Modulation Of Pain Threshold And Some Morphine Effects By Endogenous Agmatine

Agmatine in mammalian tissues, which is formed by the decarboxylation of L-Arginine by the enzyme L-arginine decarboxylase(L-ADC), has been postulated to be an endogenous ligand for imidazoline receptors. It has been proved by many experiments that agmatine could enhance morphine analgesia, inhibit tolerance to morphine and prevent substance dependence on morphine in mice and rats in vivo. All these effects of agmatine are based on activation of I-R and inhibition of NOS. Idazoxan, an I-R selective antagonist, itself could decrease the pain threshold, inhibit the analgesic effect of morphine, promote the development of tolerance to morphine and induce the abstinence syndrome of morphine-dependent mice and rats. These experimental data led to the hypothesis that endogenous agmatine may play a physiological role in the complex modulation of pain sensitivity and the action of morphine. Ornithine decarboxylase (ODC) and diamine oxdiase (DAO) are two enzymes involved in the metabolism of endogenous agmatine in mammalian tissues. a-Difiuoromethyl-ornithine (DFMO) and aminoguanidine (AMG) can inhibit the activities of the two enzymes, respectively. It is possible that the level of endogenous agmatine and the effects of agmatine on the pharmacological effects of morphine might be modulated through inhibition of the two enzymes by DFMO andAMG. Our present research was aimed at investigating the possible effects of DFMO and AMG on pain threshold and morphine action in mice. On the other hand, we also investigated if DFMO and AMG can influence the activity of NOS in brain and the level of cAMP in NG-108 cells, because the two molecules play a promoting role in nociceptive transmission and opioid action. These data will help to validate the hypothesis that endogenous agmatine modulates the pain sensitivity and the action of morphine.Method:Antinociceptive tests were evaluated by the methods of acetic acid writhing test, heat radiation tail-flick assay, and 55@ hot plate assay. NOS activity in mice brain was determined following the instruction of the NOS assay kit. A competitive protein binding assay method determined the concentration of the cAMP in cells.Results:1. DFMO and AMG have weak analgesic effect. In the acetic acid writhing test, both DFMO and AMG increased the pain threshold in a dose-dependent manner, and the effects of them may be mediated through imidazole receptors because the effects can be antagonized by idazoxan. In heat radiation tail-flick assay, AMG 0.25mg.kg-1 can also increase the pain threshold.2. DFMO and AMG enhance morphine analgesia. DFMO can enhance morphine analgesia in acetic acid writhing test, heat radiation tail-flick test and 55癈 hot plate test. And in acetic acid writhing test the effects can be antagonized by idazoxan. In heat radiation tail-flick test, DFMO (5mg.kg-1 i.v) have the same effects. AMG can enhance the morphine analgesia in heat radiation tail-flick test, but these effects can not be prevented by idazoxan, indicating that imidazoline receptors may not beinvolved.3. In heat radiation tail-flick assay, single dose of morphine (100 mg.kg-1 s.c.) induced acute tolerance indicated by the decrease in analgesic effect of 5mg/kg morphine. The PMAP of morphine (5 mg.kg-1 s.c.) was reduced from 42% to 16% (n=15, P<0.05). Administration of DFMO (0.25 mg.kg-1 i.c.v.) or AMG (0.125 mg.kg-1 i.c.v.) prior to administration of morphine (100 mg.kg-1 s.c.) blocked this declination. In these groups, the PMAP of morphine (5 mg.kg-1 s.c.) before pretreatment of the mice with single dose of morphine had no significant difference compared with that after pretreatment4. DFMO and AMG have no effect on activity of NOS. In our study, we found that at the same dose as L-NAME (0.5mg.kg-1 i.c.v.), an inhibitor of NOS, DFMO can promote but not inhibit the activity of NOS significantly in mice cerebellum from 16.2±3.0 U.mg.pro-1 to 18.8±1.1 U.mg.pro-1 (n=5, P<0.05). AMG had no effect on the activity of NOS. Thus the influ...