Extracellular signal-regulated kinase (ERK), a mitogen-activated protein kinase (MAPK), transduces a broad range of extracellular stimuli into diverse intracellular responses. It has been reported that ERK is involved in the modulation of nociceptive information and central sensitization produced by intense noxious stimuli or peripheral tissue inflammation. Our previous studies showed that the spinal neurons sensitization was involved in morphine withdrawal response. This study was to investigate the role of the spinal ERK in morphine dependence and naloxone-precipitated withdrawal response. To set up morphine-dependent model, rats were subcutaneously injected with morphine (twice a day, for 5 d). The dose of morphine was 10 mg/kg on the first day and was increased by 10 mg/kg each day. On day 6, 4 h after the injection of morphine (50 mg/kg), morphine withdrawal syndrome was precipitated by an injection of naloxone (4 mg/kg, i.p.). Using anti-phospho-ERK (pERK) antibody, the time course of pERK expression was detected by Western blot. U0126, a mitogen-activated protein kinase kinase (MEK) inhibitor, or phosphorothioate-modified antisense oligonucleotides (ODN) was intrathecally injected 30 min or 36, 24 and 12 h before naloxone-precipitated withdrawal. The scores of morphine withdrawal symptom and morphine withdrawal-induced allodynia were observed. One hour after naloxone-precipitated withdrawal, pERK expression in the spinal dorsal horn was assessed by immunohistochemical analysis and Western blot was used to detect the expression of cytosolic and nuclear fraction of pERK in the rat spinal cord. The results showed that the expression of cytosolic and nuclear fraction of pERK, not non-phospho-ERK, in the spinal cord was gradually increased following the injection of morphine. When morphine withdrawal was precipitated with naloxone, the expression of the spinal pERK further increased. Intrathecal administration of U0126 or antisense ODN against ERK decreased the scores of morphine withdrawal, attenuated morphine withdrawal-induced allodynia and also inhibited the increase of pERK expression in the spinal cord of morphine withdrawal rats. These results suggest that activation of the spinal ERK is involved in morphine-dependent and naloxone-precipitated withdrawal response.
Animals ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Male ; Morphine Dependence ; enzymology ; physiopathology ; Naloxone ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Spinal Cord ; enzymology ; Substance Withdrawal Syndrome ; enzymology ; physiopathology

The mechanism of morphine dependent is a complex Procedure. It involves in many complex mechanisms such as the ultra-structure of synapse of special brain areas, neurotransmitter, enzymology, and so on. These mechanisms have closely correlation. In this paper we reveiwed the development in enzymological mechanism of morphine dependent enzymes including protein kinase (PK), nitric oxide synthase (NOS), superoxide dismutase (SOD), adenylate cyclase (AC), Succinate dehydrogenase (SDH)and 3beta-Hydroxy steroid dehydrogenase (3beta-HSD).
Adenylyl Cyclases/metabolism* ; Animals ; Brain/enzymology* ; Morphine Dependence/pathology* ; Nitric Oxide Synthase/metabolism* ; Protein Kinases/metabolism* ; Substance Withdrawal Syndrome/metabolism* ; Succinate Dehydrogenase/metabolism* ; Superoxide Dismutase/metabolism* ; Synapses/enzymology*

OBJECTIVE: To study the changes and actions of neuronal constructive nitric oxide synthase(ncNOS) in heroin drug abuse. METHODS: The expression of ncNOS and ncNOS mRNA in neurons of cerebral cortex, periaqueductal gray matter and the ventral tegmental area was observed by immunohistochemistry, in situ hybridization and image analysis technique after heroin dependence and spontaneous withdrawal in rats. RESULTS: The quantity of ncNOS and ncNOS mRNA rised clearly and the number of ncNOS and ncNOS mRNA positive cells increased greatly in heroin dependence and withdrawal. The changes of ncNOS and ncNOS mRNA in spontaneous withdrawal were more clear than ones of dependence. Heroin dependence and withdrawal led to alterations in ncNOS and ncNOS mRNA expression in important regions implicated in the physical tolerance and dependence. CONCLUSION The ncNOS plays an important role in heroin dependence and withdrawal. The ncNOS immunohistochemical changes observed in the present study might be useful for the forensic pathological diagnosis of heroin drug abuse.
Animals ; Cerebral Cortex/enzymology* ; Forensic Medicine ; Heroin Dependence/enzymology* ; Immunohistochemistry ; Male ; Neurons/enzymology* ; Nitric Oxide Synthase/metabolism* ; Nitric Oxide Synthase Type I ; RNA, Messenger/metabolism* ; Rats ; Rats, Sprague-Dawley ; Substance Withdrawal Syndrome/enzymology*

