Long-Term Spinal Cord Stimulation Alleviates Mechanical Hypersensitivity and Increases Peripheral Cutaneous Blood Perfusion in Experimental Painful Diabetic Polyneuropathy.

OBJECTIVES: This study utilizes a model of long-term spinal cord stimulation (SCS) in experimental painful diabetic polyneuropathy (PDPN) to investigate the behavioral response during and after four weeks of SCS (12 hours/day). Second, we investigated the effect of long-term SCS on peripheral cutaneous blood perfusion in experimental PDPN. METHODS: Mechanical sensitivity was assessed in streptozotocin induced diabetic rats (n = 50) with von Frey analysis. Hypersensitive rats (n = 24) were implanted with an internal SCS battery, coupled to an SCS electrode covering spinal levels L2-L5. The effects of four weeks of daily conventional SCS for 12 hours (n = 12) or Sham SCS (n = 12) were evaluated with von Frey assessment, and laser Doppler imaging (LDI). RESULTS: Average paw withdrawal thresholds (PWT) increased during long-term SCS in the SCS group, in contrast to a decrease in the Sham group (Sham vs. SCS; p = 0.029). Twenty-four hours after long-term SCS average PWT remained higher in the SCS group. Furthermore, the SCS group showed a higher cutaneous blood perfusion during long-term SCS compared to the Sham group (Sham vs. SCS; p = 0.048). Forty-eight hours after long-term SCS, no differences in skin perfusion were observed. DISCUSSION: We demonstrated that long-term SCS results in decreased baseline mechanical hypersensitivity and results in increased peripheral blood perfusion during stimulation in a rat model of PDPN. Together, these findings indicate that long-term SCS results in modulation of the physiological circuitry related to the nociceptive system in addition to symptomatic treatment of painful symptoms.

Functional recovery, serotonergic sprouting, and endogenous progenitor fates in response to delayed environmental enrichment after spinal cord injury.

Environmental enrichment (EE) is a way to induce voluntary locomotor training that positively affects locomotor recovery after acute spinal cord injury (SCI). The beneficial effect on SCI outcome is thought to be based on enhanced plasticity in motor pathways, triggered by locomotor-specific sensory feedback to the spinal cord circuitry for locomotion (central pattern generators [CPGs]). In view of chronic SCI, we tested the hypothesis that EE improves motor outcome after SCI in the rat when started after a clinically relevant delay of 3 weeks. At the CPG level (i.e., the spinal L1-L2 level), where EE-related sensory feedback is processed, two key mechanisms of anatomical plasticity were examined: (1) serotonergic innervation, and (2) survival and differentiation of spinal cord progenitor cells. Delayed EE improved interlimb coordination, which was associated with an increased serotonergic innervation of the ventro-lateral grey matter within the L1-L2 segments. Although spinal cord progenitor cells were found to differentiate into both neurons and glial cells, EE did not affect their survival. These results show that EE induces a substantial improvement of motor outcome after SCI when commenced after a clinically-relevant delay. Increased serotonergic innervation of the lumbar CPG area is therefore suggested to play an important role in the EE-induced recovery of interlimb coordination.

Preoperative housing in an enriched environment significantly reduces the duration of post-operative pain in a rat model of knee inflammation.

The influence of the environment on clinical post-operative pain received recently more attention in human. A very common paradigm in experimental pain research to model the effect of housing conditions is the enriched environment (EE). During EE-housing, rats are housed in a large cage (i.e. social stimulation), usually containing additional tools like running wheels (i.e. physical stimulation). Interestingly, only postsurgical housing effect on post-operative pain was developed during clinical and experimental studies while little is known on the influence of preoperative housing. In this study, our aim was to investigate the influence of housing conditions prior to an operation on the development of post-operative pain, using a rat model of carrageenan-induced inflammatory pain. Four housing conditions were used: a 3-week pre-housing in standard conditions (S-) followed by a post-housing in an EE; a 3-week pre-housing in EE followed by a post-operation S-housing; a pre- and post-housing in EE; a pre- and post-S-housing. The development of mechanical allodynia was assessed by the means of the von Frey test, preoperatively and at day post-operative (DPO) 1, 3, 7, 10, 14, 17, 21, 24 and 28. Our results show that a 3-week preoperative exposure to EE leads to a significant reduction in the duration of the carrageenan-induced mechanical allodynia, comparable with a post-operative exposure to EE. Strikingly, when rats were housed in EE prior to as well as after the carrageenan injection into the knee, mechanical allodynia lasted only 2 weeks, as compared to 4 weeks in S-housed rats.

