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S Waldegger

Publications and source records attributed to S Waldegger.

At least 55 records · Page 3Linked to original sources
PubMed1996

Expression of a renal type I sodium/phosphate transporter (NaPi-1) induces a conductance in Xenopus oocytes permeable for organic and inorganic anions.

Two distinct molecular types (I and II) of renal proximal tubular brush border Na+/Pi cotransporters have been identified by expression cloning on the basis of their capacity to induce Na+-dependent Pi influx in tracer experiments. Whereas the type II transporters (e.g., NaPi-2 and NaPi-3) resemble well known characteristics of brush border Na+/Pi cotransport, little is known about the properties of the type I transporter (NaPi-1). In contrast to type II, type I transporters produced electrogenic transport only at high extracellular Pi concentrations (> or =3 mM). On the other hand, expression of NaPi-1 induced a Cl- conductance in Xenopus laevis oocytes, which was inhibited by Cl- channel blockers [5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) > niflumic acid >> 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid]. Further, the Cl- conductance was inhibited by the organic anions phenol red, benzylpenicillin (penicillin G), and probenecid. These organic anions induced outwardly directed currents in the absence of Cl-. In tracer studies, we observed uptake of benzylpenicillin with a Km of 0.22 mM; benzylpenicillin uptake was inhibited by NPPB and niflumic acid. These findings suggest that the type I Na+/Pi cotransporter functions also as a novel type of anion channel permeable not only for Cl- but also for organic anions. Such an apical anion channel could serve an important role in the transport of Cl- and the excretion of anionic xenobiotics.

Animals↗
PubMed1996

Blockade of HERG channels expressed in Xenopus oocytes by the histamine receptor antagonists terfenadine and astemizole.

The widely used histamine receptor antagonists terfenadine and astemizole were shown to prolong the QT interval in electrocardiographic recordings in cases of overdose or inappropriate co-medications, indicating a possible interaction with cardiac K+ channels. Here, terfenadine and astemizole both inhibited the human ether-a-go-go related gene (HERG) encoded channels expressed in Xenopus oocytes at nanomolar concentrations in a use- and voltage-dependent fashion. In contrast, inhibition of other delayed rectifier (Kv1.1 and IsK) or inward rectifier K+ channels (IRK1) was much weaker and occurred only at high micromolar concentrations. These results suggest that blockade of HERG channels by terfenadine and astemizole might contribute to the cardiac side effects of these compounds.

Animals↗
PubMed1996

Effect of isosorbiddinitrate on exogenously expressed slowly activating K+ channels and endogenous K+ channels in Xenopus oocytes.

1. The effects of isosorbiddinitrate (ISDN) were tested on membrane currents and resting potential in Xenopus laevis oocytes which were either uninjected or injected with cRNA encoding for K+ channels from three distinct families (slowly activating IsK channels, delayed-rectifying Kv1.1 or inwardly rectifying IRK1 K+ channels). 2. In uninjected oocytes ISDN (1 mM) resulted in a decrease of the holding current at potentials more positive than -100 mV and in an increase at potentials below -100 mV. Increasing extracellular K+ to 100 mM shifted the reversal potential for ISDN-mediated effects to approximately -12 mV, suggesting an inhibition of a K+ conductance by ISDN. 3. In current clamp studies ISDN (1 mM) and Ba2+ (3 mM) depolarized cell membrane. ISDN and Ba2+ had no additive effects on membrane potential when applied simultaneously. In voltage clamp studies, corresponding results were observed for the effects of ISDN and Ba2+ on the holding current with an apparent K(m) of 0.21 and 0.08 mM, respectively. 4. In contrast to ISDN, the nitric oxide (NO) donors isosorbidmononitrate (ISMN) and S-nitrosocysteine (SNOC) had no effects on the holding currents in Xenopus oocytes. Moreover, the guanylate inhibitor LY 83583 did not affect ISDN-mediated holding current alterations, suggesting that ISDN acts independently of the second messenger NO. 5. ISDN inhibited exogenously expressed IsK channels with an apparent K(m) of 0.15 mM, but at 1 mM only weakly inhibited Kv1.1 and IRK1 channels. 6. It is concluded that ISDN inhibits an endogenous K+ conductance in Xenopus oocytes with a similar potency to that shown by expressed IsK channels. These effects are independent of the second messenger NO.

