Pellet Retrieval Research Articles

Compensatory adaptation of parallel motor pathways promotes skilled forelimb recovery after spinal cord injury.

Skilled forelimb patterning is regulated by the corticospinal tract (CST), with support from brainstem regions. When the CST is lesioned, there is a loss of forelimb function; however, if indirect pathways remain intact, rehabilitative training can facilitate recovery. Following spinal cord injury, rehabilitation is thought to enhance the reorganization and plasticity of spared supraspinal-propriospinal circuits, aiding functional recovery. This study focused on the roles of cervical propriospinal interneurons (PNs) and rubrospinal neurons (RNs) in the recovery of reaching and grasping behaviors in rats with bilateral lesions of the CST and dorsal columns at C5. The lesions resulted in a 50% decrease in pellet retrieval, which normalized over four weeks of training. Silencing PNs or RNs after recovery resulted in reduced retrieval success. Notably, silencing both pathways corresponded to greater functional loss, underscoring their parallel contributions to recovery, alongside evidence of CST fiber sprouting in the spinal cord and red nucleus.

Chronic activation of corticospinal tract neurons after pyramidotomy injury enhances neither behavioral recovery nor axonal sprouting.

Modulation of neural activity is a promising strategy to influence the growth of axons and improve behavioral recovery after damage to the central nervous system. The benefits of neuromodulation likely depend on optimization across multiple input parameters. Here we used a chemogenetic approach to achieve continuous, long-term elevation of neural activity in murine corticospinal tract (CST) neurons. To specifically target CST neurons, AAV2-retro-DIO-hM3Dq-mCherry or matched mCherry control was injected to the cervical spinal cord of adult Emx1-Cre transgenic mice. Pilot studies verified efficient transgene expression in CST neurons and effective elevation of neural activity as assessed by cFos immunohistochemistry. In subsequent experiments mice were administered either DIO-hM3Dq-mCherry or control DIO-mCherry, were pre-trained on a pellet retrieval task, and then received unilateral pyramidotomy injury to selectively ablate the right CST. Mice then received continual clozapine via drinking water and weekly testing on the pellet retrieval task, followed by cortical injection of a viral tracer to assess cross-midline sprouting by the spared CST. After sacrifice at eight weeks post-injury immunohistochemistry for cFos verified elevated CST activity in hM3Dq-treated animals and immunohistochemistry for PKC-gamma verified unilateral ablation of the CST in all animals. Despite the chronic elevation of CST activity, however, both groups showed similar levels of cross-midline CST sprouting and similar success in the pellet retrieval task. These data indicate that continuous, long-term elevation of activity that is targeted specifically to CST neurons does not affect compensatory sprouting or directed forelimb movements.

1011 A Rare Case of Left Sided Hemiparesis Following a Pneumatic Gun-Shot Injury to Left Anterior Chest Wall in a 13-Year-Old Boy

Abstract Introduction Even though pneumatic weapons rarely cause severe direct trauma, pellet embolisation can lead to unexpected, catastrophic events. Case Report A 13-year-old, previously healthy boy was injured by an accidentally fired pneumatic gun, while playing with his twin brother. He sustained a penetrating wound to left anterior chest wall and resultant bleeding was self-limiting. He was hospitalised within 15minutes. On admission, he was hemodynamically stable with normal Respiratory, Cardiovascular and Neurological system findings. Entry wound measuring 5*5 mm was present at 5th intercostal space just left to the sternum. No exit wound was detected. Initial chest radiograph and ultra-sound abdomen and chest were normal. 2D echocardiogram revealed only a very thin layer(3mm) of pericardial effusion. Four hours later, He developed a focal seizure followed by left sided hemiparesis. Urgent NCCT brain and carotid artery duplex showed a high dense pellet in right internal carotid artery at the level of petrous temporal bone and hypo-dense Middle Cerebral Artery (MCA) suggestive of right MCA thrombosis. He was transferred to a tertiary care centre. Interventional radiological and endo-vascular attempts of pellet retrieval have been failed. Ischemic stroke pursued with worsening cerebral oedema; thus decompression craniotomy was performed on day 3. He is currently being rehabilitated in a specialised centre. Conclusions High degree of suspicion should be maintained on the possibility of projectile embolisation, in pneumatic gun-shot injuries, especially in the absence of an exit wound. Extensive early investigations should be performed to localise the projectile and retrieval should be planned.

