The Regulation Mechanism Of Peripheral Tissue Injury On The Synaptic Expression Of AMPA Receptors In The Brain

The α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid(AMPA)receptor is an ionotropic glutamate receptor that is mediate fast neurotransmission in central nervous system and is assembled from four subunits(GluA1-4).The AMPA receptors have emerged as key elements of long-lasting changes in the efficacy of central synapses,which are believed to constitute cellular correlates of complex behaviors such as learning and memory,nociception,and reward.Many studies shown that dysregulated cell surface trafficking of AMPA receptors plays an important role in the development of inflammatory and neuropathic pain in spinal level.However,the cell surface trafficking of pain facilitating receptors at supraspinal level received little attention.The periaqueductal grey(PAG)and nucleus accumbens(NAc)is a complex region in brain structure in the transmission and/or modulation of nociceptive information.This present study used the method of pharmacological behavior and western blot from overall level to investigate the influence cell trafficing of GluA2 in peripheral tissue injury induced pathological pain and to explore its molecular mechanism.The main methods of this experiment is:Part one,①intra-PAG injection of 0.15 pmol,1.5 pmol,7.5 pmol,15 pmol of GluA2-3y in intact rats,The Randall Selittlo test and the hot-plate test were used as behavion.test after10,15,20,30,45,60 minute intra-PAG injection of GluA2-3y;② 10 days after sciatic nerve ligation was performed simultaneously to produce mononeurophy,intra-PAG injection of 0.15 pmol,1.5 pmol,7.5 pmol,15 pmol of GluA2-3y in nerve injury rats.The Randall Selittlo test and the hot-plate test were used as behavior test after10,15,20,30,45,60 minute intra-PAG injection of GluA2-3y;③ Three hours after the injection of carrageenan into the intraplantar region of a hind paw produced acute inflammation,intra-PAG injection of 0.15 pmol,1.5 pmol,7.5 pmol,15 pmol of GluA2-3y in inflammation rats.The Randall Selittlo test and the hot-plate test were used as behavior test after10,15,20,30,45,60 minute intra-PAG injection of GluA2-3y;④ Intra-PAG injection of 1 1 10mg/ml morphine in intact rats,in nerve injury rats or in inflammation rats.HWLs was tested after 30 minute.Part two:10 male SD rats are seperated two goulp,one is sham group(n=5),another is CCI group(n=5).CCI group rats were sciatic nerve ligation,while sham group was performed following the same surgical procedure for sciatic nerve injury except for nerve ligation.After CCI group produced mononeurophy,rats were decapitated immediately after extinction or reinstatement and the brain was removed.The regions of the NAc and PAG were dissected at-20 ℃,using western blot to test the change of AMPA receptor in rats with neuropathic.All the results showed as follows:① Intra-PAG injection of GluA2-3y induced dose-independent increases in hindpaw withdrawal latencies(HWLs)to noxious thermal and mechanical stimulations in intact rats,in nerve injury rats and in inflammation rats,suggesting that GluA2 cell surface trafficking in PAG is involved in pathological pain modulation.② GluA2-3y had much stronger anti-nociceptive effect in rats with neuropathic pain induced by sciatic nerve ligation and in rats with inflammation.Interestingly,we showed that intra-PAG injection of 15 pmol GluA2-3y had similar analgesic effect with 10 μg(35 nmol)morphine in rats with neuropathic pain and in rats with inflammation,while morphine induced much stronger anti-nociceptive effect in intact rats,suggesting that the anti-nociceptive effects of morphine and GluA2-3y have different mechanisms.③ The results of western blot shown that the expression of GluA2 in PAG and the expression of GluA1 in the NAc showed no significant changes in sciatic nerve injury rats compared with sham group tested by western blot.Interestingly,it was found that there were significant increases in the expression of GluA2 in NAc in rats with neuropathy compared with sham group.Taken together,our results found that GluA2 trafficking in PAG played a critical role in pain modulation and a drug GluA2-3y inhibiting GluA2 endocytosis produced potent analgesic effect in rats with neuropathic pain and in rats with inflammation,strongly support the recent hypothesis that targeting receptor trafficking could be a new therapeutic target for treatment of peripheral tissue injury induced pathological pain.The western blot results indicate that GluA2 in NAc may be involved in nociceptive modulation in rat with neuropathic pain.