Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Flufenamic Acid”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 469 records · Page 26Linked to original sources
PubMed1987

Fluorescence energy transfer study of the relationship between the lone tryptophan residue and drug binding sites in human serum albumin.

The relationship between the lone tryptophan residue at position 214 and drug binding sites (Sites I and II) in human serum albumin (HSA) was studied by fluorescence energy transfer. The distance between the lone tryptophan residue and ligands bound to HSA was estimated by Förster's equation, taking into consideration the degree of ligand binding at these sites, as determined from binding parameters (binding constant, k, and the number of binding sites, n). For all ligands investigated, the distance in each case appeared to asymptotically decrease when the occupation ratio of the binding sites increased with ligand concentration. When the primary binding site of each ligand in HSA was almost saturated, the distance attained a constant value, making possible a somewhat more exact determination of the distance. The distance ranged from approximately 22 to 23 A for ligands typical of Site I (warfarin, dansylamide, dansylglutamine), and approximately 16.1 to 17.5 A for ligands typical of Site II (dansylsarcosine, dansylproline, dansylglycine, diazepam, flufenamic acid).

Amino Acids↗
PubMed1995

Plasma concentrations and pharmacokinetic parameters of nitrofenac using a simple and sensitive HPLC method.

An accurate and sensitive HPLC method has been developed for the determination of nitrofenac, a new, original diclofenac derivate showing a good tolerability and a wide anti-inflammatory profile, diclofenac, and its metabolites in plasma. This method has been applied to evaluate the pharmacokinetic parameters of the drugs, using a noncompartmental model, after the oral administration of 5 mg/kg nitrofenac to rats. Nitrofenac and the internal standard flufenamic acid were dissolved in acetonitrile, and diclofenac was dissolved in methanol. The drugs were eluted from a 5 microns LC-8 column with a mobile phase consisting of acetonitrile/water (50/50 v/v) adjusted to pH 3.3 with glacial acetic acid, at a flow rate of 2 mL/min with UV detection at 280 nm for diclofenac and 275 nm for nitrofenac. The detection limit for the drugs in plasma was 25 ng/mL. The peak concentration of nitrofenac was reached 7 h after drug administration, while with diclofenac we observed three peaks at 2, 5, and 10 h; the mean residence time and the elimination rate constant for nitrofenac were 6.18 +/- 0.09 h and 0.37 +/- 0.03 h-1 respectively, while those for diclofenac were 12.24 +/- 0.11 h and 0.11 +/- 0.04 h-1. Under our conditions, the metabolism of nitrofenac produced 23% diclofenac and other metabolites: the plasma concentrations and kinetic characteristics of diclofenac are enough to induce an anti-inflammatory activity, while the clinical importance of the other metabolites remains to be elucidated.

Animals↗
PubMed2000

Chloride secretion in kidney distal epithelial cells (A6) evoked by cadmium.

The effect of Cd(2+) on chloride secretion was examined in A6 renal epithelia cells by chloride-sensitive fluorescence (SPQ probe) and by the short-circuit-current (I(sc)) technique. Depleting the cells of Cl(-) suggests that the Cd(2+)-activated I(sc) (DeltaI(sc(Cd))) is dependent on the presence of Cl(-) ions. Among the Cl(-)-channel inhibitors the fenemates, flufenamic acid (FFA) and niflumic acid (NFA), and 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) significantly lowered DeltaI(sc(Cd)) compared with control level. In SPQ-loaded A6 cells, Cd(2+) evoked an increase in Cl(-) secretion ([DeltaCl(-)](Cd)), which significantly exceeded the basal Cl(-) transport and was blockable by FFA and NFA. The closely related metals, Zn(2+) or Ni(2+), were also able to activate Cl(-) secretion. Preexposure of Zn(2+) or Ni(2+) completely prevented [DeltaCl(-)](Cd), suggesting that Zn(2+) and Ni(2+) probably use similar mechanisms. Like Cd(2+), thapsigargin (TG), an inhibitor of intracellular Ca(2+)-ATPase and the Ca(2+)-ionophore A23187, induced an increase in I(sc). Moreover, TG and Cd(2+) were able to neutralize the responses of the counterparts as also observed in I(sc) measurements, which indicates that Cd(2+) activates Cl(-) secretion in a Ca(2+)-dependent manner. Hence, this study supports the idea that basolateral Cd(2+) (possibly also Zn(2+) and Ni(2+)), probably through a Ca(2+)-sensing receptor, causes calcium mobilization that activates apical fenemate-sensitive chloride channels leading to chloride secretion in A6 cells.

