Effects of amlodipine on TGF-β-induced Smad2, 4 expressions in adriamycin toxicity of rat mesangial cells

Transforming growth factor-β (TGF-β) is closely associated with progressive renal fibrosis. A central component of TGF-β-stimulated mesangial cell fibrogenesis is the TGF-β family-specific Smad signal transduction pathway. This study investigated the expression of TGF-β-receptor–activated Smad2, its common partner Smad4, and the phosphorylated Smad2 (p-Smad2) in adriamycin-induced toxicity of cultured rat mesangial cells. This in vitro study showed that amlodipine (10⁻⁹ to 10⁻⁵ mol/l) had no effect on the toxicity of rat mesangial cells induced by adriamycin in the absence of TGF-β1. However, amlodipine (10⁻⁷ to 10⁻⁵ mol/l) reduced the toxicity of rat mesangial cells induced by TGF-β1 in the absence of adriamycin; moreover, amlodipine (10⁻⁸ to 10⁻⁵ mol/l) significantly reduced adriamycin-induced cytotoxicity when it was given in combination with TGF-β1; amlodipine (10⁻⁶, 10⁻⁵ mol/l) had no effect on Smad2 mRNA and protein expression induced by adriamycin + TGF-β1, but it (10⁻⁶, 10⁻⁵ mol/l) dramatically inhibited the down-regulation of p-Smad2 protein expression as well as Smad4 mRNA and protein expression induced by adriamycin + TGF-β1 in rat mesangial cells. Present study shows that amlodipine exerts a significant inhibition on adriamycin-induced toxicity in rat mesangial cells by affecting the expression of TGF-β/Smad signaling intermediates p-Smad2 and Smad4.     

Valve movement response of the mussel Mytilus galloprovincialis to metals (Cu, Hg, Cd and Zn) and phosphate industry effluents from Moroccan Atlantic coast

Valve activity was measured in the Mediterranean mussel Mytilus galloprovincialis in response to sublethal concentrations of four metals (Hg, Cu, Zn and Cd) and two phosphate industry effluents from the Atlantic coast of Morocco. Valve movements were monitored using a proximity inductive sensor which could display all activity figures from full closure to wide opening of the shell valves. In a 1 h exposure experiments, all metals induced a decrease in the time of normal opening and the appearance of sequences of stress behaviour, including enhanced valve adductions and complete closure at high concentrations. Mercury (tested from 5 to 75 μg Hg l⁻¹) was the most toxic to the valve activity, with a threshold effective concentration at 10 μg Hg l⁻¹ and full valve closure occurring at 50 μg Hg l⁻¹. Copper (15–150 μg Cu l⁻¹) showed a toxic effect starting at threshold concentration of 20 μg Cu l⁻¹ and induced full valve closure at 150 μg Cu l⁻¹. Zinc (100–500 μg Zn l⁻¹) was effective in reducing the time of normal opening (threshold concentration at 100 μg Zn l⁻¹) but no complete closure was recorded in any of the tested concentrations. For cadmium (1000–5000 μg Cd l⁻¹), the valve activity was insensitive for exposures under 2000 μg Cd l⁻¹. Results for the testing of several samplings of the phosphate industry effluents (Safi and Jorf Lasfar) showed that their toxicity varied over the time. The effluent of the Jorf Lasfar plant (2–9.4%) was, however, more toxic than that of Safi (1–25%). In the light of these results, the sensitivity of the valve activity of Mytilus galloprovincialis to pollutants and its usefulness for in situ monitoring of coastal pollution in Morocco are discussed.     

Effect of pertussis toxin on baclofen- and diphenhydramine-induced amnesia

The effect of pretreatment with pertussis toxin at the doses of 0.25 and 0.50 μg per mouse ICV on the amnesic effect produced by baclofen (0.1–4 mg kg⁻¹ IP), diphenhydramine (15–30 mg kg⁻¹ IP) and scopolamine (0.5–5 mg kg⁻¹ IP) was investigated in the mouse passive avoidance test. Ten days after a single injection of pertussis toxin, baclofen (2–4 mg kg⁻¹ IP) amnesia was prevented. By contrast, pertussis toxin had no effect on diphenhydramine- and scopolamine-induced amnesia. Pretreatment with pertussis toxin at both doses used did not impair motor coordination of the mice, as revealed by the rota-rod test. The present results indicate that the activation of pertussis toxin-sensitive G-proteins represents an important transduction step in memory impairment induced by GABA_B (γ-aminobutyric acid B) agonists, but not by antihistaminic and antimuscarinic drugs. Received: 3 June 1997/Final version: 16 October 1997     

