Neuropathic pain is a common cause of pain after nerve injury, but its molecular basis is poorly understood. In a post-genechip microarray effort to identify new target genes contributing to neuropathic pain development, we report here the characterization of a novel neuropathic pain contributor, thrombospondin-4 (TSP4), using a neuropathic pain model of spinal nerve ligation injury. TSP4 is mainly expressed in astrocytes and significantly upregulated in the injury side of dorsal spinal cord that correlates with the development of neuropathic pain states. TSP4 blockade either by intrathecal antibodies, antisense oligodeoxynucleotides, or inactivation of the TSP4 gene reverses or prevents behavioral hypersensitivities. Intrathecal injection of TSP4 protein into naïve rats is sufficient to enhance the frequency of excitatory postsynaptic currents in spinal dorsal horn neurons, suggesting an increased excitatory pre-synaptic input, and cause similar behavioral hypersensitivities. Together, these findings support that injury-induced spinal TSP4 may contribute to spinal pre-synaptic hypersensitivity and neuropathic pain states. Development of TSP4 antagonists has the therapeutic potential for target-specific neuropathic pain management.

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Spinal cord injury (SCI) commonly results in the development of neuropathic pain, which can dramatically impair the quality of life for SCI patients. SCI induced neuropathic pain can be manifested as both tactile allodynia (a painful sensation to a non-noxious stimulus) and or hyperalgesia (an enhanced sensation to a painful stimulus). The mechanisms underlying these pain states are poorly understood. Clinical studies have shown that gabapentin, a drug that binds to the voltage gated calcium channel alpha-2-delta-1 subunit (Cavα2δ-1) proteins is effective in the management of SCI induced neuropathic pain. Accordingly, we hypothesized that tactile allodynia post SCI is mediated by an upregulation of Cavα2δ-1 in dorsal spinal cord (DSC). To test this hypothesis, we examined if SCI-induced dysregulation of spinal Cavα2δ-1 plays a contributory role in below-level allodynia development in a rat spinal T9 contusion injury model. We found that Cavα2δ-1 expression levels were significantly increased in L4-6 dorsal, but not ventral, spinal cord of SCI rats that correlated with tactile allodynia development in the hindpaw plantar surface. Furthermore, both intrathecal gabapentin treatment and blocking SCI induced Cavα2δ-1 protein upregulation by intrathecal Cavα2δ-1 antisense oligodeoxynucleotides could reverse tactile allodynia in SCI rats. These findings support that SCI induced Cavα2δ-1 upregulation in spinal dorsal horn is a key component in mediating below-level neuropathic pain development and selectively targeting this pathway may provide effective pain relief for SCI patients.

There is increasing motivation to develop clinically relevant experimental models for cervical SCI in rodents and techniques to assess deficits in forelimb function. Here we describe a bilateral cervical contusion model in rats. Female Sprague-Dawley rats received mild or moderate cervical contusion injuries (using the Infinite Horizons device) at C5, C6, or C7/8. Forelimb motor function was assessed using a Grip Strength Meter (GSM); sensory function was assessed by the von Frey hair test; the integrity of the corticospinal tract (CST) was assessed by biotinylated dextran amine (BDA) tract tracing. Mild contusions caused primarily dorsal column (DC) and gray matter (GM) damage while moderate contusions produced additional damage to lateral and ventral tissue. Forelimb and hindlimb function was severely impaired immediately post-injury, but all rats regained the ability to use their hindlimbs for locomotion. Gripping ability was abolished immediately after injury but recovered partially, depending upon the spinal level and severity of the injury. Rats exhibited a loss of sensation in both fore- and hindlimbs that partially recovered, and did not exhibit allodynia. Tract tracing revealed that the main contingent of CST axons in the DC was completely interrupted in all but one animal whereas the dorsolateral CST (dlCST) was partially spared, and dlCST axons gave rise to axons that arborized in the GM caudal to the injury. Our data demonstrate that rats can survive significant bilateral cervical contusion injuries at or below C5 and that forepaw gripping function recovers after mild injuries even when the main component of CST axons in the dorsal column is completely interrupted.

This study was undertaken as part of the NIH "Facilities of Research-Spinal Cord Injury" project to support independent replication of published studies. Here, we repeated a study reporting that treatment with the NgR antagonist peptide NEP1-40 results in enhanced growth of corticospinal and serotonergic axons and enhanced locomotor recovery after thoracic spinal cord injury. Mice received dorsal hemisection injuries at T8 and then received either NEP1-40, Vehicle, or a Control Peptide beginning 4-5 hours (early treatment) or 7 days (delayed treatment) post-injury. CST axons were traced by injecting BDA into the sensorimotor cortex. Serotonergic axons were assessed by immunocytochemistry. Hindlimb motor function was assessed using the BBB and BMS scales, kinematic and footprint analyses, and a grid climbing task. There were no significant differences between groups in the density of CST axon arbors in the gray matter rostral to the injury or in the density of serotonergic axons caudal to the injury. Tract tracing revealed that a small number of CST axons extended past the lesion in the ventral column in some mice in all treatment groups. The proportion of mice with such axons was higher in the NEP1-40 groups that received early treatment. In one experiment, mice treated with either NEP1-40 or a Control Peptide (reverse sequence) had higher BBB and BMS scores than Vehicle-treated controls at the early post-injury testing intervals, but scores converged at later intervals. There were no statistically significant differences between groups on other functional outcome measures. In a second experiment comparing NEP-treated and Vehicle controls, there were no statistically significant differences on any of the functional outcome measures. Together, our results suggest that treatment with NEP1-40 created a situation that was slightly more conducive to axon regeneration or sprouting. Enhanced functional recovery was not seen consistently with the different functional assessments, however.