IP₃-mediated Calcium Signaling Is Involved In The Mechanism Of Fractalkine-induced Thermal Hyperalgesia In Mice

Partâ… Effect of fractalkine on cytosolic concentration of free Ca²⁺, and cytosolic effects in BV-2 microglia cellsObjective To study mechanisms of fractalkine-induced the changes of cytosolic concentration of free Calcium ion and cytosolic effects in BV-2 microglia cells. Methods BV-2 cells were incubated in different concentrations fractalkine, [Ca²⁺]i was measured by Confocal laser microscopy; Incubation of BV-2 microglia cells with fractalkine (10 nM) for 0, 30, 60, 120 and 240 min, p38MAPK and p-p38MAPK were determined by Western blot; Incubation of BV-2 microglia cells with fractalkine (10 nM) for 0, 6, 12, and 24 h, IL-1βand TNF-αwere determined by ELISA and RT-PCR; and the effects of pretreatment with anti-CX3CR1, 2-APB and SB203580 were determined. Results Confocal tests showed calcium influx which were induced by fractalkine, while pre-incubation with anti-CX3CR1 and 2-APB suppressed fractalkine-induced calcium signaling. Fractalkine significantly increased p-p38MAPK, IL-1βand TNF-αexpressions in the BV-2 cells. Anti-CX3CR1 and 2-APB showed an effective effect on decreasing fractalkine-induced p-p38MAPK, IL-1βand TNF-αexpressions. SB203580 showed an effective effect on decreasing fractalkine-induced IL-1βand TNF-αexpressions. Conclusion Fractalkine triggers BV-2 microglia to induce Ca²⁺ influx via IP₃R channels and causes p-p38MAPK production, thereby enhancing IL-1βand TNF-αsecretions.Partâ…¡The mechanism of fractalkine-induced thermal hyperalgesia in miceObjective To explore the mechanism of fractalkine triggering brain-derived microglia to cause thermal hyperalgesia. Methods We built a hyperalgesia model induced by intracerebroventricular injection (i.c.v.) of extrinsic fractalkine in male adult Kunming mice. According to injection (i.c.v.) of the different agent, divided into five groups: physiological saline control group, fractalkine group, anti-CX3CR1 + fractalkine group, 2-APB + fractalkine group and SB203580 + fractalkine group. The thermal nociceptive thresholds and the expressions of brain IL-1β, TNF-αand p-p38MAPK were evaluated. Finally, the target cells of fractalkine-induced hyperalgesia were determinded. Results Behavioral tests demonstrated that significant nociceptive response occurred in the Kunming mice following injection (i.c.v.) of fractalkine. Fractalkine significantly increased p-p38MAPK, IL-1βand TNF-αexpressions in the brain tissues. However pretreatment with anti-CX3CR1, 2-APB and SB203580 showed an effective anti-allodynia effect with decreased p-p38MAPK, IL-1βand TNF-αexpressions. Immunofluorescence indicated that fractalkine could co-staininged with microglia via binding its receptor CX3CR1 in the brain. Conclusion Fractalkine may trigger brain-derived microglia to induce Ca²⁺ influx via IP₃R channels and cause p-p38MAPK production, enhancing IL-1βand TNF-αsecretions and thus modulating hyperalgesia.