Effect of heavy metal exposure on blood haemoglobin concentration and methemoglobin percentage in Lumbricus terrestris

The earthworm haemoglobin (Hb) is a large extracellular hemoprotein flowing in a closed circulatory system. In spite of the fundamental role of this respiratory pigment in earthworm physiology, little is known about its sensitivity to environmental pollutants. The aim of the present work was to investigate the possible effect of heavy metal (cadmium, copper, mercury) exposure on Hb concentration and oxidation state (methemoglobin formation) in the earthworm Lumbricus terrestris. In addition, the tissue concentration of metallothioneins, a well-known biomarker of heavy metal exposure, was determined as an indicator of metal uptake. The animals were exposed to increasing concentrations of Cd, Cu and Hg utilizing the standard acute toxicity test, “Filter paper test” for 48 h. Exposure to heavy metals (10⁻⁵–10⁻³ M for Cd, 10⁻⁴–10⁻³ M for Hg, and 10⁻⁴–10⁻² M for Cu) was found to increase haemoglobin concentration in L. terrestris, although the magnitude of such an increase was dependent on the metal. In addition, metal exposure led to the formation of methemoglobin. Compared to other known biological responses to heavy metals, such as metallothionein induction, methemoglobin increase showed a higher sensitivity and a higher percentage variation in exposed organisms, showing to be a possible suitable biomarker of exposure/effect to be included in a multi biomarker strategy in earthworm in soil monitoring assessment.     

Effect ofAlpha-2 adrenergic agonist onBeta adrenoceptor-mediated control of blood glucose in the fasted mouse

Dose-dependent increases in blood glucose were produced by epinephrine and clonidine in fasted male mice. Isoproterenol was ineffective in increasing blood glucose at lower doses (10⁻⁸ M/kg−10⁻⁷ M/kg); with higher dose (10⁻⁶ M/kg) the glucose level was increased. The hyperglycemia induced by epinephrine was inhibited by yohimbine, prazosin and propranolol, indicating that the hyperglycemic effect of epinephrine is mediated byalpha-1,alpha-2 andbeta adrenoceptor. When clonidine (10⁻⁶ M/kg) was administered simultaneously with isoproterenol (10⁻⁶ M/kg), an enhenced hyperglycemic effect was observed. The increment produced by clonidine plus isoproterenol was higher than that by clonidine alone. These results suggest that stimulation ofalpha-2 adrenoceptor may be responsible for the exertion of the hyperglycemic effect bybeta agonists in fasted mice.     

Effect of rilmakalim on detrusor contraction in the presence and absence of urothelium

Openers of K_ATP channels are known to inhibit KCl-, carbachol- and also electrically induced contractions in detrusor muscle strips from various species. Contractions of isolated strips of urinary bladder are usually of higher amplitude when the urothelium has been removed. This has been explained by the release of an urothelium-derived relaxing factor. In this study we examined whether intact urothelium may modulate the effect of the selective K_ATP channel opener rilmakalim. Contractile responses to 85 mM KCl and 10 μM carbachol were measured in detrusor strips from mouse, pig and man. In the presence of an intact urothelium, contractions were significantly reduced in strips from all three species investigated. In preparations with urothelium rilmakalim reduced KCl contractions with similar potency and efficacy [−logIC₅₀ (M) 4.6 to 5.1; Eff_max reduction to 14-30% of control]. However, in urothelium-denuded strips rilmakalim was more potent in pig (−logIC₅₀ 5.5) than in mouse and man (−logIC₅₀ 4.7 and 4.4, respectively). The order of potency for rilmakalim to suppress carbachol-induced contractions was pig (—logIC₅₀ 6.7) > man (5.8) > mouse (4.7); contractions were significantly more reduced in pig (Eff_max reduction to 11±2%, n=10) and in mouse (21±2%, n=8) than in human detrusor (55±5%, n=5). The presence of urothelium did not affect the concentration–response curves for rilmakalim, with the exception of KCl-induced contractions in pig. Only the rilmakalim-induced relaxation of carbachol-mediated contractions in pig were prevented by the K_ATP channel blocker glibenclamide. We conclude that with this one exception, the responses to rilmakalim in detrusor contractions were not mediated by K_ATP channel opening.     