Algorithms↗
PubMed1993

The role of a PDGF-activated nonselective cation channel in the proliferative response.

Murine fibroblasts have a 28 pS calcium- and voltage-insensitive NSC that becomes quiescent at G0 arrest and is rapidly and specifically activated by PDGF. Activation is produced by the discrete loss of long channel closures. The NSC can be rapidly and reversibly blocked with the NSAID flufenamic acid, through a prostaglandin-independent mechanism. The cell cycle (not viability) is blocked concomitantly with NSC block. A somatic cell mutant with altered NSC conductance has been isolated and used to clone the genomic locus of the channel. The mutant growth phenotype adds further support to the participation of NSC conductance in cell cycle control.

Animals↗
PubMed2007

TRPM2.

TRPM2 is a cation channel enabling influx of Na+ and Ca2+, leading to depolarization and increases in the cytosolic Ca2+ concentration ([Ca2+]i). It is widely expressed, e.g. in many neurons, blood cells and the endocrine pancreas. Channel gating is induced by ADP-ribose (ADPR) that binds to a Nudix box motif in the cytosolic C-terminus of the channel. Endogenous ADPR concentrations in leucocytes are sufficiently high to activate TRPM2 in the presence of an increased [Ca2+]i but probably not at resting [Ca2+]i. Another channel activator is oxidative stress, especially hydrogen peroxide (H2O2) that may act through ADPR after ADPR polymers have been formed by poly(ADP-ribose) polymerases (PARPs) and hydolysed by glycohydrolases. H2O2-stimulated TRPM2 channels essentially contribute to insulin secretion in pancreatic beta-cells and alloxan-induced diabetes mellitus. Inhibition of TRPM2 channels may be achieved by channel blockers such as flufenamic acid or the anti-fungal agents clotrimazole or econazole. Selective blockers of TRPM2 are not yet available; those would be valuable for a characterization of biological roles of TRPM2 in various tissues and as potential drugs directed against oxidative cell damage, reperfusion injury or leucocyte activation. Activation of TRPM2 may be prevented by anti-oxidants, PARP inhibitors and glycohydrolase inhibitors. In future, binding of ADPR to the Nudix box may be targeted. In light of the wide-spread expression and growing list of cellular functions of TRPM2, useful therapeutic applications are expected for future drugs that block TRPM2 channels or inhibit their activation.

Animals↗
PubMed1995

A calcium-activated and nucleotide-sensitive nonselective cation channel in M-1 mouse cortical collecting duct cells.

We recently reported that M-1 mouse cortical collecting duct cells show nonselective cation (NSC) channel activity (Proc. Natl. Acad. Sci. USA 89:10262-10266, 1992). In this study, we further characterize the M-1 NSC channel using single-channel current recordings in excised inside-out patches. The M-1 NSC channel does not discriminate between Na+, K+, Rb+, Cs+, and Li+. It has a linear I-V relation with a conductance of 22.7 +/- 0.5 pS (n = 78) at room temperature. The Pcation/P(anion) ratio is about 60 and there is no measurable conductance for NMDG, Ca2+, Ba2+, and Mn2+. Cytoplasmic calcium activates the M-1 NSC channel at a threshold of 10(-6) M and depolarization increases channel activity (NPo). Cytoplasmic application of adenine nucleotides inhibits the M-1 NSC channel. At doses of 10(-4) M and 10(-3) M, ATP reduces NPo by 23% and 69%, respectively. Furthermore, since ADP (10(-3) M) reduces NPo by 93%, the inhibitory effect of adenine nucleotides is not dependent on the presence of a gamma-phosphoryl group and therefore does not involve protein phosphorylation. The channel is not significantly affected by 8-Br-cGMP (10(-4) M) or by cGMP-dependent protein kinase (10(-7) M) in the presence of 8-Br-cGMP (10(-5) M) and ATP (10(-4) M). The NSC channel is not sensitive to amiloride (10(-4) M cytoplasmic and/or extracellular) but flufenamic acid (10(-4) M) produces a voltage-dependent block, reducing NPo by 35% at depolarizing voltages and by 80% at hyperpolarizing voltages. We conclude that the NCS channel of M-1 mouse cortical collecting duct cells belongs to an emerging family of calcium-activated and nucleotide-sensitive nonselective cation channels. It does not contribute to amiloride-sensitive sodium absorption and is unlikely to be a major route for calcium entry. The channel is normally quiescent but may be activated under special physiological conditions, e.g., during volume regulation.

