In Vitro And In Vivo Characterizations Of The Bifunctional μ-and δ-opioid Receptors Ligand MCRT On Mouse Gastrointestinal Motility

Chimeric opioid MCRT was a novel multi-target ligand ofμandδopioid receptors（MOR and DOR）based on morphiceptin and PFRTic-NH₂,and produced potent analgesia(ED₅₀=0.03 nmol/mouse)with less upper gastrointestinal dysmotility.In this study,we sought to performe the tests to evaluate the pharmacological effects of MCRT on distal colon motility and defecation function.Moreover,opioid receptor antagonists（Naloxone、Cyprodime and Naltrindole）and neuropeptide FF（NPFF）receptor antagonists（BIBP 3226 and RF9）were utilized to explore the mechanisms.Isolated mouse colon bioassay and colonic bead expulsion were to characterize MCRT-induced inhibition of colonic motility in vitro and in vivo,respectively.Fecal pellet output was to evaluate the defecation function.We found that（1）In vitro,MCRT elicited significant contractions of distal colon muscle stripe in a dose-dependent manner.Naloxone（the non-selective opioid receptor antagonist）completely blocked the contractile responses induced by MCRT(10^-5 M).Cyprodime（a selective MOR antagonist）and Naltrindole（a selective DOR antagonist）almost completely abolished the MCRT-induced contractile responses,while BIBP3226（the mixed antagonist of NPY Y1 and NPFF receptors）and RF9（the selective antagonist of NPFF receptor）failed to influence the MCRT-induced contractions.The data indicated that MCRT-induced colonic contraction was regulated viaμ-andδ-opioid receptors.（2）In vivo,MCRT delayed colonic bead expulsion(ED₅₀=1.1 nmol/mouse).Naloxone,BIBP3226 and RF9 were 10 min i.c.v.pre-injected prior to MCRT.None of the three antagonists showed any influence to MCRT-induced abnormal colonic propulsive motility,indicating that the colonic bead expulsion of MCRT is independent of opioid and NPFF receptors.（3）In vivo,MCRT inhibited fecal number(ED₅₀=1.43 nmol/mouse)and dry weight(ED₅₀=1.63nmol/mouse).Consequently,the co-administration of antagonists and MCRT was chosen,Naltrindole partially blocked MCRT-induced fecal number and fecal dry weight inhibition.None of the two antagonists,cyprodime and RF9 showed any influence to fecal inhibitions of MCRT.The data indicated that MCRT-induced constipation was sensitive to DOR but not to MOR blockade.Based on the above results,we speculated on the existence of such an opioid receptor subtype or MOR/DOR heterodimer,which was involved in the central analgesia and the in vitro colonic contractions but not the central colonic dysmotility.In vivo,central MCRT inhibited colonic bead propulsion and fecal pellet output,with the ED₅₀ were1.1nmol/mouse and 1.43nmol/mouse respectively,far more than its analgesic dose(ED₅₀=0.03nmol),suggesting that MCRT was less prone to induce gastrointestinal dysmotility at analgesic,and provided a possibility for safer opioid analgesic.