[Effect of nitric oxide synthase inhibitor on signal transduction pathway of opiate tolerance and dependence in NG108-15 cells expressing iNOS gene].
OBJECTIVE
To investigate the role of adenylate cyclase (AC)-cAMP system and Ca2+ system and NO-cGMP signal system and the effects of a NOS inhibitor, NG-nitro-L-arginine (L-NNA) in the neuronal mechanisms of opioid tolerance and dependence.


METHODS
The experiments were performed in five groups: control group; opioid agonist group; opioid agonist + nalonoxe group; L-NNA + opioid agonist group and L-NNA + opioid agonist + nalonoxe group. The intracellular cAMP and cGMP levels were measured by 3H-cAMP protein binding assay and 3H-cGMP radioimmunoassay, respectively. NOS activity was determined by the conversion of 3H-arginine to 3H-citrulline. The change of [Ca2+]i was studied by the laser scanning confocal microscopy technique. iNOS protein expression was detected using immunohistochemistry with monoclonal antibody of iNOS, and imaging analysis was performed.


RESULTS
Long-term administration of high-selective delta-opioid receptor agonist DPDPE and precipitation of opioid withdrawal by naloxone significantly induced increase of cAMP level and [Ca2+]i in NG-LNCXiNOS cells with stable expression of iNOS gene. The cytosolic iNOS activity and cGMP generation were enhanced by DPDPE dose-dependently. 10(-4) mol/L L-NNA could block opioid agonist-induced AC-cAMP desensitization and activity of NO-cGMP second messenger pathway, but it could not reduce opioid-induced elevation of [Ca2+]i. Furthermore, L-NNA decreased iNOS-specific protein expression in DPDPE-induced tolerance and naloxone-precipited withdrawal cells.


CONCLUSIONS
NOS inhibitor may attenuate the development of opioid tolerance and withdrawal via the negative regulation of AC-cAMP system and NO-cGMP system. It can be clinically used to prevent opiate tolerance and addiction.
Zang
M
M
Department of Pharmacology, National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005.
Shen
Q
Q
Liu
J
J
D000818:Animals; D004361:Drug Tolerance; D051379:Mice; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D019335:Nitroarginine / Q000494:pharmacology*; D009293:Opioid-Related Disorders / Q000503:physiopathology* / Q000517:prevention & control; D017465:Receptors, Opioid, delta / Q000378:metabolism; D015398:Signal Transduction
D017465:Receptors, Opioid, delta; D019335:Nitroarginine; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse