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1.
Intracellular adenosine triphosphate (ATP)-inhibited K+ currents (I
K, ATP
) in canine coronary artery smooth muscle cells were characterized in the wholecell configuration using the suction pipette method. Cells dialysed internally with solutions containing 5 mM ATP (ATPi) showed little detectable whole-cell current at potentials more negative than –30 mV. However, cells dialysed with ATPi-free solutions developed a time- and voltage-independent current which reached a maximum of 132±25 pA at –40 mV about 10 min following patch rupture. After run-up, the current showed little run-down. Concentration-dependent inhibition by ATPi yielded an inhibition constant (K
i of 350 M and a Hill coefficient of 2.3. In ATPi-free solutions, the large current at –40 mV was reduced by glibenclamide with aK
i of 20 nM and a Hill coefficient of 0.95. Conversely, in 1 mM ATPi solutions, the small current at –40 mV was increased by P-1075 from 8±2 pA to 143±33 pA, with a dissociation constant (K
d) of 0.16 M and a Hill coefficient of 1.7. The effect of P-1075 was antagonized by glibenclamide. Maximal current density elicited by either ATPi depletion or external application of the channel opener P-1075 was similar with slope conductances of 81±10 pS/pF and 76±13 pS/pF respectively in the potential range of –90 to –40 mV. External Ca2+ had no effect on this current. Finally, in 1 mM ATPi, 20 and 50 M adenosine increased the current slope conductance by 36±15% and 73±10% respectively between –90 to –40 mV. TheI
K, ATP
although very small in these cells, was extremely effective in causing membrane potential hyperpolarization. 相似文献
2.
Makoto Takano Frances M. Ashcroft 《Pflügers Archiv : European journal of physiology》1996,431(4):625-631
We studied the block of whole-cell ATP-sensitive K+ (KATP) currents in mouse pancreatic-cells produced by external Ba2+. Ba2+ produced a time- and voltage-dependent block of KATP currents, both the rate and extent of the block increasing with hyperpolarization. With 5.6 mM [K+]o, the relationship between the steady-state KATP current and [Ba2+]o, was fit by the Hill equation with aK
d of 12.5 ± 2.8 M at –123 mV and of 0.18 ± 0.02 mM at –62 mV The Hill coefficient (n) was close to 1 at all potentials indicating that binding of a single Ba2+ ion is sufficient to block the channel. When [K+]o was raised to 28 mM the Kd was little changed (12.4 ± 4.1 gM at –123 mV 0.27 ± 0.05 mM at –62 mV) and n was unaffected, suggesting that K+ does not interact with the Ba2+ binding site. The kinetics of Ba2+ block were slow, 10 M Ba2+ blocking the KATP current with a time constant of 20 ms at –123 mV in 28 mM [K+]o. The blocking rate constant was calculated as 1.7 mM–1 ms–1 and the unblocking rate as 0.02 ms–1, at –123 mV The data are discussed in terms of a model in which Ba2+ binds to a site at the external mouth of the channel to inhibit the KATP channel. 相似文献
3.
Sulayma Albarwani Blair E. Robertson Piers C. G. Nye Roland Z. Kozlowski 《Pflügers Archiv : European journal of physiology》1994,428(5-6):446-454
A novel class of Ca2+-activated K+ channel, also activated by Mg-ATP, exists in the main pulmonary artery of the rat. In view of the sensitivity of these KCa,ATP channels to such charged intermediates it is possible that they may be involved in regulating cellular responses to hypoxia. However, their electrophysiological profile is at present unknown. We have therefore characterised the sensitivity of KCa,ATP channels to voltage, intracellular Ca2+ ([Ca2+]i) and Mg-ATP. They have a conductance of 245 pS in symmetrical K+ and are approximately 20 times more selective for K+ ions than Na+ ions, with a K+ permeability (P
K) of 4.6×10–13cm s–1. Ca2+ ions applied to the intracellular membrane surface of KCa,ATP channels causes a marked enhancement of their activity. This activation is probably the result of simultaneous binding of at least two Ca2+ ions, determined using Hill analysis, to the channel or some closely associated protein. This results in a shift of the voltage activation threshold to more hyperpolarized membrane potentials. The activation of KCa,ATP channels by Mg-ATP has an EC50 of approximately 50 M. Although the EC50 is unaffected by [Ca2+]i, channel activation by Mg-ATP is enhanced by increasing [Ca2+]i. One possible interpretation of these data is that Mg-ATP increases the sensitivity of KCa,ATP channels to Ca2+. It is therefore possible that under hypoxic conditions, where lower levels of Mg-ATP may be encountered, the sensitivity of KCa,ATP channels to Ca2+ and therefore voltage is reduced. This would tend to induce a depolarising influence, which would favour the influx of Ca2+ through voltage-activated Ca2+ channels, ultimately leading to increased vascular tone. 相似文献
4.