Aminoquinolines↗
PubMed1996

Inhibition of minK protein induced K+ channels in Xenopus oocytes by estrogens.

Previously it was shown that minK protein expression in uterus is regulated by estrogen. In the present study, we were interested in putative direct effects of estrogen on minK protein induced K+ currents (IminK) in Xenopus oocytes. Superfusion with 17-beta-estradiol (1 microM) resulted in an inhibition of minK-induced currents, but had no appreciable effects on the delayed rectifier and inward rectifier K+ channels Kv1.1 and Kir2.1, respectively. The inhibition of IminK by 17-beta-estradiol was concentration-dependent, with an IC50 of approximately 0.5 microM. In the presence of 17-beta-estradiol, the conductance-voltage relationship was shifted to more depolarized potentials. IminK inhibition occurred also in the presence of the estrogen-receptor antagonist tamoxifen, suggesting that a mechanism independent of estrogen receptors is involved. The synthetic estrogen diethylstilbestrol (DES) also inhibited IminK but with a lower affinity (IC50 of 4.5 microM), while cortisol and progesterone had only weak effects on IminK. In summary, the results indicate that estrogens directly inhibit IminK.

Animals↗
PubMed1996

Blockade of epithelial Na+ channels by triamterenes - underlying mechanisms and molecular basis.

The three subunits (alpha, beta, gamma) encoding for the rat epithelial Na+ channel (rENaC) were expressed in Xenopus oocytes, and the induced Na+ conductance was tested for its sensitivity to various triamterene derivatives. Triamterene blocked rENaC in a voltage-dependent manner, and was 100-fold less potent than amiloride at pH 7.5. At -90 mV and -40 mV, the IC50 values were 5 microM and 10 microM, respectively. The blockage by triamterene, which is a weak base with a pKa of 6.2, was dependent on the extracellular pH. The IC50 was 1 microM at pH 6.5 and only 17 microM at pH 8.5, suggesting that the protonated compound is more potent than the unprotonated one. According to a simple kinetic analysis, the apparent inhibition constants at -90 mV were 0.74 microM for the charged and 100.6 microM for the uncharged triamterene. The main metabolite of triamterene, p-hydroxytriamterene sulfuric acid ester, inhibited rENaC with an approximately twofold lower affinity. Derivatives of triamterene, in which the p-position of the phenylmoiety was substituted by acidic or basic residues, inhibited rENaC with IC50 values in the range of 0.1-20 microM. Acidic and basic triamterenes produced a rENaC blockade with a similar voltage and pH dependence as the parent compound, suggesting that the pteridinemoiety of triamterene is responsible for that characteristic. Expression of the rENaC alpha-subunit-deletion mutant, Delta278-283, which lacks a putative amiloride-binding site, induced a Na+ channel with a greatly reduced affinity for both triamterene and amiloride. In summary, rENaC is a molecular target for triamterene that binds to its binding site within the electrical field, preferably as a positively charged molecule in a voltage- and pH-dependent fashion. We propose that amiloride and triamterene bind to rENaC using very similar mechanisms.

Amiloride↗
PubMed1996

Inhibition of IKs in guinea pig cardiac myocytes and guinea pig IsK channels by the chromanol 293B.

The chromanol derivative 293B was previously shown to inhibit a cAMP regulated K+ conductance in rat colon crypts. Subsequent studies on cloned K+ channels from the rat demonstrated that 293B blocks specifically IsK channels expressed in Xenopus oocytes, but does not affect the delayed and inward rectifier Kv1.1 and Kir2.1, respectively. In the present study, the specificity of 293B for the cardiac K+ conductances IKs and IKr, and for the cloned guinea pig IsK channel and the human HERG channel, which underly IKs and IKr, respectively, was analyzed. 293B inhibited both the slowly activating K+ conductance IKs in cardiac myocytes and guinea pig IsK channels expressed in Xenopus oocytes with a similar IC50 (2-6 micromol/l). In contrast, high concentrations of 293B had only a negligible effect on the more rapid activating IKr. Similarly, 293B exerted no effect on HERG channels expressed in Xenopus oocytes. In summary, 293B appears to be a rather specific inhibitor of IKs and the underlying IsK channels.

