| Objective To investigate the influence of intrathecal AIP on pain behavior and expression of p-CaMKⅡand p-CREB in a rat model of tibia cancer pain. To explore the spinal mechanism that p-CaMKⅡc ontributed to bone cancer pain and the different analgesic aspect by different dosage of intrathecal AIP.Methods The study was divided into 3 parts.1. The changes of p-CaMKⅡexpression in spinal dorsal horn in a rat model of bone cancer pain.Rats were randomly divided into three groups(n=18 each):normol group(N), without any intervention;model group(M), injecting Walker256 cells (1×107/ml) 10μl into tibia medullary cavity;control group(C), injecting 1640 fluid 10μl, the other operation was the same as model group.Pain behavior was measured before and 6,12,18d after establishing model. The lumbar4-6 spinal cord of each group was removed on 6th,12nd,18th day separately for detecting p-CaMKⅡexpression changes by immunohistochemistry(6 rats every time point).2. The effect of intrathecal injection of AIP on pain behavior in a rat model of bone cancer pain.Rats were randomly divided into 6 groups(n=8 each):control group(C);model group (M);saline group(NS);AIP5μmol/L group(AIP5);AIP10μmol/L group (AIP10); AIP20μmol /L group(AIP20).The last four groups were given singley intrathecal normal saline and different dosage of AIP respectively from day 8 to day 12 after establishing model per day for 5 days. The volume was 10μl.(1)Pain behavior was tested before and 0.5,1,2,4,6,12,24h after intrathecal injection on the 8th day after establishing model.(2)Pain behavior was tested before and 8,10,12,14,15,16,18d after establishing model, when intrathecal injection was given singley per day for 5 days.3. The effect of intrathecal injection of AIP on the p-CREB expression in spinal dorsal horn in a rat model of bone cancer pain.Rats were randomly divided into 4 groups(n=6 each):control group(C),model group(M), saline group(NS),AIP group(AIP). Intrathecal injection of normal saline and AIP20μmol/L were given respectively from day 8 to day 12 daily in NS and AIP group. The lumbar4-6 spinal cord was removed on 12nd day after establishing model, and the expression of p-CaMKⅡand p-CREB were detemined by immunohistochemistry.Results1. Mechanical withdrawal thresholds gradually decreased and pairs of hindlimb weight-bearing difference gradually increased from day 6 to day 18 after establishing model in M group compared with those in N and C group.The p-CaMKⅡimmunopositive cell number(NUM) gradually increased in that period in spinal dorsal horn(P<0.05).2.(1)After intrathecal injection on 8th day, the analgesic efficacy of AIP5 group peaked at 0.5h after injection,and that was the same as before injection at 1h.The analgesic efficacy of groups AIP10 and AIP20 reached peak at 1h, and that was the same as before injection at 4h and 6h seperately.(2)Continuous intrathecal different dosage of AIP for 5 days, which could alleviate pain behavior in different degree. The anti-nociception of AIP5 group was remarkable during the period from day 10 to day 12,and pain behavior didn't have differences between M and NS group on 14th day after establishing model(P>0.05). The anti-nociception of AIP10 and AIP20 group was remarkable from day 8 to day 12 ,and pain behavior didn't have statistically differences between M and NS group on 15th and 16th day separately(P>0.05).3. Compared with C group, the num of p-CaMKⅡand p-CREB in spinal dorsal horn was significantly increased in M and NS group(P<0.05). The num of p-CaMKⅡa nd p-CREB were significantly reduced in AIP group compared with those in M and NS group(P<0.05), but there were differences between AIP and C group(P<0.05).Conclusion1. Mechanical withdrawal thresholds gradually decreased and pairs of hindlimb weight-bearing difference gradually increased from day 6 to day 18 after establishing model. The num of p-CaMKⅡgradually increased in that period and it was mainly distributied in theⅠ～Ⅱl ayer of spinal dorsal horn.2. The trends of pain threshold were in consistency with the changes of p-CaMKⅡexpression. The changes of pain behavior were related to the up-regulated expression of p-CaMKⅡi n spinal dorsal horn.3. Intrathecal administration of AIP can significantly attenuate pain behavior in a rat model of bone cancer pain, and its analgesic aspect was dose-dependent.4. Intrathecal injection of AIP can reduce the expression of p-CaMKⅡand p-CREB. CaMKⅡ/ CREB signal transduction may be involved in the occurrence and maintenance of bone cancer pain. |
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