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1.
Jill Eveloff Ekkehard Bayerdörffer Patricio Silva Rolf Kinne 《Pflügers Archiv : European journal of physiology》1981,389(3):263-270
Isolated cells were prepared from the medullary thick ascending limb of Henle's loop (TALH) and the response of oxygen consumption was correlated with the active chloride transport system found in these cells. Oxygen consumption was 31.6 l O2/mg protein·h and inhibited 50% by the absence of either sodium or chloride in the incubation medium. The absence of both sodium and chloride produced no further inhibition of oxygen consumption. Ouabain (10–4 M) inhibited oxygen consumption by 50% and the inhibitory effect depended on the presence of both sodium and chloride in the incubation medium. Further, furosemide inhibited oxygen consumption by a maximum of 50% at 10–3 M and also had no inhibitory effect if either sodium or chloride were absent. Furosemide had no effect on the Na, K-ATPase activity or ATP levels of the TALH cells. Thus, the data suggest that 50% of the oxygen consumption of the TALH cells is related to the movement of sodium and chloride into the cell and that the ions may be transported in a coupled manner.In addition the effect of various diuretics on oxygen consumption in the isolated TALH cells was tested. The diuretics could be grouped in three categories: (1) highly effective in inhibiting chloride-dependent oxygen consumption with an apparent inhibitory constant (K
i) of around 10–6 M, including the diuretics furosemide, bumetanide, ethacrynic acid-cysteine and piretanide, (2) diuretics which were less effective in inhibiting oxygen consumption with an apparentK
i of around 10–4 M, HOE 740 and ethacrynic acid, and (3) diuretics which were ineffective in inhibiting chloride-dependent oxygen consumption, amiloride and hydrochlorothiazide. 相似文献
2.
Masashi Imai 《Pflügers Archiv : European journal of physiology》1978,374(3):255-263
Ca2+ transport across the cortical thick ascending limb of Henle's loop (TALH) was studied on the isolated rabbit renal tubule
perfused in vitro. Both the efflux (Ke) and influx coefficient (Ki) of Ca2+ were determined with45Ca at three different levels of the transtubular electrical potential difference (PDt) caused by varying the transtubular
Na+ concentration gradient. The flux ratios,Ke/Ki, always exceeded those predicted from observed PDt by simple passive diffusion, and the net Ca2+ efflux was shown to occur against an electro-chemical potential. An increase in Ca2+ concentration in the perfusate was associated with a decrease inKe. Efflux of Ca2+, therefore, tended to be saturated as luminal Ca2+ concentration was increased.Ke (10−7 cm2/s) decreased from 4.13±0.56 to 2.02±0.30 (P<0.02) along with a decrease in PDt when 10−3 M NaCN was added to the bathing fluid. By contrast, 10−3 M iodoacetamide did not affectKe in spite of a significant decrease in PDt. Similarly, neither 10−5 M ouabain added to the bathing fluid nor 10−4 M ruthenium red, added either to the bathing fluid or the perfusate, affectedKe despite significant decreases in PDt. Addition of 10−5 M furosemide in the lumen caused a decrease inKe from 4.31±0.43 to 3.17±0.25 (P<0.01) in association with a decrease in PDt from 6.3±1.04 to 1.7±0.37 mV. These findings suggest; (a) Ca2+ transport across the TALH is mainly an active process requiring aerobic metabolism; (b) dissociation of Ca2+ and Cl− transport may occur under some experimental conditions. 相似文献
3.
Previous data from our laboratory have shown that active transport in the cortical thick ascending limb of Henle's loop (cTAL), as measured by the short circuit current (ISC, A · cm–2), requires the presence of Na+ and Cl–. The data were compatible with the model of secondarily active Cl– reabsorption involving the cotransport of Na+ and Cl– across the luminal membrane. The data suggested, furthermore, that 1 Na+ and 2 Cl– interact with the luminal carrier. In the present study it was tested whether this reabsorptive mechanism also requires the presence of luminal K+. Isolated cTAL segments (n=40) were perfused at high flow rates with a modified Ringer's solution. Removal of K+ from the lumen reduced ISC significantly from 215 to 133 A·cm–2. Addition of Ba2+ (10–3 mol·l–1) which blocks the K+ conductance of the luminal membrane, to the K+-containing lumen perfusate decreased ISC significantly from 234 to 141 A·cm–2. Combination of both manoeuvres: perfusion with a K+-free and Ba2+-containing solution almost abolished ISC from a control of 237 to 56 A · cm–2. The results are compatible with the view that in rabbit cTAL the luminal carrier interacts with all 3 ions, possibly 1 Na+, 2 Cl–, and 1 K+. K+ recycles across the luminal membrane through its conductive pathway.This study was supported by Deutsche Forschungsgemeinschaft Gr. 460/5-6-2 相似文献
4.