Eric Guillemare Michel Lazdunski Eric Honoré 《Pflügers Archiv : European journal of physiology》1994,428(5-6):604-609
The two-microelectrode voltage-clamp technique was used to monitor K+ channel activity in Xenopus oocyte follicular cells, which are electrically coupled to the oocyte itself by gap junctions. Endogenous vasodilators such as calcitonin gene-related peptide (CGRP), vasoactive intestinal peptide (VIP), prostaglandin E2 (PGE2) and adenosine activate glibenclamide-ATP-sensitive K+ (KATP) channels in Xenopus oocyte follicular cells. The mechanism of action of CGRP was studied in detail. CGRP effects undergo a rapid desensitization. CGRP acts via CGRPI receptors. Its effects are antagonized by the amino-truncated CGRP analog hCGRP(8–37). The second messenger for CGRP activation of KATP channels is cAMP. Phosphodiesterase inhibition by 3-isobutyl-1-methylxanthine enhances the CGRP response while adenyl cyclase inhibition by either 2,5-dideoxyadenosine or progesterone nearly completely depresses the CGRP response. Vasoconstrictors such as ACh and angiotensin II also have receptors in follicular cells. ACh strongly inhibits the CGRP activation of K+ channels as it inhibits the activation of KATP channels by P1060, but angiotensin II does not. It is concluded that as in vascular smooth muscle cells, CGRP and probably other hyperpolarizing vasodilators open KATP channels in follicular cells by protein kinase A activation.Thanks are due to C. Roulinat and F. Aguila for expert technical assistance. This work was supported by the Centre National de la Recherche Scientifique (CNRS). 相似文献
5.
Several secretagogues were used in this study, including those which enhance intracellular cyclic adenosine monophosphate (cAMP) production, as well as others which elevate intracellular Ca2+ activity and are known to increase Cl– secretion in the intact colon and in colonic carcinoma cell lines. They were examined with respect to their effects on electrophysiological properties in isolated rabbit distal colonic crypts. Crypts were dissected manually and perfused in vitro. Transepithelial voltage (V
te), transepithelial resistance (R
te), membrane voltage across the basolateral membrane (V
bl), and fractional basolateral membrane resistance (FR
bl), were estimated. Basolateral prostaglandin E2 (PGE2, 0.1 mol/l), vasoactive intestinal peptide (VIP, 1 nmol/l) and adenosine (0.1 mmol/l) induced an initial depolarisation and a secondary partial repolarisation of (V
bl). In the case of adenosine, the initial depolarization of (V
bl) was by 31±2 mV (n=47).R
te fell significantly from 16.4±3.6 to 14.2±3.7 ·cm2 (n= 6), andFR
blincreased significantly from 0.11±0.02 to 0.51±0.10 (n=6). In the second phase the repolarisation of (V
bl) amounted 11±2 mV (n=47) and a steadystate (V
bl) of –51±2 mV (n=47) was reached.R
te fell further and significantly to a steady-state value of 12.4±3.8 ·cm2 (n=6) andFR
bl fell significantly to 0.42±0.13 (n=6). In 30% of the experiments, a transient hyperpolarisation of (V
bl) by 8±2 mV (n=14) was seen during wash out of adenosine. In the presence of adenosine, but not under control conditions, lowering of luminal Cl– concentration from 120 to 32 mmol/l depolarised (V
bl) significantly by 8±1 mV (n=9). Basolateral ATP and ADP (0.1 mmol/l) led to a short initial depolarisation followed by a sustained and significant hyperpolarisation by 6±2 mV (n=27) and 5±4 mV (n=8), respectively. Carbachol (CCH) hyperpolarised (V
bl) in a concentration-dependent manner. At 100 mol/l (bath) the hyperpolarisation was by 14±2 mV (n=11) andFR
bl fell slightly. Neurotensin (10 nmol/l), isoproterenol (10 mol/l) and uridine 5-triphosphate (UTP, 0.1 mmol/l) had no effect. It is concluded that PGE2, VIP and adenosine upregulate sequentially a luminal Cl– conductance and a basolateral K+ conductance by increasing intracellular cAMP concentration. Ca2+ mobilising hormones such as ATP, ADP, and CCH increase the basolateral K+ conductance, while the effect on luminal Cl– conductance appears to be very limited. 相似文献
6.
