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  • Cl− conductance  (11)
  • K+ conductance  (9)
  • Rat  (7)
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  • 1
    ISSN: 1432-2013
    Keywords: Proximal tubule ; Kidney ; K+ conductance ; Cell membrane potential ; Ouabain temperature ; Phlorizin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract In isolated perfused segments of the mouse proximal tubule, the potential difference across the basolateral cell membrane (PDbl) was determined with conventional microelectrodes. Under control conditions with symmetrical solutions it amounted to −62±1 mV (n=118). The potential difference across the epithelium (PDte) was −1.7±0.1 mV (n=45). Transepithelial resistance amounted to 1.82±0.09 kΩ cm (n=28), corresponding to 11.4±0.6 Ω cm2. Increasing bath potassium concentration from 5 to 20 mmol/l depolarized PDbl by +24±1 mV (n=103), and PDte by +1.6±0.1 mV (n=19). Thus, the basolateral cell membrane is preferably conductive to potassium. Rapid cooling of the bath perfusate from 38°C to 10°C led to a transient hyperpolarization of PDbl from −60±1 to −65±1 mV (n=21) within 40 s followed by gradual depolarization by +18±1% (n=14) within 5 min. The transepithelial resistance increased significantly from 1.78±0.11 kΩ cm to 2.20±0.21 kΩ cm (n=15). Rapid rewarming of the bath to 38°C caused a depolarization from −61±2 mV (n=17) to −43±2 mV (n=16) within 15 s followed by a repolarization to −59±2 mV (n=10) within 40 s. Ouabain invariably depolarized PDbl. During both, sustained cooling or application of ouabain, the sensitivity of PDbl to bath potassium concentration decreased in parallel to PDbl pointing to a gradual decrease of potassium conductance. Phlorizin hyperpolarized the cell membrane from −59±2 to −66±1 mV (n=13), virtually abolished the transient hyperpolarization under cooling, and significantly reduced the depolarization after rewarming from +17±2 mV (n=16) to +9±3 mV (n=9). The present data indicate that the contribution of peritubular potassium conductance to the cell membrane conductance decreases following inhibition of basolateral (Na++K+)-ATPase. Apparently, cooling from 37° to 10°C does not only reduce (Na−+K+)-ATPase activity but in addition luminal sodium uptake mechanisms such as the sodium glucose cotransporter. As a result, cooling leads to an initial hyperpolarization of the cell followed by depolarization only after some delay.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-2013
    Keywords: Human sweat duct ; Cl− conductance ; Cl− channel blockers ; Cystic fibrosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract To characterize the chloride conductance of human sweat duct the effect of various analogues of diphenylamine-2-carboxylate was investigated on the transepithelial potential difference (PDT) and resistance (R T ) of isolated microperfused sweat ducts. Although the most powerful analogues which block Cl− channels in various secretory and absorptive epithelia were ineffective, a number of analogues (in particular Cl substituted ones) were found which at high concentrations significantly and reversibly increased PDT andR T . The data suggest that the main chloride conductance pathway of sweat duct epithelium resides in the cell membranes rather than in the tight junctions. In addition the different blocking spectra of the chloride conductances of sweat duct and tracheal epithelium (Welsh MJ, Science 232:1648, 1986) suggest that the combined impairment of both conductances in cystic fibrosis does not result from a molecular defect in the Cl− channels.
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 357 (1975), S. 201-207 
    ISSN: 1432-2013
    Keywords: Allantoin ; Uricase ; Kidney ; Clearance ; Micropuncture ; Rat
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary Renal excretion of allantoin was measured by tracer techniques. After injection of 2-C14 urate and H3 inulin, clearances of allantoin and inulin were measured and both proximal and distal tubules were micropunctured. In confirmation of earlier results 2-C14 urate injected into an intact animal is very rapidly converted to C14 allantoin: after 15 min more than 90% of urinary tracer is present as allantoin. It was further observed that 1) allantoin clearance is essentially identical with inulin clearance over a wide range of urine flows; 2) no net transport of allantoin occurs in either proximal or distal tubules. Clearly allantoin is handled by the rat kidney like inulin. The total excretion of filtered allantoin unlike that of filtered urate provides an easy and effective mechanism for animals possessing the enzyme uricase to dispose of their purine loads.