Comparison of Skin Esterase Activities from Different Species

Purpose Many topically applied drugs contain esters that are hydrolyzed in the skin. Minipigs have emerged as potential models of human dermatology and, in some aspects, may be superior to commonly used rat skin. The aims of this study were to evaluate the suitability of minipig and rat skin as in vitro models of human epidermal esterase activity. Methods Naphthyl acetate and para-nitrophenyl acetate were tested as prototypical substrates of carboxylesterases from skin, plasma, and liver. Reaction products were monitored by high-performance liquid chromatography/ultraviolet analysis. Results Hydrolysis efficiency in skin was higher than plasma, but lower than liver. The esterase efficiency of rat skin microsomes (580–1100 min⁻¹ mg⁻¹) was two to three orders of magnitude higher than human (1.3–4.2 min⁻¹ mg⁻¹) and minipig microsomes (1.2–4.2 min⁻¹ mg⁻¹). Rat skin cytosol (80–100 min⁻¹ mg⁻¹) was 2- to 10-fold more efficient than human (2.4–67 min⁻¹ mg⁻¹) or minipig cytosol (18–61 min⁻¹ mg⁻¹). Most importantly, human skin fractions displayed kinetics of hydrolysis very similar to minipig skin. Conclusions These studies show minipig skin as an appropriate, potentially valuable model for human epidermal ester metabolism and support the use of minipig skin in preclinical development of topically applied compounds.     

Inhibitory effects of L- and T-type calcium antagonists on contractions of human detrusor muscle

The inhibitory and relaxant effects of the L-type calcium antagonists nifedipine, nimodipine, verapamil and diltiazem, and of the T-type calcium antagonist mibefradil, on contractions of isolated human detrusor muscle were investigated. The tissue was obtained from 10 patients undergoing cystectomy due to bladder cancer. Effects of the calcium antagonists at different concentrations on the concentration-response curves for carbachol were investigated. Furthermore, concentration-relaxation curves were performed using potassium-precontracted muscle strips. All L-type calcium antagonists suppressed the mean concentration-response curve of carbachol significantly at a concentration of 10⁻⁶ M. Mibefradil up to 10⁻⁵ M did not significantly suppress it. Nifedipine significantly reduced the carbachol-induced maximum contraction to 75% and 44%, verapamil to 75% and 67% of the appropriate control value at concentrations of 10⁻⁷ and 10⁻⁶ M, respectively. Diltiazem reduced it insignificantly to 96% and 71% at the above-mentioned concentrations. The concentration-relaxation experiments revealed following pD2-values and maximum relaxations of nifedipine, nimodipine, verapamil and diltiazem, respectively: 6.23, 6.37, 5.66, 5.81 and 85%, 83%, 82%, 90%. Maximum relaxations and pD2-values were not significantly different from each other. The lowest concentration, for which a significant effect compared to control in Student`s t-test was found, amounted to 10⁻¹⁰ M, 10⁻⁹ M, 10⁻⁷ M, 10^−6.5 M and 10⁻⁴ M for nimodipine, nifedipine, diltiazem, verapamil and mibefradil, respectively. L-type calcium antagonists are very potent relaxant agents of the human detrusor muscle in vitro.     

Tachykinin NK2 receptors in the hamster urinary bladder: in vitro and in vivo characterization

