Use of quadrupedal step training to re-engage spinal interneuronal networks and improve locomotor function after spinal cord injury.

Can lower limb motor function be improved after a spinal cord lesion by re-engaging functional activity of the upper limbs? We addressed this issue by training the forelimbs in conjunction with the hindlimbs after a thoracic spinal cord hemisection in adult rats. The spinal circuitries were more excitable, and behavioural and electrophysiological analyses showed improved hindlimb function when the forelimbs were engaged simultaneously with the hindlimbs during treadmill step-training as opposed to training only the hindlimbs. Neuronal retrograde labelling demonstrated a greater number of propriospinal labelled neurons above and below the thoracic lesion site in quadrupedally versus bipedally trained rats. The results provide strong evidence that actively engaging the forelimbs improves hindlimb function and that one likely mechanism underlying these effects is the reorganization and re-engagement of rostrocaudal spinal interneuronal networks. For the first time, we provide evidence that the spinal interneuronal networks linking the forelimbs and hindlimbs are amenable to a rehabilitation training paradigm. Identification of this phenomenon provides a strong rationale for proceeding toward preclinical studies for determining whether training paradigms involving upper arm training in concert with lower extremity training can enhance locomotor recovery after neurological damage.

Sub-threshold spinal cord stimulation facilitates spontaneous motor activity in spinal rats.

BACKGROUND: Epidural stimulation of the spinal cord can be used to enable stepping on a treadmill (electrical enabling motor control, eEmc) after a complete mid-thoracic spinal cord transection in adult rats. Herein we have studied the effects of eEmc using a sub-threshold intensity of stimulation combined with spontaneous load-bearing proprioception to facilitate hindlimb stepping and standing during daily cage activity in paralyzed rats. METHODS: We hypothesized that eEmc combined with spontaneous cage activity would greatly increase the frequency and level of activation of the locomotor circuits in paralyzed rats. Spontaneous cage activity was recorded using a specially designed swivel connector to record EMG signals and an IR based camcorder to record video. RESULTS AND CONCLUSION: The spinal rats initially were very lethargic in their cages showing little movement. Without eEmc, the rats remained rather inactive with the torso rarely being elevated from the cage floor. When the rats used their forelimbs to move, the hindlimbs were extended and dragged behind with little or no flexion. In contrast, with eEmc the rats were highly active and the hindlimbs showed robust alternating flexion and extension resulting in step-like movements during forelimb-facilitated locomotion and often would stand using the sides of the cages as support. The mean and summed integrated EMG levels in both a hindlimb flexor and extensor muscle were higher with than without eEmc. These data suggest that eEmc, in combination with the associated proprioceptive input, can modulate the spinal networks to significantly amplify the amount and robustness of spontaneous motor activity in paralyzed rats.

Transcutaneous Cervical Spinal Cord Stimulation Combined with Robotic Exoskeleton Rehabilitation for the Upper Limbs in Subjects with Cervical SCI: Clinical Trial.

(1) Background: Restoring arm and hand function is a priority for individuals with cervical spinal cord injury (cSCI) for independence and quality of life. Transcutaneous spinal cord stimulation (tSCS) promotes the upper extremity (UE) motor function when applied at the cervical region. The aim of the study was to determine the effects of cervical tSCS, combined with an exoskeleton, on motor strength and functionality of UE in subjects with cSCI. (2) Methods: twenty-two subjects participated in the randomized mix of parallel-group and crossover clinical trial, consisting of an intervention group (n = 15; tSCS exoskeleton) and a control group (n = 14; exoskeleton). The assessment was carried out at baseline, after the last session, and two weeks after the last session. We assessed graded redefined assessment of strength, sensibility, and prehension (GRASSP), box and block test (BBT), spinal cord independence measure III (SCIM-III), maximal voluntary contraction (MVC), ASIA impairment scale (AIS), and WhoQol-Bref; (3) Results: GRASSP, BBT, SCIM III, cylindrical grip force and AIS motor score showed significant improvement in both groups (p ≤ 0.05), however, it was significantly higher in the intervention group than the control group for GRASSP strength, and GRASSP prehension ability (p ≤ 0.05); (4) Conclusion: our findings show potential advantages of the combination of cervical tSCS with an exoskeleton to optimize the outcome for UE.

