Increased serum nitrite/nitrate (NOx) and malondialdehyde (MDA) levels during alcohol withdrawal in alcoholic patients.

Serum nitrite/nitrate (NOx) and malondialdehyde (MDA) levels of 40 male alcoholic patients and 14 healthy male controls were investigated. Severity of alcohol withdrawal in alcoholic patients was evaluated by using the Clinical Institute Withdrawal Assessment for Alcohol (CIWA-Ar) scale just before taking the blood samples. NOx levels in serum samples were determined based on the reduction of nitrate to nitrite by vanadium chloride. MDA levels were also determined spectrophotometrically at 540 nm. The total CIWA-Ar score of alcohol-withdrawn patients was found to be 17.7. NOx and MDA levels were significantly increased in alcoholics during alcohol withdrawal as compared to control subjects. In conclusion, we observed increased serum NOx levels and lipid peroxidation during alcohol withdrawal in alcoholic patients.

Synthesis and antidepressant activities of some 3,5-diphenyl-2-pyrazolines.

Ten new 3,5-diphenyl-2-pyrazoline derivatives were synthesised by reacting 1,3-diphenyl-2-propen-1-one with hydrazine hydrate. The chemical structures of the compounds were proved by means of their IR, 1H-NMR spectroscopic data and microanalyses. The antidepressant activities of these compounds were evaluated by the 'Porsolt Behavioural Despair Test' on Swiss-Webster mice. 3-(4-Methoxyphenyl)-5-(3,4-dimethoxyphenyl)-2-pyrazoline, 3-(4-methoxyphenyl)-5-(2-chloro-3,4-dimethoxyphenyl)-2-pyrazoline and 3-(4-chlorophenyl)-5-(2-chloro-3,4-dimethoxyphenyl)-2-pyrazoline reduced 41.94-48.62% immobility times at 100 mgkg(-1) dose level. In addition, it was found that 4-methoxy and 4-chloro substituents on the phenyl ring at position 3 of the pyrazoline ring increased the antidepressant activity; the replacement of these groups by bromo and methyl substituents decreased activity in mice.

Agmatine potentiates the analgesic effect of morphine by an alpha(2)-adrenoceptor-mediated mechanism in mice.

The effects of agmatine, which is an endogenous polyamine metabolite formed by decarboxylation of L-arginine, and a combination of agmatine and morphine on tail-flick test have been investigated in mice. Adult male Swiss-Webster mice were used in the study. Agmatine (10, 20 and 40 mg/kg), clonidine (0.15 mg/kg), yohimbine (0.625 and 1.25 mg/kg), or saline were injected into mice intraperitoneally. Morphine (1 and 2 mg/kg) was given subcutaneously. Agmatine alone did not produce any significant change on radiant tail-flick latencies, but it potentiated significantly and dose-dependently morphine-induced (1 mg/kg) analgesia. The potentiating effect of agmatine (40 mg/kg) on morphine-induced analgesia was blocked completely by yohimbine (0.625 mg/kg), a selective alpha(2)-adrenoceptor antagonist, pretreatment. Clonidine (0.15 mg/kg), an alpha(2-)adrenergic receptor agonist, caused a significant increase of the tail-flick latencies of the mice. Yohimbine (0.625 mg/kg) also blocked clonidine-induced analgesia. In addition, yohimbine (0.625 mg/kg) was ineffective on the tail-flick test and did not produce any significant change on the morphine-induced analgesia. Our results indicate that cotreatment of agmatine with morphine produces antinociceptive enhancement via an alpha(2-)adrenergic receptor-mediated mechanism and agmatine-morphine combination may be an effective therapeutic strategy for medical treatment of pain.

L-NAME precipitates catatonia during ethanol withdrawal in rats.