We have characterized the contractile responses produced by stimulation of the tachykinin NK₂ receptor in the hamster urinary bladder in vitro and in vivo. In isolated bladder strips, neurokinin A (NKA, pD ₂ 7.40, E_max 71% of the response to 80 mM KCl) and the synthetic tachykinin NK₂ receptor selective agonist [βAla⁸]NKA(4–10) (pD ₂ 7.48, E_max 77% of the response to KCl) both induced a concentration-dependent contraction, whereas the tachykinin NK₁ and NK₃ receptor selective agonists, [Sar⁹]substance P sulfone and senktide, respectively, produced a negligible contractile effect. The bicyclic peptide antagonists MEN 11420 and MEN 10627 behaved as competitive antagonists of the response to [βAla⁸]NKA(4–10) with apparent pK _B values of 9.3 and 9.7, respectively. Comparable apparent pK _B values were estimated against NKA (pK _B 9.2 and 9.4 for MEN 11420 and MEN 10627, respectively). Under isovolumetric recording of the intravesical pressure, the nicotinic receptor agonist DMPP (0.6 μmol/kg i.v.) produced a phasic contraction of the hamster bladder in vivo that was abolished by hexamethonium (110 μmol/kg i.v.) or by surgical ablation of pelvic ganglia. In vivo [βAla⁸]NKA(4–10) (10 nmol/kg i.v.) induced a tonic-type sustained bladder contraction with superimposed high frequency and small amplitude (<12 mmHg) phasic contractions and, in about 70% of cases examined, a few high amplitude (>20 mmHg) phasic contractions. Hexamethonium abolished the high amplitude phasic contractions, indicating their reflex origin. In animals subjected to the ablation of pelvic ganglia, the urinary bladder response to [βAla⁸]NKA(4–10) was comparable to that observed after administration of hexamethonium. Moreover, hexamethonium did not affect the contractile responses to [βAla⁸]NKA(4–10) in ganglionectomized animals. MEN 10627 and MEN 11420 produced a dose-dependent and long-lasting inhibition of the contractile response to [βAla⁸]NKA(4–10): the least effective doses of the two antagonists were 30 and 3 nmol/kg i.v. for MEN 10627 and MEN 11420, respectively. An almost complete and long-lasting inhibition of the response to the agonist was produced at doses of 10 and 100 nmol/kg i.v. of MEN 11420 and MEN 10627. In urethane-anaesthetized hamsters the non-stop intravesical infusion of saline (50 μl/min) produced repetitive micturition cycles which were abolished by hexamethonium (110 μmol/kg i.v.) or by surgical removal of the pelvic ganglia. MEN 11420 (100 nmol/kg) had no significant effect on the volume-evoked micturition reflex in anaesthetized hamsters. In conclusion, the hamster urinary bladder is a suitable preparation for studying the action of tachykinin NK₂ receptor antagonists in vivo: in this species, the stimulation of tachykinin NK₂ receptors induces bladder contractions. Blockade of tachykinin NK₂ receptors does not appreciably modify the volume-evoked micturition reflex in this species. Received: 22 April 1998 / Accepted: 12 June 1998     

Effects of an anionic surfactant (FFD-6) on the energy and information flow between a primary producer (<Emphasis Type="Italic">Scenedesmus obliquus</Emphasis>) and a consumer (<Emphasis Type="Italic">Daphnia magna</Emphasis>)

The effects of a commercially available anionic surfactant solution (FFD-6) on growth and morphology of a common green alga (Scenedesmus obliquus) and on survival and clearance rates of the water flea Daphnia magna were studied. The surfactant-solution elicited a morphological response (formation of colonies) in Scenedesmus at concentrations of 10–100 μl l⁻¹ that were far below the No Observed Effect Concentration (NOEC) value of 1,000 μl l⁻¹ for growth inhibition. The NOEC-value of FFD-6 for colony-induction was 3 μl l⁻¹. Daphnia survival was strongly affected by FFD-6, yielding LC_50–24h and LC_50–48h of 148 and 26 μl l⁻¹, respectively. In addition, clearance rates of Daphnia feeding on unicellular Scenedesmus were inhibited by FFD-6, yielding a 50% inhibition (EC_50–1.5h) at 5.2 μl l⁻¹ with a NOEC of 0.5 μl l⁻¹. When Daphnia were offered FFD-6-induced food in which eight-celled colonies (43 × 29 μm) were most abundant, clearance rates (~0.14 ml ind.⁻¹ h⁻¹) were only 25% the rates of animals that were offered non-induced unicellular (15 × 5 μm) Scenedesmus (~0.56 ml ind.⁻¹ h⁻¹). As FFD-6 concentrations in the treated food used in the experiments were far below the NOEC for clearance rate inhibition, it is concluded that the feeding rate depression was caused by the altered morphology of the Scenedesmus moving them out of the feeding window of the daphnids. The surfactant evoked a response in Scenedesmus that is similar to the natural chemically induced defensive reaction against grazers and could disrupt the natural information conveyance between these plankton organisms.     

