Kristen J. Nicholson

Upregulation of GLT‐1 by treatment with ceftriaxone alleviates radicular pain by reducing spinal astrocyte activation and neuronal hyperexcitability

Cervical nerve root injury commonly leads to radicular pain. Normal sensation relies on regulation of extracellular glutamate in the spinal cord by glutamate transporters. The goal of this study was to define the temporal response of spinal glutamate transporters (glial glutamate transporter 1 [GLT‐1], glutamate‐aspartate transporter [GLAST], and excitatory amino acid carrier 1) following nerve root compressions that do or do not produce sensitivity in the rat and to evaluate the role of glutamate uptake in radicular pain by using ceftriaxone to upregulate GLT‐1. Compression was applied to the C7 nerve root. Spinal glutamate transporter expression was evaluated at days 1 and 7. In a separate study, rats underwent a painful root compression and were treated with ceftriaxone or the vehicle saline. Glial glutamate transporter expression, astrocytic activation (glial fibrillary acidic protein [GFAP]), and neuronal excitability were assessed at day 7. Both studies measured behavioral sensitivity for 7 days after injury. Spinal GLT‐1 significantly decreased (P < 0.04) and spinal GLAST significantly increased (P = 0.036) at day 7 after a root injury that also produced sensitivity to both mechanical and thermal stimuli. Within 1 day after ceftriaxone treatment (day 2), mechanical allodynia began to decrease; both mechanical allodynia and thermal hyperalgesia were attenuated (P < 0.006) by day 7. Ceftriaxone also reduced (P < 0.024) spinal GFAP and GLAST expression, and neuronal hyperexcitability in the spinal dorsal horn, restoring the proportion of spinal neurons classified as wide dynamic range to that of normal. These findings suggest that nerve root‐mediated pain is maintained jointly by spinal astrocytic reactivity and neuronal hyperexcitability and that these spinal modifications are associated with reduced glutamate uptake by GLT‐1.

Time-dependent mechanics and measures of glial activation and behavioral sensitivity in a rodent model of radiculopathy.

Nerve root compression induces persistent behavioral hypersensitivity and spinal glial reactivity. Viscoelastic properties of neural tissues suggest that physiologic outcomes may depend on the duration of an applied nerve root compression. This study evaluated the time-dependent properties of the root under compression in the context of pain-related behavioral and physiologic outcomes. The decrease in applied load measured by load relaxation under compression was quantified for rat cervical (C6-C8) roots in situ for durations of 30 sec, 3 min, or 15 min (n = 6). Immediately following compression, the change in the root width relative to its original width was quantified as a measure of its structural recovery. Both load relaxation and structural recovery were significantly (p < 0.05) correlated with duration of compression. After 30 sec of compression, load relaxed by 22 +/- 10%; increasing to 36 +/- 18% and 56 +/- 20% at 3 and 15 min, respectively. Following 30 sec, 3 min, and 15 min of compression, the root recovered to 91 +/- 5%, 88 +/- 5 and 72 +/- 13% of its original width, respectively. A companion in vivo study imposed these same compression durations and sham procedures to the C7 root to evaluate pain symptoms and spinal glial reactivity. Allodynia was assessed for 7 days to measure behavioral sensitivity. Immunohistochemistry and quantitative densitometry detected GFAP and OX-42 in the dorsal horn at day 7. Significant correlations were detected between compression duration and allodynia (p < 0.03), and astrocyte and microglial activation (p < 0.01). These biomechanical and glial results imply that a similar duration of compression may modulate both sustained pain and spinal glial reactivity.

TRANSIENT NERVE ROOT COMPRESSION LOAD AND DURATION DIFFERENTIALLY MEDIATE BEHAVIORAL SENSITIVITY AND ASSOCIATED SPINAL ASTROCYTE ACTIVATION AND MGLUR5 EXPRESSION

Injury to the cervical nerve roots is a common source of neck pain. Animal models of nerve root compression have previously established the role of compression magnitude and duration in nerve root-mediated pain and spinal inflammation; yet, the response of the spinal glutamatergic system to transient nerve root compression and its relationship to compression mechanics have not been studied. The glutamate receptor, mGluR5, has a central role in pain, and its expression by neurons and astrocytes in the spinal cord may be pivotal for neuronal-glial signaling. This study quantified spinal GFAP and mGluR5 expression following nerve root compressions of different magnitudes and durations in the rat. Compression to the C7 nerve root was applied for a duration that was either above (10 min) or below (3 min) the critical duration for mediating afferent discharge rates during compression. To also test for the effect of the magnitude of the compression load, either a 10 gf or a 60 gf was applied to the nerve root for each duration. Mechanical allodynia was assessed, and the C7 spinal cord was harvested on day 7 for immunofluorescent analysis. Double labeling was used to localize the expression of mGluR5 on astrocytes (GFAP) and neurons (MAP2). Seven days after injury, 10 min of compression produced significantly greater behavioral sensitivity (P<0.001) and spinal GFAP expression (P=0.002) than 3 min of compression, regardless of the compression magnitude. Nerve root compression at 60 gf produced a significant increase (P<0.001) in spinal mGluR5 for both of the durations studied. There was no difference in the distribution of mGluR5 between astrocytes and neurons following nerve root compression of any type. The glutamatergic and glial systems are differentially modulated by the mechanics of nerve root compression despite the known contribution of glia to pain through glutamatergic signaling.