Animals↗
PubMed1996

Coexpression and stimulation of parathyroid hormone receptor positively regulates slowly activating IsK channels expressed in Xenopus oocytes.

Expression of the IsK protein in Xenopus oocytes induced the characteristically slow, voltage-dependent outward currents. Superfusion with the parathyroid hormone (PTH) peptide 1-34 had no effect on IsK when expressed alone, but increased IsK when IsK was coexpressed with the PTH-receptor. PTH receptor stimulation caused a shift of IsK conductance-voltage relationship to more negative potentials, and a decrease of both the rate of IsK activation and deactivation. IsK regulation by PTH was independent of extracellular Ca2+, and was also present IsK protein mutants lacking the protein kinase C consensus site. However, regulation of IsK by PTH was mimicked by activators of protein kinase A (PKA) and greatly reduced in the presence of the kinase inhibitors staurosporine and H89. These results suggest that PTH regulates IsK by a mechanism involving phosphorylation independent of protein kinase C (PKC). Such regulation may play a role in proximal tubule cells of the kidney, where both PTH receptor and the IsK protein are expressed.

Animals↗
PubMed1996

Heavy metals inhibit Pi-induced currents through human brush-border NaPi-3 cotransporter in Xenopus oocytes.

Heavy metal intoxication with Hg2+, Pb2+ and Cd2+ commonly leads to phosphaturia. In this study, we examined the effects of these heavy metals on Pi-induced currents (Ip) through NaPi-3, the human renal cotransporter for Na+ and Pi. Hg2+ inhibited Ip in a dose- and time-dependent fashion. Hg2+ decreased the extrapolated maximal current but did not alter the apparent affinity for Pi. This inhibition was also observed with the membrane-permeable oxidizing agent 2,2'-dithio-bis(5-nitropyridine) (DTNP) but not with the membrane-impermeable 5,5'-dithiobis(2-nitrobenzoic acid). Hg(2+)- and DTNP-mediated inhibition of Ip was reversible only in the presence of the reducing agent 2,3-dihydroxybutane-1,4-dithiol. Cd2+ and Pb2+ also inhibited Ip. However, while CD2+ did not significantly alter the apparent affinity for Pi, the apparent concentration needed for half-maximal current (Km) for Pi was increased by Pb2+. In contrast to Hg2+, the inhibition of Ip by Cd2+ and Pb2+ was rapidly reversible upon washout. In the presence of the Na(+)-K(+)-adenosinetriphosphatase inhibitor ouabain, Ip was not reduced, and the effects of the heavy metals were maintained. In summary, the three heavy metals Hg2+, Cd2+, and Pb2+ inhibit Ip through the Na+/Pi cotransporter NaPi-3 by distinct mechanisms. Heavy metal-mediated inhibition of NaPi-3 may be responsible for the phosphaturia observed after intoxication with these compounds.

2,2'-Dipyridyl↗
PubMed1996

Function and dysfunction of renal transport molecules: lessons from electrophysiology.

The availability of cloned transport molecules achieved by efforts in expression cloning has allowed their electrophysiological analysis in the Xenopus oocyte expression system. We describe the electrogenic uptake of various substrates by their corresponding transport molecules originally expressed in brush border membranes of proximal tubules. The functional properties of the following transport molecules are discussed: the sodium-coupled glucose transporter, the sodium-coupled phosphate transporter, the sodium-coupled sulfate transporter and the sodium-independent transporter of neutral and dibasic amino acids. Additionally, functional consequences of naturally occurring disease-causing mutations in some of these transport molecules are described.

Animals↗
PubMed1995

Effect of channel modulation and pH on IsK inhibition by the novel class III antiarrhythmic azimilide (NE-10064).

Inhibition of human IsK channels expressed in Xenopus oocytes by the novel class III antiarrhythmic azimilide was studied under distinct treatments known to increase IsK (hypotonic solution, A23187 and isoproterenol) Azimilide inhibited IsK under all conditions with similar potency. Reduction of ionic strength or pH changes from pH 6.5 to 8.5 did not alter IsK amplitude. However, inhibition of IsK by azimilide was decreased by reduced pH, but not by reduced ionic strength. Further, the apparent affinity of azimilide was increased more than tenfold by increasing pH from 6.5 to 8.5. The data suggest that the neutral form of azimilide, a weak base, inhibits IsK via a lipophilic protein-drug interaction. pH-dependence of azimilide may significantly alter its effects on IsK under distinct pathophysiological conditions (acidosis vs. alkalosis) and in distinct locations (heart vs. kidney).