Functional characterization of α1-adrenoceptor subtypes in the rabbit spleen

Phenylephrine and (±)N-[5-(4,5-dihydro-1-H-imidazol-2yl)-2-hydroxy-5,6,7,8-tetrahydronaphthalen-1-yl] methanesulphonamide hydrobromide (A 61603) evoked concentration-dependent contractions of the rabbit spleen. These contractions were antagonized by prazosin (10^–8–10^–7 M) with pA ₂ values of 8.34±0.11 and 8.15±0.10 against phenylephrine and A 61603, respectively. In both cases, the slopes of the Schild plots were not significantly (P>0.05) different from 1.0, indicating competitive antagonism. The effects of subtype-selective antagonists WB 4101 [2-(2-6-dimethoxyphenoxyethyl)aminomethyl-1,4-benzodioxane hydrochloride] and 5-methylurapidil on agonist-induced contractions were also examined. WB 4101 competitively antagonized agonist-induced contractions; pA ₂ values were 8.13±0.10 and 8.10±0.03 against phenylephrine and A 61603, respectively. Corresponding values for 5-methylurapidil were 8.28±0.17 and 7.93±0.02 against phenylephrine and A 61603, respectively. Tamsulosin and Rec 15/2739 [(8-3-[4-(2-methoxyphenyl)-1-piperazinyl]-propylcarbamoyl)-3-methyl-4-oxo-2-phenyl-4H-1-benzopyran dihydrochloride] also antagonized phenylephrine- and A 61603-induced contractions with pA ₂ values of 9.38±0.13 and 9.18±0.06 (tamsulosin) and 8.41±0.12 and 8.34±0.11 (Rec 15/2739) against phenylephrine and A 61603, respectively. HV 723 (α-ethyl-3,4,5-trimethoxy-α-(3-((2-(2-methoxyphenoxyethyl)-amino)-propyl)benzene-aceto-nitrile) fumarate) competitively antagonized phenylephrine-induced contractions with a pA ₂ value of 8.57±0.06. Chloroethylclonidine (CEC; 10^–4 M) shifted phenylephrine and A 61603 concentration-response curves to the right, reducing their potencies approximately two- to threefold, while the maximum response was reduced by 8% in both cases. It was therefore concluded that contractions of the rabbit spleen induced by α₁-adrenergic agonists were mediated predominantly by a relatively CEC-insensitive α₁-adrenoceptor subtype, possibly the α_1L-subtype. Received: 14 April 1998 / Accepted: 17 June 1998     

Biological effects of the dihydroorotate dehydrogenase inhibitor polyporic acid, a toxic constituent of the mushroom Hapalopilus rutilans , in rats and humans

Inhibitors of dihydroorotate dehydrogenase (DHO-DH), such as brequinar or leflunomide, have been intensively tested for their antitumour and immunomodulating effects. Polyporic acid (PA) from the mushroom Hapalopilus rutilans (H. r.) also is a DHO-DH inhibitor (50% inhibitory concn., IC₅₀, 10⁻⁴–10⁻³ M). As three people had been poisoned following ingestion of H. r. we wanted to investigate the effects of PA in rats and in cell cultures. Rats given PA via probang (100–800 mg/kg) within 24 h developed strongly reduced locomotor activity, depressed visual placing response and impaired wire manoeuvre. Laboratory investigation of blood revealed hepatorenal failure, metabolic acidosis as well as hypokalaemia and hypocalcaemia. All symptoms closely paralleled the effects seen in the poisoned people. Proliferation of cultured cells (including rat brain neurons and glia, fibroblasts, tumour cells) was depressed at 10⁻⁴–10⁻³ M PA. We conclude that the intoxication of people poisoned with H. r. is due to the high content of the DHO-DH inhibitor PA. Received: 6 July 1998 / Accepted: 7 September 1998     

The effect of the menstrual cycle on the pharmacokinetics of caffeine in normal, healthy eumenorrheic females

  Objective: Hormonal fluctuations of estrogen and progesterone in eumenorrheic women may be capable of altering the pharmacokinetics of certain agents. The objective of this study was to determine the effect of the luteal, ovulatory and follicular phases of the menstrual cycle on the pharmacokinetics of caffeine, a low clearance, flow-independent drug. Methods: Subjects were ten healthy, non-smoking, eumenorrheic females who were not pregnant and had not used oral contraceptives for a minimum of 3 months prior to the study. Blood samples were collected during one menstrual cycle for the determination of estradiol and progesterone concentrations during the follicular (days 2–6 post-onset of menses), ovulatory (days 13–16 post-onset of menses) and luteal (days 22–26 post-onset of menses) phases. Caffeine was administered over a single menstrual cycle during the follicular, ovulatory and luteal phases. Each subject was administered a single oral dose of caffeine (300 mg) in 100 ml of lemonade during each phase of the menstrual cycle. A venous catheter was used to collect blood samples at pre-dose and at the following time points: 0.25, 0.5, 0.75, 1, 1.5, 2, 4, 6, 8, 10, 12 and 24 h. Plasma caffeine concentrations were determined using a validated ultraviolet high-performance liquid chromatography method. Results: There were no significant (P < 0.05) differences in the pharmacokinetic parameters of caffeine across the menstrual cycle phases. The average area under the plasma concentration–time curve (AUC_inf) was 93.01 mg l^−1.h and the absorption rate constant (k _a) was 2.88 h⁻¹ during the ovulatory phase, 83.0 mg l⁻¹ h and 2.06 h⁻¹, respectively, during the luteal phase and 84.7 mg l^−1.h and 1.84 h⁻¹, respectively, during the follicular phase. Conclusions: These findings suggest that the menstrual cycle does not significantly alter the pharmacokinetics of caffeine. Received: 16 November 1998 / Accepted in revised form: 19 April 1999     

An in vitro approach for demonstrating the critical role of serum albumin in the detoxication of the carbamate carbaryl at in vivo toxicologically relevant concentrations

The hydrolysis of carbaryl by bovine serum albumin (BSA) was studied at toxicologically relevant concentrations (range 15–300 μM) in order to determine the role of this protein in the detoxication of the carbamate in vivo. The 1-naphthol released during the hydrolysis of carbaryl was monitored using gas chromatography coupled with mass spectrometry. BSA hydrolyzed carbaryl in a time-progressive way. The hydrolysis was also dependent of enzyme (1.0, 2.5, 5.0 and 7.0 mg ml⁻¹) and substrate (range between 15 and 1,000 μM) concentration. The estimated turnover number and Michaelis–Menten constant were 1.6 × 10⁻⁴ s⁻¹ and 430 μM, respectively. Thus, the second order rate constant was 0.37 M⁻¹ s⁻¹. At enzyme concentrations of 7.0 mg ml⁻¹ and substrate concentrations ranging between 50 and 300 μM about 80% of substrate was hydrolyzed in 3 h. At lower substrate concentrations (15 and 30 μM carbaryl) also significant hydrolysis was detected at the highest enzyme concentration, even when these substrate concentrations were 30 and 15 times lower than the Michaelis–Menten constant. Although the efficacy of the enzymatic hydrolysis is low, the extrapolation of our results to the physiological albumin high concentrations (around 40 mg ml⁻¹) suggests that the hydrolysis of carbaryl by serum albumins plays a critical role in the detoxication of this carbamate at in vivo toxicologically relevant concentrations. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Effect of selective phosphodiesterase inhibitors on synaptic transmission in the guinea-pig ileum

The effect of selective phosphodiesterase (PDE) inhibitors was studied on neural transmission within the enteric nervous system employing a two-compartment bath (containing the oral and the anal end of a segment of guinea-pig ileum, respectively). Ascending excitatory enteric nerve pathways were activated by electrical field stimulation (10 Hz for 2 s, 45 mA, 0.5 pulse duration) in the anal compartment and the resulting contraction of the intestinal circular muscle in the oral compartment was recorded. The partitioned bath enables PDE inhibitors and other drugs to be applied to enteric nerve pathways (in the anal compartment) without interfering with the recording of the smooth muscle contraction in the oral compartment. The PDE 4 inhibitors rolipram (0.01–10 μM) and Ro-20-1724 (0.01–10 μM) significantly (P<0.01) inhibited (10–91% and 9–83%, respectively) the nerve-mediated contractions. When both rolipram and Ro-20-1724 were tested after phentolamine (1 μM) or yohimbine (0.1 μM), they were significantly (P<0.01) less effective. By contrast prazosin (1 μM) was ineffective. Vinpocetine (50 μM), milrinone (30 μM) and zaprinast (100 μM), which inhibit PDE 1, 3 and 5, respectively, did not modify the nerve-mediated contractions. 8-Bromoadenosine 3’,5’-cyclic monophosphate (8-Br-cyclic AMP) or N⁶,2’-O-dibutyryladenosine 3’,5’ cyclic monophosphate (dibutyryl cyclic AMP), two analogues of cyclic AMP, at lower concentrations (0.1–1 μM) significantly (P<0.01) inhibited (15–73% and 5–49%, respectively) the nerve-mediated contractions, while at higher concentrations (10–100 μM) they caused a significant (P<0.01) potentiating (48–68% and 77–78%, respectively) effect. These results indicate that inhibition of PDE 4 (but not PDE 1, PDE 3 or PDE 5) produces a depression of neural transmission within the enteric nervous system, possibly by releasing noradrenaline acting at α₂-adrenoceptors on enteric neurons. Received: 21 November 1997 / Accepted: 11 March 1998     