Animals↗
PubMed1993

Effects of forskolin on crypt cells of rat distal colon. Activation of nonselective cation channels in the crypt base and of a chloride conductance pathway in other parts of the crypt.

We recently showed that prostaglandin E2 (PGE2) causes depolarization in cells at the base of isolated crypts from rat distal colon by activating nonselective cation channels. In order to investigate whether PGE2 acts via intracellular cyclic adenosine monophosphate (cAMP), the effect of forskolin on cell potential and on whole-cell current was investigated using the slow whole-cell patch-clamp method with nystatin. In addition, effects of forskolin in cells at other sites along the crypt were investigated. At the crypt base, the unstimulated cells had a resting potential of -70.6 +/- 1.3 mV (n = 25). When forskolin was added to the bath, the cells depolarized to -21.1 +/- 1.5 mV (n = 25). This depolarization was inhibited by substitution of all Na+ in the bath solution by N-methyl-D-glucamine (NMDG+) or by addition of flufenamic acid (50 mumol/l), a blocker of nonselective cation channels, to the bath. In contrast, the Cl- channel blocker 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB, 50 mumol/l) did not affect the depolarization. Moving along the crypt, the resting potential was -66.8 +/- 1.8 mV (n = 11) in the mid-crypt and -48.1 +/- 2.9 mV (n = 9) in cells of the upper part of the crypt. Forskolin caused a strong depolarization to about -20 mV in all parts of the crypt. In contrast to cells at the base, this depolarization was only partly diminished by substitution of Na+ by NMDG+, whereas substitution of bath Cl- by gluconate caused an initial further depolarization, followed by a repolarization to the cell's resting potential.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

A calcium-activated nonselective cationic channel in the basolateral membrane of outer hair cells of the guinea-pig cochlea.

The patch-clamp technique was used to investigate ion channels in the basolateral perilymph-facing membrane of freshly isolated outer hair cells (OHCs) from the guinea-pig cochlea. These sensory cells probably determine, via their motile activity, the fine tuning of sound frequencies and the high sensitivity of the inner ear. A Ca(2+)-activated nonselective cationic channel was found in excised inside-out membrane patches. The current/voltage relationship was linear with a unit conductance of 26.3 +/- 0.3 pS (n = 15) under symmetrical inger conditions. The channel excluded anions (PNa/PCl = 18 where PNa/PCl denotes the relative permeability of Na to Cl); it was equally permeant to the Na+ and K+ ions and exhibited a low permeability to N-methyl-D-glucamine and Ba2+ or Ca2+. Channel opening required a free Ca2+ concentration of about 10(-6) mol/l on the internal side of the membrane and the open probability (Po) was maximal at 10(-3) mol/l (Po = 0.72 +/- 0.06, n = 12). Adenosine 5'mono-, tri- and di-phosphate reduced Po to 29 +/- 14 (n = 5), 42 +/- 10 (n = 8) and 51 +/- 12 (n = 5) % of control Po, respectively, when they were added at a concentration of 10(-3) mol/l to the internal side. The channel was partially blocked by flufenamic acid (10(-4) mol/l) and 3',5'-dichlorodiphenylamine-2-carboxylic acid (DCDPC, 10(-5) mol/l). This type of channel, together with Ca(2+)-activated K+ channels, might participate in the control of membrane potential and modulate the motility of OHCs.

Adenine Nucleotides↗
PubMed1982

Relationship of gastric mucosal damage induced in pigs by antiinflammatory drugs to their effects on prostaglandin production.

