OBJECTIVE Toestablisharecombinantcelllinethatcanstablyexpressratpurinergic P2receptorP2X4(rP2X4R).METHODS ToconstructgreenfluorescentproteinandrP2X4recombinant plasmid (pEGFP-N1-rP2X4),lipofectamine was used to transfect pEGFP-N1-rP2X4 into human embry-onic kidney (HEK293)cells that were screened with G41 8 (1 g·L-1 ).The quantitative expression of rP2X4 receptor was verified by qRT-PCR and Western blotting analysis.Whole-cell patch clamp record-ing was used to investigate the function of the stably expressed rP2X4 receptor. RESULTS The sequence of plasmid pEGFP-N1-rP2X4 was verified by PubMed Blastn comparison.qRT-PCR and Western blotting analysis demonstrated that the expression of rP2X4 receptor in HEK293-pEGFP-N1-rP2X4 cell lines remained stable after 25 generations (P1 ,P3,P5,P10,P15,P20 and P25).Whole-cell patch clamp recording experiments showed that the rP2X4 receptor agonist,purine-5′-triphosphate (ATP,3.0 μmol·L-1 ),could activate rP2X4 receptors in HEK293-pEGFP-N1-rP2X4 cell lines.Specific activating current could be blocked by non-selective rP2X4 receptor antagonist TNP-ATP (30.0μmol·L-1).CONCLUSION rP2X4receptorisstablyexpressedinHEK293-pEGFP-N1-rP2X4cell line and maintains stable expression and function within 25 continuous generations.The establish ment of HEK293-pEGFP-N1-rP2X4 cell line can contribute to further investigations of the roles of rP2X4 receptors in neuropathic pain.

Aims To establish several behavioral paradigms to characterize the psychotropic effects of hallucinogens which ze-brafish was utilized as a model animal, and then to investigate the effects of potent hallucinogenic drugs on these models. Methods With the video record and track system, the behavior was recorded and quantified automatically. In the experiments, the bottom dwelling test, social behavior and mirror test were performed to test the hallucinogenic effects of drugs. Metham-phetamine (METH, 2 mg·L-1) and ketamine (20 mg·L-1) were selected as experimental challenges. The 30 min pre-treat-ment time was chosen based on our prior experience in zebrafish models. Results Compared to the normal group, in dwelling test, acute exposure of zebrafish to METH and ketamine de-creased transitions significantly, and in mirror reflection test, the drug-treated fish changed the preference for mirror zone, and ex-hibited aggressive for their mirror images. The pretreatment of METH and ketamine significantly reduced the contact durations, and the ketamine inhibited the contact frequency each other, the results indicated that the social interaction of zebrafish was im-paired. Conclusion The results confirm high sensitivity of ze-brafish models to hallucinogenic compounds with complex behav-ioral and physiological effects.

As an important messenger molecule, NO exists in both peripheral and central nervous system (CNS). Recently, NO was proved to play a critical role in CNS, especially in synaptic plasticity. As a kind of retrograde messenger, it plays some roles in the induction and maintenance of LTP through activation of downstream pathway or molecules, such as NO-cGMP-PKG, ADPRT and C-FOS.

Opioid peptides and their receptors play an important r ole in the pathophysiologic process of stroke. The concentration of endogenous o pioid peptides elevates, while the density of opioid receptors decreases in CNS during cerebral ischemia; on the other hand, opioids influence the development o f stroke through regulating the contraction of cerebral vessel, immuno-endocrin e system and neuroprotection against injury. Thus, drugs that regulate the opioi d system may be a new therapeutic approach for prevention and treatment of cereb ral ischemia.

25% signified analgesic effect. Part II (the effect of agmatine on analgesic effect of morphine). Fifty-six animals with neuropathic pain were randomly divided into 4 groups ( n = 14 each) . Each group received NS or agmatine 20, 40 or 60 mg? kg-1 . 30 min later each group received either NS or morphine 1 mg?kg-1(n=7 each). PWT was measured and PMAP was calculated Part III (the effects of agmatine on the development of morphine tolerance). Morphine tolerance was incuced by morphine 10 mg?kg-1, 3 times a day for 5 days. Thirty-five animals with neuropathic pain were randomly divided into 5 groups ( n = 7 each): group A received NS: group B received NS + morphine; grouop C, D, E received agmatine 5, 10, 20 mg ? kg-1 + morphine 3 times a day for 5 days. PWT was measured on the 5th day and PMAP was calculated. Results PMAP of agmatine (40, 60 mg?kg-1 i.p.) and morphine (1 mg?kg-1 s.c.) were lower than 40% . Agmatine 60 mg?kg-1 increased the PMAP of morphine 1 mg?kg-1 to 84% compared with 34% for morphine alone (P

Psychostimulant is a type of psychoactive substances that stimulate central and peripheral nervous system as well as induce drug dependence.A series of studies indicate that monoamine especially dopamine plays an important role in behavior and drug dependence,moreover,dopamine transporter(DAT)controls homeostasis of dopamine in neuron and transmission of dopamine pathways.Thus DAT might play an important role in the reward and behavioral stimulation of psychostimulant.

