In this study, we describe a novel injury device, the LISA-Vibraknife, which can produce a precise SCI. Our surgical techniques result in consistent and reproducible lesions, and our comprehensive behavioral analysis identified specific sensory, sensorimotor, and motor deficits along a dorsal-to-ventral gradient.
Stabilization of the spinal column is vital to SCI models. This was previously emphasized by Kuhn and Wrathall (1998
) and Scheff and colleagues (2003
), but was not experimentally documented. Our results suggest that supporting the segment at which the lesion is being induced () greatly reduces vertebral motion. Two additional advantages to the laceration model of SCI generated by the LISA-Vibraknife are the precise control of the lesion displacement and the elimination of accompanied contusive components at the lesion site by compression of the tissue that is often seen following microscissor lacerations (Bartholdi and Schwab, 1997
). We attribute the observed reduction in the actual lesion depth compared to the desired depth seen in to flexion or resistance in the spinal cord tissue and/or displacement of cerebrospinal fluid during the laceration procedure. One mechanical limitation of this study is the method used for determining dural contact of the blade with the spinal cord, which was visually confirmed through a dissecting microscope. An alternative may be to use an electrical circuit (Constantini and Young, 1994
; Gruner, 1992
; Huang and Young, 1994
) or laser detection to monitor blade contact with the spinal cord (Zhang et al., 2007a
In contrast to a laceration SCI in the rat, the mouse laceration does not generate a gap between the cord stumps. At 42
dpi, the rostral-caudal extent of the lesion was minimal and there was no cavitation observed. The increased interface proximity of the two cord stumps may contribute to the elimination of cavitation in this injury model. It has been previously shown that the lesion pathology seen following SCI in mice is different compared to other rodent models of SCI (Balantine, 1978
; Farooque, 2000
; Inman and Steward, 2003a
; Kuhn and Wrathall, 1998
; Sroga et al., 2003
). We anticipated that this model would exhibit a lesser degree of secondary injury by inducing less edema and tissue destruction.
A variety of tests were used (Goldberger, 1991
; Metz et al., 2000
; Muir and Webb, 2000
) to look at specific and detailed components of motor, sensorimotor, and sensory function. They enable the detection of subtle changes in the behavior of the animals, functional differentiation of the levels of injury severity, and the establishment of temporal recovery profiles for each of the injury groups. The association of behavioral/electrophysiological deficits with our laceration lesions () demonstrates that particular assessments are more sensitive at detecting functional loss for a particular lesion depth, and also demonstrates that different tests differentiate specific deficits. The higher the deficit sensitivity measure (DSM), the more sensitive that test was at detecting deficits for that injury level. This also depended on the number of deficits being detected. For example, in the 0.5-mm group, the BMS detected 7.4%, whereas the beam walk detected 21.4% of the total deficit. In the 1.1-mm group, the amount of deficit detected by each of the tests is more widely and more evenly distributed. Several of the measures still stand out with large DSMs, such as paw and tail position, beam walk, and tcMMEP, thus demonstrating the importance of these measures in detecting deficits for this level of injury.
Lesion Depth and Behavioral/Electrophysiological Deficit Sensitivity
This study does not, however, give us enough information to correlate precise laceration depths with specific white matter fiber tract loss. The two fiber tracts that we did trace, show complete transection with a 0.5-mm lesion. However, neither of these pathways carries information whose loss is detectable by the behavioral tests we performed. Further studies using anterograde and retrograde tracing of tracts involved with locomotor function (Loy et al., 2002b
; Schucht et al., 2002
) would be required to make more definitive conclusions about specific damage to tract(s) in relationship to deficits.
The BMS scores differentiated each group and independently verified the sensitivity of the BMS scale (Basso et al., 2006
) in an alternate model of mouse SCI. Footprint analysis shows a significant reduction in the BOS in the 0.5-, 0.8-, and 1.1-mm groups. These changes may be related to damage to the propriospinal system (Kunkel-Bagden et al., 1993
). This result supports previous studies that place the location of these propriospinal tracts in the dorsolateral white matter (Beaumont et al., 2006
), suggesting that the loss of these axons may contribute to the reduction in the BOS.
By modifying the beam walk test used previously on rats (Kunkel-Bagden et al., 1993
), we detected significant changes in the groups over time and significant differences between the groups. Even the mild 0.5-mm lesioned group exhibited significant deficits in performing this task. Therefore, the gradient beam walk may be an optimal approach to assess behavioral outcomes for mice that receive any type of mild SCI. A similar test, the grid walking test, has been shown to be related to propriospinal and rubrospinal pathway function (Bresnahan et al., 1987
; Whishaw et al., 1992
). Based on results from the beam walk test, the contribution of the rubrospinal pathway may be more significant than the propriospinal axons in the dorsolateral funiculus. The 0.5-mm lesion should only partially damage the rubrospinal pathway (if at all), whereas the lesions below that level would completely sever all the fibers. The significant amount of functional recovery and the fact that the 0.5-mm lesioned mice scored significantly higher than the 0.8- or 1.1-mm groups, suggests that the spared rubrospinal pathway and dorsal lateral funiculus (DLF
) are contributing to the performance of this behavioral task. In the 0.8- and 1.1-mm groups, where the rubrospinal pathway has been completely severed, we see some recovery of function, suggesting a potential contribution to performance of this task from the spared propriospinal and DLF pathways.
Results from the Hargreaves' test suggest that an animal with a lesion depth greater than 0.5
mm will show increased thermal sensitivity in the hindpaws due to a gradual dorsal-ventral loss of spinothalamic tract fibers (Willis and Westlund, 1997
). However, the reduced response time seen in the forepaws of the 1.4-mm group was unexpected. A possible explanation for this may be the increased reliance on the forepaws in these animals for support of their body due to the degree of hindlimb paralysis.
The tcMMEP results replicate in the mouse previous work done in the rat showing a correlation between loss of the ventrolateral funiculus and the tcMMEP response (Cao et al., 2005
; Loy et al., 2002a
). The small-amplitude responses detected in the 1.4-mm lesioned mice may be due to a small number of spared fibers in the ventral-most part of the ventrolateral funiculus.
The varied degrees of deficit and recovery seen in this study suggest spinal plasticity below the level of the lesion contributing to the recovery of hindlimb motor and sensory function. The sprouting of sensory fibers below the level of a spinal transection has been shown in previous studies (Goldberger et al., 1993
; Helgren and Goldberger, 1993
; Krenz and Weaver, 1998
; Wong et al., 2000
). A possible mechanism for this spontaneous recovery of function may be a reactive reorganization of the circuitry below the lesion, thus compensating for the loss of ascending and descending inputs. In future studies, we may be able to increase the amount of functional recovery seen following thoracic SCI by promoting sensory fiber sprouting.
In summary, we demonstrate that the LISA-Vibraknife can be used in future studies requiring precise, consistent, reproducible laceration SCIs. Importantly, we conclude that the combination of specific outcome measures can optimally detect sensory and motor functional deficits for particular SCI severities.