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1.
HCO3- exit across the basolateral membrane of the kidney proximal tubule cell is mediated via an electrogenic Na+:HCO3- cotransporter. We have studied the effect of pH on the activity of this cotransport system in basolateral membrane vesicles isolated from rabbit renal cortex. At constant internal pH 6.0, increasing the external pH and [HCO3-] increased the rate of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid-sensitive 22Na+ influx into the vesicles. To determine the role of internal pH on the activity of the Na+:HCO3- cotransport system, the influx of 22Na+ via HCO3-dependent Na(+)-Na+ exchange was measured in the absence of an initial pH and [HCO3-] gradient (pH(i) = pH(o), 5% CO2). Increasing the pH from 6.8 to 7.2 increased whereas, increasing the pH from 7.4 to 8.0 decreased the rate of 22Na+ influx via this exchange. Increasing pH at constant [HCO3-] (pH(i) = pH(o) = 8.0, 1.5% CO2 versus pH(i) = pH(o) = 7.2, 10% CO2) reduced the influx of 22Na+ via HCO3-dependent Na(+)-Na+ exchange. Increasing pH at constant [CO3(2-)](pH(i) = pH(o) = 8.0, 1.5% CO2 versus pH(i) = pH(o) = 7.2, 60% CO2) was associated with reduced 22Na+ uptake. Decreasing the pH (pH(i) = pH(o) = 6.3, 60% CO2 versus pH(i) = pH(o) = 7.2, 5% CO2) was associated with a reduced rate of HCO3(-)-dependent Na(+)-Na+ exchange. We conclude that the Na+:HCO3- cotransporter displays a significant pH sensitivity profile with the cotransporter being more functional at pH 7.0-7.4 and less active at more acid or alkaline pH. In addition, the results suggest that the pH sensitivity arises at the inner surface of the basolateral membrane. 相似文献
2.
T J Jentsch I Janicke D Sorgenfrei S K Keller M Wiederholt 《The Journal of biological chemistry》1986,261(26):12120-12127
Using the pH-sensitive absorbance of 5 (and 6)-carboxy-4',5'- dimethylfluorescein, we investigated the regulation of cytoplasmic pH (pHi) in monkey kidney epithelial cells (BSC-1). In the absence of HCO3-, pHi is 7.15 +/- 0.1, which is not significantly different from pHi in 28 mM HCO3-, 5% CO2 (7.21 +/- 0.07). After an acid load, the cells regulate pHi in the absence of HCO3- by a Na+ (or Li+)-dependent, amiloride-inhibitable mechanism (indicative of Na+/H+ antiport). In 28 mM HCO3-, while still dependent on Na+, this regulation is only blocked in part by 1 mM amiloride. A partial block is also observed with 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) (1 mM). With cells pretreated with DIDS, 1 mM amiloride nearly totally inhibits this regulation. Cl- had no effect on pHi regulation in the acidic range. In HCO3(-)-free saline, Na+ removal leads to an amiloride-insensitive acidification, which is dependent on Ca2+. In 28 mM HCO3-, Na+ (and Ca2+) removal led to a pronounced reversible and DIDS-sensitive acidification. When HCO3- was lowered from 46 to 10 mM at constant pCO2 (5%), pHi dropped by a DIDS-sensitive mechanism. Identical changes in pHo (7.6 to 6.9) in the nominal absence of HCO3- led to smaller changes of pHi. In the presence but not in the absence of HCO3-, removal of Cl- led to a DIDS-sensitive alkalinization. This was also observed in the nominal absence of Na+, which leads to a sustained acidification. It is concluded that in nominally bicarbonate-free saline, the amiloride-sensitive Na+/H+ antiport is the predominant mechanism of pHi regulation at acidic pHi, while being relatively inactive at physiological values of pHi. In bicarbonate saline, two other mechanisms effect pHi regulation: a DIDS-sensitive Na+-HCO3- symport, which contributes to cytoplasmic alkalinization, and a DIDS-sensitive Cl-/HCO3- exchange, which is apparently independent of Na+. 相似文献
3.