Jürgen Schrader Gert Baumann Eckehart Gerlach 《Pflügers Archiv : European journal of physiology》1977,372(1):29-35
Summary Infusion of adenosine into the coronary arteries of isolated guinea pig hearts produced a dosedependent inhibition of dP/dtmax caused by bolus injections of isoproterenol (4×10–11 moles). Threshold concentration of adenosine was 10–7 M and maximal inhibition (90%) occurred at 10–5 M. Coronary dilation induced by, papaverine did not influence the contractile response to catecholamines. In addition to its influence on cardiac performance, adenosine (10–5 M) effectively inhibited the isoproterenol (10–7 M) induced initial rise in myocardial levels of cyclic 35-AMP, glucose-1-phosphate and glucose-6-phosphate. Adenosine also antagonized the effect of isoproterenol on adenylate cyclase activity in a crude membrane preparation from guinea pig ventricles; it was without effect on the activity of the membrane phosphodiesterase. Theophylline inhibited the actions of adenosine both on adenylate cyclase activity and on contractile force development.-Upon infusion of isoproterenol (3×10–7 M) into the coronary arteries of the isolated heart (perfusion at constant pressure), the adenosine concentration in the effluent perfusate increased within 45 s from 10–8 M to about 10–6 M. It thus appears conceivable that in ventricular myocardium endogenously formed adenosine may serve 2 functions: dilation of the coronary arteries and limitation of the inotropic and metabolic effects of catecholamines.A preliminary report of these studies was presented at the 47th Meeting of the German Physiological Society in Regensburg, Germany [Pflügers Arch.365, R4 (1976)] 相似文献
7.
J. L. Saumet G. Leftheriotis A. Dittmar G. Delhomme C. S. Degoute 《European journal of applied physiology》1986,54(6):574-577
Summary The effect of general anaesthesia on skin blood flow in the left hand, measured by a new non-invasive probe using the thermal clearance method was examined. A mercury silastic gauge was placed around the third left finger and the plethysmographic wave amplitude was recorded to measure changes in finger pulse amplitude. Heart rate (HR), mean arterial blood pressure (MABP) and skin temperature were also recorded. General anaesthesia was induced by droperidol and phenoperidine injection and propanidid infusion in eight female patients. Skin thermal clearance, plethysmographic wave amplitude, HR, MABP and skin temperature were 0.40±0.02 w · m–1 C–1, 9±1 mm, 98±5 beats · min–1, 12.50±0.93 kPa and 33.3±3.4 C respectively. The minimal value of MABP was 9.58 ±1.06 kPa, whereas skin thermal clearance, plethysmographic wave amplitude, HR and skin temperature increased to 0.45±0.02 w · m–1 C–1, 29±3 mm, 110±4 beats · min–1 and 34.4±0.4 C. Changes in skin thermal clearance correlated well with plethysmographic wave amplitude. Statistically significant changes in these two parameters occurred before significant change in HR, MABP or skin temperature. The results show that the new noninvasive probe using the thermal clearance method appears to be a useful device for measuring cutaneous microcirculation in anaesthetized humans, and responds more quickly than change in skin temperature, which is a delayed effect of skin blood flow change. Our results also show that the intensity of cutaneous vasodilatation induced by general anaesthesia did not relate to the vascular tone before anaesthesia. 相似文献
8.
Klaus Turnheim Rudolf Donath Michael Weissel Norbert Kolassa 《Pflügers Archiv : European journal of physiology》1976,365(2-3):197-202
Summary In isolated K+ (16.2 mM)-arrested cat hearts perfused at constant pressure adenosine infusions (0.8 moles · min–1 · 100 g–1 for 10 min) caused an increase in myocardial14C-glucose uptake and release of14CO2+H14CO
–
3
and14C-lactate simultaneously with a rise in coronary flow. The ratio of the release of14CO2+H14CO
–
3
to that of14C-lactate and the specific activity of lactate in the effluate were not altered. In K+-arrested hearts perfused with constant volume neither glucose uptake nor glucose breakdown were influenced by 0.8 or 100 moles · min–1 · 100 g–1 adenosine with 0.1–5 mM glucose in the perfusion medium. It is concluded that adenosine does not affect directly the myocardial glucose carrier system, aerobic or anaerobic glucose breakdown or glycogenolysis, but enhances glucose uptake secondarily by increasing coronary flow. This interpretation is substantiated by the finding that mechanically produced increases in perfusion volume caused similar increases in myocardial glucose uptake as were observed with comparable adenosine-induced coronary flow increments.Parts of this work were presented at the 44th Meeting of the German Physiological Society in Bochum, FRG, March 18–21, 1975 [Pflügers Arch.355, R 16 (1975)] 相似文献
9.