Membrane transport in the proximal tubule and thick ascending limb of Henle's loop: Mechanisms and their alterations 总被引:1,自引:0,他引:1
Summary Over the past few years, our knowledge on renal tubular transport mechanisms has increased considerably. Due to new technical developments, it is now possible to understand in part transepithelial transport and its pathological and pharmacological alterations at the level of the cell membranes. Different membrane transport mechanisms are discussed in this article, whereby sodium coupled solute transport in the proximal tubule and sodium chloride transport in the thick ascending limb of Henle's loop are taken as examples. It is indicated that an altered function of the kidney can often be equated with an alteration of the membrane transport. 相似文献
5.
Monika Wittner Claudia Weidtke Eberhard Schlatter Antonio di Stefano Rainer Greger 《Pflügers Archiv : European journal of physiology》1984,402(1):52-62
Isolated segments of cortical thick ascending limbs (cTAL) of rabbit kidney were perfused in vitro and the equivalent short circuit current (Isc) was measured. In a first series all substrates were removed on either side. Isc fell rapidly to 50±12% after 3 min and to 27±6% (n=5) after 10 min. This indicates that in cTAL segments Isc is strictly dependent on the presence of substrates. In series two it was tested what substrates can be utilized by the cTAL segment, and from which epithelial side [bath (b) or lumen (l)] the substrates are taken up. From the l-side only butyrate (10 mmol · l–1) sustained the Isc at 95±2% (n=7). All other tested substrates (10 mmol · l–1): pyruvate, acetate, -OH-butyrate,d-glucose, andl-lactate lead to a marked decline in Isc. From the b-side several substrates (5–10 mmol · l–1) sustained the Isc:d-glucose,d-mannose, butyrate, -OH-butyrate, acetoacetate,l-lactate, acetate and pyruvate. Other compounds (1–10 mmol · l–1): citrate, -ketoglutarate, succinate, glutamate, glutamine, propionate, caprylate and oleate did not sustain Isc. In the third series the mechanism of substrate utilization from the basolateral cell side was studied. It was shown that the Isc is a saturable function of thed-glucose,l-lactate, acetate, pyruvate or -OH-butyrate concentration with apparentK
m's between 0.05–1.0 mmol · l–1. Several known inhibitors of sugar and of anion transport were tested at the bath side: phlorrhizin was without effect. Phloretin (500 mol · l–1) inhibited Isc by 96%, yet its effect was not dependent on the presence of substrates on the b-side since inhibition ocurred also if the b-perfusate contained no substrate and Isc was driven by luminal butyrate. Also SITS (5 mmol · l–1) exerted only a small inhibitory effect which was not specific since it was also observed with luminal butyrate. -Cyano-m-OH-cinnamate (10 mmol · l–1) inhibited the Isc specifically whenl-lactate was the bath substrate. Probenecid (1 mmol · l–1) had a similar yet less marked inhibitory effect. Thed-glucose uptake from the b-side was specifically inhibited by cytochalasin B at 5 · 10–6 mol · l–1. We conclude that the cTAL segment of the rabbit utilizesd-glucose and/or small anions such as pyruvate orl-lactate or acetate to energize salt reabsorption. The link between substrate availability and salt reabsorption is extremely close in this nephron segment. Substrate uptake occurs from the blood side. Sugar uptake can be inhibited by cytochalasin B andl-lactate uptake by probenecid and -cyano-m-OH-cinnamat. These data suggest that substrate uptake at the basolateral cell side occurs probably via carrier systems.Parts of this study have been presented at the 57th and 58th Tagung Deutsche Physiologische Gesellschaft, 17th Meeting Europ. Soc. Clin. Investigation, and XXIX Int. Congress Physiol. Sciences. This study was supported by Deutsche Forschungsgemeinschaft Gr 480/5-7 相似文献
6.
Rainer Greger Claudia Weidtke Eberhard Schlatter Monika Wittner Brigitte Gebler 《Pflügers Archiv : European journal of physiology》1984,401(1):52-57
The Na+2Cl–K+ cotransporter in the apical membrane of the cortical thick ascending limb of the Henle's loop (cTAL) of rabbit nephron utilizes the electrochemical gradient for Na+ to transport K+ and Cl– against an unfavorable electrochemical gradient from lumen to cell interior. In the present study attempts are made to measure intracellular K+ activity (
) under control conditions and after inhibition of the cotransport system by furosemide (50·10–6 mol·l–1). 70 cTAL segments of 55 rabbits were perfused in vitro. Conventional Ling-Gerard and K+-selective microelectrodes were used to measure the PD across the basolateral membrane (PDbl) as well as the PD sensed by the single barrelled K+-selective electrode (
). PDbl was –64±1 (n=65) mV and
+15±1 (n=32) mV under control conditions. The positive
value, significantly different from zero, indicates that
is higher than predicted for passive distribution. The estimate for
obtained from PDbl and
was 113±8 mmol·l–1. Furosemide lead to the previously reported hyperpolarization of PDbl by 17±4 (n=13) mV and to a reduction of
from 15±1 to 5±1 (n=20) mV. The
, obtained from this set of data, was 117±9 mmol·l–1, and was not different from the control value. The present data indicate that
is significantly above Nernst equilibrium under control conditions. The source for this above equilibrium accumulation of K+ stems from the carrier mediated uptake of Na+2Cl– and K+. Consequently, the electrochemical gradient for K+ is rapidly reduced when the carrier is blocked by furosemide. The electrochemical gradient for K+, under control conditions, energizes the back leak of K+ from cell to lumen. This K+ flux is one component responsible for the lumen positive transepithelial PD.Parts of this study have been presented at the 58th Tagung Deutsche Physiologische und Deutsche Pharmakologische Gesellschaft, Mainz 1983; 67th Federation Meeting, Chicago 1983. This study was supported by Deutsche Forschungsgemeinschaft Gr. 480/5-7 相似文献
7.