Refined movement analysis in the staircase test reveals differential motor deficits in mouse models of stroke

Accurate assessment of post-stroke deficits is crucial in translational research. Recent advances in machine learning offer precise quantification of rodent motor behavior post-stroke, yet detecting lesion-specific upper extremity deficits remains unclear. Employing proximal middle cerebral artery occlusion (MCAO) and cortical photothrombosis (PT) in mice, we assessed post-stroke impairments via the Staircase test. Lesion locations were identified using 7 T-MRI. Machine learning was applied to reconstruct forepaw kinematic trajectories and feature analysis was achieved with MouseReach, a new data-processing toolbox. Lesion reconstructions pinpointed ischemic centers in the striatum (MCAO) and sensorimotor cortex (PT). Pellet retrieval alterations were observed, but were unrelated to overall stroke volume. Instead, forepaw slips and relative reaching success correlated with increasing cortical lesion size in both models. Striatal lesion size after MCAO was associated with prolonged reach durations that occurred with delayed symptom onset. Further analysis on the impact of selective serotonin reuptake inhibitors in the PT model revealed no clear treatment effects but replicated strong effect sizes of slips for post-stroke deficit detection. In summary, refined movement analysis unveiled specific deficits in two widely-used mouse stroke models, emphasizing the value of deep behavioral profiling in preclinical stroke research to enhance model validity for clinical translation.

Development and characterization of an automated behavioral assessment platform for the Göttingen minipig

The Göttingen minipig is fast becoming the standard for assessing dermal chemical hazards because, like most swine, its skin is predictive of human skin response and because this strain’s smaller size makes laboratory manipulations and husbandry easier. Unfortunately, standard behavioral tests and apparatus have not been developed for behavioral assessments of this swine strain. Indeed, computer-controlled automated behavioral testing procedures are much needed. The present research advanced this goal by producing a home-cage behavioral testing system that could accommodate minipigs of various sizes (ages). An aluminum frame housed three levers for recording operant responses, and LEDs above and below each lever served as discriminative stimuli. A commercially available food pellet dispenser was attached to a specialized pellet receptacle capable of measuring pellet retrieval. Two behavioral tests were selected and adapted from our commonly used non-human primate behavioral assessments: delayed match-to-sample (a memory test) and temporal response differentiation (a time-estimation test). Minipigs were capable of learning both tests and attaining stable performance. Next, scopolamine was used to validate the sensitivity of the behavioral tests for gauging behavioral perturbations in this swine strain. Scopolamine dose-effect functions were comparable to those observed in other species, including non-human primates, wherein 37.5 µg/kg of scopolamine (administered intramuscularly) reduced responding approximately 50%. Thus, we were successful in developing the apparatus and automated operant behavioral tests necessary to characterize drug safety in this swine strain. This capability will be valuable for characterizing chemical agent toxicity as well as the safety and efficacy of medical countermeasures.

Enriched Rehabilitation Reduces Abnormal Motor Synergies and Enhances Motor Plasticity Following Severe Stroke in Rats.

