European Journal of Pharmacology: Environmental Toxicology and Pharmacology最新文献

Mini-osmotic pumps containing solutions of either 0.9% NaCl (infused at the rate of 0.5 μl/h) or nicotine (infused in doses of 0.224, 1.03 or 1.88 mg/kg per day) were implanted s.c. into rats 12 days before experimentation. The alkaloid increased solid food consumption, but fluid intake and average weight gain were similar among the animals given saline or nicotine. Chronic nicotine treatment dose dependently intensified cold (4°C)-restraint stress-induced ulceration and increased mast cell degranulation. Oral administration of 40% ethanol to nicotine-treated animals also produced greated mucosal damage; mast cell degranulation by ethanol was significantly worsened after alkaloid treatment. These findings show that the stress ulcer-intensifying action of the alkaloid is mainly through a systemic mechanism. In the case of ethanol-evoked mucosal damage, in addition to a topical effect, stimulation of the stomach wall ganglia is likely to participate in the exaggerated post-vagal ulcerogenic responses as seen in stress.

The hepatotoxicity of diethyldithiocarbamate was examined using an in vitro rat liver slice system. Concentration- and time-dependent losses of intracellular K⁺ and adenosine triphosphate (ATP) levels were observed in rat liver slices incubated with diethyldithiocarbamate at concentrations between 1 and 10 mM over a 4-h period. Histological study revealed perivenous hepatocyte damage. To examine the involvement of Kupffer cells in diethyldithiocarbamate-induced cytotoxicity, rats were injected intravenously with 10 mg/kg of gadolinium chloride (GdCl₃) which diminishes Kupffer cell function. Incubation of liver slice preparations from the GdCl₃-treated rats with diethyldithiocarbamate showed marked inhibition of the cytotoxicity induced by diethyldithiocarbamate. Moreover, in vitro addition of manganese-superoxide dismutase, a superoxide anion scavenger, or dimethyl sulfoxide (DMSO), a hydroxyl radical scavenger, also showed potent inhibition. However, dexamethasone, an inhibitor of tumor necrosis factor, and N,N′-diphenyl-p-phenylenediamine (DPPD), an antioxidant, showed partial prevention of cytotoxicity. Formazan deposits formed as a result of nitro blue tetrazolium reduction were found in Kupffer cells at an early stage after diethyldithiocarbamate treatment, while lipid peroxidation occurred after 3 h. Both pretreatment with GdCl₃ in vivo and addition of DMSO in vitro prevented the increase in lipid peroxidation within the liver slice preparations induced by diethyldithiocarbamate. These findings suggest that Kupffer cell function may be involved in the pathogenesis of diethyldithiocarbamate hepatotoxicity.

We investigated the regulation of the protein expression of the α isozymes of Na⁺,K⁺-ATPase in reference to the enzyme activity in the heart, brain skeletal muscle of rats during deoxycorticosterone acetate (DOCA)-salt hypertension. Treatment of rats with DOCA and salt for 28 days produced a significant increase in systolic blood pressure compared to the control groups which remained normotensive. Rats treated with DOCA expressed greater amounts of the immunoreactive α-1 isoform than untreated controls in whole heart membranes. However, the DOCA-induced increase in the α-1 isoform did not occur during DOCA-salt hypertension. There was a parallel change in the enzyme activity of the Na⁺,K⁺-ATPase and the protein expression of the α-1 isoform as a result of these treatments. We have also demonstrated that the hearts of DOCA-salt hypertensive rats expressed less of the α-2 isoform compared to the controls. We could not detect any alteration in the α-1 and α-2 isoforms of the skeletal muscle and α-1, α-2 and α-3 isoforms of the whole brain Na⁺,K⁺-ATPase during salt or DOCA treatments alone or DOCA-salt hypertension. Furthermore, the Na⁺,K⁺-ATPase activity was unaltered in these tissues during these treatments. In conclusion, cardiac Na⁺,K⁺-ATPase α-subunit protein expression appears to be regulated during DOCA-salt hypertension. In the skeletal muscle and brain, tissues not subjected directly to increased pressure, this regulation of the Na⁺,K⁺-ATPase was not apparent.