M. Wittner A. di Stefano P. Wangemann R. Nitschke R. Greger C. Bailly C. Amiel N. Roinel C. de Rouffignac 《Pflügers Archiv : European journal of physiology》1988,412(5):516-523
The effect of antidiuretic hormone (arginine vasopressin, AVP) on transepithelial Na+, Cl–, K+, Ca2+ and Mg2+ net transports was investigated in medullary (mTAL) and cortical (cTAL) segments of the thick ascending limb (TAL) of mouse nephron, perfused in vitro. Transepithelial net fluxes (J
Na
+,J
Cl
–,J
K
+,J
Ca
2+,J
Mg
2+) were determined by electron probe analysis of the collected tubular fluid. Transepithelial potential difference (PDte) and transepithelial resistance (Rte) were measured simultaneously. cTAL segments were bathed and perfused with isoosmolal, HCO
3
–
containing Ringer solutions, mTAL segments were bathed and perfused with isoosmolal HCO
3
–
free Ringer solutions. In cTAL segments, AVP (10–10 mol·l–1) significantly increasedJ
Mg
2+ andJ
Ca
2+ from 0.39±0.08 to 0.58±0.10 and from 0.86±0.13 to 1.19±0.15 pmol·min–1 mm–1 respectively. NeitherJ
Na
+ norJ
Cl
–, (J
Na
+: 213±30 versus 221±28 pmol·min–1 mm–1,J
Cl
–: 206±30 versus 220±23 pmol·min–1 mm–1) nor PDte (13.4±1.3 mV versus 14.1±1.9 mV) or Rte (24.6±6.5 cm2 versus 22.6±6.4 cm2) were significantly changed by AVP. No significant effect of AVP on net K+ transport was observed. In mTAL segments, Mg2+ and Ca2+ net transports were close to zero and AVP (10–10 mol·l–1) elicited no effect. However NaCl net reabsorption was significantly stimulated by the hormone,J
Na
+ increased from 107±33 to 148±30 andJ
Cl
– from 121±33 to 165±32 pmol·min–1 mm–1. The rise inJ
NaCl was accompanied by an increase in PDte from 9.0±0.7 to 13.5±0.9 mV and a decrease in Rte from 14.4±2.0 to 11.2±1.7 cm2. No K+ net transport was detected, either under control conditions or in the presence of AVP.To test for a possible effect of HCO
3
–
on transepithelial ion fluxes, mTAL segments were bathed and perfused with HCO
3
–
containing Ringer solutions. With the exception ofJ
Ca
2+ which was significantly different from zero (J
Ca
2+: 0.26±0.06 pmol·min–1 mm–1), net transepithelial fluxes of Na+, Cl–, K+ and Mg2+ were unaffected by HCO
3
–
. In the presence of AVP,J
Mg
2+ andJ
Ca
2+ were unaltered whereasJ
NaCl was stimulated to the same extent as observed in the absence of HCO
3
–
. In conclusion our results indicate heterogeneity of response to AVP in cortical and medullary segments of the TAL segment, since AVP stimulates Ca2+ and Mg2+ reabsorption in the cortical part and Na+ and Cl– reabsorption in the medullary part of this nephron segment.This study was supported by the Commission des communautés européennes, grant no. ST2J 00951 F(CD), and by Wissenschafts-ausschuß der Nato über den DAAD 相似文献
10.
E. Lohrmann I. Burhoff R. B. Nitschke H. J. Lang D. Mania H. C. Englert M. Hropot R. Warth W. Rohm M. Bleich R. Greger 《Pflügers Archiv : European journal of physiology》1995,429(4):517-530
Previously we have shown that arylamino-benzoates like 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB), which are very potent inhibitors of NaCl absorption in the thick ascending limb of the loop of Henle, are only poor inhibitors of the cAMP-mediated secretion of NaCl in rat colon. This has prompted our search for more potent inhibitors of NaCl secretion in the latter system. The chromanole compound 293 B inhibited the equivalent short-circuit current (I
sc) induced by prostaglandin E2 (n=7), vasoactive intestinal polypeptide (VIP,n=5), adenosine (n=3), cholera toxin (n=4) and cAMP (n=6), but not by ionomycin (n=5) in distal rabbit colon half maximally (IC50) at 2 mol/l from the mucosal and at 0.7 mol/l from the serosal side. The inhibition was reversible and paralleled by a significant increase in transepithelial membrane resistance [e.g. in the VIP series from 116±16 ·cm2 to 136±21 ·cm2 (n=5)]. A total of 25 derivatives of 293 B were examined and structure activity relations were obtained. It was shown that the racemate 293 B was the most potent compound with-in this group and that its effect was due to the enantiomer 434 B which acted half maximally at 0.25 mol/l. Further studies in isolated in vitro perfused colonic crypts revealed that 10 mol/l 293 B had no effect on the membrane voltage across the basolateral membrane (V
bl) in non-stimulated crypt cells: –69±3 mV versus –67±3 mV (n=10), whilst in the same cells 1 mmol/l Ba2+ depolarised (V
bl) significantly. However, 293 B depolarised (V
bl) significantly in the presence of 1 mol/l forskolin: –45±4mV versus –39±5 mV (n=7). Similar results were obtained with 0.1 mmol/l adenosine. 293 B depolarised (V
bl) from –40±5 mV to –30±4 mV (n=19). This was paralleled by an increase in the fractional resistance of the basolateral membrane. VIP had a comparable effect. The hyperpolarisation induced by 0.1 mmol ATP was not influenced by 10 mol/l 293 B: –75±6 mV versus –75±6 mV (n=6). Also 293 B had no effect on basal K+ conductance (n=4). Hence, we conclude that 293 B inhibits the K+ conductance induced by cAMP. This conductance is apparently relevant for Cl– secretion and the basal K+ conductance is insufficient to support secretion. 相似文献
11.