Chondroitinase ABC combined with neurotrophin NT-3 secretion and NR2D expression promotes axonal plasticity and functional recovery in rats with lateral hemisection of the spinal cord.

Elevating spinal levels of neurotrophin NT-3 (NT3) while increasing expression of the NR2D subunit of the NMDA receptor using a HSV viral construct promotes formation of novel multisynaptic projections from lateral white matter (LWM) axons to motoneurons in neonates. However, this treatment is ineffective after postnatal day 10. Because chondroitinase ABC (ChABC) treatment restores plasticity in the adult CNS, we have added ChABC to this treatment and applied the combination to adult rats receiving a left lateral hemisection (Hx) at T8. All hemisected animals initially dragged the ipsilateral hindpaw and displayed abnormal gait. Rats treated with ChABC or NT3/HSV-NR2D recovered partial hindlimb locomotor function, but animals receiving combined therapy displayed the most improved body stability and interlimb coordination [Basso-Beattie-Bresnahan (BBB) locomotor scale and gait analysis]. Electrical stimulation of the left LWM at T6 did not evoke any synaptic response in ipsilateral L5 motoneurons of control hemisected animals, indicating interruption of the white matter. Only animals with the full combination treatment recovered consistent multisynaptic responses in these motoneurons indicating formation of a detour pathway around the Hx. These physiological findings were supported by the observation of increased branching of both cut and intact LWM axons into the gray matter near the injury. ChABC-treated animals displayed more sprouting than control animals and those receiving NT3/HSV-NR2D; animals receiving the combination of all three treatments showed the most sprouting. Our results indicate that therapies aimed at increasing plasticity, promoting axon growth and modulating synaptic function have synergistic effects and promote better functional recovery than if applied individually.

Long-term rehabilitation reduces task error variability in cervical spinal cord contused rats.

To promote skilled forelimb function following a spinal cord injury, we have evaluated whether long-term voluntary sensorimotor rehabilitation can promote substantial reaching and grasping recovery. Long-Evans rats were trained to reach single pellets and then received a moderate 100 kdyn contusion to the C5 lateral funiculi. During the first eight months post-injury, a group of animals was enrolled in daily skilled reaching rehabilitation consisting of grabbing and manipulating seeds from the bottom of a grid. Single-pellet reaching and grasping recovery was tested biweekly throughout the functional follow-up and the recovery was compared to a second group of contused but non-rehabilitated animals. Following the injury, reaching and grasping success dropped to zero in both groups and remained absent for three months post-injury, followed by a slight recovery that remained constant until the end of the follow-up. No differences in reaching success were found between groups. Nevertheless, the type of gesture errors in the failed attempts were categorized and scored. The errors ranged from the animal's inability to lift the paw and initiate the movement to the final stage of the attempt, in which the pellet falls during grasping and retraction of the paw towards the mouth. Both groups of animals exhibited similar types of errors but the animals with rehabilitation showed less error variability and those that occurred at the latest stages of the attempt predominated compared to those performed by the non-trained animals. Histological examination of the injury showed that injury severity was similar between groups and that the damage was circumscribed to the site of impact, affecting mainly the dorsal and medial region of the lateral funiculi, with preservation of the dorsal component of the corticospinal tract and the interneurons and motoneurons of the spinal segments beyond the site of injury. The results indicate that activity-dependent plasticity driven by voluntary rehabilitation decreases task error variability and drives the recovery of the movement gestures. However, the plasticity achieved is insufficient to attain full functional recovery to successfully reach, grasp and release the pellets in the mouth, indicating the necessity for additional interventional therapies to promote repair.

Role of chondroitin sulfate proteoglycans in axonal conduction in Mammalian spinal cord.

Chronic unilateral hemisection (HX) of the adult rat spinal cord diminishes conduction through intact fibers in the ventrolateral funiculus (VLF) contralateral to HX. This is associated with a partial loss of myelination from fibers in the VLF (Arvanian et al., 2009). Here, we again measured conduction through the VLF using electrical stimulation while recording the resulting volley and synaptic potentials in target motoneurons. We found that intraspinal injection of chondroitinase-ABC, known to digest chondroitin sulfate proteoglycans (CSPGs), prevented the decline of axonal conduction through intact VLF fibers across from chronic T10 HX. Chondroitinase treatment was also associated with behavior suggestive of an improvement of locomotor function after chronic HX. To further study the role of CSPGs in axonal conduction, we injected three purified CSPGs, NG2 and neurocan, which increase in the vicinity of a spinal injury, and aggrecan, which decreases, into the lateral column of the uninjured cord at T10 in separate experiments. Intraspinal injection of NG2 acutely depressed axonal conduction through the injected region in a dose-dependent manner. Similar injections of saline, aggrecan, or neurocan had no significant effect. Immunofluorescence staining experiments revealed the presence of endogenous and exogenous NG2 at some nodes of Ranvier. These results identify a novel acute action of CSPGs on axonal conduction in the spinal cord and suggest that antagonism of proteoglycans reverses or prevents the decline of axonal conduction, in addition to stimulating axonal growth.

When Spinal Neuromodulation Meets Sensorimotor Rehabilitation: Lessons Learned From Animal Models to Regain Manual Dexterity After a Spinal Cord Injury.

Electrical neuromodulation has strongly hit the foundations of spinal cord injury and repair. Clinical and experimental studies have demonstrated the ability to neuromodulate and engage spinal cord circuits to recover volitional motor functions lost after the injury. Although the science and technology behind electrical neuromodulation has attracted much of the attention, it cannot be obviated that electrical stimulation must be applied concomitantly to sensorimotor rehabilitation, and one would be very difficult to understand without the other, as both need to be finely tuned to efficiently execute movements. The present review explores the difficulties faced by experimental and clinical neuroscientists when attempting to neuromodulate and rehabilitate manual dexterity in spinal cord injured subjects. From a translational point of view, we will describe the major rehabilitation interventions employed in animal research to promote recovery of forelimb motor function. On the other hand, we will outline some of the state-of-the-art findings when applying electrical neuromodulation to the spinal cord in animal models and human patients, highlighting how evidences from lumbar stimulation are paving the path to cervical neuromodulation.

Cervical Electrical Neuromodulation Effectively Enhances Hand Motor Output in Healthy Subjects by Engaging a Use-Dependent Intervention.

Electrical enabling motor control (eEmc) through transcutaneous spinal cord stimulation is a non-invasive method that can modify the functional state of the sensory-motor system. We hypothesize that eEmc delivery, together with hand training, improves hand function in healthy subjects more than either intervention alone by inducing plastic changes at spinal and cortical levels. Ten voluntary participants were included in the following three interventions: (i) hand grip training, (ii) eEmc, and (iii) eEmc with hand training. Functional evaluation included the box and blocks test (BBT) and hand grip maximum voluntary contraction (MVC), spinal and cortical motor evoked potential (sMEP and cMEP), and resting motor thresholds (RMT), short interval intracortical inhibition (SICI), and F wave in the abductor pollicis brevis muscle. eEmc combined with hand training retained MVC and increased F wave amplitude and persistency, reduced cortical RMT and facilitated cMEP amplitude. In contrast, eEmc alone only increased F wave amplitude, whereas hand training alone reduced MVC and increased cortical RMT and SICI. In conclusion, eEmc combined with hand grip training enhanced hand motor output and induced plastic changes at spinal and cortical level in healthy subjects when compared to either intervention alone. These data suggest that electrical neuromodulation changes spinal and, perhaps, supraspinal networks to a more malleable state, while a concomitant use-dependent mechanism drives these networks to a higher functional state.

Transcutaneous Electrical Neuromodulation of the Cervical Spinal Cord Depends Both on the Stimulation Intensity and the Degree of Voluntary Activity for Training. A Pilot Study.

Electrical enabling motor control (eEmc) through transcutaneous spinal cord stimulation offers promise in improving hand function. However, it is still unknown which stimulus intensity or which muscle force level could be better for this improvement. Nine healthy individuals received the following interventions: (i) eEmc intensities at 80%, 90% and 110% of abductor pollicis brevis motor threshold combined with hand training consisting in 100% handgrip strength; (ii) hand training consisting in 100% and 50% of maximal handgrip strength combined with 90% eEmc intensity. The evaluations included box and blocks test (BBT), maximal voluntary contraction (MVC), F wave persistency, F/M ratio, spinal and cortical motor evoked potentials (MEP), recruitment curves of spinal MEP and cortical MEP and short-interval intracortical inhibition. The results showed that: (i) 90% eEmc intensity increased BBT, MVC, F wave persistency, F/M ratio and cortical MEP recruitment curve; 110% eEmc intensity increased BBT, F wave persistency and cortical MEP and recruitment curve of cortical MEP; (ii) 100% handgrip strength training significantly modulated MVC, F wave persistency, F/M wave and cortical MEP recruitment curve in comparison to 50% handgrip strength. In conclusion, eEmc intensity and muscle strength during training both influence the results for neuromodulation at the cervical level.

Onset Detection to Study Muscle Activity in Reaching and Grasping Movements in Rats.

EMG signals reflect the neuromuscular activation patterns related to the execution of a certain movement or task. In this work, we focus on reaching and grasping (R&G) movements in rats. Our objective is to develop an automatic algorithm to detect the onsets and offsets of muscle activity and use it to study muscle latencies in R&G maneuvers. We had a dataset of intramuscular EMG signals containing 51 R&G attempts from 2 different animals. Simultaneous video recordings were used for segmentation and comparison. We developed an automatic onset/offset detector based on the ratio of local maxima of Teager-Kaiser Energy (TKE). Then, we applied it to compute muscle latencies and other features related to the muscle activation pattern during R&G cycles. The automatic onsets that we found were consistent with visual inspection and video labels. Despite the variability between attempts and animals, the two rats shared a sequential pattern of muscle activations. Statistical tests confirmed the differences between the latencies of the studied muscles during R&G tasks. This work provides an automatic tool to detect EMG onsets and offsets and conducts a preliminary characterization of muscle activation during R&G movements in rats. This kind of approaches and data processing algorithms can facilitate the studies on upper limb motor control and motor impairment after spinal cord injury or stroke.

Proteoglycans in the central nervous system: plasticity, regeneration and their stimulation with chondroitinase ABC.

After injury to the mammalian central nervous system (CNS), neurons are not able to regenerate their axons and recovery is limited by restricted plasticity. Axon regeneration is inhibited by the presence of the various inhibitory molecules, including chondroitin sulfate proteoglycans (CSPGs) which are upregulated around the injury site. Plasticity after the end of critical periods is restricted by extracellular matrix changes, particularly the formation of CSPG-containing perineuronal nets. Enzymatic removal of chondroitin sulfate (CS) chains with chondroitinase ABC promotes axon regeneration and reactivates plasticity. This review details the structures and properties of the different CSPGs in the normal and damaged CNS, the use of the enzyme chondroitinase ABC to promote neural regeneration and plasticity, and discusses mechanisms of action and possible therapeutic uses of this enzyme.

Electrical neuromodulation of the cervical spinal cord facilitates forelimb skilled function recovery in spinal cord injured rats.

Enabling motor control by epidural electrical stimulation of the spinal cord is a promising therapeutic technique for the recovery of motor function after a spinal cord injury (SCI). Although epidural electrical stimulation has resulted in improvement in hindlimb motor function, it is unknown whether it has any therapeutic benefit for improving forelimb fine motor function after a cervical SCI. We tested whether trains of pulses delivered at spinal cord segments C6 and C8 would facilitate the recovery of forelimb fine motor control after a cervical SCI in rats. Rats were trained to reach and grasp sugar pellets. Immediately after a dorsal funiculus crush at C4, the rats showed significant deficits in forelimb fine motor control. The rats were tested to reach and grasp with and without cervical epidural stimulation for 10weeks post-injury. To determine the best stimulation parameters to activate the cervical spinal networks involved in forelimb motor function, monopolar and bipolar currents were delivered at varying frequencies (20, 40, and 60Hz) concomitant with the reaching and grasping task. We found that cervical epidural stimulation increased reaching and grasping success rates compared to the no stimulation condition. Bipolar stimulation (C6- C8+ and C6+ C8-) produced the largest spinal motor-evoked potentials (sMEPs) and resulted in higher reaching and grasping success rates compared with monopolar stimulation (C6- Ref+ and C8- Ref+). Forelimb performance was similar when tested at stimulation frequencies of 20, 40, and 60Hz. We also found that the EMG activity in most forelimb muscles as well as the co-activation between flexor and extensor muscles increased post-injury. With epidural stimulation, however, this trend was reversed indicating that cervical epidural spinal cord stimulation has therapeutic potential for rehabilitation after a cervical SCI.

