EFFECT OF DIPYRIDAMOLE ON BLOOD GLUCOSE AND LIVER CYCLIC AMP LEVELS AND PLATELET COUNT DURING ENDOTOXAEMIA IN MICE

1. Blood glucose, liver cyclic AMP (cAMP) levels, and platelet count were studied at different times during a 12 h interval after intravenous (i.v.) endotoxin injection (40 mg/kg) in mice. 2. Blood glucose and liver cAMP levels showed a progressive decrease in endotoxic mice and the decrease was significant after 8 and 12 h. 3. Dipyridamole (50 mg/kg) administered intraperitoneally (i.p.) 7 h post-endotoxin improved blood glucose and liver cAMP levels. 4. Platelet counts were decreased after 1 h in endotoxic mice and remained decreased up to 8 h. Dipyridamole administered soon after endotoxin improved platelet counts in endotoxic mice and increased survival rate to 100%.     

Melatonin attenuates lipopolysaccharide (LPS)-induced apoptotic liver damage in D-galactosamine-sensitized mice

D-Galactosamine (GalN) depletes UTP primarily in liver, resulting in decreased RNA synthesis in hepatocytes. When given together with a sublethal dose of lipopolysaccharide (LPS), GalN highly sensitizes animals to produce apoptotic liver injury with severe hepatic congestion, resulting in rapid death. Melatonin is a cytokine modulator, antioxidant and anti-apoptotic agent. In the present study, we investigated the effect of melatonin on LPS-induced apoptotic liver damage in GalN-sensitized mice. Female CD-1 mice were intraperitoneally (i.p.) injected with melatonin (5.0mg/kg) 30min before GalN/LPS (700mg10microg/kg, i.p.), another two doses of melatonin (2.5mg/kg, i.p.) being administered 1 and 2h after GalN/LPS. Results showed that serum alanine aminotransferase (ALT) activities were markedly increased 8h after GalN/LPS treatment, massive hemorrhage being observed in histological sections of liver from GalN/LPS-treated mice. Melatonin significantly attenuated GalN/LPS-induced elevation of serum ALT. In parallel, melatonin distinctly improved GalN/LPS-induced congestion. Additional experiment showed that melatonin significantly attenuated GalN/LPS-induced hepatic apoptosis, measured by inhibition of caspase-3 activities and attenuation of DNA laddering. Furthermore, melatonin markedly increased hepatic Se-dependent glutathione peroxidase (GSH-Px) and glutathione reductase (GSH-Rd) activities and attenuated hepatic glutathione (GSH) depletion in GalN/LPS-treated mice. Increases in serum tumor necrosis factor alpha (TNF-alpha), which were observed in GalN/LPS-treated mice, were significantly reduced by melatonin. However, melatonin had no effect on LPS-evoked nitric oxide production in GalN-sensitized mice. Taken together, these results indicate that melatonin protected against LPS-induced liver damage in GalN-sensitized mice through its strong ROS-scavenging, antiinflammatory and antiapoptotic effects.     

Preventive effects of a biscoclaurine alkaloid, cepharanthine, on endotoxin or tumor necrosis factor-alpha-induced septic shock symptoms: involvement of from cell death in L929 cells and nitric oxide production in raw 264.7 cells

