Expression of metabotropic glutamate receptor mRNAs in the human spinal cord: implications for selective vulnerability of spinal motor neurons in amyotrophic lateral sclerosis

To elucidate the relevance of metabotropic glutamate receptors (mGluRs) to the selective vulnerability of motor neurons in the spinal cord in patients with amyotrophic lateral sclerosis (ALS), we investigated the distribution of mRNAs coding mGluR1-5 in the normal human spinal cord. The mRNAs for mGluR1, 4 and 5 were observed in the spinal gray matter, whereas mGluR2 mRNA was absent in the spinal cord and mGluR3 mRNA was displayed only on glial cells in the white matter. Signals for mGluR1 and mGluR5 were enriched in the dorsal horn, while mGluR4 mRNA was abundant in the ventral horn. Since agonists to group I mGluRs (mGluR 1 and 5) have been demonstrated to have neuroprotective effects on spinal motor neurons, less expression of mRNAs coding mGluR1 and mGluR5 in the ventral horn than in the dorsal horn may be implicated in the selective susceptibility of spinal motor neurons in ALS.     

Temporal plasticity of dorsal horn somatosensory neurons after acute and chronic spinal cord hemisection in rat

Unilateral T13 hemisection of the rat spinal cord produces a model of chronic spinal cord injury (SCI) that is characterized by bilateral hyperexcitability of lumbar dorsal horn neurons, and behavioral signs of central pain. While we have demonstrated that responsiveness of multireceptive (MR) dorsal horn neurons is dramatically increased at 28 days after injury, the effects of acute hemisection are unknown and predicted to be different than observed chronically. In the present study, the consequences of T13 hemisection are examined acutely at 45 min in MR neurons both ipsilateral and contralateral to the site of injury, and compared to the same class of cells at 28 days after injury (n=20 cells total per group: 2–3 cells/side of the cord from n=5 animals). Acutely, ipsilateral to the hemisection, both spontaneous and evoked activity of MR neurons were significantly increased, whereas contralaterally, only evoked activity was significantly increased. In animals 28 days after hemisection, spontaneous activity of MR neurons was comparable to intact levels ipsilaterally, and cells exhibited hyperexcitability to evoked stimuli bilaterally. Expansion of cutaneous receptive fields was observed only in hindpaws ipsilateral to the lesion, acutely. These results demonstrate dynamic plasticity in properties of dorsal horn somatosensory neurons after SCI.     

Effects of inflammation on the ultrastructural localization of spinal cord dorsal horn group I metabotropic glutamate receptors

Inflammatory pain is thought to induce functional plasticity of spinal dorsal horn neurons and may produce changes in glutamate receptor expression. Plasticity of group I metabotropic glutamate receptors (mGluR1 and mGluR5) is important in various neuronal systems, and these receptors are also known to modulate nociceptive neurotransmission in the spinal dorsal horn. The present study aimed at determining whether persistent inflammatory pain produces alterations in intracellular and plasma membrane-associated mGluR1alpha and mGluR5 in spinal cord dorsal horn. Persistent inflammation was induced in male Long Evans rats by a unilateral intraplantar injection of 100 muL of complete Freund's adjuvant (CFA). Three days after the CFA injection thermal withdrawal latencies were obtained prior to processing of transverse spinal cord sections for preembedding immunogold labeling after incubation in primary antibody for mGluR1alpha or mGluR5. Using electron microscopy, we quantified immunogold-labeled mGluR1alpha and mGluR5 profiles, located in lamina V and I-II, respectively, of both CFA-treated rats and untreated control rats. Compared to untreated rats, CFA-treated rats had a significant increase in the number of plasma membrane-associated mGluR5 immunogold-labeled particles in lamina I-II neurons of the spinal cord. Although no changes to mGluR1alpha expression were found in CFA-treated rats, plasma membrane-associated mGluR1alpha was significantly closer to the synapse. Therefore, in CFA-treated rats there was a specific increase in the ratio of plasma membrane-associated versus intracellular immunogold-labeled particles for mGluR5, and lateral movement of mGluR1alpha toward the synapse, indicating that peripheral inflammation-induced trafficking of group I mGluRs in spinal dorsal horn neurons may be an important factor in the development of plastic changes associated with inflammation-induced chronic pain.     