Experiments were performed in pigs to examine the relationship between the effects of various nonsteroid antiinflammatory drugs on gastric (fundic) mucosal content of prostaglandin (PG)E2 and 6-keto-PGF1 alpha, and the development of damage to the fundic mucosa under acute and chronic dosage conditions. Oral administration of a single dose of indomethacin (5 mg/kg) caused an almost immediate reduction in mucosal potential difference, followed at 5-15 min by ultrastructurally observed damage to mucosal capillaries, mucous, and parietal cells; efflux of Na+, K+, and Cl- ions into the gastric lumen with an apparent loss of luminal H+ ions; and a statistically significant reduction (from 10-60 min) in fundic prostaglandin content. Thus, under acute dosage conditions, development of mucosal damage by indomethacin was paralleled by the reduction in prostaglandin production. Repeated oral dosage of aspirin, indomethacin, sulindac, or diclofenac to pigs for 10 days significantly reduced gastric mucosal as well as plasma prostaglandin levels, coincident with the development of severe gastric mucosal damage. The relatively less irritant drugs, flufenamic acid, azapropazone, and fenclofenac, failed to significantly decrease gastric mucosal content of prostaglandins, although these drugs have been reported to inhibit prostaglandin synthesis in vitro and also were found to reduce the prostaglandin plasma levels in animals receiving these drugs. Another drug with low irritancy, meseclazone, markedly decreased both mucosal and plasma levels of prostaglandins. The results show that while ulcerogenic drugs reduce the mucosal and plasma prostaglandins levels in parallel with their ulcerogenicity, this relationship does not always hold for drugs with low ulcerogenic activity.

Animals↗
PubMed1979

Comparative effects of drugs on four paw oedema models in the rat.

The development of novel anti-inflammatory drugs (AID) has been claimed to be dependent on the discovery of models of inflammation that differ from those currently used for drug screening, e.g. carrageenen paw oedema and u.v. erythema. We have thus evaluated the effect of a variety of drugs in a number of novel models of inflammation in the rat produced in the hind paw. We have utilized kaolin, zymosan, anti-rat IgG (anti-IgG) and the Reversed Passive Arthus (RPA) reaction to produce these oedema models. We found that the non-steroidal AID's, e.g. aspirin, flufenamic acid, indomethacin, naproxen, and phenylbutazone, were active in all four tests. Of the nine novel AID examined, levamisole and tetramisole demonstrated considerable activity in all four tests and dapsone was especially active in the anti-IgG and RPA tests. In contrast, the anti-rheumatic d-penicillamine was inactive in all four models. Each of the ten compounds tested which has been claimed to influence complement function, was active in the RPA but not in the kaolin model. These results are discussed in the context of the aetiology of each oedema and the suspected mode of action of the various drugs.

Animals↗
PubMed1978

Effect of non-steroidal anti-inflammatory drugs of Moloney sarcoma virus inoculated mice.

Chronic administration of phenylbutazone, flufenamic acid and a new, potent non-steroidal anti-inflammatory agent ITF (3-methyl-5-benzoyl-amino-isothiazole-4-carboxy-p-ethoxyphenylamine) to BALB/c mice inoculated with a Moloney sarcoma virus resulted in a stimulation of tumor growth and increased severity of disease. This treatment, however, had no effect on the spontaneous regression of tumors. Indomethacin in a dose of 5.0 or 2.5 mg per kg suppressed the MSV-induced tumor growth, but this effect was apparently connected with the high toxicity of this drug for mice.

Animals↗
PubMed1990

Diphenylamine-2-carboxylate (DPC) inhibits both Cl- conductance and cyclooxygenase of canine tracheal epithelium.

Diphenylamine-2-carboxylate (DPC) decreases Cl- conductance (GCl) in epithelia and cells of several tissues, an effect which has been ascribed to blockade of conductive Cl- channels. However, one DPC derivative, flufenamic acid, is a clinically useful non-steroidal anti-inflammatory agent, the mechanism of action of which involves the blockade of arachidonic acid metabolism by cyclooxygenase. Because GCl in canine tracheal epithelium is stimulated by exogenous prostaglandins and induction of cyclooxygenase activity, we tested the hypothesis that DPC inhibits dog tracheal epithelium GCl through inhibition of cyclooxygenase. DPC inhibited the short circuit current of amiloride-pretreated tissues by 50% at 0.138 mmol/l and by more than 95% at 3 mmol/l. Isoproterenol reversed the inhibition seen at 0.1 mmol/l DPC and stimulated current above control (indomethacin-pretreated) levels. Higher concentrations of DPC diminished the stimulation of current by subsequent exposure to isoproterenol, such that there was little effect of isoproterenol in the presence of 3 mmol/l DPC. DPC, 0.1 mmol/l, also blocked stimulation of current by exogenous arachidonic acid, but not of exogenous prostaglandins PGE or PGD. The metabolism of 3H-arachidonic acid to 3H-PGD2, monitored by HPLC, was completely blocked by 0.1 mmol/l DPC. We conclude that the isoproterenol/prostaglandin reversible blockade of GCl by DPC can be attributed to inhibition of arachidonic acid metabolism.