The preventive effects of cepharanthine, a biscoclaurine alkaloid isolated from Stephania cepharantha Hayata, on the lethality and cell death caused by endotoxin or tumor necrosis factor (TNF)-alpha-induced syndrome in septic shock were investigated. In these experiments, we estimated the survival of mice treated with a lethal dose of endotoxin (50 mg/kg, i.p.) or recombinant human (rh) TNF-alpha (10,000 units/mouse, i.v.) together with a sublethal dose (1 mg/kg, i.p.) of endotoxin. Cepharanthine clearly protected mice from endotoxin-induced and endotoxin/rhTNF-alpha-induced lethal shock. In in vitro experiments, cepharanthine (3 micro g/ml) definitely inhibited cell death in mouse L929 fibroblast cells incubated with rhTNF-alpha (100 units/ml) at 37 degrees C for 24 h. On the other hand, non-apoptotic programmed death of cells was observed by fluorescence microscopy in rhTNF-alpha (100 units/ml)-treated L929 cells. In the 3-(4,5-Dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide (MTT) assay after 48-h drug exposure, the cell proliferation of L929 cells was significantly increased by the addition of cepharanthine (1 and 3 micro g/ml). It seems that the preventive effect of cepharanthine on rhTNF-alpha-induced cytotoxicity in fibroblast cells occurs through an increase of cell proliferation by the drug. In addition, cepharanthine suppressed nitric oxide (NO) production by endotoxin-stimulated Raw 264.7 mouse macrophage cells. These findings suggest that cepharanthine prevents lethality or cytotoxicity through suppression of endotoxin-induced NO in macrophages and that its effects are possibly mediated by the enhancement of the proliferation of fibroblast cells. Cepharanthine may therefore protect against some of the various disturbances caused by endotoxin through its ability to inhibit NO production in septic shock.     

The involvement of central cholinergic system in (+)-matrine-induced antinociception in mice

The antinociceptive effect of (+)-matrine was examined in mice by writhing, tail-pressure and hot-plate tests. (+)-Matrine (5, 10 and 20 mg/kg s.c.) produced antinociception in a dose-dependent manner. In hot-plate test, the antinociception produced by (+)-matrine (10 mg/kg s.c.) was attenuated by muscarinic receptor antagonists atropine (5 mg/kg i.p.) and pirenzepine (0.1 mug/mouse i.c.v.) and acetylcholine depletor hemicholinium-3 (HC-3) (1 mug/mouse i.c.v.), but not by opioid receptor antagonist naloxone (2 mg/kg i.p.), dopamine D(2) receptor agonist (-)-quinpirole (0.1 mg/kg i.p.) or catecholamine depletor reserpine (2.5 mg/kg i.p.). Radioligand binding assay demonstrated that (+)-matrine had no affinity for mu-, kappa- or delta-opioid receptors in a wide concentration range (1 x 10(-11)-1 x 10(-3) M). The results suggest that (+)-matrine exerts its antinociceptive effect through multiple mechanism(s) such as increasing cholinergic activation in the CNS rather than acting on opioid receptors directly.     

1,8-cineole protects against liver failure in an in-vivo murine model of endotoxemic shock

The effects of 1,8-cineole on D-galactosamine/lipopolysaccharide (GalN/LPS)-induced shock model of liver injury was investigated in mice. The co-administration of GalN (700 mg kg(-1), i.p.) and LPS (5 microg kg(-1), i.p.) greatly elevated serum concentrations of tumour necrosis factor-alpha (TNF-alpha), alanine aminotransferase and aspartate aminotransferase, and induced massive hepatic necrosis and lethality in 100% of control mice. Pretreatment with 1,8-cineole (400 mg kg(-1), p.o.) and dexamethasone (1 mg kg(-1), s.c.), 60 min before GalN/LPS, offered complete protection (100%) against the lethal shock and acute elevation in serum TNF-alpha and serum transaminases. Hepatic necrosis induced by GalN/LPS was also greatly reduced by both 1,8-cineole and dexamethasone treatment. The results indicate that 1,8-cineole protects mice against GalN/LPS-induced liver injury through the inhibition of TNF-alpha production, and suggest that 1,8-cineole may be a promising agent to combat septic-shock-associated pathologies.     

Antagonism of ethanol's central stimulation in mice by small doses of catecholamine-receptor agonists