R. N. Weiner R. Greger E. Schlatter F. Papavassiliou K. J. Ullrich 《Pflügers Archiv : European journal of physiology》1982,394(3):271-273
The present study examines directly the effect of a cytotoxin of Pseudomonas aeruginosa on the in vitro perfused rabbit cortical thick ascending limb of the loop of Henle (cTAL). 25 cTAL segments were perfused at high rate. The open circuit transepithelial electrical PD (PDte) and the specific electrical transepithelial resistance (Rt) were recorded continuously. From PDte/Rt the equivalent short circuit current (Isc) was calculated. The Isc was 214±30 A·cm–2 under control conditions, and decreased significantly to 74±34 A·cm–2 60 s after the addition of toxin (2 mg·l–1) to the lumen perfusate. Microscopic observation and photographs taken at that time clearly indicated swelling of the cTAL cells. Thereafter inhibition of active transport proceeded further, Rt fell progressively, and cells started to desquamate from the basement membrane. This effect of the toxin was dose dependent, and was half maximal at approximately 1.2 mg·l–1. From the bath side the effect was less marked and higher doses of toxin had to be used (half maximal effect at 5 mg·l–1). We conclude that this toxin of Pseudomonas aeruginosa exerts its toxic effect on the cTAL segment by increasing primarily the permeability of the lumen membrane.Part of this study has been presented at Spring meering Dt. Pharmakol. Ges., Mainz, 1982. This work has been supported by the Schutzkommission beim Bundesminister des Inneren, Bonn-Bad Godesberg, and by Deutsche Forschungsgemeinschaft, Gr 480/5-7 相似文献
8.
The effect of antidiuretic hormone (arginine vasopressin, AVP, 10−10mol.l−1), parathyroid hormone (PTH, 10−8 mol.l−8) and glucagon (10−8 mol.l−1) on the transepithelial potential difference (PDte) and the transepithelial resistance (Rte) were tested in in vitro perfused cortical (cTAL) and medullary (mTAL) thick ascending limbs of Henle's loop of the mouse
nephron. When compared with mTAL segments (PDte: 8.5±0.4 mV,n=16), cTAL segments displayed a high PDte of 15.7±0.9 mV (n=11) at the beginning of perfusion experiments which reached a value of 9.4±0.6 mV (n =11) after 38±4 min perfusion. Simultaneously Rte increased significantly from 24±3 to 28±1 Ω cm2 (n=11). When PTH, AVP or glucagon were added to the bath solution, PDte increased with PTH from 10.3±0.8 to 15.2±0.8 mV (n=13), with AVP from 10.2±0.5 to 15.0±0.7 mV (n=24) and with glucagon from 11.3±1.9 to 15.3±2.1 mV (n=8). At the same time Rte decreased from 30±3 to 23±2 Ω cm2, from 28±1 to 23±1 Ω cm2 and from 23±2 to 18±2 Ω cm2, respectively. In mTAL segments, AVP and glucagon increased PDte from 8.4+0.5 to 13.5±0.9 mV (n=11) and from 8.8±0.6 to 12.8±0.6 mV (n=8) respectively, while Rte decreased significantly from 23±1 to 20±1 Ω cm2 and from 27±3 to 21±3 Ω cm2. PTH, on the other hand, had no effect on PDte and Rte. Since the response to PTH appeared to be specific to cTAL segments, paired experiments were performed, in which AVP or glucagon
were successively tested with PTH on cTAL and mTAL segments, to ascertain the specificity of the hormonal response. In cTAL
segments, PTH and AVP increased the equivalent short-circuit current (Isc=PDte/Rte) by 82% and 86% respectively, while PTH and glucagon, in another series, increased Isc by 95% and 81% respectively. In mTAL segments, Isc was increased in the presence of AVP and glucagon by 88%, and 93% respectively, whereas PTH had no effect. These results
indicate that Nacl reabsorption in cTAL segments is stimulated by AVP, PTH and glucagon and in mTAL segments by AVP and glucagon.
