Dissecting The Mechanisms Underlying Bee Venom-induced Pain And Inflammation In Rats: The Roles Of Peripheral MAPK Signal Transduction Pathways

Background: Inflammation caused by peripheral tissue injures produces pain-related behavior (persistent spontaneous nociception and hyperalgesia) and inflammatory reaction such as redness, edema. Our previous studies have showed that intraplantar injection of bee venom (BV) to hind paw of rats did not only produce persistent spontaneous flinching reflex and hyperalgesia, but also the obvious inflammatory reaction of the injected paw, such as redness and edema of the injected paw. A growing evidence has suggested a role for ERK1/2 and P38 signal transduction pathways in inflammation and pain induced by tissue and nerve injuries.Objective: To investigate the role of peripheral MAPK such as MEK1/2, ERK1/2 and P38 signal transduction pathways in bee venom-induced inflammation and pain, the effect of pre-treatment with subcutaneous injection of ERK1/2, P38 and MEK1/2 inhibitors on bee venom-induced spontaneous flinching reflexes, mechanical hyperalgeisa, and edema was determined in the present study.Methods: We use BV-induced pain rat model to investigate the roles of MAPK in the PSN, mechanical hyperalgesia and inflammation reaction of the injected hind paw. The experiments were performed on Sprague-Dawley albino male rats weighing from 250~300g. PD98059(inhibitor of ERK), SB202190 (a inhibitor of P38) and U0126( a inhibitor of MEK) were used in the present study. Intraplantar injection of PD98059 (1ug/50ul, 10ug/50ul and 100ug/50ul), SB202190 (1ug/50ul, 10ug/50ul and 100 ug/50ul) and U0126 (0.1 ug/50ul, 1ug/50ul and 10ug/50ul) were made 10 min prior to BV(0.2mg/50　l) injection. As control, vehicle (50ul/rat,DMSO:PBS=3:7) was administrated. To exclude the systemic effect of drugs, the drugs with the highest dose were intraplantarly injected into the contralateral hind paw to BV injection. The spontaneous nociceptive behavioral response of rats was determined by counting the number of paw flinches during each 5 min interval of 1 h time course following injection of BV. Paw withdrawal mechanical threshold (PWMT) of the injected hind paws by von Frey filaments was examined at time prior to drugs administration and 2h after BV injection. Change in paw volume was used as a determinant of the BV-induced inflammatory responses. Based on the principle of Archimedes, the paw volume was measured by water displacement using a plethysmometer (Plethysmometer ZH-YLS-7A, Zheng Hua Bioinstruments Inc., China). Measurements were taken at 10 min prior to and 3 h after the BV injection.Results:(1) The effect of MAPK inhibitors upon BV-induced spontaneous painCompared with control group, pre-treatment with s.c. injection of P38 inhibitor SB202190 of different doses, either into ipsilateral hind paw or contralateral hind paw had no effect on BV-induced spontaneous nociception. Pre-treatment with s.c. injection of PD98059 with 1ug into ipsilateral hind paw produced a significant inhibition of BV-induced spontaneous flinching reflex over the observed 1 hour, compared with control group. Pre-treatment with s.c. injection of PD98059 with 10ug or 100ug into ipsilateral hind paw only produced a significant inhibition of BV-induced spontaneous flinching reflex in the late phase (26-60 min), compared with control group. Pre-treatment with s.c. injection of U0126 with 0.1ug facilitate the BV-induced spontaneous flinching reflex. Pre-treatment with s.c. injection of U0126 with 1ug had no influence on BV-induced spontaneous flinching reflex, while pre-treatment with s.c. injection of U0126 with 10ug only produced a significant inhibition of BV-induced spontaneous flinching reflex in the late phase (26-60 min), compared with control group.(2) The effect of MAPK inhibitors upon BV-induced hyperalgesiaCompared with control group, pre-treatment with s.c. injection of PD98059 and U0126 of different doses had no effect on BV-induceddecrease in PWMT.However, pre-treatment with s.c. injection of SB202190 (0.1ug-10ug) completely prevented BV-induced decrease in PWMT, compared with control group, without dose-dependent effect. All contralateral drugs treatment had no effect on BV-induced hyperalgesia, ruling out the systemic effect of these drugs.(3) The effect of MAPK inhibitors upon BV-induced inflammationCompared with control group, pre-treatment with s.c. injection of PD98059 and U0126 of different doses had no effect on BV-induced increase in paw volume. Similar to the results of PD98059 and U0126, pre-treatment with s.c. injection of SB202190 (0.1ug-1ug) had no effect on BV-induced increase in paw volume, while s.c. injection of 10 ug SB202190 produced a significant inhibition of BV-induced increase in PV, compared with control group. All contralateral drugs treatment had no effect on BV-induced BV-induced increase in PV, ruling out the systemic effect of these drugs.Conlusion(1) Peripheral MEK1/2-MAPK signal transduction pathway may mediate BV-induced spontaneous flinching reflex. MEK1/2 expression level may play a different role in BV-induced spontaneous flinching reflex: low expression level may inhibit, but not facilitate BV-induced spontaneous pain behavior, while high expression level may facilitate BV-induced spontaneous pain behavior. P38—MAPK signal transduction pathway was not involved in BV-induced spontaneous flinching reflex. ERK1/2—MAPK signal transduction pathway may play a role in BV-induced spontaneous flinching reflex.(2) Peripheral P38—MAPK signal transduction pathway may contribute to BV-induced mechanical hyperalgesia. MEK1/2-ERK1/2 signal transduction pathway did not play a role in BV-induced mechanical hyperalgesia.(3) ERK1/2-MEK1/2 signal transduction pathway wasn't involved in BV-induced inflammation, while P38-MAPK signal transduction pathway may contribute to BV-induced inflammation.