Small doses of apomorphine (0.03–0.25 mg/kg i.p.) caused a dose-dependent suppression of the ethanol-induced (2.36 g/kg, 15% v/v, i.p.) locomotor stimulation in mice and higher doses (0.5–2 mg/kg i.p.) caused a delayed suppression. The delay increased with increased doses. Very small doses of clonidine (0.025 or 0.05 mg/kg i.p.), which per se did not or only slightly affect the activity of control mice, also markedly suppressed the ethanol-induced motor stimulation. Ethanol alone (2.36 g/kg i.p.) did not significantly affect the amount of Dopa accumulating in various mouse brain regions (limbic system, corpus striatum, hemispheres and brain stem) during 30 min following administration of 3-hydroxybenzylhydrazine (NSD 1015), an inhibitor of aromatic amino-acid decarboxylase. Both the hypothermia and the locomotor stimulation by ethanol were antagonized by NSD 1015. The reduction in Dopa accumulation induced by a small dose of apomorphine (0.25 mg/kg i.p.) in the dopamine-rich regions in the mouse brain was slightly enhanced by ethanol, whereas the clonidine-induced decrease in Dopa accumulation was, if anything, reduced.In conclusion, ethanol's behavioural stimulant action in mice can be largely suppressed by apomorphine or clonidine, drugs which in the small doses used probably inhibit central catecholamine (CA) neurons.     

Possible involvement of mu-opioid receptors in effect of lithium on inhibitory avoidance response in mice

In the present study, effects of intracerebroventricular (i.c.v.) injections of mu-opioid receptor agonist and antagonist on lithium state-dependency were investigated. For memory assessment, a one-trial step-down inhibitory avoidance task was used in adult male NMRI mice. Intraperitoneal (i.p.) administration of lithium (10 mg/kg) after training impaired memory when retrieval was tested 24 h later. The memory impairment was reversed by pretest administration of the same dose of lithium, suggesting state-dependency induced by lithium. In addition, i.c.v. administration of both lithium (2 and 4 microg/mouse, i.c.v.) and morphine (3 and 6 microg/mouse, i.c.v.) before the test reversed memory impairment induced by post-training lithium (10 mg/kg, i.p.). On the other hand, pretest administration of naloxone (1 and 2 mg/kg) which had no effects alone on inhibitory avoidance response, prevented the improving effects of both morphine (3 microg/mouse, i.c.v.) and lithium (2 microg/mouse, i.c.v.) on memory retrieval. The results suggest that the mu-opioid receptors in the central nervous system may be involved in the retrieval of lithium state-dependent learning.     

Inhibition and induction of drug metabolism by psoralens: alterations in duration of sleep induced by hexobarbital and in clearance of caffeine and hexobarbital in mice.

1. Hexobarbital (100 mg/kg i.p.) sleeping times in male CD-1 mice pretreated (-1 h) with a single i.p. injection of 150 mumol/kg of psoralen or coumarin analogues were increased, most markedly (6-fold) by linear, methoxy-substituted psoralens. 2. Hexobarbital sleeping times of mice which received three daily injections (231 mumol/kg; 50 mg/kg) of 8-methoxypsoralen (8-MOP) were 44% of controls (corn oil). 3. The whole-body half-life of caffeine (1 mg) in mice was 10.2, 1.2, and 0.37 h following 8-MOP (50 mg/kg per day) x 1, vehicle, and 8-MOP x 3 respectively. 4. The whole-body concentrations of hexobarbital (100 mg/kg dose) in mice 30 min after dosing were 14.3 +/- 0.9, 8.4 +/- 0.3, and 5.2 +/- 0.5 micrograms/ml (1 mouse = 150 ml) following 8-MOP (50 mg/kg per day) x 1, vehicle, and 8-MOP x 3 respectively. 5. It is concluded that, administered acutely, psoralen analogues inhibit hexobarbital metabolism in mice; and 8-MOP administered acutely inhibits the metabolism of caffeine and hexobarbital, but administered repeatedly increases their metabolism.     

