The Changes Of Amino Acids Levels In Brain Regions Of Morphine Dependent And Relapsed Rats

Objective: To investigate the changes of amino acids levels in brain regions of morphine psychological dependent and relapsed rats by using conditioned place preference paradigm and footshock priming.Methods: Sixty-four male Sprague Dawley rats were randomly assigned to one of seven groups with 8-11 animals in each. Rats received conditioned place preference (CPP) training as follows: they were placed in drug-pairing room of CPP apparatus for 5 minutes before being injected intraperitoneally (ip) with saline (1 ml/kg, two groups) or morphine hydrochloride (4 mg/kg, five groups), and then were placed back for 25 minutes. The training was performed once a day for 10 days. Twenty-four hours after the last training, rats were given free access to both rooms of the, apparatus for 10 min (CPP test). Total time (min) the rats spent in the drug-pairing room was recorded as the result of the test. CPP was established when morphine-trained rats spent significantly longer time in the drug-pairing room than that saline-trained rats did, and it was considered that the psychological dependence had developed. After the CPP test, two groups of rats (one saline-trained group and onemorphine-trained group) were decapitated (dependent groups). The seven days after the last training was the period of spontaneous extinction. Rats were not trained in this peroid and were given another CPP test at the seventh day. We considered that the CPP and psychological dependence extincted if there was no significant difference between morphine- and saline-trained groups in CPP test. Constant-current, intermittent, inescapable footshock (FS, an effective form of stress) was used as the priming stimuli for relapse. CPP test was performed right after FS or placement in the footshock chamber (no current). CPP was reinstated if morphine-trained rats spent more time in drug-pairing room than saline-trained rats did. Then two groups of morphine-trained rats (after FS and placement respectively) were decapitated. CPP test was performed again 1 hour and 4 hours after FS respectively to investigate the maintenance of the reinstatement. The other two morphine-trained groups were decapitated immediately after the 1 hour and 4 hour test respectively.The groups in which rats were decapitated after FS were named relapsed groups.Brain tissues were dissected out and homogenized. Proteins were precipitated and pelleted by centrifugation. The supernatants were frozen for detection. The nucleus we selected were amygdala, frontal cortex, hippocampus, hypothalamus, necleus accumbens , striatum, thalamus and cerebellum. FTPLC, with precolumn derivatization andelectrochemical detector was used to quantitate the contents of glutamic acid (GLU) and Y -aminobutyric acid (GABA) in each nucleus. Data of CPP test and amino acids contents of dependent groups were analyzed with t test. Data of CPP test 1 hour and 4 hours after FS were analyzed with / test too. Data of amino acids contents and CPP test after training, extinction and right after FS of relapsed groups were analyzed with one-way analysis of variance (ANOVA) followed by Student-Newman-Keuls test.The accepted level of statistical significance was P<0.05.Result: After training with morphine for 10 days, the time rats spent in the drug-pairing room was significantly more than that of saline trained rats (F<0.05).The contents of both GLU and GABA significantly decreased in frontal cortex, hypothalamus and nucleus accumbens in brain of trained rats, especially in nucleus accumbens (P < 0.05). While the ratio of GLU/GABA significantly increased only in nucleus accumbens (P<0.05).There was no significant difference in CPP test results between morphine-trained and saline-trained rats after extinction, it was not the case after FS. The time morphine-trained and footshocked rats spent in drug-pairing room was longer than that saline-trained and footshocked rats did (P< 0.05), but not that morphine-trained and nonfootshocked rats did. Compared to saline-trained rats the difference in CPPte...