Effect of hyposerotonin on NMDA receptor phosphorylation in rat trigeminal pathway induced by dural inflammation

Abstract

This study aims to investigate the effect of serotonin depletion on the phosphorylation of NMDA receptor and NMDA receptor expression in trigeminal nucleus caudalis (TNC), and on trigeminal nociception evoked by dural inflammation. Male Wistar rats were separated into low serotonin and normal serotonin groups. Serotonin was depleted by peritoneal injection with para-chlorophenylalanine, a tryptophan hydroxylase inhibitor, 3 days prior to the experiment. To induce trigeminal nociception, inflammatory soup (IS) containing a mixture of inflammatory mediators (histamine, serotonin, bradykinin, each at 1 mM, and 0.1 mM prostaglandin E₂, pH 5.5) or low-pH artificial cerebrospinal fluid (low-pH CSF; pH 4.7) was applied on exposed dural surface for thirty minutes. In control groups, artificial cerebrospinal fluid was applied instead. Thirty minutes and two hours after induction, the brainstem and cervical cord were removed for serine-896 phosphorylated NMDA receptor NR1 subunit (pNR1), NMDA receptor NR1 subunit (NR1), and Fos immunohistochemical studies. Trigeminal nociception was determined by the number of Fos-immunoreactive neurons in the TNC. The results showed that dural application of IS, not low-pH CSF, led to the activation of trigeminal nociceptive system as well as phosphorylation of NR1 receptor. The numbers of pNR1- and Fos-immunoreactive cells were significantly greater in rats receiving IS than those of the controls. Dural inflammation-induced NR1 receptor phosphorylation as well as trigeminal nociception was enhanced in the low serotonin condition. The numbers of pNR1- and Fos-immunoreactive cells were significantly greater in serotonin-depleted rats than those of the normal serotonin rats, especially in IS group. Neither meningeal inflammation nor serotonin depletion altered NR1 receptor expression. The number of NR1-immunoreactive cells was not significantly different in all groups. There was a relationship between NR1 receptor phosphorylation and trigeminal nociception. Strong correlation between numbers of pNR1- and Fos-immunoreactive cells in the TNC was demonstrated. In normal serotonin rats, this correlation was able to represent by the linear equation of y = 0.520x (r²= 0.957, P < .001). However, the slope of equation was elevated in low serotonin rats and the equation became y = 0.706x (r²= 0.941, P < .001). In conclusion, the process of meningeal inflammation induced trigeminal nociception is enhanced in the low serotonin condition. The mechanism of nociceptive facilitation may involve the increase in NR1 receptor phosphorylation, not the increase in NR1 expression. This study further supports the role of serotonin in the control of trigeminal nociception.