Stroke results in loss of upper motor neuron control over voluntary movements and emergence of abnormal synergies. Presently, it is unclear to what extent poststroke recovery reflects true recovery (restitution), compensation, or some combination of these processes. Here, we investigated this question using behavioral and kinematic analyses of skilled reaching in rats subjected to severe stroke that affected both the forelimb motor cortex and dorsolateral striatum. After stroke, male rats either spontaneously recovered or received enriched rehabilitation. We assessed forelimb motor recovery using behavioral and kinematic outcome measures. To provide insights into the mechanisms underlying the effects of rehabilitation on behavior, we used intracortical microstimulation and FosB (protein fosB) immunostaining techniques. Enriched rehabilitation significantly improved food pellet retrieval in the staircase-reaching task. Rehabilitation resulted in several poststroke flexion synergies returning to prestroke patterns, and across subjects, these changes correlated with the intensity of rehabilitation. Enriched rehabilitation increased the proportion of distal movement representation in the perilesional cortex and increased use-dependent activation in the ipsilesional red nucleus. These results provide evidence that enriched rehabilitation enhances recovery, at least in part, by restitution of forelimb function following severe stroke. Furthermore, the restitution of function is associated with changes in multiple motor-related structures at different levels of the central nervous system. A better understanding of the processes that underlie improved motor performance, along with the identification of midbrain circuits activated by rehabilitation, represent new insights and potential targets for optimizing poststroke recovery.

Compounds co-targeting kinases in axon regulatory pathways promote regeneration and behavioral recovery after spinal cord injury in mice

Recovery from spinal cord injury (SCI) and other central nervous system (CNS) trauma is hampered by limits on axonal regeneration in the CNS. Regeneration is restricted by the lack of neuron-intrinsic regenerative capacity and by the repressive microenvironment confronting damaged axons. To address this challenge, we have developed a therapeutic strategy that co-targets kinases involved in both extrinsic and intrinsic regulatory pathways. Prior work identified a kinase inhibitor (RO48) with advantageous polypharmacology (co-inhibition of targets including ROCK2 and S6K1), which promoted CNS axon growth in vitro and corticospinal tract (CST) sprouting in a mouse pyramidotomy model. We now show that RO48 promotes neurite growth from sensory neurons and a variety of CNS neurons in vitro, and promotes CST sprouting and/or regeneration in multiple mouse models of spinal cord injury. Notably, these in vivo effects of RO48 were seen in several independent experimental series performed in distinct laboratories at different times. Finally, in a cervical dorsal hemisection model, RO48 not only promoted growth of CST axons beyond the lesion, but also improved behavioral recovery in the rotarod, gridwalk, and pellet retrieval tasks. Our results provide strong evidence for RO48 as an effective compound to promote axon growth and regeneration. Further, they point to strategies for increasing robustness of interventions in pre-clinical models.

Markerless tracking enables distinction between strategic compensation and functional recovery after spinal cord injury

Injuries to the cervical spinal cord represent around 60% of all spinal cord injuries (SCIs). A major priority for patients with cervical SCIs is the recovery of any hand or arm function. The similarities between human and rodent “reach-to-eat movements” indicate that analyzing mouse forelimb reaching behavior may be a method of identifying clinically relevant treatments for people with cervical SCIs. One popular behavioral measure of forelimb functional recovery comprises the Single Pellet Retrieval Task (SPRT). The most common outcome measure for this task, however (percentage of pellets successfully retrieved), cannot readily distinguish between recovery of pre-injury motor patterns and strategic compensation. Our objective was to establish outcome measures for the SPRT that are readily adopted by different investigators and capable of measuring recovery of limb function after SCI. We used a simple semi-automated approach to high-speed tracking of mouse forepaw movements during pellet retrieval. DeepLabCut™, a machine learning based computer vision software package, was used to track individual features of the mouse forepaw, allowing a more detailed assessment of reaching behavior after SCI. Interestingly, kinematic analysis of movements pre- and post-injury illuminated persistent deficits in specific features of the reaching motor patterns, namely pronation and paw trajectory, that were poorly correlated with recovery of the ability to successfully retrieve pellets. Thus, we have developed an inexpensive method for detailed analysis of mouse reach-to-eat behavior following SCI. Further, our results suggest that binary success/fail outcome measures primarily assess an animal's ability to compensate rather than a restoration of normal function in the injured pathways and networks.

