Unique Sensory and Motor Behavior in Thy1-GFP-M Mice before and after Spinal Cord Injury

Abstract

Sensorimotor recovery after spinal cord injury (SCI) is of utmost importance to injured individuals and will rely on improved understanding of SCI pathology and recovery. Novel transgenic mouse lines facilitate discovery, but must be understood to be effective. The purpose of this study was to characterize the sensory and motor behavior of a common transgenic mouse line (Thy1-GFP-M) before and after SCI. Thy1-GFP-M positive (TG+) mice and their transgene negative littermates (TG-) were acquired from two sources (in-house colony, n = 32, Jackson Laboratories, n = 4). C57BL/6J wild-type (WT) mice (Jackson Laboratories, n = 10) were strain controls. Moderate-severe T9 contusion (SCI) or transection (TX) occurred in TG+ (SCI, n = 25, TX, n = 5), TG- (SCI, n = 5), and WT (SCI, n = 10) mice. To determine responsiveness to rehabilitation, a cohort of TG+ mice with SCI (n = 4) had flat treadmill (TM) training 42-49 days post-injury (dpi). To characterize recovery, we performed Basso Mouse Scale, Grid Walk, von Frey Hair, and Plantar Heat Testing before and out to day 42 post-SCI. Open field locomotion was significantly better in the Thy1 SCI groups (TG+ and TG-) compared with WT by 7 dpi (p < 0.01) and was maintained through 42 dpi (p < 0.01). These unexpected locomotor gains were not apparent during grid walking, indicating severe impairment of precise motor control. Thy1 derived mice were hypersensitive to mechanical stimuli at baseline (p < 0.05). After SCI, mechanical hyposensitivity emerged in Thy1 derived groups (p < 0.001), while thermal hyperalgesia occurred in all groups (p < 0.001). Importantly, consistent findings across TG+ and TG- groups suggest that the effects are mediated by the genetic background rather than transgene manipulation itself. Surprisingly, TM training restored mechanical and thermal sensation to baseline levels in TG+ mice with SCI. This behavioral profile and responsiveness to chronic training will be important to consider when choosing models to study the mechanisms underlying sensorimotor recovery after SCI.

Keywords: locomotor function; rehabilitation; sensory function; spinal cord injury.

FIG. 1.

Animal enrollment and use. Flow diagram illustrates all mice included in the current studies. Animals included in behavioral outcomes testing are noted on the left. Timing and reason for exclusions are noted on the right. TG, transgenic; SCI, spinal cord injury; WT, wild type; TX, transection.

FIG. 2.

Transgenic (TG)+ and TG- mice have increased tissue sparing after spinal cord injury (SCI). Representative images of eriochrome cyanine staining at the injury epicenter show increased tissue sparing and decreased lesion size in Thy1-GFP-M mice (TG+, TG-) with SCI (A). Quantification of epicenter percent sparing and lesion size, respectively, is expressed as percent area (B, C; *p < 0.05; one-way analysis of variance). GFP, green fluorescent protein; WT, wild type; WMS, white matter sparing; SEM, standard error of the mean.

FIG. 3.

Transgenic (TG)+ and TG- mice have improved gross motor function but impaired precise motor control after spinal cord injury (SCI). TG+ and TG- SCI animals have significantly greater locomotor recovery as early as seven days post-injury (dpi) that persisted to chronic time points (42 days) (A; *p < 0.05, **p < 0.01 vs. wild type (WT) SCI; repeated measures analysis of variance [ANOVA]). All SCI performed better than transection (TX) mice after one dpi (A; ^^p < 0.01 vs. all groups; repeated measures ANOVA). In mice with SCI, open field performance was strongly correlated with epicenter sparing (B; *p < 0.01; Pearson r). Despite significantly greater overground locomotion, the TG+ and TG- groups did not exhibit better grid walking, suggesting that proprioception and precise motor control were markedly impaired (C; p > 0.05; one-way ANOVA). After contusion, fine motor performance was not correlated to either open field performance or sparing (D, E; p > 0.05; Pearson r). SEM, standard error of the mean; BMS, Basso Mouse Scale; WMS, white matter sparing.

FIG. 4.

Thy1 mice demonstrate divergent sensory recovery after spinal cord injury (SCI). Mechanical and thermal sensations were assessed using the von Frey Hair Test and Plantar Heat Test, respectively. Thy1 groups had hypersensitivity to mechanical stimuli before SCI that became paresthesia-like after SCI in a severity-dependent manner (A; ***p < 0.001 Main Effect of Time; ^^p < 0.01 vs. all groups; repeated measures analysis of variance [ANOVA]). In TG- mice, greater paresthesia developed than in TG+ mice (A; *p < 0.05; repeated measures ANOVA). All groups, regardless of background, green fluorescent protein expression, or injury severity become hypersensitive to thermal stimuli at 42 days post-injury (B; ***p < 0.001; repeated measures ANOVA). SEM, standard error of the mean; WT, wild type.

FIG. 5.

Late treadmill training normalizes sensation in transgenic (TG)+ mice. A subset of TG+ mice received eight consecutive days of treadmill training from 42–49 days post-injury. The eight-day rehabilitation paradigm restored mechanical and thermal sensitivity to pre-injury levels (A, B; **p < 0.01; repeated measures analysis of variance). Gross motor performance was less responsive to rehabilitation (C; p = 0.13; paired samples t test). Fine motor performance, however, showed a trend toward improvement after training (D; p = 0.07; paired samples t test). Compared with historic wild type (WT) mice that received chronic rehabilitation (Hansen and associates, 2013), TG+ mice showed a trend toward greater open field recovery (E; p = 0.17; independent samples t test) but significantly greater fine motor recovery (F; p < 0.01; independent samples t test). SEM, standard error of the mean; BMS, Basso Mouse Scale.