EphrinB2 induces pelvic-urethra reflex potentiation via Src kinase-dependent tyrosine phosphorylation of NR2B.
ABSTRACT Recently, the role of EphB receptor (EphBR) tyrosine kinase and their ephrinB ligands in pain-related neural plasticity at the spinal cord level have been identified. To test whether Src-family tyrosine kinase-dependent glutamatergic N-methyl-d-aspartate receptor NR2B subunit phosphorylation underlies lumbosacral spinal EphBR activation to mediate pelvic-urethra reflex potentiation, we recorded external urethra sphincter electromyogram reflex activity and analyzed protein expression in the lumbosacral (L(6)-S(2)) dorsal horn in response to intrathecal ephrinB2 injections. When compared with vehicle solution, exogenous ephrinB2 (5 μg/rat it)-induced reflex potentiation, in associated with phosphorylation of EphB1/2, Src-family kinase, NR2B Y1336 and Y1472 tyrosine residues. Both intrathecal EphB1 and EphB2 immunoglobulin fusion protein (both 10 μg/rat it) prevented ephrinB2-dependent reflex potentiation, as well as protein phosphorylation. Pretreatment with PP2 (50 μM, 10 μl it), an Src-family kinase antagonist, reversed the reflex potentiation, as well as Src kinase and NR2B phosphorylation. Together, these results suggest the ephrinB2-dependent EphBR activation, which subsequently provokes Src kinase-mediated N-methyl-d-aspartate receptor NR2B phosphorylation in the lumbosacral dorsal horn, is crucial for the induction of spinal reflex potentiation contributing to the development of visceral pain and/or hyperalgesia in the pelvic area.
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ABSTRACT: Patients with inflammatory gynecological/obstetrical problems often complain of irritable bowel syndrome. The authors examined whether acute uterus irritation reflexively provokes colonic motility in rat preparations. A modified colon manometry and striated abdominal muscle electromyogram activity in response to mustard oil (MO) instillation into the uterine horn were continuously recorded in anesthetized rats. The lumbosacral (L6-S1) dorsal horn was dissected to assess the level and the cellular location of phosphorylated NR2B subunit using Western blotting and immunofluorescence analysis, respectively. Finally, the uterine transient receptor potential A1 or spinal NR2B subunit was pharmacologically blocked to elucidate its roles. MO (0.1%, 0.2 ml) injected into the lower uterine horn dramatically provoked colonic hypermotility characterized by rhythmic colonic contractions (about 3-4 contractions per 10 min, n = 7) accompanied by synchronized electromyogram firing in the abdominal muscle (about 4-5 folds of control, n = 7). In addition to provoking colonic hypermotility, MO administration also up-regulated phosphorylated (about 2-3 folds of control, n = 7), but not total, NR2B expression in the dorsal horn neurons. Both intrathecal Ro 25-6981 (a selective NR2B subunit antagonist; 10 μM, 10 μl) and intrauterine HC-030031 (a selective transient receptor potential A1 receptor antagonist; 30 mg/kg, 0.2 ml) injected before the MO instillation attenuated the MO-induced colonic hypermotility and spinal NR2B phosphorylation. The comorbidity of gynecological/obstetrical and gastrointestinal problems is not coincidental but rather causal in nature, and clinicians should investigate for gynecological/urological diseases in the setting of bowel problems with no known pathological etiology.Anesthesiology 08/2013; · 6.17 Impact Factor