The amplitude of the response to the hormones is similar in the two segments. The residual stimulation in cTAL segments, however,
persists longer than in mTAL segments. 相似文献
9.
Eberhard Schlatter Rainer Greger 《Pflügers Archiv : European journal of physiology》1985,405(4):367-376
The effect of cAMP on transepithelial and transmembrane potential differences and resistances was examined in isolated in vitro perfused mouse medullary thick ascending limbs of Henle's loop (mTAL). The effects of furosemide and barium were tested. Stimulation of NaCl transport by ADH 10–9+dbcAMP 4·10–4+forskolin 10–6 mol·l–1 (paired experiments) resulted in: a) an increase in transepithelial potential difference, referenced to the grounded bath, from +6.7±0.3 mV to +12.0±0.4 mV (n=47); b) a decrease in transepithelial resistance from 25±1 cm2 to 20±1 cm2 (n=47); c) a depolarization of the basolateral membrane by 12 mV and of the apical membrane by 7 mV (n=36); d) a decrease in the fractional resistance of the basolateral membrane from 0.27±0.005 to 0.15±0.06 (n=12). Furosemide (10–4 mol·l–1) abolished the active transepithelial transport potential and hyperpolarized the basolateral membrane potential to values which were similar in both control and cAMP treated mTAL segments. Barium increased the transepithelial resistance and depolarizedPD
bl to similar values in both functional states. An increase in the fractional conductance of the basolateral membrane was also seen, if, prior to the cAMP treatment, the luminal Na+2Cl–K+ contransport was inhibited by furosemide. Thus, we propose that stimulation of active NaCl reabsorption in the mTAL segment of the mouse by ADH, mediated via cAMP, increases primarily the basolateral chloride conductance.Supported by Deutsche Forschungsgemeinschaft Gr 480/6-2Parts of this study have been presented at the 59th Meeting of the German Physiological Society in Dortmund 1984 and at the 69th FASEB Meeting in Anaheim 1985 相似文献
10.
Marie-Marcelle Trinh-Trang-Tan Olivier Levillain Lise Bankir 《Pflügers Archiv : European journal of physiology》1988,411(6):676-680
It has recently been reported that branched-chain amino acid aminotransferase (BCAATase) is inhomogeneously distributed in the kidney. BCAATase activity is several-fold higher in the medullary thick ascending limb (MTAL) than in other nephron segments. The present work was designed to determine whether leucine, a branched-chain amino acid (AA), is used as metabolic fuel by this nephron segment. MTAL were isolated from the inner stripe of the outer medulla of adult Sprague Dawley rats by mild enzymatic digestion and appropriate sieving. Leucine aminotransferase activity measured in homogenates of MTAL was 653±52 pmol -ketoglutarate formed/g protein per hour, a value threefold higher than that observed in the renal cortex or muscle in the same rats. Substrate oxidation was assessed by measuring14CO2 production from tracer amounts of uniformly labeled14C-amino acids or glucose in isolated MTAL incubated in modified Earle balanced salt solution. When each substrate was offered at a concentration of 1 mM, leucine oxidation was much higher than that of unbranched AA, but fivefold lower than that of glucose. With 1 mM glucose and 1 mM leucine in the medium, leucine oxidation was close to that of glucose (123±8 versus 177±15 pmol CO2/g protein per hour), probably because glucose contributed to the formation of -ketoglutarate, a cosubstrate for leucine transamination. Inhibition of salt transport by furosemide (0.1 mM) decreased oxidation of both substrates by 60–70%. Inhibition of salt transport by ouabain (1 mM) decreased glucose oxidation markedly. However, it doubled leucine oxidation when glucose was absent from the medium and decreased leucine oxidation by only 28% when glucose was present. This might be due to an ouabain-dependent alteration in membrane permeability to AA. These findings show that leucine is oxidized in rat MTAL and may contribute to supporting active transport in this segment. This contribution could be important after a protein meal or on high protein diet, situations in which plasma level of branched-chain AA is elevated. 相似文献
11.