Ionic mechanism of Na+-HCO3- cotransport in rabbit renal basolateral membrane vesicles 总被引:5,自引:0,他引:5
The exit of HCO3- across the basolateral membrane of the proximal tubule cell occurs via the electrogenic cotransport of 3 eq of base per Na+. We have used basolateral membrane vesicles isolated from rabbit renal cortex to identify the ionic species transported via this pathway. Media of varying pH and pCO2 were employed to evaluate the independent effects of HCO3- and CO3(2-) on 22Na transport. Na+ uptake was stimulated when [CO3(2-)] was increased at constant [HCO3-], indicating the existence of a transport site for CO3(2-). In the presence of HCO3-, Na+ influx was stimulated more than 3-fold by an inward SO3(2-) gradient. SO3(2-)-stimulated Na+ influx was stilbene-sensitive, confirming that it occurs via the Na+-HCO3- cotransport system. Na+-SO3(2-) cotransport was demonstrated and found to have a 1:1 stoichiometry. Increasing [CO3(2-)] at constant [HCO3-] reduced the stimulation of Na+ influx by SO3(2-), suggesting competition between SO3(2-) and CO3(2-) at a common divalent anion site. Additional divalent anions that were tested, such as SO4(2-), oxalate2-, and HPO4(2-), did not interact at this site. SO3(2-) stimulation of Na+ influx was absolutely HCO3-(-)dependent and was increased as a function of [HCO3-], indicating the presence of a separate HCO3- site. Lastly, we tested whether Na+ interacts via ion pair formation with CO3(2-) or binds to a distinct site. Na+, which has lower affinity than Li+ for ion pair formation with CO3(2-), was found to have greater than 5-fold higher affinity than Li+ for the Na+-HCO3- cotransport system. Moreover, when its inhibition was studied as a function of [Na+], harmaline was found to be a competitive inhibitor of Na+ influx, indicating the existence of a distinct cation site. Our data are compatible with a model in which base transport across the basolateral membrane of the proximal tubule cell takes place via 1:1:1 cotransport of CO3(2-), HCO3-, and Na+ on distinct sites. 相似文献
4.
Intracellular pH regulation in the renal proximal tubule of the salamander. Basolateral HCO3- transport 总被引:31,自引:4,他引:27
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We have used pH-, Na-, and Cl-sensitive microelectrodes to study basolateral HCO3- transport in isolated, perfused proximal tubules of the tiger salamander Ambystoma tigrinum. In one series of experiments, we lowered basolateral pH (pHb) from 7.5 to 6.8 by reducing [HCO3-]b from 10 to 2 mM at a constant pCO2. This reduction of pHb and [HCO3-]b causes a large (approximately 0.35), rapid fall in pHi as well as a transient depolarization of the basolateral membrane. Returning pHb and [HCO3-]b to normal has the opposite effects. Similar reductions of luminal pH (pHl) and [HCO3-]l have only minor effects. The reduction of [HCO3-]b and pHb also produces a reversible fall in aiNa. In a second series of experiments, we reduced [Na+]b at constant [HCO3-]b and pHb, and also observed a rapid fall in pHi and a transient basolateral depolarization. These changes are reversed by returning [Na+]b to normal. The effects of altering [Na+]l in the presence of HCO3-, or of altering [Na+]b in the nominal absence of HCO3-, are substantially less. Although the effects on pHi and basolateral membrane potential of altering either [HCO3-]b or [Na+]b are largely blocked by 4-acetamido-4- isothiocyanostilbene-2,2'-disulfonate (SITS), they are not affected by removal of Cl-, nor are there accompanying changes in aiCl consistent with a tight linkage between Cl- fluxes and those of Na+ and HCO3-. The aforementioned changes are apparently mediated by a single transport system, not involving Cl-. We conclude that HCO3- transport is restricted to the basolateral membrane, and that HCO3- fluxes are linked to those of Na+. The data are compatible with an electrogenic Na/HCO3 transporter that carries Na+, HCO3-, and net negative charge in the same direction. 相似文献
5.
Subcellular localization of prostaglandin E2 binding sites in bovine adrenal medulla 总被引:1,自引:0,他引:1
Effect of changing [K+], [Na+] and [Cl-] in nutrient solution on potential difference (PD) and resistance was studied in bullfrog antrum with and without nutrient HCO3(-) but with 95% O2/5% CO2 in both cases. In both cases, changing from 4 to 40 mM K+ gave about the same initial PD maximum (anomalous response) which was followed by a decrease below control level. Latter effect was much less with zero than with 25 mM HCO3(-). Changing from 102 to 8 mM Na+ gave initial normal PD response about the same in both cases. However, 10 min later the change in PD with zero HCO3(-) was insignificant but with 25 mM HCO3(-) the PD decreased (anomalous response of electrogenic NaCl symport). PD maxima due to K+ and Na+ were largely related to (Na+ + K+)-ATPase pump. Changes in nutrient Cl- from 81 to 8.1 mM gave only a decrease in PD (normal response). Initial PD increases are explained by relative increases in resistance of simple conductance pathways and of parallel pathways of (Na+ + K+)-ATPase pump and Na+/Cl- symport. Removal of HCO3(-) and concurrent reduction of pH modify resistance of these pathways. 相似文献
6.