Preventive effects of a verapamil against tumor necrosis factor-alpha-induced shock symptoms: approached from lipoprotein metabolic disorders

We examined the role of intracellular Ca2+ in the mechanism of the preventive effects of the Ca2+-channel blocker verapamil against lipoprotein disturbances during tumor necrosis factor (TNFa)-induced shock syndrome. The heparin-releasable lipoprotein lipase (LPL) activity in plasma of TNFalpha (5 X 10(4) units/mouse, i.v.)-injected mice was markedly lower at 4 h post-intoxication than in the controls. In mice treated with verapamil (10 mg/kg, s.c.), the activity of LPL 4 h after TNFalpha injection was significantly higher than in mice treated with TNFalpha alone. On the other hand, on polyacrylamide gel disk electrophoresis, very low density lipoprotein (VLDL) and high density lipoprotein (HDL) fractions in the sera of TNFalpha-injected mice were increased and reduced, respectively, relative to the controls. The administration of verapamil clearly prevented the lipoprotein damage arising from TNFalpha challenge. We investigated whether verapamil could suppress TNFalpha generation in endotoxin-treated J774A.1 cells. Treatment with verapamil (30 microM) markedly inhibited endotoxin (1 microg/ml)-induced TNFalpha production in these cells. These findings suggest that the concentration of intracellular Ca2+ may contribute to the extent of lipoprotein disturbances in plasma, which results from LPL suppression in TNFalpha-induced shock syndrome. Verapamil may, therefore, protect against some of the various disturbances caused by changes in Ca2+ mobilization through its ability to inhibit TNFalpha production in septic shock.     

Effects of the new eburnamenine derivative RU 24722 on EEG recovery and cerebral energy metabolism after complete ischemia

The influence of a new eburnamenine derivative RU 24722 [(3 beta, 14 alpha, 16 alpha)-(+/-)-14,15-dihydro-20,21-dinoreburnamenin -14-ol] on post-ischemic EEG recovery was studied in N2O anesthetized rats subjected to 1 min of global-compression cerebral ischemia. RU 24722 was compared with vincamine, dihydroergotoxine mesylate and nicergoline. Treatment with RU 24722 (2 mg/kg i.v.) significantly decreased the EEG recovery time and increased the electrocortical activity during the first phase of the post-ischemic recovery. Vincamine (2 mg/kg i.v.), dihydroergotoxine mesylate (0.5 mg/kg i.v.) and nicergoline (0.5 mg/kg i.v.) were devoid of activity. In an attempt to elucidate its mechanism of action, the influence of RU 24722 on changes in the cerebral metabolic energy reserves was studied in mouse brain after different periods of decapitation ischemia. The changes occurring during the first 10 s of ischemia were used to calculate the baseline cerebral metabolic rate (CMR). The activity of RU 24722 was compared with that of vincamine and pentobarbital. RU 24722 (10 mg/kg i.p.) significantly retarded glucose, phosphocreatine and adenosine triphosphate utilisation and lactate production. Vincamine (10 mg/kg i.p.) had no effect on cerebral energy substrates. Pentobarbital (100 mg/kg i.p.) markedly increased the tissue concentration of glucose and phosphocreatine and decreased lactate levels before and after ischemia. The improvement of EEG recovery suggests that RU 24722 may be therapeutically effective in cerebral insufficiency, and the decreased brain energy demand may be one of the mechanisms by which RU 24722 has a protective effect against cerebral ischemic damage.     

Pharmacological effects in mice of anandamide and its related fatty acid ethanolamides, and enhancement of cataleptogenic effect of anandamide by phenylmethylsulfonyl fluoride.