The effect of N(G)-nitro-L-arginine methyl ester (L-NAME), a non-specific inhibitor of nitric oxide (NO) synthase, on catatonia in ethanol dependent rats was investigated. Ethanol was given to rats by a modified liquid diet. An isocaloric liquid diet without ethanol was also given to control rats. L-NAME (50, 100 and 200 mg/kg) and saline were injected intraperitoneally to ethanol-dependent rats 30 min before ethanol withdrawal. Then, catatonia was evaluated by vertical wire test at the 30th min, 2nd, 4th and 6th h of ethanol withdrawal. The injections were repeated 30 min before the observation of 6 h. Locomotor activity was also recorded for 5 min in the same observation intervals. L-NAME (200 mg/kg) or saline were also injected to ethanol non-dependent control rats. L-NAME (50 and 100 mg/kg) inhibited both incidence and intensity of the audiogenic seizures which appeared at 6 h of ethanol withdrawal. L-NAME (200 mg/kg) produced a significant augmentation in both incidence and intensity of the catatonia in ethanol dependent rats. This dose of L-NAME also reduced the locomotor activity of both ethanol dependent and non-dependent rats. The locomotor inhibitory effect was more prominent in ethanol-dependent group. The catatonia precipitating effect of L-NAME was not prevented by L-arginine (1 g/kg, i.p.), a NO precursor, pretreatment. In the naive rats, L-NAME also did not produce catatonia. These results indicate that L-NAME has a catatonia precipitating effect during ethanol withdrawal in rats and this effect seems to be independent from NO mediated mechanisms.

Investigation of extracellular L-citrulline concentration in the striatum during alcohol withdrawal in rats.

In this study, changes in striatal extracellular L-citrulline concentrations were investigated hourly for 5 h following alcohol withdrawal in chronic alcohol feeding Wistar rats. Alcohol (7.2% ethyl alcohol, v/v) was given to rats as modified liquid diet for 20 days. Signs of alcohol withdrawal appeared from the 1st h of alcohol withdrawal and the total alcohol withdrawal scores remained higher during the course of experiments. The mean of basal levels of L-citrulline in the microdialysis samples collected in conscious rat model from the striatum of control and alcoholized rats were found to be 1.28 +/- 0.48 microM and 0.35 +/- 0.08 microM, respectively. L-citrulline levels in the striatum of alcoholized rats increased by 4 folds significantly within 1 h following alcohol withdrawal. The increased striatal L-citrulline concentration was blocked by NG-nitro-L-arginine methyl ester (L-NAME; 60 mg/kg), a nitric oxide synthase inhibitor, pretreatment. Our results indicate an increased L-citrulline level in the rat striatum during early alcohol withdrawal and this situation may be related to an increased nitric oxide production.

Effects of chronic ethanol administration on serotonin metabolism in the various regions of the rat brain.

Changes in serotonin (5-HT) and 5-hydroxy indole acetic acid (5-HIAA), its major metabolite, in cerebral cortex, corpus striatum and hippocampus were investigated at 10th and 21st days of chronic ethanol ingestion in Wistar rats. Ethanol (7.2% v/v) was given to rats in a modified liquid diet. Biochemical analysis was performed in two groups of ethanol-treated and control rats (n = 6 for each group). Rats in each group were decapitated at the 10th and 21st days of ethanol consumption. Brains were removed and cerebral cortex, corpus striatum and hippocampus were dissected. 5-HT and 5-HIAA levels were measured in respective brain regions by using high performance liquid chromatography. In cerebral cortex and corpus striatum, 5-HT levels were significantly lower than control at the 10th day of ethanol consumption. At the 21st day, the levels tended to remain low, but not significantly different statistically. In hippocampus, 5-HIAA levels were significantly higher than control at 10th day of ethanol consumption. Increased 5-HIAA level returned to control values at the 21 st day of ethanol consumption. Our results suggest that, 5-HT clearly seems to play a critical role in the brain at the 10th day of chronic ethanol consumption.

Effects of NMDA receptor blockers on cocaine-stimulated locomotor activity in mice.

Effects of MK-801 and ketamine, N-methyl-D-aspartate (NMDA) receptor blockers, on cocaine-stimulated locomotor activity were investigated in male Swiss-Webster mice. MK-801 (0.25, 0.5, 1.0 and 2.5 mg/kg), ketamine (10, 25 and 50 mg/kg) or saline was injected 20 min before cocaine (5, 10 and 20 mg/kg i.p.). Locomotor activity was measured for 30 min immediately following cocaine treatment. All doses of the drugs were also tested for ability to depress or stimulate locomotor activity in the naive (no cocaine-treated) mice. Cocaine produced a dose-dependent increase in locomotor activity that was blocked dose-dependently by MK-801 or ketamine. The blockade by MK-801 was more prominent than by ketamine. Our results may suggest that cocaine-induced locomotor stimulation in mice is modulated via NMDA receptor mediated mechanisms.