Intracerebral brain microdialysis was performed in awake, freely moving rats to study the effect of acute inhalation exposure of toluene (2000 ppm, 2 h) on extracellular levels of γ-aminobutyric acid (GABA) within the globus pallidus and the striatum. GABA within the globus pallidus decreased (20%) during and after (26%) exposure to toluene, while no reduction was seen in the striatal GABA level during exposure. After the exposure there was a tendency towards an increase (maximally 37%) in striatal GABA. 2 h of perfusion with tetrodotoxin (10⁻⁶ M) decreased (32%) the extracellular GABA levels within the globus pallidus. The results suggest that the effect of acute toluene exposure varies with brain region and that the GABA output from the striatum to globus pallidus is more affected by the exposure than the GABA release within the striatum.

The antiinflammatory, antioxidant activity of taurine and niacin against cyclophosphamide-induced early lung injury in rats was investigated. A single intraperitoneal injection of cyclophosphamide markedly altered the levels of several biomarkers in bronchoalveolar lavage fluid: total protein, albumin, angiotensin converting enzyme, lactate dehydrogenase, lactate, N-acetyl-β-d-glucosaminidase, alkaline phosphatase, acid phosphatase and lipid peroxidation product were significantly elevated. In contrast, decreased levels of total reduced glutathione (GSH) and ascorbic acid were observed. Cyclophosphamide significantly increased malondialdehyde levels in serum and lung. Significant increases in lung content of lipid hydroperoxides were seen that paralleled the decreased levels of total reduced glutathione and total sulfhydryl groups. Pretreatment of rats with daily intraperitoneal injection of taurine plus niacin 7 days prior to and 2 days after cyclophosphamide insult significantly inhibited the development of lung injury, prevented the alterations in lavage fluid biomarkers associated with inflammatory reactions, with less lipid peroxidation and restoration of antioxidants. In conclusion, our results suggest that taurine and niacin in combination is efficient in blunting cyclophosphamide-induced pulmonary damage.

Triphenyltin induces a contracture of the mouse phrenic nerve-diaphragm preparation. This contracture was not inhibited by (+)-tubocurarine, high magnesium or the absence of electrical stimulation. Triphenyltin (0.1 mM) reduced the muscle membrane potential, the amplitude of the muscle action potential and the muscle membrane input resistance. Pretreatment with high K⁺ (25 mM) or veratridine (1.5 μM; a Na⁺ channel activator) briefly shortened the onset of the contracture and increased the peak tension of the contracture. Pretreatment with tetrodotoxin (0.3 μM; a Na⁺ channel blocker) or glycerol (a T tubule uncoupler), however, significantly reduced the triphenyltin-induced contracture. Removing Ca²⁺ from external solution and prolonged treatment with either caffeine (20 mM) or ryanodine (2 μM) inhibited the triphenyltin-induced contracture. However, a brief treatment with a lower concentration of caffeine (10 mM) potentiated the contracture. ⁴⁵Ca²⁺ uptake studies showed that triphenyltin caused the muscle to accumulate Ca²⁺ which entered from external solution. Pretreatment with trypsin and dithiothreitol (a sulfhydryl-containing reducing agent) blocked the contracture induced by triphenyltin. These results suggest that triphenyltin initially interacts with the sulfhydryl groups of membrane bound proteins (possibly the Na⁺ channel) to cause depolarization of the muscle fibres. This depolarization triggers the release of Ca²⁺ from sarcoplasmic reticulum through the mechanism of Ca²⁺ inducing Ca²⁺ release, activates the contractile filaments and causes the muscle to contract.

