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

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.

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Regulation of intracellular pH in avian renal proximal tubules

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1997.272.1.r341 ◽

1997 ◽

Vol 272 (1) ◽

pp. R341-R349 ◽

Cited By ~ 3

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.

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

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1997.273.6.r1845 ◽

1997 ◽

Vol 273 (6) ◽

pp. R1845-R1854 ◽

Cited By ~ 3

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).

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Intracellular pH in snake renal proximal tubules

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1995.269.4.r822 ◽

1995 ◽

Vol 269 (4) ◽

pp. R822-R829 ◽

Cited By ~ 1

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.

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

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1993.265.4.g677 ◽

1993 ◽

Vol 265 (4) ◽

pp. G677-G685 ◽

Cited By ~ 18

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)

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Depolarization-induced alkalinization in proximal tubules. I. Characteristics and dependence on Na+

AJP Renal Physiology ◽

10.1152/ajprenal.1989.256.2.f342 ◽

1989 ◽

Vol 256 (2) ◽

pp. F342-F353 ◽

Cited By ~ 4

Author(s):

A. W. Siebens ◽

W. F. Boron

Keyword(s):

Intracellular Ph ◽

Initial Rate ◽

Basolateral Membrane ◽

Ph Sensitive ◽

Proximal Tubules ◽

Ambystoma Tigrinum ◽

Tiger Salamander ◽

Additive Effects ◽

Basolateral Membranes ◽

Dependent Processes

We used intracellular pH-sensitive and voltage microelectrodes to examine the effects of depolarization on intracellular pH (pHi) in isolated perfused proximal tubules from the tiger salamander Ambystoma tigrinum. Tubules were depolarized by raising [K+] in the bath (b) or lumen (l), or by adding Ba2+ (1 mM) to the bath or lumen, always in nominally HCO3-free solutions. Increasing [K+]b from 2.5 to 50 mM caused the basolateral membrane to depolarize by an average of 45 mV, and pHi to increase by 0.23 over 3 min. Similar alkalinization was observed when basolateral Ba2+ (1 mM) was used to depolarize the cell at constant extracellular [K+], suggesting that the alkalinization observed during exposure to elevated [K+]b results from depolarization rather than an increase in [K+]b. The initial rate of depolarization-induced alkalinization (DIA) was proportional to the magnitude of the depolarization, regardless of whether tubules were depolarized by elevated [K+]b, elevated [K+]l, or by basolateral Ba2+. An exception was the initial rate of the alkalinization caused by 1 mM luminal Ba2+, which was more than 10-fold greater than that predicted from the depolarization. The voltage and pHi responses to basolateral Ba2+ were smaller in some tubules than others, as were the responses to elevated [K+]l. Tubules with small responses to 1 mM [Ba2+]b had large responses to 50 mM [K+]l, whereas tubules with large responses to 1 mM [Ba2+]b had small responses to 50 mM [K+]l. This variability can be accounted for by differences in the luminal K+ conductance. The DIA was partially inhibited by removal of Na+ from only the lumen or only the bath, but completely inhibited by bilateral Na+ removal. We conclude that the depolarization-induced alkalinization results from additive effects of Na+-dependent processes at both the luminal and basolateral membranes.

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LONG TERM EFFECTS OF THYROID STIMULATING HORMONE AND INSULIN ON INTRACELLULAR pH IN FRTL-5 CELLS

Journal of Endocrinology ◽

10.1677/joe.0.133r009 ◽

1992 ◽

Vol 133 (2) ◽

pp. R9-R11

Author(s):

A.M. Wood ◽

S.P. Bidey ◽

J. Soden ◽

W.R. Robertson

Keyword(s):

Intracellular Ph ◽

Small Groups ◽

Thyroid Stimulating Hormone ◽

Ph Sensitive ◽

Long Term Effects ◽

Bicarbonate Ions ◽

Chronic Effects ◽

Petri Dishes ◽

Presence And Absence

ABSTRACT We have studied the chronic effects of TSH (100μU/ml) and insulin (10μg/ml) on intracellular pH (pHi) in FRTL-5 cells using the pH sensitive probe 2′7-bis (2-carboxyethyl-5′-6′) carboxyfluorescein. FRTL-5 cells were cultured on Petri dishes either in the presence of 4H, ie. Coons F-12 containing cortisol (10nM), transferrin (0.5μg/ml), glycyl-histidyl lysine acetate (10ng/ml) and somatostatin (10μg/ml), or with 4H+insulin (5H), 4H+TSH, or 4H+TSH+insulin (6H). pHi was measured in small groups of cells by microspectrofluorimetry both in the presence and absence of bicarbonate ions after cells had been deprived of serum for at least a day. In

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Studies on retinal acid-base behaviour with glass pH sensitive microelectrode in the intact mammalian eye

Experimental Eye Research ◽

10.1016/0014-4835(76)90088-9 ◽

1976 ◽

Vol 22 (3) ◽

pp. 293

Author(s):

M. Tsacopoulos ◽

S. Lévy

Keyword(s):

Ph Sensitive ◽

Acid Base

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Effect of systemic acid-base disorders on ileal intracellular pH and ion transport

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1986.250.5.g588 ◽

1986 ◽

Vol 250 (5) ◽

pp. G588-G593 ◽

Cited By ~ 4

Author(s):

J. D. Wagner ◽

P. Kurtin ◽

A. N. Charney

Keyword(s):

Intracellular Ph ◽

Respiratory Acidosis ◽

Strong Correlations ◽

Sprague Dawley ◽

Acid Base ◽

Ileal Mucosa ◽

Arterial Pco2 ◽

Co2 Partial Pressure ◽

Arterial Ph ◽

Acute Metabolic Acidosis

We previously reported that changes in ileal net Na absorption correlated with arterial pH, changes in net HCO3 secretion correlated with the plasma HCO3 concentration, and changes in net Cl absorption correlated with arterial CO2 partial pressure (PCO2) during the systemic acid-base disorders. To determine whether changes in intracellular pH (pHi) and HCO3 concentration [( HCO3]i) mediated these effects, we measured pHi and calculated [HCO3]i in the distal ileal mucosa of anesthetized, mechanically ventilated Sprague-Dawley rats using 5,5-[14C]dimethyloxazolidine-2,4,-dione and [3H]inulin. Rats were studied during normocapnia, acute respiratory acidosis, and alkalosis, and uncompensated and pH-compensated acute metabolic acidosis and alkalosis. When animals in all groups were considered, mucosal pHi was not altered, but there were strong correlations between mucosal [HCO3]i and both arterial PCO2 (r = 0.97) and [HCO3] (r = 0.61). When we considered the rates of ileal electrolyte transport that characterized these acid-base disorders [A. N. Charney and L.P. Haskell, Am. J. Physiol. 245 (Gastrointest. Liver Physiol. 8): G230-G235, 1983], we found strong correlations between mucosal [HCO3]i and both net Cl absorption (r = 0.88) and net HCO3 secretion (r = 0.82). These findings suggest that the systemic acid-base disorders do not affect ileal mucosal pHi but do alter mucosal [HCO3]i as a consequence of altered arterial PCO2 and [HCO3]. The effects of these disorders on ileal net Cl absorption and HCO3 secretion may be mediated by changes in [HCO3]i. Arterial pH does not appear to alter ileal Na absorption through changes in the mucosal acid-base milieu.

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