Animals↗
PubMed1995

Electrophysiological effects of progesterone on hepatocytes.

The addition of progesterone (1-100 mumol/l) to the extracellular fluid bathing rat hepatocytes led to a rapid and fully reversible depolarization of the cell membrane. The progesterone-induced depolarization was paralleled by a decrease of potassium selectivity and an increase of cell membrane resistance and was abolished in the presence of the potassium channel blocker barium. Accordingly, in whole cell recordings, progesterone led to a decrease of the cell membrane conductance. 17 alpha-Hydroxyprogesterone and beta-estradiol were less effective by a factor of 10, whereas cholesterol, corticosterone and hydrocortisone did not significantly alter the potential difference across the cell membrane. In conclusion, acute administration of progesterone depolarized rat hepatocytes by decreasing the potassium conductance of the cell membrane.

Animals↗
PubMed1995

Molecular basis of IsK protein regulation by oxidation or chelation.

Slowly activating IsK channels were expressed in Xenopus oocytes and exposed to oxidative agents. Oxidative treatment reduced the resulting current IsK, while no inhibition was observed for IsK protein mutants carrying a Ser mutation instead of a highly conserved Cys residue in the intracellular domain. In contrast, Hg2+, which may not only oxidize thiol groups but also form chelates with dibasic amino acids, caused a use-dependent, positive regulation of IsK. This effect was reversed in an IsK protein mutant with a deletion in the extracellular domain. These data suggest opposite effects of peroxides and Hg2+ on IsK, a peroxide-mediated IsK inhibition by intracellular oxidation and a Hg(2+)-mediated IsK increase, caused by extracellular Hg2+ chelation of the IsK protein.

Animals↗
PubMed1995

The nitroso-donor S-nitroso-cysteine regulates IsK expressed in Xenopus oocytes via a c-GMP independent mechanism.

In Xenopus oocytes expressing slowly activating IsK channels superfusion with the nitroso-donor S-Nitroso-Cysteine (SNOC) resulted in an increase of IsK, which was greatly enhanced when the amino acid-exchanger rBAT was coexpressed. The effects of SNOC on IsK could not be prevented by the guanylate cyclase inhibitor LY-83,583 and the cGMP kinase inhibitor H8, but was abolished in the presence of staurosporine. SNOC also increased the currents induced by the expression of protein mutants lacking intracellular sites, previously described to be involved in IsK regulation by oxidation and phosphorylation. These data suggest that the NO-donor SNOC regulates IsK indirectly via a cGMP independent, but staurosporine sensitive, pathway.

Alkaloids↗
PubMed1995

Heavy metal mediated inhibition of rBAT-induced amino acid transport.

rBAT, a protein which is located in the brush border membranes of intestine and renal proximal tubule cells, was recently shown to induce electrogenic countertransport of neutral and dibasic amino acids after its expression in Xenopus oocytes. Here, we studied the effects of heavy metals on rBAT induced amino acid transport in Xenopus oocytes to clarify a possible involvement of rBAT in heavy metal-induced aminoaciduria. The heavy metals Hg2+ and Pb2+ inhibited rBAT-induced amino acid transport with a different profile of action. The Pb2+ mediated inhibition occurred rapidly upon superfusion and was readily reversible upon washout. The maximal inhibition caused by Pb2+ was about 50% of the amino acid-induced currents at an apparent affinity (Km) of about 10 microM. In contrast, the Hg(2+)-mediated inhibition occurred rather slowly, depending on its concentration, and was not reversible during washout with control solution. However, the Hg(2+)-mediated amino acid transport inhibition could be reversed with Hg2+ chelating agents and reducing compounds. Other oxidative agents, such as the membrane permeable 2,2'-Dithio-bis(5-Nitropyridine) (DTNP), but not the membrane impermeable 5,5'-Dithio-bis (2-Nitrobenzoic acid) (DTNB), mimicked the effect of Hg2+, and their effect could similarly be reversed with 2,3-Dihydroxybutane-1,4-dithiol (DTE). In conclusion, Pb2+ and Hg2+ inhibit rBAT-induced amino acid transport in a noncompetitive, allosteric fashion. Blockade of rBAT-induced amino acid transport may be involved in aminoaciduria following mercury or lead intoxication.