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 351 (1974), S. 323-330 
    ISSN: 1432-2013
    Keywords: Uricase ; Urate ; Allantoin ; Liver ; Kidney ; Microperfusion ; Rat
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary 1. In vivo uricase activity was tested in rats by injection of 2-C14 urate and measurement of the total C14 activity and the fractional activities of allantoin, allantoic acid and urea in samples of blood and urine. In control animals, 5 min after the injection, 70% of the plasma tracer was already present in the form of allantoin. No allantoic acid and urea were produced. Intestinectomy had no measurable influence on uricase activity. On the other hand, hepatectomy or ligation of the hepatic artery combined with subtotal viscerectomy did abolish uricase activity almost completely. 2. Following microinjections into proximal tubules of Ringer solution containing 2-C14 urate, urine samples during early recovery mainly contained labelled urate, whereas in later samples the fraction of labelled allantoin increased. About 12 min after the microinjection the urine of both kidneys contained equal amounts of tracer mainly in the form of allantoin. 3. When segments of proximal tubules were perfused with an equilibrium solution containing tracer amounts of C 14 urate, no urate was metabolized during its passage through the proximal tubule. 4. C 14 urate was offered from the peritubular capillaries and samples of tubular fluid were analyzed, Again, all the tracer in the tubular fluid was in the form of urate, indicating that urate is not oxidized when it is transported across the tubular cell. It is concluded from these results that: 1. The rat kidney has no significant uricase activity. 2. Urate transport in the kidney is not influenced by this enzyme. 3. The degradation of urate to allantoin takes place at extrarenal sites, mainly in the liver.
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 411 (1988), S. 546-553 
    ISSN: 1432-2013
    Keywords: Pancreas ; Isolated perfused ducts ; Luminal membrane ; Cl− conductance ; Cl−/HCO 3 − antiport ; cAMP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The aim of the present study was to investigate by what transport mechanism does HCO 3 − cross the luminal membrane of pancreatic duct cells, and how do the cells respond to stimulation with dibytyryl cyclic AMP (db-cAMP). For this purpose a newly developed preparation of isolated and perfused intra-and interlobular ducts of rat pancreas was used. Responses of the epithelium to inhibitors and agonists were monitored by electrophysiological techniques. Addition of HCO 3 − /CO2 to the bath side of nonstimulated ducts depolarized the PD across the basolateral membrane (PDbl) by about 9mV, as also observed in a previous study [21]. This HCO 3 − effect was abolished by Cl− channel blockers or SITS infused into the lumen of the duct: i. e. 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB, 10−5 M) hyperpolarized PDbl by 8.2±1.6 mV (n=13); 3′,5-dichlorodiphenylamine-2-carboxylic acid (DCl-DPC, 10−5 M) hyperpolarized PDbl by 10.3±1.7 mV (n=10); and SITS hyperpolarized PDbl by 7.8±0.9 mV (n=4). Stimulation of the ducts with dbcAMP in the presence of bath HCO 3 − /CO2 resulted in depolarization of PDbl, the ductal lumen became more negative and the fractional resistance of the luminal membrane decreased. Together with forskolin (10−6 M), db-cAMP (10−4 M) caused a fast depolarization of PDbl by 33.8±2.5 mV (n=6). When db-cAMP (5×10−4 M) was given alone in the presence of bath HCO 3 − /CO2, PDbl depolarized by 25.3±4.2 mV (n=10). In the absence of exogenous HCO 3 − , db-cAMP also depolarized PDbl by 24.7±3.0 mV (n=10). The present data suggest that in the luminal membrane of pancreatic duct cells there is a Cl− conductance in parallel with a Cl−/HCO 3 − antiport. Dibutyryl cyclic AMP increases the Cl− conductance of the luminal membrane. Taking together our present results, and the recent data obtained for the basolateral membrane [21], a tentative model for pancreatic HCO 3 − transport is proposed.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 421 (1992), S. 224-229 
    ISSN: 1432-2013