R. Nitschke U. Fröbe R. Greger 《Pflügers Archiv : European journal of physiology》1991,417(6):622-632
The effect of antidiuretic hormone ([Arg]vasopressin, ADH) on intracellular calcium activity [Ca2+]i of isolated perfused rabbit cortical thick ascending limb (cTAL) segments was investigated with the calcium fluorescent dye fura-2. The fluorescence emission ratio at 500–530 nm (R) was monitored as a measure of [Ca2+]i after excitation at 335 nm and 380 nm. In addition the transepithelial potential difference (PD
te) and transepithelial resistance (R
te) of the tubule were measured simultaneously. After addition of ADH (1–4 nmol/l) to the basolateral side of the cTAL R increased rapidly, but transiently, from 0.84±0.05 to 1.36±0.08 (n = 46). Subsequently, within 7–12 min R fell to control values even in the continued presence of ADH. The increase in R evoked by the ADH application corresponded to a rise of [Ca2+]i from a basal level of 155±23 nmol/l [Ca2+]i up to 429±53 nmol/l [Ca2+]i at the peak of the transient, as estimated by intra- or extracellular calibration procedures. The electrical parameters (PD
te and R
te) of the tubules were not changed by ADH. The ADH-induced Ca2+ transient was dependent on the presence of Ca2+ on the basolateral side, whereas luminal Ca2+ had no effect. d(CH2)5[Tyr(Me)2]2,Arg8vasopressin, a V1 antagonist (Manning compound, 10 nmol/l), blocked the ADH effect on [Ca2+]i completely (n = 5). The V2 agonist 1-desamino-[d-Arg8]vasopressin (10 nmol/l, n=4), and the cAMP analogues, dibutyryl-cAMP (400 mol/l, n = 4), 8-(4-chlorophenylthio)-cAMP (100 mol/l, n = 1) or 8-bromo-cAMP (200 mol/1, n = 4) had no influence on [Ca2+]i. The ADH-induced [Ca2+]i increase was not sensitive to the calcium-channel blockers nifedipine and verapamil (100 mol/l, n = 4). We conclude that ADH acts via V1 receptors to increase cytosolic calcium activity transiently in rabbit cortical thick ascending limb segments, possibly by an initial Ca2+ release from intracellular stores and by further Ca2+ influx through Ca2+ channels in the basolateral membrane. These channels are insensitive to L-type Ca2+ channel blockers, e.g. nifedipine and verapamil.Supported by DFG GR 480/10 相似文献
12.
Patricio Silva Barbara Koenig Stephanie Lear Jill Eveloff Rolf Kinne 《Pflügers Archiv : European journal of physiology》1987,409(1-2):74-80
Because the medullary thick ascending limb of the loop of Henle is the target of several polypeptide hormones that stimulate adenyl cyclase in this nephron segment, we examined the effects of cyclic AMP on thick ascending limb of the loop of Henle cells isolated by enzymatic digestion and density gradient centrifugation from the outer medulla of the rabbit kidney. The functional parameter that was measured was transport dependent oxygen consumption. Oxygen consumption was measured using a polarographic oxygen electrode in a constant temperature chamber. We found that dibutyryl cyclic AMP inhibited oxygen consumption in a dose dependent way. Maximal inhibition was observed at a concentration of 10–5M. The effect of dibutyryl cyclic AMP was not present in the absence of either sodium, chloride or both implying that its effect is restricted to the sodium and chloride dependent oxygen consumption. The effect of dibutyryl cyclic AMP was additive to that of furosemide·10–4M while that of furosemide was not additive to that of cyclic AMP suggesting that the site of action of cyclic AMP is distal to that of furosemide. The effect of dibutyryl cyclic AMP was not additive to that of ouabain and was absent in cells where oxygen consumption was stimulated with amphotericin B in the absence of chloride indicating that it has no effect on Na–K-ATPase. On the basis of these results, and by analogy with other epithelia where the effect of cyclic AMP on chloride transport has been demonstrated to be at the chloride conductance, the effect of cyclic AMP on transport dependent oxygen consumption can be most probably localized to the chloride conductance. The hormone responsible for the primary step was not identified in these experiments. 相似文献
13.
To examine whether sodium reabsorption in the thick ascending limb of Henle's loop (TALH) in the dog kidney has a passive component, the ratios between reductions in sodium reabsorption and oxygen consumption (ΔNa/ΔQo 2 ratio) were measured by inhibiting tubular transport with bumetanide (30 μg kg-1) and ouabain (120 ng kg-1 intrarenally). Clearance studies were performed in volume expanded dogs treated with acetazolamide (100 mg kg-1) or maleate (400 mg kg-1). In five acetazolamide-treated dogs, bumetanide gave a ΔNa/ΔQo 2 ratio of 29.9±2.5, whereas the combination of bumetanide and ouabain gave 19.0±0.6. When ouabain was given before bumetanide, ouabain gave a ΔNa/ΔQo 2 ratio of 19.2±1.1 and the combination gave 19.9±1.2. In the maleate-treated dogs, bumetanide gave a ΔNa/Qo 2 ratio 30.3±1.7, and the combination of bumetanide and ouabain gave 27.1±1.5. To localize the metabolic effect of bumetanide and ouabain, local heat production was measured at 18 places in four kidneys with copper-constantan thermocouples. Bumetanide reduced metabolic rate in the outer medulla by 51±4%, and in the cortex by 16±6%. Subsequent infusion of ouabain reduced metabolic rate in the outer medulla by only 9±3%, whereas cortical metabolism was reduced by 33±4%. The results show that bumetanide mainly acts in the outer medullar where TALH is located, whereas the additional effect of ouabain is mainly located in cortical segment of the nephron including the proximal tubules. Bumetanide inhibits the reabsorption of 30 mol sodium for each mole oxygen consumed, which show that for each 18 mol sodium that are transported through the cells in the TALH in dog kidneys, 12 mol (40%) are transported along the paracellular route without additional requirement of energy. 相似文献
14.