Differential motor and electrophysiological outcome in rats with mid-thoracic or high lumbar incomplete spinal cord injuries.

We have investigated the motor changes in rats subjected to a moderate photochemical injury on mid-thoracic (T8) or high lumbar (L2) spinal cord segments. Fourteen days after surgery, L2 injured animals presented gross locomotor deficits (scored 10+/-2.8 in the BBB scale), decreased amplitude of motor-evoked potentials (MEPs) recorded on tibialis anterior (TA) and plantar (PL) muscles (24% and 6% of the preoperative mean values, respectively), reduced M wave amplitudes (75%, 62%), and also facilitated monosynaptic reflexes evidenced by an increase of the H/M amplitude ratio (158% and 563%). On the other hand, T8 injured animals had only slight deficits in locomotion (18+/-0.6 in the BBB scale), a minimal reduction in MEP amplitudes (78% and 71% in TA and PL muscles), normal M wave amplitudes, and a milder increase of the H/M ratio in the TA muscle (191%) but less pronounced in the PL muscle (172%). The percentage of spared tissue at the site of injury was similar in both experimental groups (L2: 79% and T8: 82%). Taken together, these results indicate that lumbar spinal injuries have more severe consequences on hindlimb motor output than injuries exerted on thoracic segments. The causes of this anatomical difference may be attributed to damage inflicted on the central pattern generator of locomotion resulting in dysfunction of lumbar motoneurons and altered spinal reflexes modulation.

Longitudinal Evaluation of Residual Cortical and Subcortical Motor Evoked Potentials in Spinal Cord Injured Rats.

We have applied transcranial electrical stimulation to rats with spinal cord injury and selectively tested the motor evoked potentials (MEPs) conveyed by descending motor pathways with cortical and subcortical origin. MEPs were elicited by electrical stimulation to the brain and recorded on the tibialis anterior muscles. Stimulation parameters were characterized and changes in MEP responses tested in uninjured rats, in rats with mild or moderate contusion, and in animals with complete transection of the spinal cord. All injuries were located at the T8 vertebral level. Two peaks, termed N1 and N2, were obtained when changing from single pulse stimulation to trains of 9 pulses at 9 Hz. Selective injuries to the brain or spinal cord funiculi evidenced the subcortical origin of N1 and the cortical origin of N2. Animals with mild contusion showed small behavioral deficits and abolished N1 but maintained small amplitude N2 MEPs. Substantial motor deficits developed in rats with moderate contusion, and these rats had completely eliminated N1 and N2 MEPs. Animals with complete cord transection had abolished N1 and N2 and showed severe impairment of locomotion. The results indicate the reliability of MEP testing to longitudinally evaluate over time the degree of impairment of cortical and subcortical spinal pathways after spinal cord injuries of different severity.

Plasticity of subcortical pathways promote recovery of skilled hand function in rats after corticospinal and rubrospinal tract injuries.

