The Role Of P2X4 Receptor In Chronic Morphine Tolerance

Opioids receptor agonist, such as morphine, are widely used as, effective analgesic drugs for acute and chronic pain in clinical practice, especially for neuropathic pain and cancer pain. However, repeated use of morphine could lead to the development of tolerance to analgesia including the attenuated analgesic efficacy, the shorted analgesic duration, hyperalgesia and withdrawal-induced pain enhancement. Therefore, its long-trem therapy is limited by the development of tolerance. In recent years, researchers have approached for the mechanisms of morphine tolerance on corpuscular and molecular level and gained much outcome. However, the mechanism is not fully understood in the present.Recently, researches had confirmed that role of P2X4 receptor and glia in the CNS played the role of neuropathic pain and P38MAPK signal channel might be involved in it. Cytokines such as tumor necrosis factor-alpha, interleukinlβand neurotransmitters such as brain-derived neurotrophic factors participated in the development of neuropathic pain and morphine tolerance. Moreover, the latest literature reported that morphine enhances microglial migration through modulation of P2X4 receptor signaling.Because morphine tolerance and neuropatic pain share similar mechanisms, we may assume that P2X4R/P38MAPK pathway might be involved in chronic morphine tolerance. To confirm the above hypothesis, we design to investigate:1) the effect of morphine tolerance on P2X4 receptor and microglial activation in the spinal cord; 2) the effect of P2X4 receptor antagonists TNP-ATP and PPADS on the morphine tolerance and the mechanisms. The data from our studies will provide new evidence for the mechanisms of morphine tolerance, moreover, it will provide theory for novel analgesic drugs exploitation.ObjectivesTo examine the effects of morphine tolerance on pain threshold of the ratsMethods12 male Sprague-Dawley (SD) rats (250-300g) fitted with intrathecal (i.t.)Catheters, Randomly, rats were divided into 2 groups(n=6), NS group:rats were intrathecally administrated saline; MOR group:rats were intrathecally administrated morphine 10μg.3 days after intrathecal catheters implantation, rats were administrated NS or morphine. And 5 day after administration, the model of morphine tolerance was established. The rat hind paw withdrawal threshold (WT) to mechanical stimuli and withdrawal latency (WL) to radiant heat were determined from day 1 to 7 after administration.ResultsCompared with NS group, a significant increase on day 1 in WT and WL was observed in MOR group. From 3d to 7d WT decreased, and day 7 reached the lowest point.ConclusionConsecutive administration of morphine induced morphine tolerance. Objectives1. To examine spinal microglial activation induced by morphine tolerance in rats.2. To examine spinal microglial activation induced by morphine tolerance in rats.Methods36 male Sprague-Dawley (SD) rats (250-300g) fitted with intrathecal (i.t.)catheters. Randomly, rats were divided into 2 groups(n=18):normal control group, NS group:rats were intrathecally administrated saline; MOR group:rats were intrathecally administrated morphine 10μg.3 days after intrathecal catheters implantation, rats were administrated NS or morphine. And 5 day after administration, the model of morphine tolerance was established. The spinal cord around lumbar enlargement and dorsal root ganglion was removed. Spinal microglial activation was evaluated with OX-42 immunoreactivity and spinal P2X4R expression were determined by immunohistochemisty.ResultsCompared with NS group, spinal P2X4R,OX-42 expression were significantly increased, While P2X4R in DRG didn't changed.ConclusionConsecutive administration of morphine induced spinal P2X4R,OX-42 expression increase, which suggested that spinal glial P2X4R receptor may be involved in the development of morphine tolerance. Objective1. To examine the effects of morphine tolerance on pain threshold of the rats.2. To examine spinal microglial activation and spinal P2X4R expression induced by morphine tolerance in rats.Methods114 male Sprague-Dawley(SD) rats(250-300g) fitted with intrathecal (i.t) catheters. Randomly, rats were divided into 7 groups(n=18):NS group:rats underwent were intrathecally administrated saline(NS); MOR group:rats were intrathecally morphine; TNP-ATP group:rats were intrathecally 30nmol TNP-ATP and morphine; TNP-ATP+NS group:rats were intrathecally TNP-ATP and NS; PPADS group:rats were intrathecally 30nmol PPADS and morphine; PPADS group+NS group: rats were intrathecally PPADS and NS.3 days after intrathecal catheters implantation, rats were administrated various intrathecal medicine. And 5 day after administration, the model of morphine tolerance was established. The rat hind paw withdrawal threshold(WT) to mechanical stimuli and withdrawal latency(WL) to radiant heat were determined from day 1 to 7.7 days after administration, the spinal cord around lumbar enlargement was removed. Spinal microglial activation was evaluated with OX-42 immunoreactivity and spinal P2X4R,p-p38MAPK,BDNF expression were determined by immunohistochemisty, P2X4R,BDNF,p-p38MAPK spinal expression were assessed by western blotting, spinal P2X4R,BDNF mRNA expression was assessed by reverse transcriptase polymerase chain reation (RT-PCR).ResultsCompared with MOR group, a significant increase in WT and WL was observed and spinal P2X4R,BDNF,OX42,p-p38MAPK expression were significantly decreased in TNP-ATP group On day 3 and day 7, while group PPADS did not.ConclusionIntrathecal TNP-ATP attenuated the morphine tolerance in rats, and suppressed spinal P2X4R,p-p38MAPK,BDNF expression and microglial activation, while PPADS could not. Therefore, the activation of spinal P2X4R/p38MAPK/BDNF pathway may be involved in the development of morphine tolerance.