Properties of the basolateral membrane of the cortical thick ascending limb of Henle's loop of rabbit kidney 总被引:10,自引:2,他引:8
Rainer Greger Eberhard Schlatter 《Pflügers Archiv : European journal of physiology》1983,396(4):325-334
The present study utilizes the transepithelial and transmembrane electrophysiological approach to study the properties of the basolateral membrane of the in vitro perfused cortical thick ascending limb of Henle's loop (cTAL) of rabbit kidney. Eight different series were performed in a total of 119 tubules. The key observations are: 1. A K+ concentration upward step in the bath from 3.6 to 18.6 mmol · l?1 depolarizes the basolateral membrane by 19±2 mV. 2. This depolarization can be abolished when Ba2+ (3 mmol · l?1) is added to the bath: The depolarization by Ba2+ alone is equal to that by Ba2+ plus the K+ concentration upward step (21.7 versus 22.4 mV). 3. This effect of Ba2+ is not accompanied by any change in transepithelial resistance nor in the fractional resistance of the basolateral membrane. 4. A Cl? concentration downward step in the bath from 150 to 50 mmol · l?1 leads to a depolarization between 8–15 mV. We conclude that the K+ exit at the basolateral membrans is mainly electroneutral and that Cl? leaves the cell both electroneutrally (KCl) and diffusionally. The present data, together with previous findings from our laboratory, are used to draw a tentative model for the NaCl reabsorption in the cTAL segment. In this model the (Na++K+)-ATPase provides the primary driving force. Na+, 2 Cl?, K+ are cotransported luminally, K+ recycles across the lumen membrane. Cl? leaves the cell in part in conjunction with K+, and thus utilizing the chemical gradient for K+, and the remainder leaves the cell through the Cl? conductive pathway. The discrepancy of the conductivity properties of both cell membranes, the lumen membrane K+ conductive, and the basolateral membrane Cl? conductive, is the main source for the lumen positive transepithelialPD. ThePD, in turn, drives a seizable fraction of the Na+ through the paracellular shunt pathway. 相似文献
15.
Rainer Greger Eberhard Schlatter 《Pflügers Archiv : European journal of physiology》1983,396(4):315-324
Previous data suggest the cotransport of Na+, Cl?, and K+ across the lumen membrane of the cortical thick ascending limb (cTAL) of rabbit nephron. For this cotransporter to operate K+ recycling across the lumen membrane has to be postulated. The present data focus on the conductivity properties of the lumen membrane. Methods for impaling individual cells of in vitro perfused cTAL segments are described. The meanPD across the lumen membrane (PD 1) is 76 mV (lumen positive). Rapid increase in lumen perfusate K+ concentration (3.6→18.6 mmol · l?1) leads to a depolarization ofPD 1 by 20 mV. Ba2+ (3 mmol · l?1) added to the lumen perfusate inhibits the K+ conductive pathway of the lumen membrane, and consequently increases the voltage divider ratio (current pulse induced voltage deflection across the lumen membrane divided by that across the basolateral membrane) from 2 to 36 as well as transepithelial resistance from 34 to 46 Ω cm2. From these changes, and with the use of simultaneous equations of the VDR and of Kirchhoff's law, the resistances of the lumen membrane (88 Ω cm2), of the basolateral membrane (47 Ω cm2), and of the paracellular shunt pathway (47 Ω cm2) can be calculated. Using these estimates of the individual resistances and using the observed change inPD 1 in the K+ concentration step experiments (29±3 mV per decade in K+ concentration change) an apparent transference number of the lumen membrane for K+ in the order of 0.9–1.0 can be calculated. This indicates that the lumen membrane is essentially K+ conductive. This conclusion is strengthened further by the results of another series in which no evidence for a Cl? conductive pathway in the lumen was obtained. The data of this study can be used to calculate the intracellular K+ activity of some 90–100 mmol ·l?1. For this K+ activity the K+ diffusion from cell to lumen equals the carrier mediated uptake from lumen to cell. This indicates that an essentially complete recycling of K+ across the lumen membrane of the cTAL segment is feasible. 相似文献
16.