Anandamide (N-arachidonoylethanolamine) and six fatty acid ethanolamides were synthesized and their pharmacological effects in mice were assessed using catalepsy, hypothermia and pentobarbital-induced sleep prolongation as indices. The effects of phenylmethylsulfonyl fluoride (PMSF) pretreatment on anandamide effects were also evaluated and discussed in relation to inhibition of anandamide amidohydrolase in mouse brain and liver. The cataleptogenic effect of anandamide (ED50=6.0 mg/kg, i.v.) was 4 to 6 times more active than those of N-oleoyl- (ED50=26.5 mg/kg, i.v.) and N-linoleoylethanolamine (ED50=37.5 mg/kg, i.v.), although the peak time in the effect was observed within 1 min after i.v. administration. None of the saturated fatty acid ethanolamides (N-myristoyl-, N-palmitoyl-, N-stearoyl- and N-arachidoylethanolamine) showed a positive response in the cataleptogenic effect even at a dose up to 40 mg/kg i.v. Anandamide, N-linoleoyl-, N-oleoyl- and N-myristoylethanolamine (10 mg/kg, i.v.) produced a significant hypothermia (0.19 to 0.59 degrees C) at 5 to 15 min after administration. The duration of the effects of these ethanolamides was also relatively short. Anandamide, N-linoleoyl-, N-oleoyl- and N-palmitoylethanolamine (5 or 10 mg/kg, i.v.) significantly prolonged pentobarbital-induced sleeping time by 148-207% of control sleeping time. The cataleptogenic effect of anandamide was markedly potentiated by pretreatment of mice with PMSF (100 mg/kg, i.p.). The ED50 (mg/kg, i.v.) of anandamide was 0.48 (0.24-0.96) in PMSF-pretreated mice. The pretreatment of mice with PMSF significantly decreased the metabolic clearance rate of anandamide in microsomal fractions of liver and brain. Thus, the Vmax/Km values of brain and hepatic microsomes were 26 and 10%, respectively, as compared with those of control mice. The present study demonstrated that anandamide and N-acylethanolamines of unsaturated fatty acids exhibited cannabinoid-like effects in mice, and that anandamide amidohydrolase has an important role in the pharmacological effects of anandamide in vivo.     

Depressed isoprenaline vascular response in endotoxic rats

The injection of a sublethal dose of E. coli endotoxin (0127 B8) to intact rats (2 mg/kg i.v.) or adrenalectomized rats (0.01 mg/kg i.v.) depressed hypotensive responses to isoprenaline (0.2, 0.4, 0.8 μg/kg i.v.) for at least 4 h following endotoxin injection. These findings evidence an early (under 1 h after administration) and long-lasting (over 4 h) depression of the vascular response to isoprenaline in endotoxic rats. This depression was not related to catecholamine adrenal discharge or to the hypotensive and lethal effects of endotoxin.     

Effects of probenecid and acivicin on potassium dichromate-induced acute nephrotoxicity in mice.

Male OF1 mice were injected subcutaneously with 80 mg/kg potassium dichromate (K2Cr2O7). Examination of cryostat kidney sections stained for alkaline phosphatase (APP) revealed damage to about 40-70% of the proximal tubules after 8 h. Pretreatment with the organic anionic transport inhibitor probenecid (i.p., 3 x 0.75 mmol/kg) reduced the number of damaged tubules by 60% in mice treated with potassium dichromate. Pretreatment with the gamma-glutamyltranspeptidase (gamma-GT) inactivator acivicin (AT-125, 50 mg/kg p.o., plus 50 mg/kg i.p.) failed to prevent chromate-induced renal toxicity. These results support the conclusion that a probenecid-sensitive transport process, but not a gamma-GT-catalyzed degradation, is involved in the mouse renal toxicity of potassium dichromate.     

Central effect of histamine and peripheral effect of histidine on the formalin-induced pain response in mice