Charney AN Alexander-Chacko J Gummaconda R Egnor RW 《Biochimica et biophysica acta》2002,1573(2):141-148
Carbonic anhydrase (CA) inhibition reduces NaCl absorption in rat distal ileum, a pH-sensitive, low CA activity tissue, and in distal colon, a CO(2)-sensitive, high CA activity tissue. We hypothesized that CA plays a non-catalytic role in NaCl absorption in these segments. Unidirectional fluxes of Na(+) and Cl(-), and total HCO(3)(-) generation (estimated as the sum of radiolabeled HCO(3)(-) and CO(2) produced from glucose) were measured in Ussing chambers in nominally CO(2), HCO(3)(-)-free HEPES Ringer. Measurements were made in the presence and absence of 0.1 mM methazolamide, a membrane-permeant CA inhibitor. Ringer pH reduction from 7.6 to 7.1 stimulated ileal but not colonic Na(+) and Cl(-) absorption. In the ileum, methazolamide reduced J(ms)(Na) and J(ms)(Cl) and caused net Cl(-) secretion at pH 7.6, and prevented the stimulatory effect of lowering pH. In the colon, methazolamide reduced Na(+) and Cl(-) absorption at pH 7.6. Total HCO(3)(-) generation was minimal in HEPES at pH 7.6 and 7.1 in both segments, was minimally affected by methazolamide, and did not account for the changes in Cl(-) absorption caused by pH or methazolamide. We conclude that CA plays a role in ileal and colonic NaCl absorption independent of its catalytic function. 相似文献
7.
T J Jentsch P Schwartz B S Schill B Langner A P Lepple S K Keller M Wiederholt 《The Journal of biological chemistry》1986,261(23):10673-10679
BSC-1 kidney epithelial cells derived from the African green monkey are known to express a Na+HCO3- symport (Jentsch, T. J., Schill, B. S., Schwartz, P., Matthes, H., Keller, S. K., and Wiederholt, M. (1985) J. Biol. Chem. 260, 15554-15560). In the present work, 4,4-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS)-sensitive 22Na+ uptake into confluent monolayers of BSC-1 is measured in the presence of ouabain (10(-4) M) and amiloride (10(-3) M) to define the interactions between Na+ and HCO3- binding and pH. Dependence of DIDS-sensitive 22Na+ fluxes on either Na+ or HCO3- can be described by Michaelis-Menten kinetics. External apparent Km for HCO3- decreases with increasing Na+ concentration (Km app (HCO3-) = 36 +/- 10, 18 +/- 5, and 9 +/- 3 mM at 20, 45, and 151 mM Na+o, respectively (pHo = 7.4)). Similarly, external apparent Km for Na+ decreases with increasing HCO3- concentration (Km app (Na+) = 73 +/- 22, 28 +/- 8, and 14 +/- 4 mM at 6, 17, and 56 mM HCO3o-, respectively (pHo = 7.4)). Vmax app remains constant within the experimental error. When data are replotted as a function of calculated NaCO3- concentration, they can be approximated by a single Michaelis-Menten equation. DIDS-sensitive uptake at constant Na+ and HCO3- displays a broad pH optimum in the range between 7.2 and 7.6. The data are compatible with the ion pair model in which the transported species, NaCO3-, binds to the transport site with Km = 15.3 +/- 4 microM. However, the data may also be fitted by either a random or ordered bireactant system. Sets of parameters necessary for these fits are given. 相似文献
8.