Activation of p38 MAP kinase is involved in central neuropathic pain following spinal cord injury

Recent work regarding chronic central neuropathic pain (CNP) following spinal cord injury (SCI) suggests that activation of key signaling molecules such as members of the mitogen activated protein kinase (MAPK) family play a role in the expression of at-level mechanical allodynia. Previously, we have shown that the development of at-level CNP following moderate spinal cord injury is correlated with increased expression of the activated (and thus phosphorylated) forms of the MAPKs extracellular signal related kinase and p38 MAPK. The current study extends this work by directly examining the role of p38 MAPK in the maintenance of at-level CNP following spinal cord injury. Using a combination of behavioral, immunocytochemical, and electrophysiological measures we demonstrate that increased activation of p38 MAPK occurs in the spinal cord just rostral to the site of injury in rats that develop at-level mechanical allodynia after moderate SCI. Immunocytochemical analyses indicate that the increases in p38 MAPK activation occurred in astrocytes, microglia, and dorsal horn neurons in the spinal cord rostral to the site of injury. Inhibiting the enzymatic activity of p38 MAPK dose dependently reverses the behavioral expression of at-level mechanical allodynia and also decreases the hyperexcitability seen in thoracic dorsal horn neurons after moderate SCI. Taken together, these novel data are the first to demonstrate causality that increased activation of p38 MAPK in multiple cell types play an important role in the maintenance of at-level CNP following spinal cord injury.     

Glutamate and metabotropic glutamate receptors associated with innervation of the uterine cervix during pregnancy: receptor antagonism inhibits c-Fos expression in rat lumbosacral spinal cord at parturition

Dorsal root ganglia (DRG) neurons connect the spinal cord and uterine cervix, and are activated at parturition with subsequent stimulation of secondary neurons in the spinal dorsal horn and autonomic areas. Neuropeptide neurotransmitters and receptors have been studied in these areas, but amino acid transmitters, e.g., glutamate, an excitatory neurotransmitter involved in sensory and nociceptive processing, have not been characterized. To determine if glutamate is involved in innervation of the cervix, rats were examined for markers of glutamatergic neurons in the L6-S1 spinal cord, DRG and cervix. Metabotropic glutamate receptors mGluR5 in the spinal dorsal horn and their expression over pregnancy were examined in pregnant rats and pregnant rats treated continuously with an antagonist of mGluR5, 2-methyl-6-(phenylethynyl) pyridine (MPEP). Rats were allowed to deliver pups to determine if the antagonist altered the expression of an early response gene protein, Fos, in the L6-S1 cord. Immunohistochemistry showed glutamate- and vesicular glutamate transporter1 (VGluT1)-positive fibers in the cervix, glutamate- and VGluT1-expressing neurons in the DRG, some of which also exhibited retrograde tracer from cervical injections, and VGluT1 and mGluR5 immunoreactivities in the L6-S1 spinal dorsal horns. Expression of mGluR5 receptors increased over pregnancy. Fos-positive neurons were present among mGluR5-immunoreactivity in the spinal dorsal horn. Parturition-induced Fos-positive neurons in the spinal cords were abundant in control rats, but were reduced by 70% in MPEP-treated animals. These results suggest that glutamate is likely involved in the transmission of sensory signals, possibly pain, from the cervix to the spinal cord at parturition.     