OBJECTIVE: To observe the changes of adenylate cyclase(AC) on cerebral regions related to morphine dependence in rats and investigate the relationship between the enzymological changes and the mechanism of morphine dependence. METHODS: The technique of enzyme-histochemistry was used to detect the variations of AC of special seven cerebral regions including frontalis cortex, lenticula, corpus amygdaloideun, substantia nigra, hippocampus, periaqueductal gray and locus coerleus in morphine dependent rats. The enzymological changes were observed by optical microscope. Changes of gray degree of these cerebral regions were also observed by using the image analysis system. RESULTS: Compared with those in control group, the contents of AC in morphine dependent groups were increased. CONCLUSION The contents of AC are increase in those regions. The mechanism of morphine dependence close related to the increasing of AC. The correlation of the mechanism of morphine dependence and up-regulation of AC/cAMP-PKA system is discussed.
Adenylyl Cyclases/metabolism* ; Animals ; Brain/pathology* ; Cerebral Cortex/enzymology* ; Disease Models, Animal ; Female ; Hippocampus/enzymology* ; Male ; Morphine Dependence/pathology* ; Periaqueductal Gray/enzymology* ; Rats ; Rats, Sprague-Dawley ; Substance Withdrawal Syndrome/metabolism* ; Time Factors

Our previous studies showed that spinal neurons sensitization was involved in morphine withdrawal response. This study was to investigate the roles of spinal protein kinase C (PKC) alpha, gamma in morphine dependence and naloxone-precipitated withdrawal response. To set up morphine dependence model, rats were subcutaneously injected with morphine (twice a day, for 5 d). The dose of morphine was 10 mg/kg in the first day and was increased by 10 mg/kg each day. On day 6, 4 h after the injection of morphine (50 mg/kg), morphine withdrawal syndrome was precipitated by an injection of naloxone (4 mg/kg, i.p.). Chelerythrine chloride (CHE), a PKC inhibitor, was intrathecally injected 30 min before the administration of naloxone. The scores of morphine withdrawal symptom and morphine withdrawal-induced allodynia were observed. One hour after naloxone-precipitated withdrawal, Fos protein expression was assessed by immunohistochemical analysis and Western blot was used to detect the expression of cytosol and membrane fraction of PKC alpha and gamma in the rat spinal cord. The results showed that intrathecal administration of CHE decreased the scores of morphine withdrawal, attenuated morphine withdrawal-induced allodynia and also inhibited the increase of Fos protein expression in the spinal cord of morphine withdrawal rats. The expression of cytosol and membrane fraction of PKC alpha was significantly increased in the spinal cord of rats with morphine dependence. Naloxone-precipitated withdrawal induced PKC alpha translocation from cytosol to membrane fraction, which was prevented by intrathecal administration of CHE. During morphine dependence, but not naloxone-precipitated withdrawal, PKC gamma in the spinal cord translocated from cytosol to membrane fraction, and intrathecal administration of CHE did not change the expression of PKC gamma in the spinal cord of naloxone-precipitated withdrawal rats. It is suggested that up-regulation and translocation of PKC in the spinal cord contribute to morphine dependence and naloxone-precipitated withdrawal in rats and that PKC alpha and gamma play different roles in the above-mentioned effect.
Animals ; Male ; Morphine Dependence ; physiopathology ; Naloxone ; pharmacology ; Protein Kinase C ; metabolism ; physiology ; Protein Kinase C-alpha ; metabolism ; physiology ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Spinal Cord ; metabolism ; physiopathology ; Substance Withdrawal Syndrome ; enzymology ; physiopathology