Involvement of nitric oxide in the potentiation of neurogenic contraction by manganese and nickel ions in mouse urinary bladder

Repetitive electrical field stimulation evoked muscle contractions of the isolated mouse detrusor strips, which could be abolished by tetrodotoxin (TTX). Both Mn²⁺ and Ni²⁺ (0.01–0.06 mM) enhanced neurogenic detrusor contractions in high Ca²⁺ (5 mM) medium. The non-cholinergic component of the evoked detrusor contractions (in the presence of atropine) was specifically sensitive to this enhancing effect by either Mn²⁺ or Ni²⁺. In contrast, the cholinergic component in α,β-methylene ATP-treated detrusors remained unaffected. As compared with Mn²⁺ and Ni²⁺, other metal ions, Cu²⁺, Co²⁺, Cd²⁺ and UO₂ ²⁺, failed to enhancing the non-cholinergic component of neurogenic detrusor contractions. Nitric oxide synthase (NOS) inhibitors, N^G-nitro-l-arginine methyl ester and 7-nitroindazole (a selective neuronal NOS inhibitor) markedly inhibited the enhancing effect by either Mn²⁺ or Ni²⁺ on the neurogenic detrusor contractions. Moreover, Mn²⁺ and Ni²⁺ did not affect the contractions induced by either carbachol or α,β-methylene ATP. It is concluded from these findings that the neuronal NOS-NO generation pathway is apparently involved in the enhancing effect of Mn²⁺ and Ni²⁺ on the muscle contractions elicited by excitatory non-cholinergic neurotransmission in the mouse detrusor strips. Received: 16 July 1998 / Accepted: 7 September 1998     

Comparison of the pharmacokinetics of tirilazad mesylate in healthy volunteers and stable subjects with mild liver cirrhosis

Objective: The pharmacokinetics of tirilazad mesylate and an active reduced metabolite, U89678, were studied in 7 volunteers with mild cirrhosis of the liver and seven age, sex, weight and smoking status matched healthy normal volunteers. Subjects received a single intravenous infusion of 2.0 mg ⋅kg⁻¹ tirilazad mesylate over 10 min. Results: Mean tirilazad AUC_0–∞ was 8.83 μmol h⋅l⁻¹ and 18.6 μmol h⋅l⁻¹ in healthy volunteers and cirrhotic subjects, respectively. Mean tirilazad clearance in cirrhotics (12.7 l⋅h⁻¹) was approximately 2.1 fold lower than in healthy volunteers (27.8 l⋅h⁻¹). The differences were statistically significant. Mean U-89678 AUC_0–∞ in cirrhotic subjects (3.88 μmol h⋅l⁻¹) was 2.5 fold higher than in healthy controls (1.53 μmol h⋅l⁻¹), but the difference was marginally significant. Conclusion: These results indicate that clearance of both tirilazad mesylate and U89678 is decreased in subjects with hepatic impairment. This observation may be attributed either to decreases in liver blood flow and/or intrinsic clearance. The results of this study thus suggest that increased monitoring and or a reduction in tirilazad dosing may be necessary in patients with hepatic impairment. Received: 10 August 1995/Accepted in revised form: 10 July 1995     

Urotensin II acutely increases myocardial length and distensibility: potential implications for diastolic function and ventricular remodeling