Synaptic relationship of the neurons containing a metabotropic glutamate receptor, MGluR5, with nociceptive primary afferent and GABAergic terminals in rat spinal superficial laminae

Recent pharmacological evidence showed that metabotropic glutamate receptors (mGluRs), particularly mGluRs1/5, had a potential role in spinal nociceptive processing. However, previous morphological studies on mGluRs have been limited mainly to their distribution in the spinal cord. In the present study, electron microscopic immunocytochemistry was employed to identify the synaptic relationship of the neurons containing mGluR5, with nociceptive primary afferent and gamma-aminobutyric acid-ergic (GABAergic) terminals in the superficial dorsal horn of the spinal cord. Nociceptive C- and A(delta)-primary afferent terminals selectively labeled with horseradish peroxidase conjugated to wheat-germ agglutinin were in asymmetric synaptic contacts with or in direct apposition to mGluR5 positive dendritic profiles. The double-labeling studies revealed that mGluR5 immunoreactive dendrites also received symmetric synaptic contacts from axon terminals labeled with immunogold particles indicating GABA. The present demonstration of mGluR5 neurons receiving inputs from both nociceptive primary afferents and GABAergic terminals of presumed interneurons further supports the involvement of mGluR5 in the transmission and modulation of nociceptive information in the spinal cord.     

Spinal cord injury triggers sensitization of wide dynamic range dorsal horn neurons in segments rostral to the injury

A spinal cord injury (SCI) was produced in adult rats by complete spinal cord transection at L6-S1. Neuropathic pain behaviors similar to the chronic central pain (CCP) syndrome in human, such as thermal hyperalgesia, mechanical allodynia and autotomy, were present in these rats after spinal cord injury. Meanwhile, wide dynamic range (WDR) neurons recorded in the spinal dorsal horn rostral to the lesion responded as high frequency of spontaneous activities, long duration of after-discharges to noxious electrical stimuli and an augmented wind-up to 0.5 Hz stimuli. By using bupivacaine powder, a sodium channel blocker, at the locus of transection immediate after nerve injury, the chronic pain behaviors were prevented; the hyperexcitability of WDR neurons was also substantially reduced. It is suggested that spinal cord transection induces the CCP syndromes, which may be evoked and maintained by the hyperexcitability in WDR neurons rostrally. Reducing the neuronal activity at the site of lesion following injury may prevent the development of CCP after SCI.     

Changes in metabotropic glutamate receptor expression following spinal cord injury

Spinal cord injury (SCI) initiates biochemical events that lead to an increase in extracellular excitatory amino acid concentrations, resulting in glutamate receptor-mediated excitotoxic events. These receptors include the three groups of metabotropic glutamate receptors (mGluRs). Group I mGluR activation can initiate a number of intracellular pathways that increase neuronal excitability. Group II and III mGluRs may function as autoreceptors to modulate neurotransmission. Thus, all three groups may contribute to the mechanisms of central sensitization and chronic central pain. To begin evaluating mGluRs in SCI, we quantified the changes in mGluR expression after SCI in control (naive), sham, and impact injured adult male Sprague-Dawley rats (200-250 g). SCI was produced at spinal segment T10 with a New York University impactor (12.5-mm drop, 10-g rod of 2-mm diameter). Expression levels were determined by Western blot and immunohistochemistry analyses at the epicenter of injury, as well as segments rostral and caudal. The group I subtype mGluR1 was increased over control levels in segments rostral and caudal by postsurgical day (PSD) 7 and remained elevated through PSD 60. The group I subtype mGluR5 was unchanged in all segments rostral and caudal to the injury at every time point measured. Group II mGluRs were decreased compared to control levels from PSD 7 through PSD 60 in all segments. These results suggest that different subtypes of mGluRs have different spatial and temporal expression patterns following SCI. The expression changes in mGluRs parallel the development of mechanical allodynia and thermal hyperalgesia following SCI; therefore, understanding the expression of mGluRs after SCI may give insight into mechanisms underlying the development of chronic central pain.     