The corticospinal and rubrospinal tracts are the predominant tracts for controlling skilled hand function. Injuries to these tracts impair grasping but not gross motor functions such as overground locomotion. The aim of the present study was to determine whether or not, after damage to both the corticospinal and rubrospinal tracts, other spared subcortical motor pathway can mediate the recovery of skilled hand function. Adult rats received a bilateral injury to the corticospinal tract at the level of the medullar pyramids and a bilateral ablation of the rubrospinal axons at C4. One group of rats received, acutely after injury, two injections of chondroitinase-ABC at C7, and starting at 7days post-injury were enrolled in daily reaching and grasping rehabilitation (CHASE group, n=5). A second group of rats received analogous injections of ubiquitous penicillinase, and did not undergo rehabilitation (PEN group, n=5). Compared to rats in the PEN group, CHASE rats gradually recovered the ability to reach and grasp over 42days after injury. Overground locomotion was mildly affected after injury and both groups followed similar recovery. Since the reticulospinal tract plays a predominant role in motor control, we further investigated whether or not plasticity of this pathway could contribute to the animal's recovery. Reticulospinal axons were anterogradely traced in both groups of rats. The density of reticulospinal processes in both the normal and ectopic areas of the grey ventral matter of the caudal segments of the cervical spinal cord was greater in the CHASE than PEN group. The results indicate that after damage to spinal tracts that normally mediate the control of reaching and grasping in rats other complementary spinal tracts can acquire the role of those damaged tracts and promote task-specific recovery.

Evaluation of optimal electrode configurations for epidural spinal cord stimulation in cervical spinal cord injured rats.

BACKGROUND: Epidural spinal cord stimulation is a promising technique for modulating the level of excitability and reactivation of dormant spinal neuronal circuits after spinal cord injury (SCI). We examined the ability of chronically implanted epidural stimulation electrodes within the cervical spinal cord to (1) directly elicit spinal motor evoked potentials (sMEPs) in forelimb muscles and (2) determine whether these sMEPs can serve as a biomarker of forelimb motor function after SCI. NEW METHOD: We implanted EMG electrodes in forelimb muscles and epidural stimulation electrodes at C6 and C8 in adult rats. After recovering from a dorsal funiculi crush (C4), rats were tested with different stimulation configurations and current intensities to elicit sMEPs and determined forelimb grip strength. RESULTS: sMEPs were evoked in all muscles tested and their characteristics were dependent on electrode configurations and current intensities. C6(-) stimulation elicited more robust sMEPs than stimulation at C8(-). Stimulating C6 and C8 simultaneously produced better muscle recruitment and higher grip strengths than stimulation at one site. COMPARISON WITH EXISTING METHOD(S): Classical method to select the most optimal stimulation configuration is to empirically test each combination individually for every subject and relate to functional improvements. This approach is impractical, requiring extensively long experimental time to determine the more effective stimulation parameters. Our proposed method is fast and physiologically sound. CONCLUSIONS: Results suggest that sMEPs from forelimb muscles can be useful biomarkers for identifying optimal parameters for epidural stimulation of the cervical spinal cord after SCI.

Increased expression of cyclo-oxygenase 2 and vascular endothelial growth factor in lesioned spinal cord by transplanted olfactory ensheathing cells.

Olfactory ensheathing cells (OECs) were transplanted in adult rats after photochemical injury of the spinal cord. Rats received either 180,000 OECs suspended in DMEM or DMEM alone. Locomotor ability scored by the BBB-scale, pain sensibility, and motor and somatosensory evoked potentials were evaluated during the first 14 days post-surgery. At 3, 7, and 14 days, 5 rats per day of both groups were perfused and transverse sections from proximal, lesioned and distal spinal cord blocks were stained for COX-2, VEGF, GFAP and lectin. The BBB-score and the amplitude of motor and somatosensory evoked potentials were significantly higher in OEC- than in DMEM-injected animals throughout follow-up, whereas the withdrawal latency to heat noxious stimulus was lower in OEC- than in DMEM-injected rats. The area of preserved spinal cord and the levels of COX-2 and VEGF staining were significantly higher in OEC- than in DMEM-injected rats. GFAP- but no LEC-positive cells expressed COX-2 staining in OEC-transplanted rats. The density of blood vessels was also significantly increased in OEC- with respect to DMEM-injected rats. Our results show that OECs promote functional and morphological preservation of the spinal cord after photochemical injury, increasing neoangiogenesis and up-regulation of COX-2 and VEGF expression in astrocytes.

Functional and electrophysiological characterization of photochemical graded spinal cord injury in the rat.

