| Abusage and prevalence of drugs and dopes is one of the important social problems. Up to now, the mechanism of tolerance, addiction, and withdrawal of opioid is not thoroughly clear, so there is no effective approach to radical cure the addition of it.The purpose of our study is to reveal effect of heroin on gene expression of iNOS and COX-2 on rat C6 glioma,which may further provide evidences for the mechanism of tolerance, addiction, and withdrawal of opioid.NO is synthesized from L-arginine by the NO synthase (NOS), together with a stoechionetric production of L-citrulline. The reaction requires molecular oxygen. It is getting more and more attribute importance to the relationship between morphine dependence and NOS. Nitric oxide synthase (NOS) is the key enzyme to catalystic and synthesize NO in vivo. NOS was documented with three isoforms, the neuronal NOS (nNOS), the inducible NOS (iNOS), and the endothelial NOS (eNOS). Nitric oxide synthase (NOS) inhibitors attenuate or block morphine action. NO/NOS system has an effect on morphine-induced place preference. Action of NO/NOS system are involved in the activation of opioid receptors. The effect of NO/NOS system on the expression of morphine withdrawal symptoms may be partially mediated by noradrenergic neuron system. Prostanoids are shown to be important lipid mediators, not only in periphery but also in the brain, where they appear to modulate synaptic transmission. Recent studies have demonstrated that cyclooxygenase (COX) pathway might modulate the neurotransmission ofγ-aminobutyric acid and dopamine in the central nervous system. COX is the rate-limiting enzyme involved in the synthesis of prostanoids from arachidonic acid. The former is constitutively expressed in most cells. In contrast, COX-2 is normally not present but may be induced by certain serum factors, cytokines, and growth factors. Operating through G-protein linked cell-surface receptors they act as regulators of second messengers such as cAMP, inositol phosphates and diacylglycerol, in turn affecting a variety of cellular processes including calcium mobilisation, protein kinase activity and protease activity.In this study, we investigate the effect of morphine on gene expression of iNOS and COX-2 on C6 glioma.1. Methods C6 glioma cells were cultured in DMEM containing 5% FBS and divided into 5 groups:①normal Control group: PBS was added for 24h;②LPS control group: LPS(10mg/L) was added for 24h;③LPS + Morphine group: Morphine (10ug/ml) was given 0.5h before activatione with LPS (10mg/L) for 24h;④Morphine group: Morphine (10ug/ml) was added for 24h;⑤Naloxone + Morphine: Morphine (10ug/ml) was given 0.5h before activatione with naloxone (1umol/L) for 24h. The activity of iNOS was measured by KIT. The expression of iNOS and COX-2 mRNA and protein were detected by Western blotting and RT-PCR , respectively.2.Result LPS induces a pronounced increase the activity of iNOS and the expression of iNOS and COX-2 in C6 glioma. Morphine was able to significantly inhibit activity of iNOS and expression of iNOS and COX-2 when sdded after LPS challenge. Compared with that in the control group, the activity of iNOS and expression of iNOS and COX-2 had no significant change in C6 glioma treated simply with morphine for 24h.3.Conclusion Morphine added for 24h had no significant effect on the activity iNOS and expression of iNOS and COX-2 in C6 glioma, but morphine induces a pronounced reduction in LPS-induced activity of iNOS and expression of iNOS and COX-2. |