    Keywords: Cl− conductance ; HT29 ; P2 receptor ; Colon ; Cl− secretion ; cAMP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The colonic carcinoma cell line HT29 was used to examine the influence of agonists increasing cytosolic cAMP and Ca2+ activity on the conductances and the cell membrane voltage (V m). HT29 cells were grown on glass cover-slips. Cells were impaled by microelectrodes 4–10 days after seeding, when they had formed large plaques. In 181 impalements V m was −51±1 mV. An increase in bath K+ concentration from 3.6 mmol/l to 18.6 mmol/l or 0.5 mmol/l Ba2+ depolarized the cells by 10±1 mV (n=49) or by 9±2 mV (n=3), respectively. A decrease of bath Cl− concentration from 145 to 30 mmol/l depolarized the cells by 11±1 mV (n=24). Agents increasing intracellular cAMP such as isobutylmethylxanthine (0.1 mmol/l), forskolin (10 μmol/l) or isoprenaline (10 μmol/l) depolarized the cells by 6±1 (n=13), 15±3 (n=5) and 6±2 (n=3) mV, respectively. In hypoosmolar solutions (225 mosmol/l) cells depolarized by 9±1 mV (n=6). Purine and pyrimidine nucleotides depolarized the cells dose-dependently with the following potency sequence: UTP 〉 ATP 〉 ITP 〉 GTP 〉 TIP 〉 CTP = 0. The depolarization by ATP was stronger than that by ADP and adenosine. The muscarinic agonist carbachol led to a sustained depolarization by 27±6 mV (n=5) at 0.1 mmol/l, and to a transient depolarization by 12±4 mV (n=5) at 10 μmol/l. Neurotensin depolarized with a half-maximal effect at around 5 nmol/l. The depolarization induced by nucleotides and neurotensin was transient and followed by a hyperpolarization. We confirm that HT29 cells possess Cl−- and K+-conductive pathways. The Cl− conductance is regulated by intracellular cAMP level, cytosolic Ca2+ activity, and cell swelling. The K+ conductance in HT29 cells is regulated by intracellular Ca2+ activity.
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  • 7
    ISSN: 1432-2013
    Keywords: Vascular smooth muscle cell ; K+ conductance ; Big Ca2+-dependent K+ channel ; Patch clamp ; Verapamil ; Protein kinase C
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Vascular smooth muscle cells were obtained from rabbit aorta and were studied in primary culture on days 1–7 after seeding with electrophysiological techniques. In impalement experiments a mean membrane potential difference (PD) of −50±0.3 mV (n=387) was obtained with Ringer-type solution in the bath. PD was depolarized by 6±0.3 mV (n=45) and 16±2 mV (n= 5) when the bath K+ concentration was increased from the control value of 3.6 mmol/l to 13.6 and 23.6 mmol/l, respectively. Ba2+ (0.1–1 mmol/l) depolarized PD. Tetraethylammonium (TEA, 10 mmol/l) depolarized PD only slightly but significantly. Verapamil (0.1 mmol/l) and charybdotoxin (10 nmol/l) had no effect on PD. The conductance properties of these cells were further examined with the patch-clamp technique. K+ channels were spontaneously present in cell-attached patches. When the pipette was filled with 145 mmol/l KCl, a mean conductance (g K) of 209.6±4.6 mV (n=17) was read from the current/voltage curves at a clamp voltage (V c) of 0 mV. After excision K+ channels were found in 129 patches with inside-out and in 50 with outside-out configuration. With KCl on one and NaCl on the other side the mean g K at a V c of 0 mV was 134.6±3.9 pS (n=179). The mean permeability was 0.89±0.03×10−12 cm3/s. With symmetrical KCl solution the mean g K was 227±6 pS (n=17). The conductance sequence was g K≫ g Rb= g Cs=g Na=0. TEA blocked dose-dependently only from the outside.(1–10 mmol/l). Lidocaine (5 mmol/l) quinidine (0.01–1 mmol/l) and quinine (0.01–1 mmol/l) blocked from both sides. Charybdotoxin (0.5–5 nmol/l) blocked only from the extracellular side. Ba2+ blocked from the cytosolic side and the inhibition was increased by depolarization and reduced by hyperpolarization. At a V c of 0 mV a half-maximal inhibition (IC50) of 2 μmol/l was obtained. Verapamil and diltiazem blocked from both sides, verapamil with an IC50 of 2 μmol/l and diltiazem with an IC50 of 10 μmol/l. The open probability of this channel was increased by Ca2+ on the cytosolic side at activities 〉 0.1 μmol/l. Half-maximal activation occurred at Ca2+ activities exceeding 1 μmol/l. The present data indicate that the vascular smooth muscle cells of rabbit aorta in primary culture possess a K+ conductance. In excised patches only a maxi K+ channel was detected. This channel has properties different from the macroscopic K+ conductance. Hence, it is likely that the K+ conductance of the intact cell is dominated by yet another and thus far not detected K+ channel.