Amiloride↗
PubMed2004

Mechanism of apoptosis induced by diazoxide, a K+ channel opener, in HepG2 human hepatoma cells.

The effect of diazoxide, a K+ channel opener, on apoptotic cell death was investigated in HepG2 human hepatoblastoma cells. Diazoxide induced apoptosis in a dose-dependent manner and this was evaluated by flow cytometric assays of annexin-V binding and hypodiploid nuclei stained with propidium iodide. Diazoxide did not alter intracellular K+ concentration, and various inhibitors of K+ channels had no influence on the diazoxide-induced apoptosis; this implies that K+ channels activated by diazoxide may be absent in the HepG2 cells. However, diazoxide induced a rapid and sustained increase in intracellular Ca(2+) concentration, and this was completely inhibited by the extracellular Ca(2+) chelation with EGTA, but not by blockers of intracellular Ca(2+) release (dantrolene and TMB-8). This result indicated that the diazoxide-induced increase of intracellular Ca(2+) might be due to the activation of a Ca(2+) influx pathway. Diazoxide-induced Ca(2+) influx was not significantly inhibited by either voltage-operative Ca(2+) channel blockers (nifedipine or verapamil), or by inhibitors of Na+, Ca(2+)-exchanger (bepridil and benzamil), but it was inhibited by flufenamic acid (FA), a Ca(2+)-permeable nonselective cation channel blocker. A quantitative analysis of apoptosis by flow cytometry revealed that a treatment with either FA or BAPTA, an intracellular Ca(2+) chelator, significantly inhibited the diazoxide-induced apoptosis. Taken together, these results suggest that the observed diazoxide-induced apoptosis in the HepG2 cells may result from a Ca(2+) influx through the activation of Ca(2+)-permeable non-selective cation channels. These results are very significant, and they lead us to further suggest that diazoxide may be valuable for the therapeutic intervention of human hepatomas.

Apoptosis↗
PubMed1998

Maitotoxin (MTX) activates a nonselective cation channel in Xenopus laevis oocytes.

Maitotoxin (MTX) may exert its toxic effect by activating ion conductances and has been shown to elicit a fertilization-like response in Xenopus laevis oocytes. In the present study we investigated the electrophysiological response of stage V-VI Xenopus oocytes to MTX using the two-microelectrode voltage-clamp technique. Membrane voltage (Vm) measurements demonstrated that MTX (50 pM to 1 nM) depolarized the oocytes from -49+/-7 to -14+/-1 mV. Subsequent replacement of bath Na+ by the impermeant cation NMDG (N-methyl-d-glucamine) shifted Vm from -14+/-1 to -53+/-5 mV (n=29). This indicates that MTX activates a cation conductance. Indeed, current measurements at a holding potential of -60 or -100 mV showed that within 10 s of MTX application an inward current component developed which was largely abolished by extracellular Na+ removal. After a 1-min application of 1 nM MTX the NMDG-sensitive current increased more than 100-fold from 0.14+/-0.03 microA to a peak value of 21+/-3 microA (n=11). The effect of MTX was concentration dependent with an EC50 of about 250 pM but only slowly reversible. Ion substitution experiments indicated that the stimulated conductance was nonselective for monovalent cations with a slight preference for NH4+ (2.1) > K+ (1.5) > Na+ (1.0) > Li+ (0.7). Regarding divalent cations, a complex biphasic response to extracellular Na+ replacement by Ca2+ was observed, which suggests that the stimulated channels may have a small Ca2+ permeability but that exposure to high extracellular Ca2+ enhances recovery from MTX stimulation. No significant conductance for Mn2+ was observed. Application of 1 mM benzamil, 1 mM amiloride, or 100 microM 1-(beta-[3-(4-Methoxyphenyl)-propoxy]-4-methoxyphenethyl)-1H-imidazol e hydrochloride (SK&F 96365) reduced the MTX-stimulated inward current by 81%, 62%, or 65%, respectively. Gd3+ had an inhibitory effect of 29% and 38% at concentrations of 10 microM or 100 microM, respectively. Flufenamic acid, niflumic acid, (RS)-(3,4-dihydro-6, 7-dimethoxyisoquinoline-1-gamma1)-2-phenyl-N,N-di-[2-(2,3, 4-trimethoxyphenyl)-ethyl]-acetamide (LOE908), and 3', 5'-dichlorodiphenylamine-2-carboxylic acid (DCDPC), known blockers of other nonselective cation channels, had no significant effect. We conclude that MTX activates a nonselective cation conductance in Xenopus oocytes. The underlying channels may be involved in changes in Vm that occur during the early stages of fertilization.