Effects of agmatine on ethanol withdrawal syndrome in rats.

Effects of agmatine, which is an endogenous polyamine metabolite formed by decarboxylation of L-arginine, have been investigated on the ethanol withdrawal syndrome in rats. Adult male Wistar rats were used in the study. Ethanol (7.2% v/v) was given to the rats by a liquid diet for 21 days. Agmatine (20, 40, 80 and 160 mg/kg) and saline were injected to rats intraperitoneally 30 min before ethanol withdrawal testing. After 30th min, 2nd and 6th h of ethanol withdrawal, rats were observed for 5 min, and withdrawal signs which included locomotor hyperactivity, agitation, stereotyped behavior, wet dog shakes and tremor were recorded or rated. A second series of injections was given at 6 h after the first one, and subjects were then tested for audiogenic seizures. Agmatine caused dose-dependent and significant inhibitory effects on stereotyped behaviors, wet dog shakes and tremors during the observation period. It did not cause any significant change in motor coordination of naive (not ethanol-dependent) rats. Our results suggest that agmatine attenuates withdrawal syndrome in ethanol-dependent rats; thus, this drug may be beneficial in the treatment of ethanol dependence.

Nitric oxide synthase inhibition blocks amphetamine-induced locomotor activity in mice.

Effects of N(G)-nitroarginine methyl ester (L-NAME), a nonspecific inhibitor of nitric oxide (NO) synthase, on amphetamine-induced locomotor activity were investigated in Swiss-Webster mice. Locomotor activity was measured for 30 min immediately following amphetamine (1, 2 and 4 mg/kg, i.p.) or saline treatments. L-NAME (15 and 30 mg/kg) and a combination of L-arginine (1000 mg/kg) and L-NAME (30 mg/kg) were injected 30 min before amphetamine (2 mg/kg) to other groups of the mice. L-Arginine was injected 30 min before L-NAME treatment when they were combined. L-NAME (30 mg/kg) and L-arginine (1000 mg/kg) were also tested for ability to depress or stimulate locomotor activity in the absence of amphetamine. Amphetamine caused a dose-dependent increase in locomotor activity of the mice. L-NAME blocked the amphetamine-induced locomotor stimulation dose dependently. L-Arginine pretreatment prevented the inhibitory effects of L-NAME on amphetamine-induced locomotor stimulation. L-NAME and L-arginine did not cause any significant change in locomotor activity in mice not treated with amphetamine. These results suggest that amphetamine-induced locomotor stimulation in mice is modulated by NO.

Lack of tolerance to ethanol-induced motor impairment on accelerod performance in rats.

The effect of ethanol on rats was investigated at increasing rates of acceleration for bar rotation speed. Ethanol was given to rats by a liquid diet starting with 2.4% ethanol (v/v) for 3 days. Then the ethanol concentration was increased to 4.8% (v/v) for 3 days and finally to 7.2% (v/v) for 15 days. Accelerod performance was recorded before and throughout 20 days of ethanol intake. Mean blood ethanol levels were 266.34+/-13.11 and 285.20+/-9.77 mg/dl on the 7th and 15th days of ethanol (7.2% v/v) consumption, respectively, as measured in a parallel group of animals. Ethanol produced significant concentration-dependent impairments in the accelerod performance of rats. The motor impairment effect of ethanol was most prominent in the test using the greatest rate of acceleration (from 0 to 79 rpm within 2 min). The impairment effect of ethanol on accelerod performance occurred throughout the period of ethanol exposure. Our results indicate that motor impairment on the accelerod performance test produced by an ethanol liquid diet depends on the concentration of ethanol and the rate of acceleration. In addition, under free-access conditions accelerod performance may not be a suitable behavioral test for detecting tolerance development to ethanol in rats.

Analgesic effect of tianeptine in mice.

