Study On Central Analgesic Effects Of Ghrelin Active Fragments And Their Blood-brain Barrier Permeabilities

Objectives:Ghrelin is an endogenous brain-gut peptide with analgesic effect discovered in recent years.Its unique N-terminal Ser³ acylation structure enables it to pass the blood-brain barrier smoothly,but difficult synthesis,high price and short half-life greatly limit its clinical application.The active fragments of ghrelin have N-terminal Ser³ acylated structure,which have good binding abilities with ghrelin receptor GHS-R1α.They are simple in structure and easy to synthesize.In this study,the central analgesic activities and receptor mechanisms of the active fragments G（1-5）-NH₂,G（1-7）-Lys-NH₂,G（1-9）and G（1-11）were investigated,and the blood-brain barrier permeabilities were further studied.The above experiments provided a reference for clinical research and development of effective analgesics with good analgesic effect,simple structure and good permeability of blood-brain barrier.Methods:The central analgesic activities of G（1-5）-NH₂,G（1-7）-Lys-NH₂,G（1-9）and G（1-11）were explored with tail withdrawal test.The relationships between the four ghrelin active fragments and GHS-R1αand opioid receptors were investigated after intracerebroventriculary co-injection.The blood-brain barrier permeabilities of ghrelin active fragments were investigated by peripheral naloxone methiodide and central naloxone.Naloxone methiodide did not pass the blood-brain barrier.Results:1.Four ghrelin active fragments caused concentration-and time-dependent analgesic effects in the tail withdrawal test after intracerebroventriculary injection（mouse,i.c.v.）.2.The analgesic effects caused by intracerebroventriculary injection of G（1-5）-NH₂,G（1-7）-Lys-NH₂ and G（1-11）could be antagonized by[D-Lys³]-GHRP-6（DLS）which is the antagonist of GHS-R1α.This indicated that GHS-R1αwas involved in the analgesic effects caused by G（1-5）-NH₂,G（1-7）-Lys-NH₂ and G（1-11）.3.The central analgesic effect of G（1-5）-NH₂（i.c.v.）was mediated byμ-opioid receptor andκ-opioid receptor.The central analgesic effect of G（1-7）-Lys-NH₂（i.c.v.）was mediated byμ-opioid receptor andδ-opioid receptor,the partly central analgesic effect was mediated byκ-opioid receptor.The central analgesic effect of G（1-9）（i.c.v.）was mediated byμ-opioid receptor.The central analgesic effect of G（1-11）（i.c.v.）was mediated byμ-opioid receptor andδ-opioid receptor.4.After injection through the caudal vein,G（1-5）-NH₂ and G（1-9）caused concentration-and time-dependent analgesic effects in the tail withdrawal test.5.The analgesic effects caused by caudal vein injection of G（1-5）-NH₂ or G（1-9）could not be antagonized by naloxone methiodide（i.p.）which did not pass the blood-brain barrier.The analgesic effects caused by caudal vein injection of G（1-5）-NH₂ or G（1-9）could be antagonized by naloxone（i.c.v.）.The results showed that G（1-5）-NH₂ and G（1-9）had analgesic effects after being administered through the caudal vein and reached the center through the blood-brain barrier.Conclusion:Intracerebroventricular injection of ghrelin active fragments G（1-5）-NH₂,G（1-7）-Lys-NH₂,G（1-9）and G（1-11）produced obvious time and concentration-dependent analgesic effects.G（1-9）had the best analgesic effect among them.GHS-R1αwas involved in the central analgesic effects caused by G（1-5）-NH₂,G（1-7）-Lys-NH₂ and G（1-11）.The central analgesic effects of G（1-5）-NH₂,G（1-7）-Lys-NH₂,G（1-9）and G（1-11）were all mediated by different opioid receptors.The active fragments G（1-5）-NH₂ and G（1-9）can produce analgesic effects in the center through the blood-brain barrier after intravenous injection.