Sodium-dependent chloride transport in basolateral membrane vesicles isolated from rabbit proximal tubule 总被引:1,自引:0,他引:1
The mechanisms for Cl transport across basolateral membrane vesicles (BLMV) isolated from rabbit renal cortex were examined by using the Cl-sensitive fluorescent indicator 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ). The transporters studied included Cl/base exchange, Cl/base/Na cotransport, K/Cl cotransport, and Cl conductance. Initial rates of chloride influx (JCl) were determined from the measured time course of SPQ fluorescence in BLMV following inwardly directed gradients of Cl and gradients of other ions and/or pH. For a 50 mM inwardly directed Cl gradient in BLMV which were voltage and pH clamped (7.0) using K/valinomycin and nigericin, JCl was 0.80 +/- 0.14 nmol S-1 (mg of vesicle protein)-1 (mean +/- SD, n = 8 separate preparations). In the absence of Na and CO2/HCO3 in voltage-clamped BLMV, JCl increased 56% +/- 5% in response to a 1.9 pH unit inwardly directed H gradient; the increase was further enhanced by 40% +/- 3% in the presence of CO2/HCO3 and inhibited 30% +/- 8% by 100 microM dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonic acid. Na gradients did not increase JCl in the absence of CO2/HCO3; however, an outwardly directed Na gradient in the presence of CO2/HCO3 increased JCl by 31% +/- 8% with a Na KD of 7 +/- 2 mM. These results indicate the presence of Cl/OH and Cl/HCO3 exchange, and Cl/HCO3 exchange trans-stimulated by Na. There was no significant effect of K gradients in the presence or absence of valinomycin, suggesting lack of significant K/Cl cotransport and Cl conductance under experimental conditions.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
9.
Webb RA Xue L 《Comparative biochemistry and physiology. Part A, Molecular & integrative physiology》1998,119(2):553-562
Absorption of exogenous choline by the cestode Hymenolepis diminuta was found to be both Na+- and HCO3--dependent and, at pH 6 to 7, accounted for up to 65% of the total choline uptake. Na+/HCO3- dependent choline uptake was activated at approximately 6 mM HCO3- (EC50 approximately 9 mM), and, above 100 mM Na+, the rate of uptake was directly proportional to the Na+ concentration. Atempts to uncouple Na+-dependent uptake from HCO3--dependent uptake were not successful: K+-depolarization was without effect on HCO3--dependent choline uptake, and use of valinoomycin to hyperpolarize the brush-border membrane resulted in inhibition of uptake. Na-/HCO3--dependent choline uptake was not associated with solvent drag. The Na+/HCO3--dependent choline uptake displayed a Q10 of 6.4 (27 degrees to 37 degrees) and a relatively high activation energy of 126 kJ x mol(-1). At pH 6.0 and 7.0, Na-/HCO3--dependent choline uptake rates were similar, but Na+/HCO3--dependent choline uptake was reduced at pH 5.0. The Na+/HCO3--dependent choline uptake, at pH 7.0, displayed a Kt of approximately 500 microM and a Vmax of 4.01 pmol x mg wet weight(-1) x min(-1). The Na+/HCO3--dependent choline uptake was hemicholinium-3 sensitive, but not significantly inhibited by 200 microM bumetanide, 100 microM amiloride, benzamil, or EIPA or by 1 mM 4,4'-diisothiocyano-2,2'-stilbene disulfonate (DIDS) or 4-acetamido-4'-isothiocvanostilbene-2,2'-disulfonic acid (SITS). Although it remains to be shown that HCO3- uptake is coupled directly to both choline and Na+ uptake, the data suggest that choline up take occurs via choline/Na+/HCO3--co-trans porter. 相似文献
10.
J A Zanghi A E Schmelzer T P Mendoza R H Knop W M Miller 《Biotechnology and bioengineering》1999,65(2):182-191
Accumulation of CO(2) in animal cell cultures can be a significant problem during scale-up and production of recombinant glycoprotein biopharmaceuticals. By examining the cell-surface polysialic acid (PSA) content, we show that elevated CO(2) partial pressure (pCO(2)) can alter protein glycosylation. PSA is a high-molecular-weight polymer attached to several complex N-linked oligosaccharides on the neural cell adhesion molecule (NCAM), so that small changes in either core glycosylation or in polysialylation are amplified and easily measured. Flow-cytometric analysis revealed that PSA levels on Chinese hamster ovary (CHO) cells decrease with increasing pCO(2) in a dose-dependent manner, independent of any change in NCAM content. The results are highly pH-dependent, with a greater decrease in PSA at higher pH. By manipulating medium pH and pCO(2), we showed that decreases in PSA correlate well with bicarbonate concentration ([HCO(3)(-)]). In fact, it was possible to offset a 60% decrease in PSA content at 120 mm Hg pCO(2) by decreasing the pH from 7.3 to 6.9, such that [HCO(3)(-)] was lowered to that of control (38 mm Hg pCO(2)). When the increase in osmolality associated with elevated [HCO(3)(-)] was offset by decreasing the basal medium [NaCl], elevated [HCO(3)(-)] still caused a decrease in PSA, although less extensive than without osmolality control. By increasing [NaCl], we show that hyperosmolality alone decreases PSA content, but to a lesser extent than for the same osmolality increase due to elevated [NaHCO(3)]. In conclusion, we demonstrate the importance of pH and pCO(2) interactions, and show that [HCO(3)(-)] and osmolality can account for the observed changes in PSA content over a wide range of pH and pCO(2) values. 相似文献
11.