1. The present study was designed to investigate the role of brain histamine in modulating pain transmission in mice. 2. In conscious mice implanted with an intracerebroventricular (i.c.v.) cannula, the effects of i.v.c. injections of normal saline (control) and low and high doses histamine (2 and 40 microg/mouse, respectively) were investigated on the duration of paw licking and biting induced by subcutaneous (s.c.) injection of formalin (20 microL; 5%) into the plantar surface of the left hindpaw. 3. To clarify the involvement of histidine in the pain response, the effects of intraperitoneal (i.p.) injections of low and high doses of histidine (50 and 1000 mg/kg, respectively) alone or before i.c.v. injection of histamine were also examined. 4. Intraplantar injection of formalin induced a biphasic pain response (first phase: 0-5 min after injection; second phase: 20-40 min after injection). 5. Histamine (2 microg/mouse, i.c.v.) had no effect on the first phase of the pain response, but suppressed the second phase. The higher dose of histamine (40 microg/mouse, i.c.v.) suppressed both phases of the pain response. 6. Histidine, at 50 mg/kg, i.p., had no effect on the pain response, but the higher dose (1000 mg/kg, i.p.) suppressed the both phases of the pain response. 7. Pretreatment with the low dose of histidine (50 mg/kg, i.p.) prior to administration of 2 microg/mouse, i.c.v., histamine did not change the antinociception induced by low-dose histamine. However, pretreatment with the high dose of histidine (1000 mg/kg, i.p.) prior to 2 microg/mouse, i.c.v., histamine produced antinociception that resembled that seen following administration of the high dose of either histidine or histamine. Pretreatment with the low dose of histidine (50 mg/kg, i.p.) prior to administration of 40 microg/mouse, i.c.v., histamine has no effect on the pain response following high-dose histamine. Pretreatment with 1000 mg/kg, i.p., histidine prior to administration of 40 microg/mouse, i.c.v., histamine strongly suppressed both phases of the formalin-induced pain response, particularly the second phase. 8. The results of the present study indicate that: (i) activation of brain histamine produces antinociception in the mouse formalin test; (ii) peripheral loading with a high dose of histidine (1000 mg/kg, i.p.) alone exerts the same effect as that seen following 40 microg/mouse, i.c.v., histamine; and (iii) pretreatment with a high dose of histidine potentiates central histamine-induced antinociception.     

The Enhancing Effect of Tumour Necrosis Factor-α on Oxidative Stress in Endotoxemia

Abstract: The enhancing effect of tumour necrosis factor-α (TNF-α) on oxidative stress with or without a sublethal dose of endotoxin was examined. The mortality of mice treated with recombinant human TNF-α (1X10⁴ units/mouse, intravenously) and endotoxin (0.01-1 mg/kg, intraperitoneally) was dependent on the dose of endotoxin. The liver lipid peroxide level, superoxide anion generation and serum lactate dehydrogenase activity, especially serum lactate dehydrogenase-5 isozyme leakage, in mice 2-4 hr after administration of recombinant human TNF to endotoxin-pretreated mice (0.5 mg/kg, intraperitoneally) were markedly higher than in those without endotoxin, whereas the administration of recombinant human TNF significantly decreased the non-protein sulfhydryl level, superoxide dismutase and glutathione peroxide activities in the liver of endotoxin-injected mice compared with those in mice treated with recombinant human TNF or endotoxin alone. Furthermore, findings clearly demonstrated that J774A.1 cells stimulated with recombinant human TNF (1X10⁴ units/ml) can effectively produce nitric oxide in the presence of endotoxin, and the production was dependent on the dose of endotoxin (0.01-10 μg/ml). The level of lipid peroxide in mice 4 hr after administration of recombinant human TNF and lead acetate (50 mg/kg, intravenously) was markedly higher than that in the mice treated with recombinant human TNF alone. By contrast, injection of polymyxin-B (20 mg/kg, intraperitoneally, an anti-endotoxin drug) markedly decreased the lipid peroxide level in the liver of the mice treated with recombinant human TNF and lead acetate. These findings suggest that the oxidative stress caused by TNF occurs as a enhancing effect of endotoxion or by bacterial translocation from the intestinal gut under reduction of reticuloendothelial system function in various disease states, and that the effect of TNF may cause a marked increase of toxicity of oxidative stress by endotoxin.     

Scopolamine-induced convulsions in fasted mice after food intake: determination of blood glucose levels, [3H]glutamate binding kinetics and antidopaminergic drug effects