Spatial and temporal activation of spinal glial cells: role of gliopathy in central neuropathic pain following spinal cord injury in rats

In the spinal cord, neuron and glial cells actively interact and contribute to neurofunction. Surprisingly, both cell types have similar receptors, transporters and ion channels and also produce similar neurotransmitters and cytokines. The neuroanatomical and neurochemical similarities work synergistically to maintain physiological homeostasis in the normal spinal cord. However, in trauma or disease states, spinal glia become activated, dorsal horn neurons become hyperexcitable contributing to sensitized neuronal-glial circuits. The maladaptive spinal circuits directly affect synaptic excitability, including activation of intracellular downstream cascades that result in enhanced evoked and spontaneous activity in dorsal horn neurons with the result that abnormal pain syndromes develop. Recent literature reported that spinal cord injury produces glial activation in the dorsal horn; however, the majority of glial activation studies after SCI have focused on transient and/or acute time points, from a few hours to 1 month, and peri-lesion sites, a few millimeters rostral and caudal to the lesion site. In addition, thoracic spinal cord injury produces activation of astrocytes and microglia that contributes to dorsal horn neuronal hyperexcitability and central neuropathic pain in above-level, at-level and below-level segments remote from the lesion in the spinal cord. The cellular and molecular events of glial activation are not simple events, rather they are the consequence of a combination of several neurochemical and neurophysiological changes following SCI. The ionic imbalances, neuroinflammation and alterations of cell cycle proteins after SCI are predominant components for neuroanatomical and neurochemical changes that result in glial activation. More importantly, SCI induced release of glutamate, proinflammatory cytokines, ATP, reactive oxygen species (ROS) and neurotrophic factors trigger activation of postsynaptic neuron and glial cells via their own receptors and channels that, in turn, contribute to neuronal-neuronal and neuronal-glial interaction as well as microglia-astrocytic interactions. However, a systematic review of temporal and spatial glial activation following SCI has not been done. In this review, we describe time and regional dependence of glial activation and describe activation mechanisms in various SCI models in rats. These data are placed in the broader context of glial activation mechanisms and chronic pain states. Our work in the context of work by others in SCI models demonstrates that dysfunctional glia, a condition called "gliopathy", is a key contributor in the underlying cellular mechanisms contributing to neuropathic pain.     

Expression of CAPON after Spinal Cord Injury in Rats

The adaptor protein, carboxy-terminal PDZ ligand of nNOS (CAPON), regulates the distribution of neuronal nitric oxide synthase (nNOS) that increased after spinal cord injury (SCI) and produces the key signaling molecule nitric oxide (NO). But little is known about the role of CAPON in the pathological process of SCI. The main objective of the present study was to investigate expression of CAPON and nNOS in a spinal cord contusion model in adult rats. Real time-polymerase chain reaction (PCR) and Western blot analysis revealed that mRNA and protein for CAPON increased at 2 h after SCI and reached the peak at 8 h, gradually recovered to the baseline level at 14 days. The expression of nNOS mRNA and protein was similar to that of CAPON. During the peak expression, CAPON mRNA was found in the ventral horn, mediate zone, dorsal horn, and white matter by in situ hybridization. Immunofluorescence showed that CAPON was colocalized with nNOS in neurons, oligodendrocytes, and some astrocytes of spinal cord tissues within 5 mm from the epicenter. Interaction between CAPON and nNOS was also detected by co-immunoprecipitation. Thus, the transient expression of high levels of CAPON may provide new insight into the secondary response after SCI. Chun Cheng and Xin Li contributed equally to this work.     

Temporal and spatial distribution of Fos protein in the lumbar spinal dorsal horn neurons in the rat with chronic constriction injury to the sciatic nerve

The temporal and spatial expression pattern of Fos protein in spinal dorsal horn neurons was examined by immunohistochemistry in rats with chronic constriction injury (CCI) to the sciatic nerve. In normal animals, a few Fos-immunoreactive (-IR) neurons were detected in the dorsal horn of the lumbar spinal cord. Following induction of CCI, a very large number of Fos-IR neurons appeared in the spinal dorsal horn, but a significant number of Fos-IR neurons were also observed in the contralateral dorsal horn where primary afferents of the injured sciatic nerve rarely project. Sham-operated animals also had a significant number of Fos-IR neurons in the dorsal horn bilaterally. The number of Fos-IR neurons reached its maximal level 1 day following placement of the ligatures (PO 1d). The ratio of the number of Fos-IR neurons in the ipsilateral dorsal horn to the contralateral dorsal horn, however, had its peak level 3 days following CCI (3.1-fold increase compared to the contralateral dorsal horn). The number of Fos-IR neurons in the dorsal horn gradually decreased, but increased again around PO 15d. On PO 30d, the number of Fos-IR neurons decreased and became comparable to that in normal animals. The present results indicate that the induction of Fos-IR neurons in the dorsal horn caused by CCI is biphasic and reaches its maximal level on PO 3d, near the time of hyperalgesia onset.     