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  • 8
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 419 (1991), S. 76-83 
    ISSN: 1432-2013
    Keywords: Pancreas ; Ducts ; K+ conductance ; Barium ; Bicarbonate ; Cell membrane resistances ; Secretin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The aim of this study was to investigate the role of the K+ conductance in unstimulated and stimulated pancreatic ducts and to see how it is affected by provision of exogenous HCO 3 − /CO2. For this purpose we have applied electrophysiological techniques to perfused pancreatic ducts, which were dissected from rat pancreas. The basolateral membrane potential PDbl of unstimulated duct cells was between −60mV and −70mV, and the cells had a relatively large K+ conductance in the basolateral membrane as demonstrated by (a) 20–22 mV depolarization of PDbl in response to increase in bath K+ concentration from 5 mmol/l to 20mmol/l and (b) the effect of a K+ channel blocker, Ba2+ (5 mmol/l), which depolarized PDbl by 30–40mV. These effects on unstimulated ducts were relatively independent of bath HCO 3 − /CO2. The luminal membrane seemed to have no significant K+ conductance. Upon stimulation with secretin or dibutyryl cyclic AMP, PDbl depolarized to about −35 mV in the presence of HCO 3 − /CO2. Notably, the K+ conductance in the stimulated ducts was now only apparent in the presence of exogenous HCO 3 − /CO2 in the bath solutions. Upon addition of Ba2+, PDbl depolarized by 13±1 mV (n=7), the fractional resistance of the basolateral membrane, FRbl increased from 0.66 to 0.78 (n=6), the specific transepithelial resistance, R te, increased from 52±13 Ω cm2 to 59±15 Ω cm2 (n=11), and the whole-cell input resistance, R c, measured with double-barrelled electrodes, increased from 20 MΩ to 26 MΩ (n=3). These results are consistent with Ba2+ inhibition of the K+ conductance. Following removal of exogenous HCO 3 − /CO2 in the same ducts, stimulation led to a larger depolarization on PDbl to about −25 mV, and Ba2+ had a smaller effect on PDbl and no significant effect on the resistances. The individual resistances in the duct epithelium were estimated from equivalent circuit analysis. The luminal membrane resistance, R 1 decreased from about 2000 Ω cm2 to 80 Ω cm2 upon stimulation. The basolateral membrane resistance, R bl, remained at 90–120 Ω cm2, and the paracellular shunt resistance, R s, at 50–80 Ω cm2. Ba2+ increased R bl of stimulated ducts to about 200 Ω cm2, an effect present only if the ducts were provided with exogenous HCO 3 − /CO2. Taken together, the present results indicate that the basolateral K+ conductance of pancreatic ducts is sensitive to exogenous HCO 3 − /CO2, i.e. without HCO 3 − /CO2 the conductance becomes very low although the ducts are undergoing stimulation.
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 429 (1995), S. 682-690 
    ISSN: 1432-2013
    Keywords: Cl− conductance ; K+ conductance ; Brefeldin A ; Cytochalasin D ; Epithelial cells ; Actin ; Microtubules
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Activation of Cl− and K+ channels is necessary to drive ion secretion in epithelia. There is substantial evidence from previous reports that vesicular transport and exocytosis are involved in the regulation of ion channels. In the present study we examined the role of cytoskeletal elements and components of intracellular vesicle transport on ion channel activation in bronchial epithelial cells. To this end, cells were incubated with a number of different compounds which interact with either microtubules or actin microfilaments, or which interfere with vesicle transport in the Golgi apparatus. The effectiveness of these agents was verified by fluorescence staining of cellular microtubules and actin. The function was examined in 36Cl− efflux studies as well as in whole-cell (WC) patch-clamp and cell-attached studies. The cells were studied under control conditions and after exposure to (in mmol/l) ATP (0.1), forskolin (0.01), histamine (0.01) and hypotonic bath solution (HBS, NaCl 72.5). In untreated control cells, ATP primarily activated a K+ conductance whilst histamine and forskolin induced mainly a