Urotensin II (U-II) is a cyclic peptide that may be involved in cardiovascular dysfunction. In the present study, the acute effects of U-II on diastolic properties of the myocardium were investigated. Increasing concentrations of U-II (10⁻⁸ to 10⁻⁶ M) were added to rabbit papillary muscles in the absence (n = 15) or presence of: (1) damaged endocardial endothelium (EE; n = 9); (2) U-II receptor antagonist, urantide (10⁻⁵ M; n = 7); (3) nitric oxide (NO) synthase inhibitor, N^G-Nitro-l-Arginine (10⁻⁵ M; n = 9); (4) cyclooxygenase inhibitor, indomethacin (10⁻⁵ M; n = 8); (5) NO synthase and cyclooxygenase inhibitors, N^G-Nitro-l-Arginine (10⁻⁵ M) and indomethacin (10⁻⁵ M), respectively, (n = 8); or (6) protein kinase C (PKC) inhibitor, chelerythrine (10⁻⁵ M; n = 9). Passive length–tension relations were constructed before and after a single concentration of U-II (10⁻⁶ M; n = 3). U-II concentration dependently decreased inotropy and increased resting muscle length (RL). At 10⁻⁶ M, active tension decreased 13.8 ± 5.4%, and RL increased to 1.007 ± 0.001 L/L _max. Correcting RL to its initial value resulted in an 18.1 ± 3.0% decrease in resting tension, indicating decreased muscle stiffness, which was also suggested by the down and rightward shift of the passive length–tension relation. This effect remained unaffected by EE damage and PKC inhibition. In contrast, the presence of urantide and NO inhibition abolished the effects of U-II on myocardial stiffness, while cyclooxygenase inhibition significantly attenuated them. U-II decreases myocardial stiffness, an effect that is mediated by the urotensin-II receptor, NO, and prostaglandins. This represents a novel mechanism of acute neurohumoral modulation of diastolic function, suggesting that U-II is an important regulator of cardiac filling. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Presented in part at the American Heart Association Scientific Sessions conference, 2006, in Chicago, Illinois.     

Effect of diprafenone on the pharmacokinetics of digoxin

This study was conducted to investigate the effect of diprafenone on the steady-state pharmacokinetics of digoxin. Twelve healthy men, all rapid hydroxylators of debrisoquine, received digoxin (0.5 mg per day over 7 days with a loading dose of 2 × 1 mg) or digoxin and diprafenone (3 × 100 mg per day) in three different phases, without a wash-out period (phase 1, digoxin alone; phase 2, digoxin + diprafenone; phase 3, digoxin alone). Blood and urine samples were collected for pharmacokinetic analyses. Diprafenone caused a statistically significant increase in digoxin trough concentrations [1.4 (SD 0.2) vs 1.6 (0.3) ng ⋅ml⁻¹], AUC_0–24 values [41 (7) vs 48 (9) ng⋅h⋅ml⁻¹ and C_ss-max [3.9 (0.6) vs 5.5 (0.9) ng⋅ml⁻¹]. In all volunteers the parameters tended to return to the original values after administration of diprafenone was discontinued [1.4 (0.3) ng⋅ml⁻¹, 39 (11) ng⋅h⋅ml⁻¹, and 3.9 (1.1) ng⋅ml⁻¹ for trough concentration, AUC_0–24 and C_max respectively]. The mean relative magnitude of the increase in AUC_0–24 and trough concentration values corresponded to the mean relative decrease in the renal clearance of digoxin (in both cases approximately 20%). This suggests that the increase in AUC and C_ss was caused by reduced renal clearance of digoxin. Received: 10 July 1995 /Accepted in revised form: 5 October 1995     

Effect of pyridine and quinoline N-oxides on microsomal Na,K-ATPase activity

Some heteroaromatic N-oxides of pyridine and quinoline derivatives at concentrations ranging from 10⁻⁴ to 10⁻¹⁰ mole/liter inhibit Na,K-ATPase activity in cattle brain microsomes stronger than does strophanthin K (a drug used for the treatment of cardiac insufficiency). A new Na,K-ATPase activator, 2-(2,4-dimethoxystyryl)quinoline-N-oxide), has been found, which is capable of acting at concentrations within 10⁻⁶–10⁻⁹ mole/liter. Since these compounds activate the enzyme in very low concentrations, they can probably be effective for the treatment of some disorders involving violation of the Na,K-ATPase function. __________ Translated from Khimiko-Farmatsevticheskii Zhurnal, Vol. 40, No. 7, pp. 3–4, July, 2006.