Riluzole effects on behavioral sensitivity and the development of axonal damage and spinal modifications that occur after painful nerve root compression

OBJECT Cervical radiculopathy is often attributed to cervical nerve root injury, which induces extensive degeneration and reduced axonal flow in primary afferents. Riluzole inhibits neuro-excitotoxicity in animal models of neural injury. The authors undertook this study to evaluate the antinociceptive and neuroprotective properties of riluzole in a rat model of painful nerve root compression. METHODS A single dose of riluzole (3 mg/kg) was administered intraperitoneally at Day 1 after a painful nerve root injury. Mechanical allodynia and thermal hyperalgesia were evaluated for 7 days after injury. At Day 7, the spinal cord at the C-7 level and the adjacent nerve roots were harvested from a subgroup of rats for immunohistochemical evaluation. Nerve roots were labeled for NF200, CGRP, and IB4 to assess the morphology of myelinated, peptidergic, and nonpeptidergic axons, respectively. Spinal cord sections were labeled for the neuropeptide CGRP and the glutamate transporter GLT-1 to evaluate their expression in the dorsal horn. In a separate group of rats, electrophysiological recordings were made in the dorsal horn. Evoked action potentials were identified by recording extracellular potentials while applying mechanical stimuli to the forepaw. RESULTS Even though riluzole was administered after the onset of behavioral sensitivity at Day 1, its administration resulted in immediate resolution of mechanical allodynia and thermal hyperalgesia (p < 0.045), and these effects were maintained for the study duration. At Day 7, axons labeled for NF200, CGRP, and IB4 in the compressed roots of animals that received riluzole treatment exhibited fewer axonal swellings than those from untreated animals. Riluzole also mitigated changes in the spinal distribution of CGRP and GLT-1 expression that is induced by a painful root compression, returning the spinal expression of both to sham levels. Riluzole also reduced neuronal excitability in the dorsal horn that normally develops by Day 7. The frequency of neuronal firing significantly increased (p < 0.045) after painful root compression, but riluzole treatment maintained neuronal firing at sham levels. CONCLUSIONS These findings suggest that early administration of riluzole is sufficient to mitigate nerve root-mediated pain by preventing development of neuronal dysfunction in the nerve root and the spinal cord.

Salmon and Human Thrombin Differentially Regulate Radicular Pain, Glial-Induced Inflammation and Spinal Neuronal Excitability through Protease-Activated Receptor-1

Chronic neck pain is a major problem with common causes including disc herniation and spondylosis that compress the spinal nerve roots. Cervical nerve root compression in the rat produces sustained behavioral hypersensitivity, due in part to the early upregulation of pro-inflammatory cytokines, the sustained hyperexcitability of neurons in the spinal cord and degeneration in the injured nerve root. Through its activation of the protease-activated receptor-1 (PAR1), mammalian thrombin can enhance pain and inflammation; yet at lower concentrations it is also capable of transiently attenuating pain which suggests that PAR1 activation rate may affect pain maintenance. Interestingly, salmon-derived fibrin, which contains salmon thrombin, attenuates nerve root-induced pain and inflammation, but the mechanisms of action leading to its analgesia are unknown. This study evaluates the effects of salmon thrombin on nerve root-mediated pain, axonal degeneration in the root, spinal neuronal hyperexcitability and inflammation compared to its human counterpart in the context of their enzymatic capabilities towards coagulation substrates and PAR1. Salmon thrombin significantly reduces behavioral sensitivity, preserves neuronal myelination, reduces macrophage infiltration in the injured nerve root and significantly decreases spinal neuronal hyperexcitability after painful root compression in the rat; whereas human thrombin has no effect. Unlike salmon thrombin, human thrombin upregulates the transcription of IL-1β and TNF-α and the secretion of IL-6 by cortical cultures. Salmon and human thrombins cleave human fibrinogen-derived peptides and form clots with fibrinogen with similar enzymatic activities, but salmon thrombin retains a higher enzymatic activity towards coagulation substrates in the presence of antithrombin III and hirudin compared to human thrombin. Conversely, salmon thrombin activates a PAR1-derived peptide more weakly than human thrombin. These results are the first to demonstrate that salmon thrombin has unique analgesic, neuroprotective and anti-inflammatory capabilities compared to human thrombin and that PAR1 may contribute to these actions.