The present study was performed to evaluate the role(s) of hypoglycemia, changes in [(3)H]glutamate binding kinetics and dopaminergic activity in the occurrence of scopolamine-induced convulsions in fasted mice after food intake. Plasma glucose levels and density (B(max)) and affinity (K(d)) of [(3)H]glutamate binding sites in whole brain synaptic membranes were determined in animals fed ad lib or fasted for 48 h and treated intraperitoneally (i.p.) with 3 mg/kg scopolamine or saline and allowed to eat for 5 min. Fasting for 48 h decreased plasma glucose levels. After refeeding, plasma glucose concentrations increased in saline treated animals, but remained unchanged in scopolamine treated animals which consumed less food. Fasting for 48 h also produced significant changes in the kinetics of [(3)H]glutamate binding. The B(max) and K(d) of the binding sites decreased in fasted animals. These changes were partially antagonized by scopolamine treatment and food intake. For the evaluation of the contribution of dopaminergic activity, another group of mice fasted for 48 h and pretreated (i.p.) with saline or dopamine antagonists, 2 mg/kg chlorpromazine or 2 or 4 mg/kg haloperidol, were treated 10 min later with either saline or 3 mg/kg scopolamine. Then 20 min later, they were allowed to eat ad lib and were observed for 30 min for the incidence and onset of clonic convulsions. Pretreatment of both 2 mg/kg chlorpromazine and 4 mg/kg haloperidol markedly suppressed the convulsions. These results indicate that the decrease in the [(3)H]glutamate binding induced by fasting, its antagonism by scopolamine treatment and food intake, and the dopaminergic hyperactivity may be possible factors contributing to the occurrence of convulsions.     

Clonidine-induced hypoactivity and mydriasis in mice are respectively mediated via pre- and postsynaptic alpha 2-adrenoceptors in the brain

Since brain alpha 2-adrenoceptors occur both pre- and postsynaptically, experiments were carried out to determine the synaptic locations of those receptors mediating clonidine-induced hypoactivity and mydriasis. Intraperitoneal (i.p.) injection of clonidine (1-3000 micrograms/kg) to mice dose dependently induced these two responses and also decreased brain concentrations of 3-methoxy-4-hydroxyphenylglycol (MHPG). The ED50 values were: 120 micrograms/kg for hypoactivity (95% confidence limits 103-140 micrograms/kg), 54 micrograms/kg for mydriasis (95% confidence limits 40-74 micrograms/kg) and 18 micrograms/kg for MHPG reduction (95% confidence limits 8-36 micrograms/kg) suggesting that these responses could all be presynaptically mediated. However, methamphetamine which increases noradrenaline turnover was found to dose dependently produce mydriasis, but not hypoactivity, after peripheral (0.1-5 mg/kg i.p.) or central (0.5-10 micrograms i.c.v.) injection. The mydriasis produced by methamphetamine (0.5 mg/kg i.p.) was abolished by i.c.v. injection of 1 micrograms idazoxan or yohimbine, but not 2.5 micrograms prazosin or pindolol, showing this effect was mediated by central alpha 2-adrenoceptors. Methamphetamine (1-10 micrograms i.c.v.) potentiated the mydriasis induced by clonidine (50 micrograms/kg i.p.) suggesting this was a postsynaptic alpha 2-adrenoceptor response. By contrast, methamphetamine (1-10 micrograms i.c.v.) dose dependently reversed clonidine (100 micrograms/kg i.p.) hypoactivity indicating this response was mediated by presynaptic alpha 2-adrenoceptors. These hypotheses were confirmed by destruction of noradrenergic neurones using DSP-4 (100 mg/kg i.p. x 2). This treatment prevented the mydriasis response to methamphetamine (0.5 mg/kg i.p.), but not clonidine (100 micrograms/kg i.p.) and markedly attenuated clonidine (100 micrograms/kg i.p.) hypoactivity.     

Protective effect of the antioxidant 6-ethoxy-2,2-pentamethylen-1,2-dihydroquinoline (S 33113) in models of cerebral neurodegeneration