Lyall V Alam RI Phan TH Russell OF Malik SA Heck GL DeSimone JA 《The Journal of general physiology》2002,120(6):793-815
Mixture interactions between sour and salt taste modalities were investigated in rats by direct measurement of intracellular pH (pH(i)) and Na(+) activity ([Na(+)](i)) in polarized fungiform taste receptor cells (TRCs) and by chorda tympani (CT) nerve recordings. Stimulating the lingual surface with NaCl solutions adjusted to pHs ranging between 2.0 and 10.3 increased the magnitude of NaCl CT responses linearly with increasing external pH (pH(o)). At pH 7.0, the epithelial sodium channel (ENaC) blocker, benzamil, decreased NaCl CT responses and inhibited further changes in CT responses induced by varying pH(o) to 2.0 or 10.3. At constant pH(o), buffering NaCl solutions with potassium acetate/acetic acid (KA/AA) or HCO(3)(-)/CO(2) inhibited NaCl CT responses relative to CT responses obtained with NaCl solutions buffered with HEPES. The carbonic anhydrase blockers, MK-507 and MK-417, attenuated the inhibition of NaCl CT responses in HCO(3)(-)/CO(2) buffer, suggesting a regulatory role for pH(i). In polarized TRCs step changes in apical pH(o) from 10.3 to 2.0 induced a linear decrease in pH(i) that remained within the physiological range (slope = 0.035; r(2) = 0.98). At constant pH(o), perfusing the apical membrane with Ringer's solutions buffered with KA/AA or HCO(3)(-)/CO(2) decreased resting TRC pH(i), and MK-507 or MK-417 attenuated the decrease in pH(i) in TRCs perfused with HCO(3)(-)/CO(2) buffer. In parallel experiments, TRC [Na(+)](i) decreased with (a) a decrease in apical pH, (b) exposing the apical membrane to amiloride or benzamil, (c) removal of apical Na(+), and (d) acid loading the cells with NH(4)Cl or sodium acetate at constant pH(o). Diethylpyrocarbonate and Zn(2+), modification reagents for histidine residues in proteins, attenuated the CO(2)-induced inhibition of NaCl CT responses and the pH(i)-induced inhibition of apical Na(+) influx in TRCs. We conclude that TRC pH(i) regulates Na(+)-influx through amiloride-sensitive apical ENaCs and hence modulates NaCl CT responses in acid/salt mixtures. 相似文献
12.
One of the main obstacles in assigning any distinct function to histamine in health and disease was the longlasting controversy on the existence of any physiological, endogenous histamine formation in man and most of the other mammals except the rat. Using a modification of Schayer's isotope dilution method, a renewed attempt was made to identify the very low activities of an acid (specific) histidine decarboxylase in rabbit gastric mucosa capable of producing endogenous histamine in physiological conditions, to develop tests for its identification in crude enzyme extracts and to demonstrate the specificity of the enzymatic assay by excluding any relevant Dopa decarboxylase activity and also nonenzymatic decarboxylation interfering with the determination of acid (specific) histidine decarboxylase. To achieve this aim five tests were developed: In the pH profile (test 1), a pH optimum was found at 7.0 in the presence of a low substrate concentration (1.6 X 10(-6)M L-[ring-2-14C]-histidine). The apparent Michaelis concentration at the pH optimum (test 2) was 1.8 X 10(-4)M, the maximum rate 12.5pmol [14C]histamine formed X min-1. To increase the specificity of inhibition experiments with alpha-methylhistidine and alpha-methyl-L-Dopa a pH profile was determined in the presence of these two enzymatic inhibitors (test 3 and 4). alpha-Methylhistidine was used for a reliable diagnostic confirmation test, alpha-methyl-L-Dopa for a reliable exclusion test. Benzene showed no influence on either blanks or recovery rates, but inhibited the enzymic activity at pH 7.0, not however that of unspecific histidine decarboxylase and hence was very valuable as an additional diagnostic exclusion test (test 5). Although these new tests identifying acid (specific) histidine decarboxylase and demonstrating the specificity of its determination were tedious, despite the use of the modified isotope dilution method, they excluded the presence of any Dopa decarboxylase activity in mixtures with crude enzyme preparations as well as of any kind of nonspecific and nonenzymatic histidine decarboxylation. An endogenous histidine decarboxylase in rabbit gastric mucosa is postulated, capable of forming histamine in vivo. 相似文献
13.