Animals↗
PubMed2004

Inhibition of TRPM2 channels by the antifungal agents clotrimazole and econazole.

TRPM2 is a Ca(2+)-permeable non-selective cation channel that uniquely is activated by intracellular ADP-ribose. To date, only one pharmacological blocker of this channel, namely flufenamic acid (FFA), has been described. Here we demonstrate, using patch clamp electrophysiology, that the antifungal imidazoles clotrimazole and econazole inhibit ADP-ribose-activated currents in HEK-293 cells expressing recombinant human TRPM2 (hTRPM2). For both compounds, all concentrations in a range from 3 microM to 30 microM produced an essentially complete inhibition of the TRPM2-mediated current. The rate of current antagonism was dependent on the concentration applied, with higher concentrations producing faster block. In addition, decreasing extracellular pH accelerated inhibition of TRPM2 by both clotrimazole and econazole; extracellular alkalisation produced the converse effect. Additional experiments indicated hTRPM2 activation was required for the antagonism of either compound to develop, and that neither compound blocked from the intracellular face of the plasma membrane. ADP-ribose-activated whole-cell and single-channel currents in the rat insulinoma cell-line CRI-G1 were also antagonised by clotrimazole. Contrary to the observations made with hTRPM2, antagonism in CRI-G1 cells could be largely reversed following clotrimazole removal. These experiments suggest that imidazole antifungals may be useful tool antagonists for future studies of TRPM2 function.

Animals↗
PubMed2000

Mechanism of shrinkage activation of nonselective cation channels in M-1 mouse cortical collecting duct cells.

It has previously been shown that osmotic cell shrinkage activates a nonselective cation (NSC) channel in M-1 mouse cortical collecting duct cells [54] and in a variety of other cell types [20]. In the present study we further characterized the shrinkage-activated NSC channel in M-1 cells and its mechanism of activation using whole-cell current recordings. Osmotic cell shrinkage induced by addition of 100 mm sucrose to the bath solution caused a 20-fold increase in whole-cell inward currents from -10.8 +/- 1.5 pA to -211 +/- 10.2 pA (n = 103). A similar response was observed when cell shrinkage was elicited using a hypo-osmotic pipette solution. This indicates that cell shrinkage and not extracellular osmolarity per se is the signal for current activation. Cation substitution experiments revealed that the activated channels discriminate poorly between monovalent cations with a selectivity sequence NH(4) (1.2) > or = Na(+) (1) approximately K(+) (0.9) approximately Li(+) (0.9). In contrast there was no measurable permeability for Ca(2+) or Ba(2+) and the cation-to-anion permeability ratio was about 14. The DPC-derivatives flufenamic acid, 4-methyl-DPC and DCDPC were the most effective blockers followed by LOE 908, while amiloride and bumetanide were ineffective. The putative channel activator maitotoxin had no effect. Current activation was dependent upon the presence of intracellular ATP and Mg(2+) and was inhibited by staurosporine (1 microm) and calphostin C (1 microm). Moreover, cytochalasin D (10 microm) and taxol (2 microm) reduced the current response to cell shrinkage. These findings suggest that the activation mechanism of the shrinkage-activated NSC channel involves protein kinase mediated phosphorylation steps and cytoskeletal elements.