Characterization of two MDCK-cell subtypes as a model system to study principal cell and intercalated cell properties 总被引:1,自引:2,他引:1
Michael Gekle Stefan Wünsch Hans Oberleithner Stefan Silbernagl 《Pflügers Archiv : European journal of physiology》1994,428(2):157-162
Madin-Darby canine kidney (MDCK) cells originate from the renal collecting duct and consist of different cell subtypes. We cloned two MDCK cell subtypes denominated as C7 and C11 with different morphology and different function. The two clones maintained their functional differences after cloning. C7 monolayers exhibit a high transepithelial resistance (R
te=5648±206 · cm2, n=20) and secrete K+ (K+=1.31±0.08 mmol/l, n=10) into the apical medium. C11 monolayers display a low R
te (330±52 · cm2, n=20) and secrete Cl– (Cl–= 16.9±1.8 mmol/l, n=10) into the apical medium. Aldosterone (1 mol/l) stimulates K+ secretion (K+ of 3.58±0.11 mmol/l, n=7) in C7 cells and H+ secretion in C11 cells (pH=0.060±0.007, n=10). Aldosterone-induced stimulation of K+ secretion is inhibited by apical application of amiloride (1 mol/l). cAMP stimulates H+ secretion in C11 cells (pH= –0.068±0.004, n=10). Furthermore, C7 cells are peanut-lectin(PNA)-negative and exhibit an intracellular pH of 7.39±0.05 (n=7), whereas C11 cells maintain intracellular pH at 7.16±0.05 (n=8) and a major fraction of cells is PNA positive. We conclude that we have cloned two subtypes of MDCK cells which stably express different functional characteristics. The C7 subtype resembles principal cells (PC) of the renal collecting duct, whereas the C11 subtype resembles intercalated cells (ICC) of the renal collecting duct. The C11 subtype seems heterogenous and possibly consists of two subpopulations similar to the -ICC and -ICC of the renal collecting duct. The two cloned subtypes could serve as a valuable model to study PC-like cells and ICC-like cells independently. 相似文献
12.
Unlike mammalian muscle cells in culture, cultured myotomal muscle cells of Xenopus embryos express ATP-sensitive K+ (KATP) channels. The KATP channels are blocked by internal ATP (half-maximal inhibition K
0.5 = 16 M) and to a lesser extent by internal ADP, are voltage independent, have an inward rectification at positive potentials and are inhibited by glibenclamide (K
0.5 = 2 M). Surprisingly, these KATP channels are not sensitive to K+ channel openers such as cromakalim. Opening of these KATP channels does not occur under normal physiological conditions. It is elicited by metabolic exhaustion of the muscle cell and it precedes the development of an irreversible rigor state. Neither intracellular acidosis nor an increase of intracellular Ca2+ are involved in KATP channel opening. Different types of K+ channels are successively expressed after plating of myotomal muscle cells: (1) sustained delayed-rectifier K+ channels; (2) KATP channels; (3) inward-rectifier K+ channels; (4) transient delayed-rectifier K+ channels. The current density associated with KATP channels far exceeds that of voltage-dependent K+ channels. Innervation controls the expression of these KATP channels. Co-culture of muscle cells with neurons from the neural tube decreases the number of active KATP channels per patch. Similarly, in situ innervated submaxillaris muscle of tadpoles at stage 50–55 has a very low density of KATP channels. 相似文献
13.
