Effect of tramadol on expression of P2X7 receptor in the midbrain periaqueductal gray in rats with inflammatory pain

Objective To investigate whether activation of the P2X7 receptor in the ventrolateral region of the midbrain periaqueductal gray (vlPAG) is involved in the analgesic mechanism of tramadol in rats with inflammation-induced pain. Methods One hundred and twelve male Sprague-Dawley (SD) rats were randomly divided into 7 groups (n=16 each): normal control (C group), normal saline group (NS group), formalin group (F group), formalin + low dose tramadol group (TL group), formalin + middle dose tramadol group (TM group), formalin + high tramadol dose group (TH group) and formalin + high dose tramadol + A-438079 group (TH+A-438079 group). The rat models of inflammatory pain were established by subcutaneous injection of 100 μL of 5% formalin into the left thenar of the rats in F, TL, TM and TH groups. TL, TM and TH groups had an intrathecal injection of tramadol (5, 15, 25 μg/kg dissolved in 20 μL of normal saline, respectively) while F group had an intrathecal injection of equal amount of normal saline. A-438079 (100 pmol/0.3 μL), a selective P2X7 receptor antagonist, was microinjected into the vlPAG of the TH+A-438079 rats, and followed with the establishment of inflammatory pain rats which were then subjected to intrathecal injection of tramadol (25 μg/kg dissolved in 20 μL normal saline). The changes in pain behavior were observed for pain intensity scoring (PIS) at 5, 10, 15, 25, 35, 40, 50 and 60 min after formalin injection in each group. The number of P2X7 receptor positive cells and P2X7 protein expression levels in the vlPAG were detected by immunohistochemistry and Western blotting. Results (1) The PIS, the number of P2X7 receptor positive cells and the P2X7 receptor protein levels in the vlPAG in F group were significantly increased at all time points compared with C group (P<0.05). (2) Compared with F group, the PIS scores in TM and TH groups were significantly decreased respectively at all time points from 25 and 15 min after formalin injection (P< 0.05); the expression and protein levels of P2X7 receptor in the vlPAG were further significantly up-regulated (P<0.05). (3) Compared with TM group, the PIS scores in TH group were significantly decreased at all time points from 15 min after formalin injection (P<0.05); the expression and protein levels of P2X7 receptor in the vlPAG were further significantly up-regulated (P<0.05). (4) Compared with TH group, microinjection of P2X7 receptor antagonist A-438079 in the vlPAG in TH+A-438079 group significantly raised the PIS scores at all time points from 15 min after formalin injection (P<0.05). Conclusion Intrathecal injection of tramadol may induce analgesic effect on inflammatory rats by promoting the expression of P2X7 receptor in vlPAG. Key words: Inflammatory pain; Tramadol; P2X7 receptor; Midbrain periaqueductal gray