[Formula: see text] Ringer, addition of 10−6 M NE to the bath reversibly depolarized the basolateral membrane potential ( V 1), the luminal membrane potential ( V 2), and the transepithelial potential difference ( V 3) and increased pHi by 0.14 ± 0.02. These effects were mimicked by isoproterenol but were abolished after pretreatment with SITS or in the absence of CO2/[Formula: see text]. Removal of bath Na+ depolarized V 1 and V 2, hyperpolarized V 3, and decreased pHi. These effects are largely mediated by the electrogenic Na+-([Formula: see text]) n cotransporter. In the presence of NE, the effects of Na+ removal on membrane potential differences and the rate of change of pHi were significantly smaller. Reducing bath [Formula: see text] concentration from 10 to 2 mM at constant CO2 (pH 6.8) depolarized V 1 and V 2, decreased pHi, and lowered[Formula: see text]. These changes are also due to Na+-([Formula: see text]) n . In the presence of NE, reducing bath [[Formula: see text]] caused a smaller depolarizations of V 1 and V 2, and the rate of pHi decrease was significantly reduced. Our results indicate: 1) NE causes an increase in pHi; 2) the NE-induced alkalinization is mediated by a SITS-sensitive and[Formula: see text]-dependent transporter on the basolateral membrane; and 3) in the presence of NE, the reduced effects caused by basolateral[Formula: see text] changes or Na+ removal are indicative of an inhibitory effect of NE on Na+-([Formula: see text]) n cotransport.

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