Differential distribution of metabotropic glutamate receptors 1a, 1b, and 5 in the rat spinal cord

Metabotropic glutamate receptors (mGluRs) modulate somatosensory, autonomic, and motor functions at spinal levels. mGluR postsynaptic actions over spinal neurons display the pharmacologic characteristics of type I mGluRs; however, the spinal distribution of type I mGluR isoforms remains poorly defined. In this study, the authors describe a differential distribution of immunoreactivity to various type I mGluR isoforms (mGluR1a, mGluR5a,b, and mGluR1b) that suggests a correlation between specific isoforms and particular aspects of spinal cord function. Two different antisera raised against mGluR5a,b detected intense immunoreactivity within nociceptive afferent terminal fields (laminae I and II) and also in autonomic regions (parasympathetic and sympathetic). In contrast, two of three anti-mGluR1a antibodies did not immunostain lamina I or II. Laminae I and II immunostaining by a third anti-mGluR1a antibody was competed by a peptide sequence obtained from a homologous region in mGluR5, suggesting possible cross reactivity in fixed tissue. Autonomic neurons did not express mGluR1a immunoreactivity. All anti-mGluR1a antibodies strongly and specifically immunolabeled dendritic and somatic membranes of neurons in the deep dorsal horn (lamina III-V) and the ventral horn (lamina VI-IX). Somatic motoneurons expressed mGluR1a immunoreactivity but little or no mGluR5 immunoreactivity. Phrenic and pudendal motoneurons expressed the highest level of mGluR1a immunoreactivity in the spinal cord. Intense mGluR1b immunoreactivity was restricted to a few scattered neurons and a prominent group of neurons in lamina X. Lamina II neurons expressed low levels of mGluR1b immunoreactivity. Ultrastructurally, type I mGluR immunoreactivity was found mostly at extrasynaptic sites on the plasma membrane, but it was also found perisynaptically, in the body of the postsynaptic regions or in relation to intracytoplasmic structures.     

Role of the spinal cord NR2B-containing NMDA receptors in the development of neuropathic pain

Activation of N-methyl-d-aspartate (NMDA) receptors in the spinal dorsal horn has been shown to be essential for the initiation of central sensitization and the hyperexcitability of dorsal horn neurons in chronic pain. However, whether the spinal NR2B-containing NMDA (NMDA-2B) receptors are involved still remains largely unclear. Using behavioral test and in vivo extracellular electrophysiological recording in L5 spinal nerve-ligated (SNL) neuropathic rats, we investigate the roles of spinal cord NMDA-2B receptors in the development of neuropathic pain. Our study showed that intrathecal (i.t.) injection of Ro 25-6981, a selective NMDA-2B receptor antagonist, had a dose-dependent anti-allodynic effect without causing motor dysfunction. Furthermore, i.t. application of another NMDA-2B receptor antagonist ifenprodil prior to SNL also significantly inhibited the mechanical allodynia but not the thermal hyperalgesia. These data suggest that NMDA-2B receptors at the spinal cord level play an important role in the development of neuropathic pain, especially at the early stage following nerve injury. In addition, spinal administration of Ro 25-6981 not only had a dose-dependent inhibitory effect on the C-fiber responses of dorsal horn wide dynamic range (WDR) neurons in both normal and SNL rats, but also significantly inhibited the long-term potentiation (LTP) in the C-fiber responses of WDR neurons induced by high-frequency stimulation (HFS) applied to the sciatic nerve. These results indicate that activation of the dorsal horn NMDA-2B receptors may be crucial for the spinal nociceptive synaptic transmission and for the development of long-lasting spinal hyperexcitability following nerve injury. In conclusion, the spinal cord NMDA-2B receptors play a role in the development of central sensitization and neuropathic pain via the induction of LTP in dorsal horn nociceptive synaptic transmission. Therefore, the spinal cord NMDA-2B receptor is likely to be a target for clinical pain therapy.     