Impaired performance on the angle board test is induced in a model of painful whiplash injury but is only transient in a model of cervical radiculopathy.

Although clinical studies report motor impairment associated with some painful injuries of the neck, assessment of motor function in animal models has been largely limited only to studies of direct trauma to the nervous system. The incline plane test was modified to evaluate motor function in two rodent pain models of facet joint distraction (FJD) and nerve root compression (NRC) injury (n = 5/group). Sham groups were also included as controls. Motor function was measured using the modified inclined plane test with rats facing downward before surgery (baseline) and following surgery on days corresponding to when mechanical sensitivity is established and remains elevated. Mean baseline values of the board angle inducing slip for FJD (45.8 ± 3.1°) was significantly greater (p = 0.014) than that for NRC (43.5 ± 2.5°), but baseline measurements did not vary for either group over time. No changes in motor function were found for shams. Motor function after FJD significantly decreased (p < 0.001) at days 1 and 7 after injury. In contrast, at day 1 after NRC injury, slip occurred at significantly lower (p = 0.0016) incline angles, but returned to baseline levels by day 7. These results show motor function impairment is induced following painful FJD and suggest the incline plane test offers utility to evaluate functional deficits in painful injuries.

Nerve and Nerve Root Biomechanics

Together, the relationship between the mechanical response of neural tissues and the related mechanisms of injury provide a foundation for defining relevant thresholds for injury. The nerves and nerve roots are biologic structures with specific and important functions, and whose response to mechanical loading can have immediate, long-lasting and widespread consequences. In particular, when nerves or nerve roots are mechanically loaded beyond their mechanical tolerance for injury, motor and/or sensory deficits can result. The severity and persistence of the symptoms are modulated by the profile of the mechanical insult. In this chapter, the relevant anatomy, and structure are reviewed in the context of biomechanical data describing the mechanical behavior of nerves and nerve roots. While both nerves and nerve roots have components of their anatomic organization that protect the axons of their neurons, there are distinct differences in their structure and composition. These variations contribute to differences in their mechanical response to loading owing to their strength and stiffness. However, both tissues are time-dependent and exhibit viscoelastic behavior. These time-dependent characteristics in mechanical responses imply that, in addition to the magnitude of force or deformation, the rate of loading and duration of applied mechanical insult can modulate physiologic outcomes associated with mechanical loading to these tissues. This chapter reviews all of these concepts in the context of biomechanics and physiologic outcomes. In addition, a review of current work studying nerve and nerve root loading in animal models is provided in order to relate these outcomes to clinical symptoms.

Duration of Nerve Root Compressive Trauma Modulates the Subsequent Thermal Hyperalgesia and Spinal Expression of the Glutamate Transporter, GLT1

Mechanical compression of the cervical nerve roots is a common injury modality [1] and a frequent source of neck pain, affecting 30–50% of adults each year [2]. Since the nerve root is viscoelastic in compression (Fig. 1) [3,4], its response to loading from different injury scenarios is also likely a function of the duration of the applied tissue insult, which varies with the type of injury. For example, the nerve root undergoes brief periods of compression during sports and auto-related trauma, whereas a more prolonged compression occurs for a bulging disc or foraminal stenosis [1]. Similarly, mechanical sensitivity (i.e. pain) after root compression is has been shown to be duration-dependent [3,4]. Compression of the cervical nerve root is only sufficient to induce mechanical sensitivity in a rat model if the compression is applied for more than 3 minutes [3]. Yet, mechanical sensitivity is only one behavioral sequelae of radicular pain and it is not known whether the duration dependent response is similar for other types of evoked pain, such as thermal sensitivity.© 2013 ASME

The Duration of a Nerve Root Compression Modulates Evoked Neuronal Responses in a Rat Model of Painful Injury

The annual incidence for neck pain in the adult population is 30–50% [1]. The cervical nerve roots are at risk for mechanical injury due to impingement of surrounding structures which can result in pain and numbness [2]. During nerve root compression, an immediate, brief increase in spontaneous afferent activity and a gradual decrease in electrically evoked axonal conduction have been reported [3,4]. Although previous studies demonstrate that a transient cervical nerve root compression induces persistent behavioral sensitivity [5,6], it is not known how the tissue mechanics during loading modulate neuronal function or how they relate to the onset of pain. Therefore, the goal of this study was to quantify neuronal activity in the spinal cord as a function of the duration of applied compression by measuring both electrically-evoked and spontaneous afferent activity during a transient compression of the cervical nerve root in a rat model of pain [5,6].Copyright