Amino Acid Transport Systems, Basic↗
PubMed1995

Properties of electrogenic Pi transport by a human renal brush border Na+/Pi transporter.

Inorganic phosphate (Pi) induced an inward current (IP) in Xenopus oocytes expressing the human renal Na+/Pi cotransporter NaPi-3. At 100mM Na+, Pi-transport was independent of the holding potential and resulted in an apparent Km of 0.08 mM; lowering the Na+ concentration to 50 mM resulted in an increase of the apparent Km to 0.22 mM at -50 mV and to 0.31 mM at -90 mV. In contrast, the apparent Km for Na+ was not significantly influenced by the holding potential. A decrease of the pH from 7.8 to 6.8 resulted in a decrease of IP at 50 mM Na+, but not at 150 mM Na+. Arsenate induced inward currents through NaPi-3 and decreased the apparent Km in measurements of IP. Phosphonoformic acid itself induced no currents, but inhibited Pi-induced currents with an apparent Ki of 3.6 mM. In summary, NaPi-3 displays characteristic Na+/Pi cotransporter properties with relevant interactions with arsenate (transport substrate) and phosphonoformic acid (inhibitor). Monovalent and divalent Pi both appear to be transported by NaPi-3.

Animals↗
PubMed1994

Opposite directed currents induced by the transport of dibasic and neutral amino acids in Xenopus oocytes expressing the protein rBAT.

Voltage- and current-clamp studies have been performed on a renal and intestinal protein (rBAT) which induces transport for neutral and dibasic amino acids when expressed in Xenopus oocytes. In current-clamp mode, superfusion with L-leucine caused a hyperpolarization while superfusion with L-arginine depolarized the oocyte. Accordingly, in voltage-clamp experiments dibasic amino acids and neutral amino acids induced inward and outward currents, respectively. The relationship between currents and substrate concentrations could be fitted by simple Michaelis-Menten kinetics. Currents induced by L-arginine and L-leucine were also voltage-dependent. pH changes from 6.25 to 8.75 did not affect the currents induced by saturating concentrations of L-arginine and L-leucine, but reversed the direction of L-histidine-induced currents from inward to outward. The reversal potentials as well as the apparent Km for L-histidine-induced currents were altered by the ambient pH. Currents induced by individual amino acids decreased during extended superfusion periods. However, extended superfusion with neutral amino acids increased dibasic amino acid induced currents, while prior superfusion with dibasic amino acid resulted in an increase of currents induced by neutral amino acids. The reversal potentials for L-leucine- and L-arginine-induced currents were depending on their intra- (after preloading) and extracellular concentrations. In conclusion, rBAT-mediated transport of neutral and dibasic amino acids is associated with net outward or inward currents, respectively, which may be caused by an exchange of neutral with dibasic amino acids.

Amino Acid Transport Systems, Basic↗
PubMed1994

Blockade of human IsK channels expressed in Xenopus oocytes by the novel class III antiarrhythmic NE-10064.

cRNA encoding the human IsK protein was injected into Xenopus oocytes and the expressed channels were investigated using the two-microelectrode voltage-clamp method. The novel class III antiarrhythmic NE-10064 (1-[[[5-(4-chlorophenyl)-2-furanyl]methylene]-amino]-3- [4-(4-methyl-1-piperazinyl)-butyl]-2,4-imidazolidinedione dihydrochloride) was tested for its ability to block these channels. The compound displayed potent inhibitory effects with an EC50 of 5.4 microM. The block caused by NE-10064 was use-dependent, i.e. channels had to be activated for the inhibition to occur. Further, the reversal of the inhibition during the wash-out period was use-dependent. Finally, the blockade of human IsK channels by NE-10064 appeared to be voltage-dependent, being more pronounced at depolarized potentials. We conclude that this novel class III antiarrhythmic is a potent inhibitor of human IsK channels and suggest that such effects could be involved in its antiarrhythmic action.

Animals↗