The ability of three highly homologous mouse liver CYP2A enzymes to activate aflatoxin B₁ was studied by expressing them in recombinant AH22 Saccharomyces cerevisiae yeast cells. The reconstituted monooxygenase complex with CYP2A5 purified from yeast cell microsomes produced epoxide at a rate of 17.2 nmol/min per nmol P450 in the presence of 50 μM aflatoxin B₁ while CYP2A4 had about 10% and P4507α only 1.5% of this activity. However, K_m values were 530 and 10 μM and V_max values 12.5 and 14.3 nmol/min per nmol P450 for CYP2A4 and CYP2A5, respectively. When recombinant yeast cells were exposed to aflatoxin B₁ LC₅₀ concentrations were 7.5 ± 5.5 μM for CYP2A4, 0.45 ± 0.10 μM for CYP2A5 and > 320 μM for P4507α expressing yeast cells. Aflatoxin B₁-DNA adduct levels in the same yeast cells were 50, 890 pmol/mg DNA and below detection limit when 3.0 μM aflatoxin B₁ was used in the incubation mixture. Coumarin an inhibitor for CYP2A4 and a substrate for CYP2A5 diminished the toxicity of aflatoxin B₁ in a dose-dependent manner for these recombinant yeast cells. These data demonstrate that (1) highly homologous mouse CYP2A enzymes activate aflatoxin B₁ in a different manner and (2) that recombinant yeast cells expressing mammalian CYP enzymes are a useful and inexpensive system to test the role of different enzymes in aflatoxin B₁ toxicity. The data also indicate that mouse CYP2A5 and its counterpart in other species could have a significant role in aflatoxin B₁ toxicity in organs where it is expressed at high levels.

The effect of amiloride on the positive inotropic and toxic effects of ouabain in guinea-pig left atria has been studied. In atria driven at 1 Hz, amiloride (0.3 and 0.5 mM) decreased the EC₅₀ but did not affect the maximal tension developed by ouabain. At 0.1 Hz, amiloride did not change either the EC₅₀ or the maximal tension developed by ouabain. Ouabain toxicity (onset of arrhythmias) was not changed by amiloride at either frequency of stimulation. Therefore, amiloride did not antagonize either the positive inotropic or the toxic effect of ouabain. The positive inotropic effect of amiloride has been ascribed to the inhibition of the Na⁺/Ca²⁺ exchanger. Since amiloride inhibits also the Na⁺/H⁺ exchanger, 5-(N-ethyl-N-isopropyl)amiloride (EIPA), an amiloride derivative which selectively inhibits the Na⁺/H⁺ exchange, has been tested to evaluate the role of the Na⁺/H⁺ exchange in the amiloride-ouabain interaction. EIPA increased the EC₅₀ values of ouabain and decreased the maximal developed tension by the glycoside in atria driven at 0.1 and 1 Hz, but did not antagonize the toxic response (arrhythmias) of atria to ouabain. It is suggested that the inhibition of Ca²⁺ exit through the Na⁺/Ca²⁺ exchange by amiloride and ouabain may explain the observation that the positive inotropic effects of amiloride and ouabain are additive.

Mechanisms for the hypertrophy of rat pancreas induced by long-term administration of bethanechol were investigated. The administration of bethanechol, an acetylcholine receptor agonist, to male Wistar rats for 14 days induced significant increases in the pancreatic weight and contents of protein, amylase and RNA in the pancreas without altering the content of DNA and the incorporation of [³H]thymidine into DNA. Simultaneous administration of atropine with bethanechol suppressed the bethanechol-induced pancreatic hypertrophy. Long-term administration of other acetylcholine receptor agonists also showed similar effects as produced by bethanechol. CR1505 (loxiglumide; d,l-4-(3,4-dichlorobenzoyl-amino)-5-(N-3-methoxypropyl-pentylamino)-5-oxopentanoic acid), an antagonist of cholecystokinin receptors, inhibited pancreatic growth induced by long-term administration of pentagastrin, whereas pancreatic hypertrophy induced by bethanechol was not inhibited by CR1505. These results suggest that long-term administration of bethanechol induces pancreatic hypertrophy through direct activation of muscarinic receptors in the pancreas.