Adenosine Triphosphate↗
PubMed1999

Osmotic shrinkage activates nonselective cation (NSC) channels in various cell types.

Osmotic cell shrinkage activates a nonselective cation (NSC) channel in M-1 mouse cortical collecting duct cells (Volk, Frömter & Korbmacher, 1995, Proc. Natl. Acad. Sci. USA 92: 8478-8482). To see whether shrinkage-activated NSC channels are an ubiquitous phenomenon, we tested the effect of hypertonic extracellular solution on whole-cell currents of HT29 human colon carcinoma cells, BSC-1 renal epithelial cells, A10 vascular smooth muscle cells, and Neuro-2a neuroblastoma cells. Addition of 100 mm sucrose to an isotonic NaCl bath solution induced cell shrinkage of HT29 cells as evidenced by a decrease in cell diameter from 18 +/- 1 microm to 12 +/- 1 microm (n = 13). Upon cell shrinkage whole-cell currents of HT29 cells increased within 8 +/- 1 min by about 30-fold (n = 13). Cell shrinkage and current activation were reversible upon return to isotonic solution. Replacement of bath Na+ by K+ or Li+ had almost no effect on the stimulated inward current. In contrast, replacement by N-methyl-d-glucamine (NMDG) completely abolished it and shifted the reversal potential from -4.5 +/- 0.7 mV to -57 +/- 4.1 mV (n = 10). Thus, the stimulated conductance is nonselective for alkali cations but highly selective for cations over anions with a cation-to-anion permeability ratio of about 13. Flufenamic acid (100 microm) inhibited the stimulated current by 84 +/- 4.7% (n = 8). During the early phase of hypertonic stimulation single-channel transitions could be detected in whole-cell current recordings, and a gradual activation of 12 and more individual channels with a single-channel conductance of 17.6 +/- 0.9 pS (n = 4) could be resolved. In analogous experiments similar shrinkage-activated NSC channels were also observed in BSC-1 renal epithelial cells, A10 vascular smooth muscle cells, and Neuro-2a neuroblastoma cells. These findings indicate that shrinkage-activated NSC channels are an ubiquitous phenomenon and may play a role in volume regulation.

Animals↗
PubMed1999

Changes of intracellular calcium concentrations by phenylephrine in renal arterial smooth muscle cells.

In smooth muscle cells isolated from swine renal interlobar arteries, phenylephrine (PE) at concentrations of 1-10 microM produced biphasic increases of the intracellular calcium concentration. An early transient rise was followed by a maintained plateau. The maintained component was sensitive to extracellular calcium, in contrast to the early transient, which was still observed in nominally calcium-free solution. Nifedipine (1 microM) and NiCl2 (100 microM) only weakly affected the calcium signal, suggesting that voltage-sensitive calcium channels play only a minor role in the PE-induced changes in intracellular calcium. Thapsigargin (0.5 microM) elevated the intracellular calcium concentration and depressed both the early transient and the maintained component of the PE response. In calciumfree medium PE induced a transient rise of the intracellular calcium concentration with a depressed plateau. Readmission of calcium elevated the intracellular calcium concentration above the baseline. Both components of the PE-induced calcium signal were completely abolished when the cells were pretreated with the phospholipase C (PLC) inhibitor U73122 (2 microM). LaCl3 (100 microM, 1 mM), an inhibitor of calcium-release-activated current (ICRAC), had no effect on the PE-induced calcium signal. GdCl3 (50 microM), SKF 96365 (10 microM) and flufenamic acid (100 microM), reported to inhibit nonselective cation channels, blocked or transiently reduced the maintained calcium signal. Several protein kinase inhibitors such as genistein (10 microM), H7 (50 microM), H89 (1 microM) and bisindolylmaleimide (0.2 microM) reduced the maintained calcium signal. We conclude that the initial transient spike of the PE-induced calcium signal is due to release of calcium from inositol 1,4,5-trisphosphate-sensitive calcium stores evoked by alpha 1-adrenoceptor-coupled stimulation of PLC and that the maintained component is due to capacitative calcium entry, which is modulated by protein kinases.

Adrenergic alpha-Agonists↗