In a previous study Dorey et al. [Bio. Org. Chem. Lett., 10 (2000) 935] a series of novel dihydroquinoline compounds were developed, based on the potent antioxidant 6-ethoxy-1,2-dihydro-2,2,4-trimethylquinoline (ethoxyquin), and permitted the selection of the analogue 6-ethoxy-2,2-pentamethylen-1,2-dihydroquinoline (S 33113) lacking the hypothermic effects associated with ethoxyquin at equivalent doses. Herein, an extensive investigation of the neuroprotective capacity of S 33113 in different in vitro and in vivo paradigms of oxidative stress-mediated cellular degeneration was undertaken. In vitro S 33113 was a potent inhibitor (IC(50) = 0.29 microM) of Fenton-reaction-induced lipid peroxidation in mouse cortical membranes. Administration of S 33113 either intraperitoneally (< or =150 mg/kg i.p.) or orally (< or =600 mg/kg p.o.) did not significantly modify body temperature in NMRI mice. Furthermore, S 33113 (150 mg/kg i.p. or 600 mg/kg p.o.) markedly reduced the lethality induced by an intracerebroventricular injection of t-butylhydroperoxide in NMRI (naval medical research institute) mice for up to 5 h. Oral administration of S 33113, significantly attenuated alloxan-mediated hyperglycaemia in NMRI mice at 400 and 600 mg/kg (60%; P < 0.001). Administration of S 33113 (150 mg/kg i.p.) 30 min before transient global ischaemia significantly prevented delayed neuronal cell death in the CA1 region of the rat hippocampal formation, 7 days post-ischaemia (33% cell loss vs. 88% in ischaemia controls; P < 0.001). Similarly, a single pre-administration of S 33113 (150 mg/kg i.p.) prevented kainic acid-induced cell death in the CA3 hippocampal region at 7 days post-exposure (17% cell loss vs. 52% in kainate-treated controls; P < 0.01). Furthermore, D-methamphetamine-mediated dopamine depletion in the striatum of C57BL/6 mice (39-46%) was significantly prevented with S 33113 administered at either (2 x 150mg/kg i.p.) (11%; P < 0.01) or (2x150 mg/kg p.o.) (17%; P < 0.001). In conclusion, S 33113 represents a novel dihydroquinoline compound with potential for the treatment of cerebral pathologies implicating chronic neurodegeneration.     

Effect of nicotine-induced corticosterone elevation on nitric oxide production in the passive skin arthus reaction in rats

To elucidate the anti-inflammatory action of nicotine-induced corticosterone elevation on the passive skin Arthus reaction (PSAR), we investigated the inflammatory process in the PSAR. The polymorphonuclear leukocyte (PMNs) infiltration was observed just before as well as after elicitation by measuring extractable myeloperoxidase. The plasma exudation was significantly inhibited by anti-rat tumor necrosis factor (TNF)-alpha antibody (5 microg/site, i.d.) at the time of sensitization or by superoxide dismutase (52500 units/kg, i.p.) 1 h before elicitation or N(G)-nitro-L-arginine-methyl ester (100 mg/kg, i.v.) just at elicitation. Pretreatment with a single injection of nicotine (0.8 mg/kg, i.p.) 30 min before elicitation suppressed the plasma exudation but not the PMNs infiltration. This nicotine-induced decreasing effect was abolished in animals supplemented with L-arginine (300 mg/kg, i.v.) just at elicitation. The production of nitric oxide (NO) in peritoneal PMNs derived from an animal injected peritoneally with oyster glycogen was significantly suppressed by pretreatment with nicotine (0.8 mg/kg, i.v.) 30 min prior to harvesting. This inhibitory action of nicotine was abolished in animals pretreated with mifepristone (30 mg/kg, s.c.), a glucocorticoid receptor antagonist. These findings indicate that a single systematic administration of nicotine may attenuate the plasma exudation in the PSAR by suppressing the production of NO in the PMNs primed with TNF-alpha via nicotine-induced endogenous glucocorticoid.     

Role of reactive oxygen intermediates in the hepatotoxicity of endotoxin

Administration of endotoxin (2.5 micrograms/mouse, iv) to Corynebacterium parvum-pretreated (14 days earlier, 1 mg/mouse, i.v.) mice caused a rapid (90 min) decrease in liver cytochrome P450-dependent drug metabolism and an elevation of serum transaminase. The time course of the priming effect of C. parvum suggested that macrophages might be responsible for this sensitization to endotoxin. The antioxidant N-acetylcysteine (500 mg/kg) effectively protected against this depression of liver drug metabolism, thus supporting the hypothesis that liver macrophage-generated free radicals might mediate this hepatotoxic effect of endotoxin.