ro-25-6981 and Visceral-Pain

Irritable bowel syndrome (IBS) is one of the most common challenging diseases for clinical treatment. The aim of this study is to investigate whether transcranial direct current stimulation (tDCS) has analgesic effect on visceral hypersensitivity (VH) in an animal model of IBS as well as the underlying mechanism. As the activation of GluN2B in anterior cingulate cortex (ACC) takes part in VH, we examined whether and how GluN2B in ACC takes part in the effect of tDCS. Neonatal maternal deprivation (NMD), a valuable experimental model to study the IBS pathophysiology, was used to induce visceral hypersensitivity of rats. We quantified VH as colorectal distention threshold and performed patch-clamp recordings of ACC neurons. The expression of GluN2B were determined by RT-qPCR and Western blotting. The GluN2B antagonist Ro 25-6981 was microinjected into the rostral and caudal ACC. tDCS was performed for 7 consecutive days. It was found that NMD decreased expression of GluN2B, which could be obviously reversed by tDCS. Injection of Ro 25-6981 into rostral and caudal ACC of normal rats induced VH and also reversed the analgesic effect of tDCS. Our data sheds light on the nonpharmacological therapy for chronic VH in pathological states such as IBS.

Topics: Animals; Animals, Newborn; Disease Models, Animal; Excitatory Amino Acid Antagonists; Gyrus Cinguli; Hyperalgesia; Irritable Bowel Syndrome; Male; Patch-Clamp Techniques; Phenols; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Transcranial Direct Current Stimulation; Visceral Pain

We previously reported estrogen modulates spinal N-methyl-d-aspartate (NMDA) receptor processing of colorectal pain through changes in spinal GluN1 subunit phosphorylation/expression. The purpose of this study was to investigate whether spinal GluN2B containing NMDA receptors are involved in estrogen modulation of visceral pain processing.. Behavioral, molecular, and immunocytochemical techniques were used to determine spinal GluN2B expression/phosphorylation and function 48 h following subcutaneous injection of estradiol (E2) or vehicle (safflower oil, Saff oil) in ovariectomized rats in the absence or presence of colonic inflammation induced by mustard oil.. E2 increased the magnitude of the visceromotor response (VMR) to colorectal distention compared to Saff oil in non-inflamed rats. Intrathecal injection of the GluN2B subunit antagonist, Ro 25-6981, had no effect on the VMR in non-inflamed E2 or Saff oil rats. Colonic inflammation induced visceral hyperalgesia in E2, but not Saff oil rats. Visceral hyperalgesia in E2 rats was blocked by intrathecal GluN2B subunit selective antagonists. In inflamed rats, E2 increased GluN2B protein and gene expression in the thoracolumbar (TL), but not lumbosacral (LS), dorsal spinal cord. Immunocytochemical labeling showed a significant increase in GluN2B subunit in the superficial dorsal horn of E2 rats compared to Saff oil rats.. These data support the hypothesis that estrogen increases spinal processing of colonic inflammation-induced visceral hyperalgesia by increasing NMDA receptor activity. Specifically, an increase in the activity of GluN2B containing NMDA receptors in the TL spinal cord by estrogen underlies visceral hypersensitivity in the presence of colonic inflammation.

Topics: Animals; Behavior, Animal; Colitis; Estradiol; Estrogens; Female; Hyperalgesia; Immunohistochemistry; Lumbar Vertebrae; Mustard Plant; Ovariectomy; Phenols; Phosphorylation; Piperidines; Plant Oils; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Thoracic Vertebrae; Visceral Pain

The roles of spinal N-methyl-d-aspartic acid receptor 2B (NR2B) subunit in central sensitization of chronic visceral pain were investigated. A rat model with irritable bowel syndrome (IBS) was established by colorectal distention (CRD) on post-natal days 8-14. Responses of the external oblique muscle of the abdomen to CRD were measured to evaluate the sensitivity of visceral pain in rats. The sensitivity of visceral pain significantly increased in IBS-like rats. Expressions of spinal NR2B subunit and phosphorylated NR2B subunit significantly increased by 50-55% in IBS-like rats when compared with those in control rats. Ro 25-6981, a selective antagonist of NR2B subunit, has a dose-dependent anti-allodynic and anti-hyperalgesic effect without causing motor dysfunction in IBS-like rats. Furthermore, the activation mechanism of the spinal NR2B subunit in chronic visceral pain was also investigated. Spinal administration of genistein, a specific inhibitor of tyrosine kinases, also decreased the visceral pain hypersensitivity of IBS-like rats in a dose-dependent manner. In addition, the expression of phosphorylated NR2B subunit was decreased after spinal administration of Ro 25-6981 or genistein in IBS-like rats. In conclusion, tyrosine kinase activation-induced phosphorylation of NR2B subunit may play a crucial role in central sensitization of chronic visceral pain.

Topics: Animals; Animals, Newborn; Disease Models, Animal; Dose-Response Relationship, Drug; Electromyography; Excitatory Amino Acid Antagonists; Genistein; Irritable Bowel Syndrome; Male; Pain Measurement; Phenols; Phosphorylation; Piperidines; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Time Factors; Tyrosine; Visceral Pain

Pain and learning memory have striking similarities in synaptic plasticity. Activation of the N-methyl-D-aspartic acid receptors 2B subunits (NR2B-NMDAs) is responsible for the hippocampal LTP in memory formation. In our previous studies, we found the significant enhancement of CA1 hippocampal long-term potentiation (LTP) induced by high-frequency stimulation (HFS) in rats with chronic visceral pain. However, it is unclear whether the NR2B-NMDAs are required for the LTP in chronic visceral pain. In this study, a rat model with irritable bowel syndrome (IBS) was established by colorectal distention (CRD). The sensitivity of visceral pain and HFS-induced LTP at SC-CA1 synapses were significantly enhanced in IBS-like rats (p<0.05). In addition, hippocampal NR2B protein levels significantly increased in IBS-like rats (p<0.05). To test whether NR2B-NMDAs are responsible for the LTP, effects of Ro 25-6981, a selective antagonist of NR2B-NMDAs, on field potential in CA1 region were investigated in vitro. Our results demonstrated that Ro 25-6981 dose-dependently inhibited the facilitation of CA1 LTP in IBS-like rats. The plausible activation mechanism of hippocampal NR2B-NMDAs in the LTP enhancement was further explored. Western blot data indicated that expression of tyrosine phosphorylated NR2B protein in hippocampus significantly enhanced in IBS-like rats. Accordingly, genistein, a specific inhibitor of tyrosine kinases, dose-dependently blocked the facilitation of hippocampal LTP in IBS-like rats. Furthermore, EMG data revealed that intra-hippocampal injection of Ro 25-6981 dose-dependently attenuated the visceral hypersensitivity. In conclusion, hippocampal NR2B-NMDAs are responsible for the facilitation of CA1 LTP via tyrosine phosphorylation, which leads to visceral hypersensitivity.

Topics: Animals; CA1 Region, Hippocampal; Chronic Pain; Disease Models, Animal; Excitatory Amino Acid Antagonists; Genistein; Hippocampus; Hyperalgesia; Irritable Bowel Syndrome; Long-Term Potentiation; Male; Phenols; Piperidines; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Synapses; Visceral Pain