Basolateral membrane Na/base cotransport is dependent on CO2/HCO3 in the proximal convoluted tubule 总被引:12,自引:2,他引:12
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The mechanism of basolateral membrane base transport was examined in the in vitro microperfused rabbit proximal convoluted tubule (PCT) in the absence and presence of ambient CO2/HCO3- by means of the microfluorometric measurement of cell pH. The buffer capacity of the cells measured using rapid NH3 washout was 42.8 +/- 5.6 mmol.liter-1.pH unit-1 in the absence and 84.6 +/- 7.3 mmol.liter-1.pH unit-1 in the presence of CO2/HCO3-. In the presence of CO2/HCO3-, lowering peritubular pH from 7.4 to 6.8 acidified the cell by 0.30 pH units and lowering peritubular Na from 147 to 0 mM acidified the cell by 0.25 pH units. Both effects were inhibited by peritubular 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonate (SITS). In the absence of exogenous CO2/HCO3-, lowering peritubular pH from 7.4 to 6.8 acidified the cell by 0.25 pH units and lowering peritubular Na from 147 to 0 mM decreased cell pH by 0.20 pH units. Lowering bath pH from 7.4 to 6.8 induced a proton flux of 643 +/- 51 pmol.mm-1.min-1 in the presence of exogenous CO2/HCO3- and 223 +/- 27 pmol.mm-1.min-1 in its absence. Lowering bath Na from 147 to 0 mM induced proton fluxes of 596 +/- 77 pmol.mm-1.min-1 in its absence. The cell acidification induced by lowering bath pH or bath Na in the absence of CO2/HCO3- was inhibited by peritubular SITS or by acetazolamide, whereas peritubular amiloride had no effect. In the absence of exogenous CO2/HCO3-, cyanide blocked the cell acidification induced by bath Na removal, but was without effect in the presence of exogenous CO2/HCO3-. We reached the following conclusions. (a) The basolateral Na/base n greater than 1 cotransporter in the rabbit PCT has an absolute requirement for CO2/HCO3-. (b) In spite of this CO2 dependence, in the absence of exogenous CO2/HCO3-, metabolically produced CO2/HCO3- is sufficient to keep the transporter running at 30% of its control rate in the presence of ambient CO2/HCO3-. (c) There is no apparent amiloride-sensitive Na/H antiporter on the basolateral membrane of the rabbit PCT. 相似文献
14.
M J Hughes I A Coret 《Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.)》1975,148(1):127-133
The onset of burimamide inhibition of histamine stimulation of rabbit atria is rapid, and a near steady-state blockade occurs at approximately 15 min ( larger than or equal to 90% complete). The blockade is reversible but requires several washings suggesting the disassociation is slow. The administration of histamine may accelerate the decay of the burimamide effect. Reciprocal plots (rate response versus histamine concentration) of dose-response curves are linear for both rabbit and guinea pig atria. In the presence of low concentrations of burimamide; (2.4 times 10-5 M), the displacement of curves suggests a competitive type of inhibition both for rabbit and guinea guinea pig atria. The apparent association constants calculated from these curves are: K1 (rabbit) 3.7 times 10-6M and K-1 (guinea pig) 6.7 times 10-6M. These results for guinea pig atria are in satisfactory agreement with the value obtained in another laboratory (2). At higher concentrations of burimamide, inhibition curves showed distinct evidence of departure from competitive character for both guinea pig and rabbit atria. 相似文献
15.
Micropuncture analysis of the cellular mechanisms of electrolyte secretion by the in vitro rabbit pancreas 总被引:4,自引:0,他引:4
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Micropuncture techniques have been used to study electrolyte secretion by the spontaneously secreting in vitro rabbit pancreas over a wide range of environmental conditions. Pancreatic secretion does not have a strong requirement for HCO3 and secretion continues at nearly normal rates when exogenous HCO3 is replaced by acetate. Acetate concentration in the juice averages 70 meq/liter, nearly three times the environmental concentration. The similar characteristics exhibited by HCO3 and acetate secretion indicate that they are secreted by a common mechanism involving active H transport. In vitro acid-base alterations demonstrate that the secretion rate is controlled by the environmental HCO3 concentration and to a much lesser extent by the pCO2. Secretion also requires active Na transport across the mucosal membrane. The effects of ouabain and a low Na environment strongly suggest coupling between the transport of Na and H and a cellular mechanism for electrolyte secretion is proposed involving Na-H exchange mechanisms at both the mucosal and serosal membranes. 相似文献
16.