Environmental housing affects the duration of mechanical allodynia and the spinal astroglial activation in a rat model of chronic inflammatory pain.

In this study, we aimed at investigating the effect of an enriched environment (EE) on the recovery from chronic inflammatory pain. Inflammatory pain was induced by the injection of 2 mg of carrageenan (CAR) into the right knee of male Sprague-Dawley rats (n=34). Rats were housed either singly (S-housed) or in an EE (EE-housed). The EE consisted of a large cage (L x W x H=2.0 x 1.0 x 0.8 m) containing various attributes (e.g. running wheels, shelter house, climbing frame). Withdrawal response to von Frey filament was used to assess mechanical allodynia at days post-operative (DPO) -1, 1, 7, 14, 21 and 28. S-housed animals showed a marked tactile sensitivity in the ipsilateral paw from DPO1 to DPO21. Four weeks after the CAR injection, S-housed rats were no longer allodynic. In contrast, EE-housed rats showed a significantly faster recovery: already at DPO21, they were no longer allodynic. In a first attempt to analyse the possible role of astroglial cells in the EE-induced effect, histological analysis at DPO21 was performed. Immunohistochemical staining of the spinal dorsal horn at L3-L5 indeed showed that spinal levels of astroglial activation are different between the two housing groups and therefore may play a role in the EE-induced effect on the duration of mechanical allodynia. In conclusion, our results showed that EE-housing results in a reduced duration of mechanical allodynia in chronic inflammatory pain in rats. Astroglial activation is suggested to be involved in this housing effect.

Strain and locomotor speed affect over-ground locomotion in intact rats.

A variety of animal models for neurological disease and injury exist and locomotor performance is an important outcome parameter in studies employing these models. The CatWalk, an automated quantitative gait analysis method is a method to study over-ground locomotor performance in large groups of animals. In the present study, we used the CatWalk which allowed us to investigate strain differences in over-ground locomotion in three commonly used strains of laboratory rat (i.e. Lewis, Wistar and Sprague-Dawley rats) based on objective data-analysis in a large number of animals. The present results revealed marked strain differences on the static paw parameters; base-of-support, and the relative paw position. Furthermore, strain differences were noted on the static parameter stride length and the dynamic parameters stance-, swing- and stepcycle duration, which are due logically to morphological differences between strains. The parameters related to coordination did not reveal any differences between the strains. Furthermore, the swing duration and the cruciate and alternate patterns i.e. regular step patterns Ca ("cruciate" pattern type a) and Ab ("alternate" pattern type b) were shown to be differentially affected by the locomotor speed. We conclude that differences in gait traits exist between the three laboratory rat strains investigated and several of the examined gait parameters showed strain dependent interdependency with locomotor speed.

Mice lacking L1 have reduced CGRP fibre in-growth into spinal transection lesions.

Repair strategies for spinal cord injury often focus on promoting regeneration of injured axons and stimulating subsequent functional recovery. Although many of these strategies have proven their merits, less is known about potential unwanted side-effects, such as sprouting of nociceptive CGRP immunoreactive axons, which may bring about pain-related behavior. Sprouting of CGRP axons into lesion sites spontaneously occurs after spinal cord injury (SCI). Using L1-deficient mice we show a reduction of such CGRP growth response. This reduction was specific for CGRP axons since the overall neurofilament positive fibre in-growth into the spinal lesion site was not affected. Our results may have important implications on the development and assessment of repair strategies that should not only stimulate functional recovery, but also prevent the development of pain or autonomic dysreflexia.