This study characterizes by functional and electrophysiological methods changes following photochemically induced injuries to the spinal cord in adult rats. The spinal cord was exposed by laminectomy and bathed with 1.5% rose bengal solution for 10 min (T12-L1 vertebrae). The excess dye was removed by saline rinse and the spinal cord was irradiated with "cold" light for 0, 1, 2.5, 5, and 10 min in different groups of rats. During the first 15 days postlesion, locomotion activity, pain sensibility, motor and somatosensory evoked potentials, and motor and nerve action potentials were evaluated. Graded locomotor and nociceptive recovery was observed in irradiated rats depending on the photoinduction time. At 15 days, the amplitude of motor and sensory evoked potentials was significantly lower in irradiated groups with respect to control rats. The amplitude of compound muscle action potentials and of reflex H wave after sciatic nerve stimulation decreased significantly in irradiated animals with respect to control rats, while the latency did not show significant differences. In irradiated groups, significant differences were seen between pre- and postoperative values for most functional and electrophysiological parameters analyzed. A significant negative relationship was found between the area of cystic cavity of the spinal cord and the functional and electrophysiological impairment.

Morphological characterization of photochemical graded spinal cord injury in the rat.

This study characterizes the histological and immunohistochemical changes in the adult rat spinal cord following photochemically induced spinal cord lesions. The spinal cord was exposed by laminectomy (T12-L1 vertebrae) and bathed with 1.5% rose bengal solution for 10 min. The excess dye was removed by saline rinse and the spinal cord was irradiated with "cold" light for 0, 1, 2.5, 5, and 10 min in different groups of rats. After 15 days a graded loss of spinal tissue was observed according to photoinduction times. Animals irradiated for 1 min showed spinal cavities involving the dorsal funiculi. The cavity became progressively larger, involving dorsal horns in animals irradiated for 2.5 min, together with the dorsolateral funiculi in animals irradiated for 5 min and the ventrolateral funiculi in those irradiated for 10 min, with loss of gray matter in these three groups. Changes in GFAP-, CGRP-, proteoglycan- and calbindin-immunoreactivity were observed in all lesioned groups when compared with control spinal cords. Hypertrophied and heavily GFAP- and proteoglycan-stained astrocytes were seen in irradiated spinal cords. Reactive microglial cells were also found. Both astroglial and microglial reactions paralleled the severity of the spinal cord lesion. A significant loss of CGRP-immunoreactive somas was seen in animals irradiated for 10 min, whereas the wider distribution of calbindin-positive neurons was found in lesioned rats. In spinal cord sections from animals illuminated for 5 min and perfused 60 min postillumination, light and electron microscopy showed cytotoxic edema with astrocytic swelling, red blood cell extravasation, and myelin degradation.

Plastic changes in lumbar segments after thoracic spinal cord injuries in adult rats: an integrative view of spinal nociceptive dysfunctions.

PURPOSE: Spinal cord injuries (SCI) cause motor, sensory and autonomic dysfunctions as well as neuropathic pain. We investigated plastic changes occurring in cord segments caudal to the lesion to assess their potential contribution to pain states after SCI. METHODS: Different thoracic SCIs were performed in adult rats. Functional and algesimetry tests were performed along 3 months. Several elements of the spinal nociceptive circuitry were assessed by immunohistochemical analyses of lumbar segments. RESULTS: Injured animals manifested mechanical and thermal hyperalgesia. Wind-up responses and spinal reflexes were enhanced, indicating spinal hyperexcitability. We found an increase in density of nociceptive sensory afferences and in GABA inhibitory activity in dorsal horns, and increased glial reactivity. Serotoninergic descending fibers and contacts on ventral horn motoneurons were reduced. Motoneurons presented more abundant inhibitory inputs, identified by gephyrin. Not all the changes kept direct relationship to the severity of the injury. CONCLUSION: The existence of hyperalgesia despite the boost of inhibitory elements in the spinal cord confirms the dysbalance between excitatory and inhibitory mechanisms, leading to a general disinhibition. Widespread dysfunctions in remote segments after central injuries contribute to the appearance of pain, and they may be new targets for therapies aimed to modulate spinal dysfunctions after injury.