M. Wittner B. Mandon N. Roinel C. de Rouffignac A. Di Stefano 《Pflügers Archiv : European journal of physiology》1993,423(5-6):387-396
Recent studies from our laboratory have shown that in the cortical thick ascending limb of Henle's loop of the mouse (cTAL) Ca2+ and Mg2+ are reabsorbed passively, via the paracellular shunt pathway. In the present study, cellular mechanisms responsible for the hormone-stimulated Ca2+ and Mg2+ transport were investigated. Transepithelial voltages (PDte) and transepithelial ion net fluxes (J
Na, J
Cl, J
K, J
Ca, J
Mg) were measured in isolated perfused mouse cTAL segments. Whether parathyroid hormone (PTH) is able to stimulate Ca2+ and Mg2+ reabsorption when active NaCl reabsorption, and thus PDte, is abolished by luminal furosemide was first tested. With symmetrical lumen and bath Ringer's solutions, no Ca2+ and Mg2+ net transport was detectable, either in the absence or in the presence of PTH. In the presence of luminal furosemide and a chemically imposed lumen-to-bath directed NaCl gradient, which generates a lumen-negative PDte, PTH slightly but significantly increased Ca2+ and Mg2+ net secretion. In the presence of luminal furosemide and a chemically imposed bath-to-lumen-directed NaCl gradient, which generates a lumen-positive PDte, PTH slightly but significantly increased Ca2+ and Mg2+ net reabsorption. In view of the observed small effect of PTH on passive Ca2+ and Mg2+ movement, a possible interference of furosemide with the hormonal response was considered. To investigate this possibility, Ca2+ and Mg2+ transport was first stimulated with PTH in tubules under control conditions. Then active NaCl reabsorption was abolished by furosemide and the effect of PTH on J
Ca and J
Mg measured. In the absence of PDte and under symmetrical conditions, no Ca2+ and Mg2+ transport was detectable, either in the presence or absence of PTH. In the presence of a bath-to-lumen-directed NaCl gradient, Ca2+ and Mg2+ reabsorption was significantly higher in the presence than in the absence of PTH. Finally, when active NaCl transport was not inhibited by furosemide, but reduced by a bath-to-lumen-directed NaCl gradient, PTH strongly increased J
Ca and J
Mg, whereas only a small increase in PDte was noted. In conclusion, these data suggest that PTH exerts a dual action on Ca2+ and Mg2+ transport in the mouse cTAL by increasing the transepithelial driving force for Ca2+ and Mg2+ reabsorption through hormone-mediated PDte alterations and by modifying the passive permeability for Ca2+ and Mg2+ of the epithelium, very probably at the level of the paracellular shunt pathway. 相似文献
17.
Rainer Greger Eberhard Schlatter Florian Lang 《Pflügers Archiv : European journal of physiology》1983,396(4):308-314
Previously we have shown that chloride reabsorption in the isolated perfused cortical thick ascending limb of Henle's loop of rabbit (cTAL) is dependent on the presence of sodium and potassium. The present study was performed to elucidate the dependence on chloride in quantitative terms. Ninety-four cTAL segments were perfused at high rates with solutions of varying chloride concentration. Chloride was substituted by sulfate, methylsulfate or nitrate. The open circuit transepithelial electrical potential difference (PD te, mV) and the specific transepithelial resistance (R t, Ωcm2) were measured, and from both the equivalent short circuit current (I sc) was calculated. The correlation ofI sc versus Cl? concentration (in 294 observations) revealed a saturation kinetics depending on the (Cl?)2 concentration. The apparent constants wereK 1/2 50 mmol·l?1 andI sc,max 198 μA cm?2. These results are compatible with the assumption of 2 Cl? interacting with the luminal cotransport system. Although this finding in conjunction to our previous observations already is highly suggestive for a non-charged carrier this question was pursued further by recording the membranePD across both cell membranes during luminal application of furosemide. The data (n=26) indicate that furosemide (10?5 mol·l?1, lumen) produces an immediate decline inPD te to values close to zero, and a simultaneous hyperpolarization of both cell membranes by 15±2 mV for the basolateral and by 8±2 mV for the lumen membrane. These data exclude the possibility that the lumen positive transepithelialPD is generated by a net negative current flow from the lumen into the cell. It is concluded that the cotransport across the luminal membrane of the cTAL segment is electroneutral and involves 1 Na+∶2 Cl?∶1K+. The hyperpolarization of both cell membranes observed immediately after application of furosemide is caused most likely by a rapid fall in intracellular chloride activity. 相似文献
18.