The effects of tianeptine, a novel and unusual tricyclic antidepressant drug, on tail-flick and hot-plate tests, which are two thermal analgesia evaluating methods, have been investigated in mice. Tianeptine (5 and 10 mg/kg), para-chlorophenylalanine (pCPA) (100 mg/kg) and a combination of pCPA and tianeptine (10 mg/kg) or saline were injected to mice intraperitoneally. pCPA (100 mg/kg) was injected 24 h before tianeptine or saline treatment when it was combined with tinaeptine (10 mg/kg) or tested alone. The tail-flick latencies and hot-plate reaction times of the mice were measured between 15th and 180th minutes following injections. Tianeptine (10 mg/kg) exhibited a significant antinociceptive activity that could be measured by both tests as compared to groups which were treated with saline or pCPA alone between 15th and 180th min of the observation period. The lower dose of tianeptine (5 mg/kg) or pCPA (100 mg/kg) did not produce any significant changes on tail-flick latency or hot-plate reaction time of the mice. However, pretreatment with pCPA completely blocked the antinociceptive effect induced by tianeptine (10 mg/kg) in both tests used in the present study. Furthermore, tianeptine (10 mg/kg) did not cause any significant impairment effects on rotarod performance of the mice. Our results suggested that tianeptine has a prominent thermal antinociceptive activity in mice and that increased serotonergic activity may be responsible for the analgesic effect of tianeptine.

The effects of flumazenil on two way active avoidance and locomotor activity in diazepam-treated rats.

The present study was undertaken to determine the effects of chronic flumazenil treatment alone and simultaneously with diazepam on acquisition performance in an active-avoidance task and on locomotor activity in rats. Flumazenil (5, 10 and 20 mg/kg) and diazepam (0,5, 1.0 and 2.0 mg/kg) were administered intraperitoneally to rats before each daily training session for 5 days. The baseline of avoidance performance was set to approximately 50% and responses were expressed as acquisition rate. Locomotor activity of the rats was simultaneously recorded but only following the first training session. Diazepam decreased acquisition rate between the dose range used. Flumazenil had no effect on the acquisition rate of naive rats but reversed low dose diazepam-induced learning and memory impairment. Diazepam induced locomotor depression within the same dose range that decreased acquisition rate. Flumazenil had no effect on locomotor activity, but reversed the locomotor depressant effect of diazepam. The striking contradiction with previous data that flumazenil has no effect on learning-memory processing is discussed.

Gamma-vinyl-GABA potentiates the severity of naloxone-precipitated abstinence signs in morphine-dependent rats.

Effects of gamma-vinyl-GABA (GVG), an antiepileptic drug that inhibits GABA transaminase and increases extracellular GABA concentrations in the brain, were investigated on the morphine abstinence syndrome (AS) in male Wistar rats. Two morphine pellets (75 mg morphine base in each) were implanted subcutaneously on the back of the rats. Seventy-two hours after the morphine implantation, naloxone (NL, 2 mg kg-1) was injected intraperitoneally (i.p.) to induce precipitated morphine AS. GVG was administered at the doses of 250 mg kg-1 (n = 11) and 500 mg kg-1 (n = 11) i.p. 24 h prior to AS and at the dose of 500 mg kg-1 (n = 13) i.p. 6 h prior to AS. Immediately after NL injections, rats were observed for 5 min and AS signs (jumping, teeth chattering, wet dog shake, diarrhoea, ptosis and defecation) were assessed. The behavioural signs of GVG-treated rats were compared with the control groups (n = 10) during the AS. Jumping, wet dog shake, teeth chattering were found to be significantly increased in all of the GVG-treated groups. Ptosis was found to have increased in only 500 mg kg-1 GVG groups. GVG potentiated the severity of morphine AS signs. GVG does not seem to have any therapeutic potential for treatment of morphine abstinence unlike some other drugs that enhance GABAergic transmission.

Dopaminergic and serotonergic alterations in the rat brain during ethanol withdrawal: association with behavioral signs.

