| Clinically, a long-term therapy of opioids has been regarded as a effective method for management of chronic and cancer pain, but opioid tolerance occurs following chronic drug exposure which limits their usefulness. The exact neural plasticity mechanisim which contribute to morphine tolerance is not yet clear entirely, but studies have shown that morphine tolerance and neuropathic pain share the same cellular mechanism. Calcitonin gene-related peptide (CGRP), P substance (SP) and brain-derived neurotrophic factor (BDNF) may be involved in nociceptive mechanism of morphine tolerance. Meanwhile, more and more studies have focus on the transient receptor potential vanilloid subfamily 1(TRPV1;VR1) which is a member of nociceptors.There were many kinds of nociceptors and pain transmitters in dorsal root ganglion(DRG) which is a site for processing and integrating nociceptive information. In this study, based on the model of inflammation-morphine tolerance, we investigate the effects of CGRP, SP, BDNF and TRPV1 in neural plasticity changes of DRG. Electro-acupuncture(EA) is an important treat method in Chinese traditional medicine, its analgesic effect has been recognized internationally.Therefore, we investigate the effects of EA in neural plasticity changes of DRG to find a solution for controlling morphine tolerance.Part One:Establishment of arthritis-chronic morphine tolerance rat model and combined with electro-acupuncture.Objective To establish the rat model of arthritis-chronic morphine tolerance and combined with EA, and to investigate the effects of EA on the formation of morphine tolerance.Methods Thirty adult male SD rats with successful intrathecal cannulation were randomly divided into 6 groups(n=5 each):Group A:administered 10μl normal saline intrathecally after chronic inflammation pain (CIP) in hind paw induced by complete Freund's adjuvant(CFA) two times a day for 7 consecutive days. Group B: administered 10μg/kg(10μl) morphine intrathecally two times a day for 7 consecutive days. Group C:administered 10μg/kg(10μl) morphine intrathecally after CIP in hind paw induced by CFA only once. Group D:administered 10μg/kg(10μl) morphine intrathecally after CIP in hind paw induced by CFA two times a day for 7 consecutive days. Group E:based on the methods of group D,given 2Hz electroacupuncture on Yanglingquan and Zusanli for 30 min after the first administration once a day for 7 consecutive days. Group F:The frequency of electroacupuncture is 15Hz, other methods were the same with group E. The paw withdrawal latency(PWL) to noxious thermal stimulation and the paw withdrawal threshold to mechanical stimulation was measured daily.Results Morphine tolerance occurred in group B and group D, but not in group E and group F. The level of hyperalgesia in group E is lower than that in group F. Conclusion Chronic intrathecal morphine exposure can induce morphine tolerance in arthritis rats. EA inhibit the formation of morphine tolerance,2Hz is more effective than 15Hz.Part Two:Effects of opioids tolerance induced by intrathecal morphine on expression of calcitonin gene-related peptide, substance P and brain-derived neurotrophic factor mRNA in dorsal root ganglion in rats with inflammation in hind paw and effects of electroacupuncture. Objective To investigate the effects of opioids tolerance induced by intrathecal (IT) morphine on expression of calcitonin gene-related peptide(CGRP), substance P (SP)and brain-derived neurotrophic factor(BDNF) mRNA in dorsal root ganglion in rats with chronic inflammation pain(CIP) in hind paw and effects of electroacupuncture(EA). Methods The methods for group dividing and behavioral testing was the same as part one. The lumbar segment (L4-6) of DRG was removed on the 7th day after administration for determination of CGRP mRNA,SP mRNA and BDNF mRNA expression in the DRG using RT-PCR. Results Morphine tolerance occurred in group B and group D, but not in group E and group F. Among the six groups, the expression of CGRP, SP and BDNF mRNA in group D were the highest. The expression of CGRP, SP and BDNF mRNA in group E were lower than that in group F. Conclusion The changes in expression of CGRP, SP and BDNF in DRG are involved in the mechanism of morphine tolerance. EA can inhibit the formation of morphine tolerance, enhanced expression of CGRP, SP and BDNF are involved in the mechanism of that.Part Three:Effects of opioids tolerance induced by intrathecal morphine on expression of TRPV1 in dorsal root ganglion in rats with inflammation in hind paw and effects of electroacupunctureObjective To investigate the effects of opioids tolerance induced by intrathecal (IT) morphine on expression of TRPV1 in dorsal root ganglion in rats with chronic inflammation pain(CIP) in hind paw and effects of electroacupuncture(EA). Methods The methods for group dividing and behavioral testing was the same as part one.The lumbar segment(L4.6) of DRG was removed on the 7th day after administration for determination of the number of TRPV1 positive neurons by immunohistochemistry and for determination of TRPV1 expression in total protein and membrane protein of the DRG using Western blot. Results Morphine tolerance occurred in group B and group D, but not in group E and group F. TRPV1 positive immunoreaction is distributed in cell membrane area of medium or small diameter DRG neurons. Among the six groups, the expression of TRPV1 in total protein and membrane protein in group D were the highest. The expression of TRPV1 in total protein and membrane protein in group E were lower than that in group F. Conclusion The changes in expression of TRPV1 in total protein and membrane protein of DRG are involved in the mechanism of morphine tolerance. EA can inhibit the formation of morphine tolerance, attenuated expression and transport of TRPV1 are involved in the mechanism of that. |
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