Expression of acetylated histone 3 in the spinal cord and the effect of morphine on inflammatory pain in rats

In this study, a rat model of inflammatory pain was produced by injecting complete Freund’s adjuvant into the hind paw, and the expression of acetylated histone 3 in the spinal cord dorsal horn was examined using immunohistochemical staining. One day following injection, there was a dramatic decrease in acetylated histone 3 expression in spinal cord dorsal horn neurons. However, on day 7, expression recovered in adjuvant-injected rats. While acetylated histone 3 labeling was present in dorsal horn neurons, it was more abundant in astrocytes and microglial cells. The recovery of acetylated histone 3 expression was associated with a shift in expression of the protein from neurons to glial cells. Morphine injection significantly upregulated the expression of acetylated histone 3 in spinal cord dorsal horn neurons and glial cells 1 day after injection, especially in astrocytes, preventing the transient downregulation. Our results indicate that inflammatory pain induces a transient downregulation of acetylated histone 3 in the spinal cord dorsal horn at an early stage following adjuvant injection, and that this effect can be reversed by morphine. Thus, the downregulation of acetylated histone 3 may be involved in the development of inflammatory pain.     

Activation of p-38α MAPK contributes to neuronal hyperexcitability in caudal regions remote from spinal cord injury

In the present study, we examined whether activation of p-38α MAPK modulates mechanical allodynia and neuronal hyperexcitability, and if propentofylline (PPF, a glial modulator) modulates specifically localized activated p-38α MAPK expression in caudal regions remote from a low thoracic hemisection injury in rats. T13 spinal hemisection produces bilateral mechanical allodynia in hindpaws with evoked (in response to mechanical stimuli) neuronal hyperexcitability in lumbar spinal wide dynamic range (WDR) neurons compared to sham controls. The mechanical allodynia and the evoked activity of WDR neurons is attenuated by intrathecal and topical administration of SB203580, an inhibitor of p-38 MAPK activation, dose dependently (p < 0.05); however, the spontaneous activity showed no significant differences compared to sham controls. After T13 spinal hemisection, significantly increased phosphorylated (activated form) p-38α MAPK expression was present in both superficial and deep dorsal horn neurons as well as in microglia, but not in astrocytes, in the lumbar spinal cord compared to sham controls (p < 0.05). Intrathecal application of PPF significantly attenuated the expression of phosphorylated p-38α MAPK in superficial dorsal horn neurons (10 mM) and in microglia (1 and 10 mM) in the lumbar spinal cord compared to the hemisection group (p < 0.05). In conclusion, our present data demonstrate that activated neuronal and microglial, but not astrocytic, p-38α MAPK contributes to the maintenance of neuronal hyperexcitability in caudal regions following spinal cord injury.     

EphA4 deficient mice maintain astroglial–fibrotic scar formation after spinal cord injury