Changes in dopaminergic and serotonergic levels and metabolites in cerebral cortex, corpus striatum and hippocampus were investigated during the first 6-h of withdrawal in ethanol-dependent Wistar rats. Ethanol was given by a liquid diet for 21 days. The concentration of ethanol was 7.2% (v/v) for the last 15 days of the exposure. After 2, 4 and 6 h of ethanol withdrawal, and after audiogenic stimulus (100 dB for 60 s) at 6 h of ethanol withdrawal, various brain regions were assayed for levels of dopamine (DA), DOPAC, HVA, serotonin (5-HT) and 5-HIAA. Behavioral signs of ethanol withdrawal and blood ethanol levels were also evaluated in other parallel groups of ethanol-dependent rats. Significant decreases in 5-HT levels and significant increases in HVA levels in striatum were found during the first 6 h of ethanol withdrawal and after the audiogenic seizures. In hippocampus, 5-HIAA levels were significantly reduced after 2 h of ethanol withdrawal and after the audiogenic seizures. 5-HIAA levels significantly increased after 2 h of ethanol withdrawal in cerebral cortex. Significant increases in both DA and 5-HT levels were also found in cerebral cortex after the audiogenic seizures. The results suggest that the levels of DA, 5-HT and their metabolites are altered by ethanol withdrawal. Furthermore, this may suggest that DA and 5-HT may be involved in the first 6 h of ethanol withdrawal syndrome in rats.

Inhibitory effect of agmatine on naloxone-precipitated abstinence syndrome in morphine dependent rats.

Effects of agmatine, which is an endogenous polyamine metabolite formed by decarboxylation of L-arginine, were investigated on the morphine abstinence syndrome in rats. Two pellets containing 75 mg morphine base (total 150 mg) were implanted subcutaneously on the back of rats. Seventy-two hours after morphine implantation, agmatine sulphate (20, 30 and 40 mg/kg) or saline was injected intraperitoneally. Forty-five min later, naloxone (2 mg/kg) was injected intraperitoneally to induce precipitated withdrawal. Immediately after naloxone injection, rats were observed for 15 min, and abstinence syndrome signs, which included jumping, wet dog shake, writhing, defecation, ptosis, teeth chattering and diarrhea were counted or rated. Agmatine attenuated all of the signs of the morphine abstinence syndrome dose dependently and significantly. Our results suggest that agmatine prevents naloxone-precipitated abstinence syndrome in morphine dependent rats; thus, this drug may be beneficial in the treatment of opioid dependence.

Nitric oxide synthase inhibition attenuates signs of ethanol withdrawal in rats.

The effects of N(G)-nitro arginine methyl ester (L-NAME) and 7-nitroindazole (7-NI), nitric oxide synthase inhibitors, and L-arginine, a nitric oxide precursor, on ethanol withdrawal signs were investigated in rats. Ethanol (7.2% v/v) was given to rats by a liquid diet for 16 days. L-NAME (30 and 60 mg/kg), 7-NI (40 and 80 mg/kg), L-arginine (100 mg/kg), a combination of L-arginine (100 mg/kg) and 7-NI (40 mg/kg), and saline or vehicle were injected to rats intraperitoneally 30 min before ethanol withdrawal. A second series of injections was given at 6 hour after the first one, and subjects were then tested for audiogenic seizures. 7-NI (40 mg/kg), vehicle and saline were also administered to naive rats. 7-NI (40 mg/kg) did not produce any significant change in locomotor activity in naive rats. Both L-NAME and 7-NI significantly inhibited locomotor hyperactivity from the 2nd to the 6th hour of the withdrawal period. They also reduced the total ethanol withdrawal score from the 30th min to the 6th hour, and they significantly decreased audiogenic seizures. Neither drug increased locomotor activity nor total ethanol withdrawal score, which were increased significantly by L-arginine (100 mg/kg); however, L-arginine (100 mg/kg) prevented the inhibitory effects of 7-NI (40 mg/kg) on increased locomotor activity, total ethanol withdrawal score, and audiogenic seizure. Our results suggest that nitric oxide synthase inhibition by L-NAME and 7-NI alleviates the signs of ethanol withdrawal. The data also support the hypothesis that nitric oxide may take part in the neuroadaptation that develops during chronic ethanol ingestion in rats.

Heterogeneity of tolerance developed to effects of ethanol on rotarod and accelerod performances in rats.