Cl− conductance. HBS activated both K+ and Cl− conductances. Incubation of the cells with brefeldin A (up to 100 μmol/l) did not inhibit WC current activation and 36Cl− efflux. Nocodazole (up to 170 μmol/l) reduced the ATP-induced WC current, and mevastatin (up to 100 μmol/l) the cell-swelling-induced WC current. Neither had any effect on the WC current induced by forskolin and histamine. Also 36Cl− efflux induced by HBS, ATP, forskolin and histamine was unaltered by these compounds. Similarly, colchicine (10 μmol/l) and taxol (6 μmol/l) affected neither 36Cl− efflux nor WC current induced by ATP, forskolin, histamine or HBS. In contrast, depolymerisation of actin by cytochalasin D (10 μmol/l) significantly attenuated 36Cl− effluxes and WC current activation by the above-mentioned agonists. Incubation with a C2 clostridial toxin (5 nmol/l) showed similar effects on WC currents. Moreover, when cytochalasin D (10 μmol/l), C2 clostridial toxins (5 nmol/l), or phalloidin (10 μmol/l) were added to the pipette filling solution current activation was markedly reduced. However, in excised inside-out membrane patches, cytochalasin D (10 μmol/l), G-actin (10 μmol/l) and phalloidin (10 μmol/l) had no effect. These data suggest that actin participates in the activation of ion channels in 16HBE14o- epithelial cells and support the concept that exocytosis is a crucial step in the regulation of Cl− and K+ channels in these cells.
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  • 10
    ISSN: 1432-2013
    Keywords: N-Acetyl-l-cysteine-S-Carboxymethyl-l-cysteine ; Respiratory epithelial cells ; Cystic fibrosis ; CFTR ; Cl− conductance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract N-Acetyl-l-cysteine (NAC) is a widely used mucolytic drug in patients with a variety of respiratory disorders including cystic fibrosis (CF). The beneficial effects of NAC are empirical and the exact mechanism of action in the airways remains obscure. In the present study we examined the effects on whole-cell (we) conductance (G m) and voltage (V m) of NAC and the congeners S-carboxymethyl-l-cysteine (CMC) andS-carbamyl-L-cysteine (CAC) andL-cysteine in normal and CF airway epithelial cells.L-Cysteine (1 mmol/1) had no detectable effect. The increase inG m (ΔGm) by the other compounds was concentration dependent and was (all substances at 1 mmol/1) 3.8 ± 1.4 nS (NAC; n = 11), 4.2 ± 1.0 nS (CMC;n = 16) and 3.8 ± 1.6 nS (CAC;n = 18), respectively. The changes in Gm were paralleled by an increased depolarization (ΔVm) when extracellular Cl− concentration was reduced to 34 mmol/1: under control conditions = -4.1 ± 2.1 versus 10.2 ± 2.1 mV in the presence of NAC, CMC, CAC (n = 36). In the presence of NAC, CMC and CAC, the reduction in Cl− concentration was paralleled by a reduction ofG m by 2.1 ± 0.4 nS (n = 35), indicating that all substances acted by increasing the Cl− conductance. Analysis of intracellular pH did not reveal any changes by any of the compounds (1 mmol/1). A Cl− conductance was also activated in HT29 colonic carcinoma and CF tracheal epithelial (CFDE) cells but not in CFPA1 cells, which do not express detectable levels of ΔF508-CFTR, suggesting that the presence of CFTR may be a prerequisite for the induction of Cl− currents. Next we examined the ion currents in Xenopus oocytes microinjected with CFTR-cRNA. Water-injected oocytes did not respond to activation by forskolin and 3-isobutyl-l-methylxanthine (IBMX) (ΔGm = 0.08 ±0.04 μS;n = 10) and no current was activated when these oocytes were exposed to NAC or CMC. In contrast, in CFTR-cRNA-injected cocytesG m was enhanced when intracellular adenosine 3′,5′-cyclic monophosphate (cAMP) was increased by forskolin and IBMX (G m = 4.5 ± 1.3 μS;n = 8).G m was significantly increased by 0.74 ± 0.2 μS (n = 11) and 0.46 ± 0.1 μS (n = 10) when oocytes were exposed to NAC and CMC, respectively (both I mmol/1). In conclusion, NAC and its congeners activate Cl− conductances in normal and CF airway epithelial cells and hence induce electrolyte secretion which may be beneficial in CF patients. CFTR appears to be required for this response in an as yet unknown fashion.
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