1. The interrelationship of acidosis and Ca(2+) on the stimulation of gluconeogenesis by rat kidney-cortex slices was studied. 2. Ca(2+) stimulated gluconeogenesis from glutamine, glutamate, 2-oxoglutarate, succinate, malate, pyruvate, lactate and fructose, but not from galactose. 3. The [Ca(2+)] needed for optimum gluconeogenesis was about 2mm, but at this concentration, acidosis, produced in vitro by a decrease of [HCO(3) (-)] in the medium at constant pCO(2) or by an increase in pCO(2) at constant [HCO(3) (-)], did not stimulate gluconeogenesis. 4. In the absence of Ca(2+), acidosis (low [HCO(3) (-)]) stimulated gluconeogenesis from glutamine, glutamate, 2-oxoglutarate, succinate, malate, pyruvate and lactate but not from fructose or galactose. With succinate as substrate, the stimulatory effect of acidosis (low [HCO(3) (-)]) disappeared at Ca(2+) concentrations above 1.0mm. 5. The [HCO(3) (-)] was the most important determinant of the acidosis effect since a decrease of pH caused by an increase in pCO(2) did not uniformly stimulate gluconeogenesis, whereas a decrease in [HCO(3) (-)] without a change in pH consistently stimulated glucose formation in a way similar to the stimulation produced by acidosis (low [HCO(3) (-)]) in the absence of Ca(2+). 6. Acidosis in vitro inhibited the rate of decrease of activity of phosphoenolpyruvate carboxylase in slices, and Ca(2+) caused an increase in the activity of fructose 1-phosphate aldolase. 7. Respiratory acidosis in vitro caused an increase in the activity of phosphoenolpyruvate carboxylase in kidney cortex and an increase in gluconeogenesis from glutamine. 8. Possible points of interaction between Ca(2+), H(+) and HCO(3) (-) with the gluconeogenic sequence are discussed. 相似文献
17.
Microspectrofluorimetry of the fluorescent indicators 2',7'-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein and 6-methoxy-N-(3-sulfopropyl)-quinolinium was used to measure intracellular pH (pHi), intracellular Cl (Cli), and transmembrane fluxes of HCO3 and Cl in single parietal cells (PC) in isolated rabbit gastric glands incubated in HCO3/CO2-buffered solutions. Steady-state pHi was 7.2 in both resting (50 microM cimetidine) and stimulated (100 microM histamine) PCs. Transmembrane anion (HCO3 or Cl) flux rates during Cl removal from or readdition to the perfusate were the same in resting and stimulated PCs. These rates increased at alkaline pHi, though this pHi dependence was small in the physiological range. Maximum velocity (Vmax) for Cl influx or HCO3 efflux was 80-110 mM/min at pHi 7.6-7.8, and the Km for extracellular concentrations of Cl (Clo) was 25 mM; in the physiological range (pHi 7.1-7.3), Vmax for anion fluxes was approximately 50 mM/min. Steady-state Cli in the unstimulated PC was 62 +/- 5 mM, but on histamine stimulation, Cli decreased rapidly to 25 mM and then increased back to a steady-state level of 44 mM. HCO3 fluxes due to Cl removal or readdition were completely blocked by 0.5 mM 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid (H2DIDS), but Cl fluxes were only inhibited by 80%. H2DIDS did not inhibit the decrease in Cli that occurred with histamine treatment. Diphenylamine carboxylate (0.5 mM) inhibited Cl flux by only 50% and caused no additional inhibition of Cl flux when used in conjunction with H2DIDS. Transmembrane anion fluxes during solution Cl removal or readdition occurred 80% through the anion exchanger at the basal membrane and 20% through other pathway(s), presumably the Cl channel in the apical membrane. We conclude that the increase in transport activity via the Cl/HCO3 exchanger that occurs during histamine-induced increases in HCl secretion is due mostly to the decrease in Cli. In the resting cell with Cli = 62 mM, Clo = 120 mM, pHi = 7.2, and extracellular pH = 7.4, the anion exchanger is poised near its thermodynamic equilibrium. During histamine stimulation Cli drops from 62 mM to 44 mM, the thermodynamic equilibrium of the anion exchanger at the basolateral membrane is disturbed, and the anion exchanger then exchanges cellular HCO3 for extracellular Cl. Cli serves a crucial regulatory role in stimulus-secretion coupling in the PC. 相似文献
18.