A. Di Stefano R. Greger C. de Rouffignac M. Wittner 《Pflügers Archiv : European journal of physiology》1992,420(3-4):290-296
The aim of the present study was to investigate whether bicarbonate buffer (CO2 + HCO
3
–
) is required to sustain maximal NaCl transport in the cortical thick ascending limb of Henle's loop (cTAL) of the mouse. Transepithelial Na+ and Cl– net fluxes (J
Na, J
Cl, pmol min–1 mm–1), measured by electron microprobe analysis, were similar irrespective of the presence or absence of CO2 + HCO
3
–
in luminal and bathing solutions J
NaCl with CO2 + HCO
3
–
=203±25 pmol min–1 mm–1; J
NaCl without CO2 + HCO
3
–
=213±13 pmol min–1 mm–1, n=14). Furthermore the transepithelial potential difference, V
te, the transepithelial resistance, R
te, and the basolateral membrane potential, V
bl, were unaffected by CO2 + HCO
3
–
. In the absence of CO2 + HCO
3
–
, V
te was +17.0±1.7 mV(n=9) (lumen positive), R
te was 28±2 cm2 (n=9) and V
bl was –76±4 mV (n=6). In the presence of CO2 + HCO
3
–
, V
te, R
te and V
bl were +15.9±1.5 mV, 29±1 cm2 and –73±5 mV, respectively. 4-Acetamido-4-isothiocyanatostilbene-2,2-disulphonic acid (SITS; 0.1 mmol l–1) and amiloride (1 mmol l–1) added to the (CO2 + HCO
3
–
)-containing lumen perfusate were without effect on V
te and R
te. Finally, the effect of furosemide (0.1 mmol l–1) on V
te and V
bl in the presence of CO2 + HCO
3
–
was investigated. Furosemide reversibly decreased V
te from +13.7±1.1 mV to +1.7±0.7 mV (n=6) and hyperpolarized Vbl from –70±1 to –89±3 mV (n=5), suggesting passive distribution of Cl– across the basolateral membrane. In conclusion, these data suggest that active NaCl transport in the cTAL of the mouse does not require the presence of CO2 + HCO
3
–
. 相似文献
19.
A. Di Stefano M. Wittner B. Corman 《Pflügers Archiv : European journal of physiology》1991,419(3-4):327-331
The maximal urinary osmolality that can be reached by the kidney is reduced with age. This may be due to impaired NaCl transport by the medullary thick ascending limb of Henle's loop, which is part of the renal concentrating mechanism and is modulated by antidiuretic hormone (ADH). We therefore tested in vitro a possible age-related change in the transport capacity and in the response of this nephron segment to ADH in young (1–2 months) and old (20–24 months) mice. The transepithelial potential difference (V
te) was significantly higher in young mice (+8.5±0.4 mV, n=13) than in old ones (+6.6±0.5 mV, n=17). Addition of 0.1 nmol.l–1 ADH to the bath solution significantly increased V
te by 5.2±0.5 mV in the young and by 3.1±0.6 mV in the old animals. Application of dibutyryl-cAMP (0.1 mmol.1–1) did not further increase the hormonal response in both groups. The ADH-mediated increase in the corresponding equivalent short-circuit current (I
SC = V
te/Rte) was twice as great in young mice as in old, indicating that the stimulation of NaCl transport by ADH across the medullary thick ascending limb is significantly reduced with age. These results suggest that the previously reported age-related defect in the urinary concentrating ability of the kidney is partly due to a decreased response of the medullary thick ascending limb to ADH. 相似文献
20.
M. Wittner A. Di Stefano E. Schlatter J. Delarge R. Greger 《Pflügers Archiv : European journal of physiology》1986,407(6):611-614
Torasemide (1-isopropyl-(4-(3-methylphenylamino)pyrid-3-yl)urea) is a new diuretic. The present study examines the effects of this substance in the isolated perfused thick ascending limb (TAL) of mouse and rabbit kidney. In cortical TAL segments of the rabbit, torasemide added to the lumen perfusate led to a fall in equivalent short circuit current (= transepithelial voltage divided by transepithelial resistance, which corresponds to the rate of chloride reabsorption) with a half maximal inhibition concentration of 3 · 10–7 mol/l. This effect was accompanied by a hyperpolarization of the luminal and basolateral membrane from –78 to –81 mV and from –72 to –81 mV, respectively. A similar hyperpolarization of both membrane voltages was also observed in medullary TAL segments of the mouse. Torasemide, added to the basolateral perfusate of cortical TAL segments of the rabbit, also inhibited the equivalent short circuit current. However, 3 · 10–5 mol/l were necessary for a half maximal inhibition. The fall in the equivalent short circuit current was accompanied by a significant increase in transepithelial resistance from 34 to 38 cm2, by an increase in the fractional resistance of the basolateral membrane, and by a hyperpolarization mainly of the basolateral membrane. Again, similar results were obtained in the medullary TAL segment of the mouse.The strong inhibitory effect of torasemide from the lumen side can be explained by an interference with the Na+ 2Cl–K+ carrier in the luminal membrane. In fact, torasemide apparently is structurally related to furosemide. The weaker effect of torasemide from the peritubular side can, at least in part, be explained as an interference with chloride channels present in the basolateral membrane. Torasemide is also structurally related to chloride channel blockers such as diphenylamine-2-carboxylate.Supported by Deutsche Forschungsgemeinschaft DFG Gr 480/6-4 and 6-5 相似文献