W. V. Breuer E. Mack A. Rothstein 《Pflügers Archiv : European journal of physiology》1988,411(4):450-455
In dissociated MDCK cells, activators of the cyclic AMP system cause depolarization detectable by changes in fluorescence of the membrane potential sensitive dye bisoxonol. Addition of forskolin (60 M), vasopressin (2 M), 8-bromo-cyclic AMP (0.5 mM) or l-epinephrine (10 M) depolarized the cells substantially in low Cl– (5 mM) but had little effect in high Cl– (140 mM) solution. These results are consistent with cyclic AMP activation of Cl– channels. The Ca2+-ionophore ionomycin (1 M) produced a rapid hyperpolarization in low and high Cl– solutions, consistent with K+ channel opening. Using a clonal subline, MDCK-14, the magnitude of the ionomycin hyperpolarization was roughly proportional to the concomitant rise in [Ca2+]i as measured with the intracellular Ca2+ probe indo-I. Both l-epinephrine and isoproterenol appeared to activate the Cl– channels. However only l-epinephrine produced a [Ca2+]i rise and a transient hyperpolarization (due to K+ channel opening), which preceeded the depolarization due to Cl– channel opening. The l-epinephrine-induced [Ca2+]i response of the heterogeneous MDCK cell population but not of the clonal subline MDCK-14 was inhibited by removal of extracellular Ca2+. In the latter only the slow secondary phase of the [Ca2+]i rise was affected by Ca2+ removal. It is concluded that l-epinephrine activates K+ and Cl– channels in a sequential manner in MDCK cells by Ca2+ and cAMP signals, presumably via - and -adrenergic receptors located on the same cell.Abbreviations MDCK cells
Madin Darby Canine Kidney cells
- [Ca2+]i
intracellular calcium concentration
- [Cl–]i
intracellular chloride concentration
- [Cl–]o
extracellular chloride concentration
- [Na++K+]i
intracellular concentration of Na+ and K+
- [Na++K+]o
extracellular concentration of Na+ and K+
- EM
transmembrane potential
- ECl
chloride equilibrium potential
- EK
potassium equilibrium potential
- bis-oxonol
[bis(1,3-diethylthio-barbiturate)] trimethine oxonol
- DMSO
dimethylsulfoxide
- EDTA
ethylenediaminetetraacetic acid
- EGTA
ethylene glycol bis (-aminoethyl ether) N,N-tetraacetic acid
- Hepes
4-(2-hydroxyethyl)-1 piperazineethanesulfonic acid
- NMG+
N-methylglucamine
- RPMI medium
Rosewell Park Memorial Institute medium 相似文献
14.
Andreas Lückhoff Michael Horster 《Pflügers Archiv : European journal of physiology》1981,391(4):301-305
The distal colon (rat) was perfused in vivo at low rates (1–2 ml/h). Dialyzed polyethyleneglycol 4,000 (PEG) was used to vary the luminal osmotic activity (eff). Perfusate sodium concentration of 3.7 mmol l–1±0.9 (SD) resulted in an effluent Na+ of 3.3 mmol l–1±1.4. Potassium concentrations of 15 or 25 mmol l–1 remained unaltered. At this cationic steady-state, transmural net water flux (J
v
) was linearly correlated (r=–0.96,n=24) to the logarithm of eff.J
v
was zero at 634 mosm l–1.L
p
(l h–1 cm–2 mosm–1 l) was 0.195 at eff 85 mosm l–1 and 0.046 at 1,050 mosm l–1. These data characterize the distal colon as an epithelium with high capacity for salt conservation. 相似文献
15.
Takako Ohno-Shosaku Chosaburo Yamamoto 《Pflügers Archiv : European journal of physiology》1992,422(3):260-266
To determine whether membranes of mammalian central neurons contain an ATP-sensitive K+ (KATP) channel similar to that present in pancreatic cells, the patch-clamp technique was applied to cultured neurons prepared from the neonatal rat cerebral cortex and hippocampus. In whole-cell experiments with hippocampal neurons, extracellular application of 0.5 mM diazoxide (a KATP channel activator) elicited a hyperpolarization concomitant with an increase in membrane conductance, whereas application of 0.5 mM tolbutamide (a KATP channel blocker) induced a depolarization with a decrease in conductance. Similar results were obtained with cortical neurons. In outside-out patch experiments with cortical neurons, a K+ channel sensitive to these drugs was found. The channel was completely blocked by 0.5 mM tolbutamide and activated by 0.5 mM diazoxide. The single-channel conductance was 65 pS under symmetrical 145 mM K+ conditions and 24 pS in a physiological K+ gradient. In inside-out patch experiments, this channel was demonstrated to be inhibited by an application of 0.2–1 mM ATP to the cytoplasmic surface of the patch membrane. These results indicate that the membranes of rat cortical neurons contain a KATP channel that is quite similar to that found in pancreatic cells. It is also suggested that the same or a similar K+ channel may exist in membranes of hippocampal neurons. 相似文献
16.