AIMTo further elucidate the role of agmatine on the pharmacological effects of opioids. METHODSThe effect of L-arginine and L-arginine decarboxylase(L-ADC) antibodies on pain threshold, morphine ntinociception and tolerance were investigated in mouse acetic acid writhing test, mouse radiant heat tail flick test and mouse hot plate test. RESULTSIn mouse acetic acid writhing test, intracerebroventricular injection of L-arginine dose-ependently inhibited the writhing of mice compared with saline control. L-arginine did not influence the tail flick latency itself in mouse radiant heat tail flick test, but enhanced antinociceptive effect of morphine in a dose-dependent manner. The possible maximal analgesia percentage of morphine 2.5 mg*kg-1 was increased from 23% to 71%. Furthermore, L-arginine inhibited acute tolerance induced by morphine 100 mg*kg-1in mouse radiant heat tail flick test. The effect of L-arginine as mentioned above could be antagonized by idazoxan (3 mg*kg-1, ip), which is a selective antagonist of imidazoline receptors. L-ADC specific antibodies inhibited morphine antinociception and promoted the development of tolerance to morphine in mouse radiant heat tail flick test and 55℃ hot plate test. CONCLUSIONL-Arginine and L-ADC play important roles in the formation of pain threshold, morphine antinociception and tolerance.

Schizophrenia, described as the worst disease affecting mankind, is a severe and disabling mental disorder. Schizophrenia is characterized by complicated symptoms and still lacks a diagnostic neuropathology, so developing schizophrenia animal models which have quantifiable measures tested in a similar fashion in both humans and animals will play a key role in new therapeutic approaches. According to the symptoms of cognitive impairment and emotional disorder, the N-methyl-d-aspartate (NMDA)-receptor antagonist MK-801 was applied to induce schizophrenia-like behavior in mice. Locomotor activity and prepulse inhibition (PPI) were selected as indices and the effect of clozapine was also investigated in this model. The results showed that compared with the normal group, MK-801-treated mice exhibited significantly increased locomotor activity and impaired PPI, and pre-exposure to clozapine could ameliorate the abnormality and make it back to normal level. These findings suggest that the model we established could be a useful tool for antipsychotic drug screening.

OBJECTIVE To analyze impulsive-like behaviors of SD rats induced by pramipexole in Y-maze avoidance tasks. METHODS Behaviors of SD rats in Y-maze avoidance tasks were recorded with a camera and analyzed by Noldus Etho Vision XT8 software after acute subcutaneous injection of pramipexole(0.1,1 and 10 mg · kg-1),including right reaction numbers of 20 consecutive avoidance tasks,shuttle number of times between the three arms of Y-maze, distance covered in Y-maze and time spent in safe arms during 20 consecutive avoidance tasks. Then,the prepulse inhibition(PPI)of the startle reflex test was used to assess the effect of pramipexole on sensorimotor gating (SG). Effects of pramipexole on the dialyzed content of monoamine neurotransmitter and its metabolites in the striatum and amygdala of SD rats were measured by microdialysis in vivo. RESULTS Compared with normal control group,the rats of pramipexole group showed a significant increase in the shuttle number of times and distance covered in Y-maze between Y-maze avoidance tasks(P<0.01),but a statistically significant decrease in the time spent in safe arms(P<0.01),while the number of right reactions in Y-maze avoidance tasks was not changed. Such premature responses were quite similar to certain impulsive-compulsive behaviors in rodent models,such as five-choice serial reaction time tasks. In the PPI test,pramipexole displayed an impairing effect on SG(P<0.01). The microdialysis results showed that there was an increase of dopamine and 5-hydroxytryptamine in the striatum of pramipexole group, but not statistically significant. Monoamine neurotransmitters and their metabolites were not significantly changed in the amygdala. CONCLUSION Pramipexole can induce impulsive-compulsive behaviors in Y-maze avoidance tasks,which might be attributed to impaired SG.

Opioid receptors, as an important member of G protein coupled receptors (GPCR), are the binding targets of endogenous opioid peptides and exogenous opiates. The activation of opioid receptors influences the nervous system, immune physiology and endocrine system. However, prolonged activation of opioid receptors is likely to produce opioid tolerance, leading to opioid addiction. Receptor endocytosis and sorting into the recycling pathway contribute to recovery of cellular opioid responsiveness. Recent studies have revealed that GPCR can be modulated by ubiquitination which plays a unique roles in governing GPCR trafficking. Moreover, ubiquitination of the opioid receptors (μ, κand δ) is increased after stimulation of most opioid agonists. Mutation of the ubiquitin sites affects the internalization and degradation of opioid receptors, which contributes to changes in signal pathways and regulation of opioid receptors. ln this paper, ubiquitination of opioid receptors and the fundamental role of ubiquitination in trafficking of opioid receptors are reviewed.