The ethanol-induced impairment of rotarod and accelerod performances has been investigated in rats. Ethanol (7.2%, v/v) was given orally ad lib to rats in a modified liquid diet for 15 days. Rotarod and accelerod performances were recorded before and at the 2nd, 4th, 7th, and 15th days of ethanol intake. The daily ethanol consumption of the rats ranged from 12.03 to 16.4 g/kg. Mean blood ethanol level was estimated as 282.3 and 242.5 mg/dl on the 7th and 15th days of ethanol consumption, respectively. Ethanol significantly decreased (p < 0.01 rotarod performance on the 2nd and 4th days. But tolerance developed to the ethanol-induced impairment of rotarod performance from the 7th day. It also significantly decreased (p < 0.01) accelerod performance of the rats throughout the 15 days. So no tolerance was seen to this action of ethanol. Our results suggest that rotarod and accelerod performance tests seem to have differential characteristics in the context of tolerance development to ethanol in rats.

Investigation of the effects of moclobemide in chronic ethanol feeding rats.

Effects of chronic moclobemide administration on ethanol consumption, blood ethanol level, motor coordination and seizures induced by an audiogenic stimulus during ethanol withdrawal syndrome have been investigated in chronically ethanol intoxicated rats. Adult male Wistar rats (211-289 g) were used. Ethanol (7.2% v/v) was given to rats by a special liquid diet for 15 days. Moclobemide (10, 30 and 45 mg kg-1) or saline was injected to ethanol-feeding rats subcutaneously through 15 days. Daily ethanol intake and body weight gain of the rats were recorded. Blood ethanol levels were measured in rats given saline or chronic moclobemide (30 mg kg-1) just before and 6 h after ethanol withdrawal. At the sixth hour of the ethanol withdrawal, the incidence, intensity and latency of the seizures induced by an audiogenic stimulus were recorded in saline and acute (30 mg kg-1) or chronic (10, 30 and 45 mg kg-1) moclobemide-administered rats. Accelerod performances of the chronic moclobemide (45 mg kg-1) or saline injected rats were also evaluated. The mean ethanol intake of the rats ranged from 10.06 to 15.63 g kg-1 day-1 during the study. Chronic moclobemide treatment did not occur any significant effect on daily ethanol consumption, ethanol clearance and accelerod performances of the rats. No significant changes on the incidence, intensity and the latency of the audiogenic seizures were also observed. Our results suggest that there has been no adverse interaction between ethanol and moclobemide.

Effects of flumazenil on ethanol withdrawal syndrome in rats.

Effects of flumazenil (Ro 15-1788, CAS 78755-81-4) on ethanol withdrawal syndrome (EWS) has been investigated in rats. Behavioral EWS symptoms appeared during the first 6 h of ethanol withdrawal. Flumazenil (2.5 and 10 mg/kg i.p.) increased horizontal and vertical locomotor activity significantly and also precipitated abnormal gait and agitation at the beginning of EWS in a dose dependent manner. However, thereafter it reduced the severity of abnormal posture and gait, tail stiffness, agitation and stereotyped behavior in a dose dependent manner. At the 6th hour of EWS, flumazenil (10 mg/kg) reduced total EWS score significantly, but shortened the latency of audiogenic seizures and increased the severity of wet dog shakes. Flumazenil (2.5 and 10 mg/kg) did not elicit behavioral EWS symptoms and audiogenic seizures in non-dependent (control) rats. It did not cause any significant change on locomotor activities in these groups. According to those results, certain actions of flumazenil on the experimental EWS may suggest a potential beneficial effect of this drug in the treatment of EWS in alcoholics, but its enhancing effects on some behavioral EWS symptoms and a potential proconvulsant activity may be a drawback for its use in the treatment of EWS.

A modified liquid diet of chronic ethanol administration: validation by ethanol withdrawal syndrome in rats.

The validity of a modified liquid diet prepared from cow's milk, 7.2% ethanol, vitamin A and sucrose mixture has been investigated for chronic ethanol administration in rats. The rats fed by the modified liquid diet lost 3.8% of their body weight along 20 days. The mean ethanol consumption was in the range of 12.6 to 17.6 g kg-1 day-1. The blood ethanol level just before the withdrawal of ethanol was found to be 301.38 +/- 36.68 mg dl-1. A severe ethanol withdrawal syndrome was observed after 20-day exposure to ethanol in the modified liquid diet. The results suggest that the modified liquid diet can be used satisfactorily for chronic ethanol administration in rats.