| Objectives:Local anesthetic is widely used in surgery anesthesia and acute pain therapy; recent clinical and animal study indicated that intrathecal injection of local anesthetic could alleviate neuropathic pain, but the underlying mechanisms remained unclear because the effect occur at lower dose blocking voltage-gated sodium channels, the main targets of local anesthetics. Furthermore, the analgesic effects of the drugs persist for longer pharmacokinetics.In neuropathic pain, there are more evidence indicating that pain processing might be regulated by epigenetic mechanisms:1. epigenetic changes are required for long-lasting neuronal plasticity that is essential for the development of chronic pain states;2. epigenetic modifications are crucial to long-term memory formation that shares common mechanisms with pain processing.Our experiment is divided into two parts.1) the effect of continuous ropivacaine intrathecal infusion on the established neuropathic pain in CCI rats.2) investigate the pain behavior and the spinal expression of Type I histone deactylases, HDAC1and HDAC2in rat model of chronic constriction injury (CCI) and its expression after intrathecal ropivacaine interference; Besides, we explored possible epigenetic targets of HDAC1and HDAC2.Method:(1)30rats in which intrathecal catheter was successfully placed for5days were randomly divided into3groups:group Sham+NS (n=10), group CCI+NS (n=10), group CCI+Ropi (n=10). NS or Ropivacaine was infused for seven consecutive day from D7to D13. We assessed the basic mechanical and thermal pain threshold1day and3days before CCI operation. After the operation, latencies of hind-limb withdrawl response to rediant heat and mechanical stimulation is measured on3th,7th th,10th,14th15th16th17th21th day,before intrathecal injection on7th,101h day. On14th day,3rats from each group were perfused with PBS and paraformaldehyde, lumber enlargement was removed for frozen section examination.3rats from each group were sacrificed via decapitation, their spinal enlargement was harvested for western blot.(2)35male SD rats weighed200-250g were randomly divided into3groups:group I was naive group(n=5), group II sham operation(n=15), groupIII was CCI operation(n=15).Left CCI model was established according Bennet and Xie. In sham group, left sciatic nerve were only exposed without ligation. We assessed the basic mechanical and thermal pain threshold1day and3days before CCI operation. After the operation, latencies of hind-limb withdrawl response to rediant heat and mechanical stimulation is measured on 3th,7th,10th,14th,21thday. Rats from each group was sacrificed on7th,10th,14thday, their spinal enlargement and samples from the first part was harvested for western blot and immunofluoresence to detect the expression of HDAC1,HDAC2,and GDNF.Results:(1) Compared with group Sham+NS, the pain threshold of group CCI+NS was decreased, the most prominent time point was on D14(P<0.05). Compared with group CCI+NS, the mechanical pain threshold of group CCI+Ropi was increased on D14andD15(P<0.05), the themal threshold of group CCI+Ropi was increased on D10, D14andD15(P<0.05), with no significance was seen on D16, D17and D21.(2) Compare with group Sham, the pain threshold of group CCI is decreased from D7to D21(P<0.05), the most prominent time point of hyperalgesia is on D14, while no change is observed between group Sham and group naive (P>0.05). The protein expression of HDAC1and HDAC2in group CCI was upgraded while GDNF was downgraded, compared with group Sham (P<0.05). The immu-positive cells of HDAC1and HDAC2in group CCI were upgraded on D14when compared with group Sham(P<0.05), The immu-positive area of GDNF in group CCI were upgraded on D14when compared with group Sham(P<0.05). Spinal horn immu-positive cells and the protein expression of HDAC1and HDAC2On D14in group CCI+Ropi was decreased, compared with group CCI+NS(P<0.05). The protein expression and immu-positive area of GDNF in group CCI+Ropi on D14was upregulated when compared with group CCI+NS (P<0.05).Conclusions:(1)The immune expression of HDAC1, HDAC2, GDNF in lumber enlargement was correspondent with the pain behavior of CCI model.(2)The immune expression of HDAC1, HDAC2, GDNF in lumber enlargement after consecutive infusion of0.25%ropivacaine was correspondent with the alleviating pain behavior。... |
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