One important aspect of recovery and repair after spinal cord injury (SCI) lies in the complex cellular interactions at the injury site that leads to the formation of a lesion scar. EphA4, a promiscuous member of the EphA family of repulsive axon guidance receptors, is expressed by multiple cell types in the injured spinal cord, including astrocytes and neurons. We hypothesized that EphA4 contributes to aspects of cell–cell interactions at the injury site after SCI, thus modulating the formation of the astroglial–fibrotic scar. To test this hypothesis, we studied tissue responses to a thoracic dorsal hemisection SCI in an EphA4 mutant mouse line. We found that EphA4 expression, as assessed by β-galactosidase reporter gene activity, is associated primarily with astrocytes in the spinal cord, neurons in the cerebral cortex and, to a lesser extent, spinal neurons, before and after SCI. However, we did not observe any overt reduction of glial fibrillary acidic protein (GFAP) expression in the injured area of EphA4 mutants in comparison with controls following SCI. Furthermore, there was no evident disruption of the fibrotic scar, and the boundary between reactive astrocytes and meningeal fibroblasts appeared unaltered in the mutants, as were lesion size, neuronal survival and inflammation marker expression. Thus, genetic deletion of EphA4 does not significantly alter the astroglial response or the formation of the astroglial–fibrotic scar following a dorsal hemisection SCI in mice. In contrast to what has been proposed, these data do not support a major role for EphA4 in reactive astrogliosis following SCI.     

Bladder and urethral sphincter responses evoked by microstimulation of S2 sacral spinal cord in spinal cord intact and chronic spinal cord injured cats

Urinary bladder and urethral sphincter responses evoked by bladder distention, ventral root stimulation, or microstimulation of S2 segment of the sacral spinal cord were investigated under alpha-chloralose anesthesia in cats with an intact spinal cord and in chronic spinal cord injured (SCI) cats 6-8 weeks after spinal cord transection at T9-T10 spinal segment. Both SCI and normal cats exhibited large amplitude reflex bladder contractions when the bladder was fully distended. SCI cats also exhibited hyperreflexic bladder contractions during filling and detrusor-sphincter dyssynergia during voiding, neither was observed in normal cats. Electrical stimulation of the ventral roots revealed that the S2 sacral spinal cord was the most effective segment for evoking large amplitude bladder contractions or voiding in both types of cats. Microstimulation with a stimulus intensity of 100 microA and duration of 30-60 s via a single microelectrode in the S2 lateral ventral horn or ventral funiculus evoked large amplitude bladder contractions with small urethral contractions in both normal and SCI cats. However, this stimulation evoked incomplete voiding due to either co-activation of the urethral sphincter or detrusor-sphincter dyssynergia. Stimulation in the S2 dorsal horn evoked large amplitude sphincter responses. The effectiveness of spinal cord microstimulation with a single electrode to induce prominent bladder and urethral sphincter responses in SCI animals demonstrates the potential for using microstimulation techniques to modulate lower urinary tract function in patients with neurogenic voiding dysfunctions.     

Intrathecal injection of carbenoxolone, a gap junction decoupler, attenuates the induction of below-level neuropathic pain after spinal cord injury in rats

The most common type of chronic pain following spinal cord injury (SCI) is central neuropathic pain and SCI patients typically experience mechanical allodynia and thermal hyperalgesia. The present study was designed to examine the potential role of astrocyte gap junction connectivity in the induction and maintenance of “below-level” neuropathic pain in SCI rats. We examined the effect of intrathecal treatment with carbenoxolone (CARB), a gap junction decoupler, on SCI-induced bilateral thermal hyperalgesia and mechanical allodynia during the induction phase (postoperative days 0 to 5) and the maintenance phase (days 15 to 20) following T13 spinal cord hemisection. Immunohistochemistry was performed to determine potential SCI-induced changes in spinal astrocyte activation and phosphorylation of the NMDA receptor NR1 subunit (pNR1). CARB administered during the induction period dose-dependently attenuated the development of bilateral thermal hyperalgesia and mechanical allodynia. Intrathecal CARB also significantly reduced the bilateral SCI-induced increase in GFAP-immunoreactive (ir) staining and the number of pNR1-ir cell profiles in the spinal cord dorsal horn compared to vehicle-treated rats. In contrast, CARB treatment during the maintenance phase had no effect on the established thermal hyperalgesia and mechanical allodynia nor on spinal GFAP expression or the number of pNR1-ir cell profiles. These results indicate that gap junctions play a critical role in the activation of astrocytes distant from the site of SCI and in the subsequent phosphorylation of NMDA receptors in the lumbar spinal cord. Both of these processes appear to contribute to the induction of bilateral below-level pain in SCI rats.