H. Pavenstädt F. Bengen M. Späth P. Schollmeyer R. Greger 《Pflügers Archiv : European journal of physiology》1993,424(2):137-144
The effects of bradykinin (BK) and histamine (Hist) on the membrane voltage (V
m), ion conductances and ion channels of cultured human glomerular epithelial cells (hGEC) were examined with the nystatin patch clamp technique. Cells were studied between passage 3 and 20 in a bath rinsed with Ringer-like solution at 37°C. The mean value of V
m was –41±0.5 mV (n=189). BK (10–6 mol/l, n=29) and Hist (10–5 mol/l, n= 55) induced a rapid transient hyperpolarization by 15±1 mV and 18±1 mV, respectively. The hyperpolarization was followed by a long lasting depolarization by 6±1 mV (BK 10–6 mol/l) and 7±1 mV (Hist 10–5 mol/l). The ED50 was about 5×10–8 mol/l for BK and 5×10–7 mol/l for Hist. In the presence of both agonists, increases of outward and inward currents were observed. A change in the extracellular K+ concentration from 3.6 to 30 mmol/l depolarized V
m by 8±1 mV and completely inhibited the hyperpolarizing effect of both agents (n=11). Reduction of extracellular Cl– concentration from 145 to 30 mmol/l led to a depolarization by 2 ±1 mV (n=25). In 30 mmol/l Cl– the depolarizations induced by BK (10–7 mol/l) and Hist (10–6 mol/l) were augmented to 9±2 mV (n=14) and to 10±2 mV (n=11), respectively. Ba2+ (5 mmol/l) depolarized V
m by 19±5 mV (n=6) and completely inhibited the hyperpolarization induced by BK (10–6 mol/l, n=3) and reduced that of Hist (10–5 mol/l) markedly (n=3). Preincubation with the K+ channel blocker charybdotoxin (1–10 nmol/l) for 3 min had no significant effect on V
m, but reduced markedly the BK(10–6 mol/l, n=11) and Hist-(10–5 mol/l, n=6) induced hyperpolarizations. In 10 out of 31 experiments in the cell attached nystatin patch configuration big K+ channels with a conductance
of 247±17 pS were found. The open probability of these K+ channels was increased 3- to 5-fold during the hyperpolarization induced by BK (10–7 mol/l) or Hist (10–5 mol/l, both n= 4). In excised inside/out patches this K+ channel had a mean conductance of 136±8.5 pS (n=10, clamp voltage 0 mV). The channel was outwardly rectifying and its open probability was increased when Ca2+ on the cytosolic side was greater than 0.1 mol/l. The data indicate that BK and Hist activate a
and a
in hGEC. The hyperpolarization is induced by the activation of a Ca2+-dependent maxi K+ channel. 相似文献
17.
Y. Kondo T. Bührer E. Frömter W. Simon 《Pflügers Archiv : European journal of physiology》1989,414(6):663-668
A new Cl– selective microelectrode based on the ionophore 5,10,15,20-tetraphenyl-21H,23H-porphin manganese(III) chloride is presented which discriminates better against HCO
3
–
and several organic anions than electrodes containing the Corning 477913 ion-eschanger. Using a redesigned construction procedure, fine-tip double-barrelled microelectrodes were produced which had slopes of –52.4±0.6 mV (SE,n=24), resistances of about 7·1011 and a selectivity coefficient logK
Cl, HCO3
pot
of –1.40±0.03. Some electrodes showed a small unexplained sensitivity to pH>7.6. When used to puncture cells of isolated S3 segments of rabbit renal proximal tubule during perfusion with HCO
3
–
Ringer solution, the electrodes gave a membrane potential of –69.8±1.5 mV and an intracellular Cl– activity, [Cl–]i, of 35.3±2.6 mmol/l. Upon switching bath and lumen perfusions to Cl–-free solutions the residual [Cl–]i dropped to 1.20±0.03 mmol/l, while in similar measurements with ion-exchanger electrodes the residual [Cl–]i dropped only to 10.9±0.5 mmol/l. These observations demonstrate the superiority of the new electrode and prove that previously determined high [Cl–]i values in Cl–-free ambient solutions reflect interference problems rather than non-exchangeable intracellular chloride. 相似文献
18.