Limitations in transplantation of astroglia-biomatrix bridges to stimulate corticospinal axon regrowth across large spinal lesion gaps.

Regrowth of injured axons across rather small spinal cord lesion gaps and subsequent functional recovery has been obtained after many interventions. Long-distance regeneration of injured axons across clinically relevant large spinal lesion gaps is relatively unexplored. Here, we aimed at stimulating long-distance regrowth of the injured corticospinal (CS) tract. During development, an oriented framework of immature astrocytes is important for correct CS axon outgrowth. Furthermore, a continuous growth promoting substrate may be needed to maintain a CS axon regrowth response across relatively large spinal lesion gaps. Hence, we acutely transplanted poly(D,L)-lactide matrices, which after seeded with immature astrocytes render aligned astrocyte-biomatrix complexes (R. Deumens, et al. Alignment of glial cells stimulates directional neurite growth of CNS neurons in vitro. Neuroscience 125 (3) (2004) 591-604), into 2-mm long dorsal hemisection lesion gaps. In order to create a growth promoting continuum, astrocyte suspensions were also injected rostral and caudal to the lesion gap. During 2 months, locomotion was continuously monitored. Histological analysis showed that astrocytes injected into host spinal tissue survived, but did not migrate. None of the astrocytes on the biomatrices survived within the lesion gap. BDA-labeled CS axons did not penetrate the graft. However, directly rostral to the lesion gap, 120.9+/-38.5% of the BDA-labeled CS axons were present in contrast to 12.8+/-3.9% in untreated control animals. The observed anatomical changes were not accompanied by locomotor improvements as analyzed with the BBB and CatWalk. We conclude that although multifactorial strategies may be needed to stimulate long-distance CS axon regrowth, future studies should focus on enhancing the viability of cell/biomatrix complexes within large spinal lesion gaps.

Neurite outgrowth promoting effects of enriched and mixed OEC/ONF cultures.

Olfactory ensheathing cell (OEC) transplants stimulate axon regeneration and partial functional recovery after spinal cord injury. However, it remains unclear whether enriched OEC or mixed transplants of OEC and olfactory nerve fibroblasts (ONF) are optimal for stimulating axon regrowth. The neurite outgrowth stimulating effects of enriched OEC, ONF, and mixed OEC/ONF cultures on neonatal cerebral cortical neurons were compared using co-cultures. We show that (1) OEC are more neurite outgrowth promoting than ONF, and (2) ONF do not enhance the neurite outgrowth stimulating effects of OEC in mixed OEC/ONF cultures. Hence, our data indicate that there is no preference for the use of enriched OEC or mixed OEC/ONF cultures with respect to stimulation of neurite growth in vitro.

The assessment of locomotor function in spinal cord injured rats: the importance of objective analysis of coordination.

The Basso, Beattie and Bresnahan (BBB) locomotor rating scale is the most widely used open field test and has been accepted as a valid way to assess locomotor function after spinal cord contusion injury in the rat. A limitation within the BBB locomotor rating scale is the correct assessment of forelimb (FL)-hindlimb (HL) coordination. This limitation can have major implications for the final assessment of locomotor function. In the present study, we show an objective method to assess coordination based on the regularity index (RI), achieved through the use of the CatWalk method. The RI grades the degree of coordination as the result of the number of normal step sequence patterns multiplied by four and divided by the total amount of paw placements. Using the RI, single walkway crossings can be objectively analyzed on coordination. Integration of the CatWalk based coordination into the BBB scale indicates that objective analysis of coordination results in reliable and more sensitive assessment of locomotor function. This new method has been tested successfully in determination of positive effects of enriched housing on functional recovery after spinal cord injury (SCI).