The effect of changing [K+], [Na+] and [Cl-] in nutrient solution was studied in bullfrog antrum with and without HCO3- in nutrient. In 25 mM HCO3- (95% O2/5% CO2) and in zero HCO3- (100% O2), nutrient pH was maintained at 7.3. Changing from 4 to 40 mM K+ or from 81 to 8.1 mM Cl- gave a decrease 10 min later in transmucosal PD (nutrient became more negative)--a normal response. These responses were less in zero than in 25 mM HCO3-. A decrease from 102 to 8 mM Na+ decreased PD (anomalous response of electrogenic NaCl symport). This effect was attenuated or eliminated in zero HCO3-. In contrast, change from 4 to 40 mM K+ gave initial anomalous PD response and change from 102 to 8 mM Na+, initial normal PD response with either zero or 25 mM HCO3-. Both responses were associated with (Na+ + K+)-ATPase pump and were greater in zero than in 25 mM HCO3-. Initial PD increases in zero HCO3- are explained as due to increase in the resistance of passive conductance and/or NaCl symport pathways. Thus, removal of HCO3- modifies conductance pathways of nutrient membrane. 相似文献
19.
Schück O Matousovic K 《Biomedical papers of the Medical Faculty of the University Palacky, Olomouc, Czechoslovakia》2005,149(1):69-73
Acid-base balance evaluation according to the Henderson-Hasselbalch equation enable us to assess the contribution of respiratory (pCO2) and/or non-respiratory (metabolic, HCO3(-)) components to the acid-base balance status. A new approach to acid-base balance evaluation according to Stewart-Fencl, which is based on a detailed physical-chemical analysis of body fluids shows that metabolic acid-base balance disorders are characterized not only by [HCO3(-)]. According to this concept independent variables must be taken into an account. The abnormality of concentration of one or more of the independent variable(s) determines the pH of a solution. The independent variables are: 1. strong ion difference (SID); 2. total concentration of nonvolatile weak acids [A(tot)]; 3. in agreement with the Henderson-Hasselbalch concept also pCO2. Traditional evaluation of acid-base balance disorders is based on the pH of body fluids (though pH may be within normal range if several acid-base balance disturbances are present). In order to maintain this view and simultaneously to respect the Stewart-Fencl principle, we invented a new equation, which uses only the independent variables to define the pH of body fluids. This analysis shows that for a given value of pCO2, the pH of body fluids is determined by a difference between SID and [A(tot)-]. pH = 6.1 + log((SID - [A(tot)-])/(0.03pCO2)) or in itemized form: pH = 6.1 + log((([Na+] + [K+] + [Ca2+] + [Mg2+] - [Cl-] - [UA-]) - (k1[Alb] + k2[P(i)]))/(0.03 x pCO2)). Evaluation of the individual components of this equation enables us to detect, which of the independent variable (or a combination of independent variables) deviates from the normal range and therefore which one or ones is a cause of the acid-base balance disorder. At the end of this paper we give examples of a practical application of this equation. 相似文献
20.
Intracellular pH values (pHi) of Xenopus oocytes were optically measured using a fluorescent dye, 2', 7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). The oocytes were loaded with dye by incubation with a membrane-permeable form (BCECF-AM). Mean pHi of the oocytes in pH 7.6 solution was 7.69. Increasing ambient pCO2 rapidly decreased pHi and estimated buffering power was 23.8 mM/pH unit. Changing ambient HCO3- from 5 to 30 mM did not alter pHi. After incubation in a Na(+)-free solution, Na+ addition to the bath rapidly increased pHi and this response was blocked by amiloride (ED50 2 microM). The addition of NH4Cl to the bath caused an initial transient increase in PHi followed by a secondary decrease. The secondary decrease was greatly inhibited by a histidine specific reagent, diethylpyrocarbonate. It was also slightly inhibited by ouabain, Ba2+ and furosemide, but not by amiloride. These data suggest that (1), fluorescence technique is applicable to PHi measurements of Xenopus oocytes; (2), Xenopus oocytes have an amiloride sensitive Na+/H(+)-exchange, and permeabilities to CO2, NH3, and NH+4. These observation may be useful in studying the relationship between pHi and oocytes development, and the expression of acid/base transporters in Xenopus oocytes. 相似文献