Nina M. Griffiths N. L. Simmons 《Pflügers Archiv : European journal of physiology》1990,416(1-2):80-87
Specific 125I-labelled vasoactive intestinal peptide (VIP) binding was determined in feline renal cortical and medullary plasma membranes. For the cortex, Scatchard analysis of the data resulted in a curvilinear plot with a high-affinity site K
0.5 of 8.4±2.6 nmol l–1 (SE, n=6) and a second low-affinity site K
0.5 204 ± 16 nmol l–1 with binding site concentrations (B
max) of 385±44.5 and 2710±181.3 fmol mg protein–1 respectively. Conversely a similar analysis of the results obtained for outer medullary membranes gave a single site with a K
0.5 of 1.2±0.2 nmol l–1 (SE, n=4) and B
max of 157.8±24.7 fmol mg–1. Inner medullary membrane binding data. Gave a single site of lower affinity (K
0.5= 62.5±21.6 nmol l–1; n=3). Structurally related peptides, glucagon and secretin, were ineffective (up to 1 mol l–1) in displacing VIP from specific sites in both cortex and medulla. Porcine PHI 1–27 (a peptide having N-terminal histidine and C-terminal isoleucine) and a VIP antagonist [4-Cl-D-Phe6Leu17]VIP both displaced 125I-VIP from cortical and medullary membrane binding sites with IC50 values of 43.0 nmol l–1 and 1.3 mol l–1 (cortex) and 132.0 nmol l–1 and 1.5 mol l–1 (medulla) respectively. The localisation of specific VIP binding sites in feline kidney was investigated further by in vitro autoradiography. A high density of binding sites was visible at the cortico-medullary boundary as well as in the outer stripe of the outer medulla and in radial structures projecting into the cortex. There was a moderate density of binding sites in the superficial cortex. In addition the distribution of tubular VIP-sensitive adenylate cyclase was determined in microdissected nephron segments. In the presence of 10 mol l–1 GTP, 1 mol l–1 VIP effected marked stimulations over basal adenylate cyclase activity in medullary collecting tubules (4.7-fold), the bright and granular portions of the distal convoluted tubule (4.1- and 2.2-fold respectively) and in the pars recta of the proximal tubule (2.7-fold). Thus VIP-responsive adenylate cyclase has a discrete localisation in the feline nephron, which appears to correlate with specific binding sites as defined by autoradiography. 相似文献
19.
Shai D. Silberberg Cornelis van Breemen 《Pflügers Archiv : European journal of physiology》1992,420(1):118-120
K+ channels which are inhibited by intracellular ATP (ATPi) (KATP channels) are thought to be the physiological target site of the K+ channel opening drugs (2) and to underlie a variety of physiological phenomena including hypoxia induced vasodilation (3). However, electrophysiological evidence for ATPi-regulated K+ currents in smooth muscle is scarce. We, therefore, investigated the effects of one K+ channel opener, lemakalim, and metabolic inhibition on the membrane conductance of freshly dissociated rabbit mesenteric artery smooth muscle cells, using the perforated-patch whole cell recording technique (6). The cells were metabolically inhibited with 1 mM iodoacetic acid and 50 M dinitrophenol. Both lemakalim (0.1–3 M) and metabolic inhibition activated a time-independent and glyburide sensitive K+ current at physiological membrane potentials. The similarities between the lemakalim and metabolic inhibition activated currents suggest that a single class of channels underlies both currents. These results are the first whole-cell current recordings to demonstrate the activation of a smooth muscle membrane conductance by metabolic inhibition, lending support to the view that hypoxia induced vasodilation arises from the activation of KATP channels. 相似文献
20.
Ian Findlay 《Pflügers Archiv : European journal of physiology》1988,412(1-2):37-41
K+ currents were recorded from ATP-sensitive channels in inside-out membrane patches excised from isolated rat ventricular myocytes. ATP-sensitive K+ channel inhibition could be evoked by ATP in the absence of magnesium where most ATP would be present as the free acid ATP4–. Channel inhibition was enhanced when the same total concentration of ATP was applied in the presence of magnesium, where most ATP would be bound as ATP·Mg. Dose-response relationships for ATP-sensitive K+ channel inhibition evoked by ATP had a Hill coefficient of 2 andK
i of 17 and 30 M for ATP in the presence and absence of magnesium respectively. This was the obverse of the expected results if ATP4– were to be the sole form of ATP to effect channel closure. ATP-sensitive K+ channel inhibition evoked by ATPS, AMP-PNP and AMP-PCP was also enhanced in the presence of magnesium. It is concluded that the ATP-sensitive K+ channel of rat ventricular myocytes binds and is closed by both the free-acid and divalent-cationbound forms of ATP. 相似文献