Knock down of lncRNA H19 promotes axon sprouting and functional recovery after cerebral ischemic stroke

Ischemic stroke is a leading cause of irreversible brain damages and disabilities. In the past decade, much attention has been focused on exploring effective strategies to promote circuit reorganization and functional recovery post injury. Here, we showed that the expression level of a long non-coding RNA (lncRNA H19) is bilaterally increased in the sensorimotor cortex after a cerebral ischemia induced by middle cerebral artery occlusion (MCAO). Knock down of contralaterally elevated H19 robustly enhanced the midline-crossing sprouting of the intact corticospinal axons in the spinal cord. Furthermore, H19 knockdown mice showed significant improvement on the performance of the food pellet retrieval assay, a skilled, cortical dependent motor task.Mechanistically, lncRNA H19 inhibition increased IGF1R expression and activated IGF1 mediated mTOR pathway. Our research thereby provided novel insights into identifying therapeutic targets for ischemic stroke.

Reaching task performance is associated to neuromuscular junction adaptations in rats with induced diabetes mellitus.

Upper limb performance is affected by diabetes mellitus (DM). Neuromuscular junction (NMJ) is a key structure to understand the relationship between performance and morphology in DM. The aim of the study was to analyze NMJ plasticity due to DM in an animal model and its relationship with the function of forelimbs in rats. Twelve Wistar rats were divided into control (C) and DM groups. Animals were trained to perform a grasping task, following procedures of habituation, shaping, and reaching task. DM was induced using streptozotocin. Forelimb neuromuscular performance for dexterity was evaluated one day before DM induction and five weeks following induction. After that, biceps, triceps, and finger flexors and extensors were removed. Connective tissue and muscle fiber cross-sectional area (CSA) were measured. NMJ was assessed by its morphometric characteristics (area, perimeter, and maximum diameter), using ImageJ software. Motor performance analyses were made using single pellet retrieval task performance test. Student’s t-test was used for comparisons between groups. A significant decrease in all NMJ morphometric parameters was observed in the DM group compared with the C group. Results showed that DM generated NMJ retraction in muscles involved in a reaching task. These alterations are related to signs of muscular atrophy and to poor reaching task performance. In conclusion, induced DM caused NMJ retraction and muscular atrophy in muscles involved in reaching task performance. Induced DM caused significantly lower motor performance, especially in the final moments of evaluation, when DM compromised the tropism of the muscular tissue.

Chronic stability of single-channel neurophysiological correlates of gross and fine reaching movements in the rat.

While substantial task-related neural activity has been observed during motor tasks in rodent primary motor cortex and premotor cortex, the long-term stability of these responses in healthy rats is uncertain, limiting the interpretability of longitudinal changes in the specific patterns of neural activity associated with learning or motor recovery following injury. This study examined the stability of task-related neural activity associated with execution of two distinct reaching tasks in healthy rodents. A novel automated rodent behavioral apparatus was constructed and rats were trained to perform a reaching task combining a ‘gross’ lever press and a ‘fine’ pellet retrieval. In each animal, two chronic microelectrode arrays were implanted in motor cortex spanning the caudal forelimb area (rodent primary motor cortex) and the rostral forelimb area (rodent premotor cortex). We recorded multiunit spiking and local field potential activity from 10 days to 7–10 weeks post-implantation to characterize the patterns of neural activity observed during each task component and analyzed the consistency of channel-specific task-related neural activity. Task-related changes in neural activity were observed on the majority of channels. While the task-related changes in multi-unit spiking and local field potential spectral power were consistent over several weeks, spectral power changes were more stable, despite the trade-off of decreased spatial and temporal resolution. These results show that neural activity in rodent primary and premotor cortex is associated with specific phases of reaching movements with stable patterns of task-related activity across time, establishing the relevance of the rodent for future studies designed to examine changes in task-related neural activity during recovery from focal cortical lesions.

Skilled reaching deterioration contralateral to cervical hemicontusion in rats is reversed by pregabalin treatment conditional upon its early administration.

Introduction:Gabapentinoids are first-line treatments for painful traumatic and nontraumatic central nervous system disorders. Evidence from a large human study suggests that early use of gabapentinoids after spinal cord injury improves motor scores. The underlying mechanism is unknown.Objectives:We sought to examine the effects of early pregabalin (PGB, a gabapentinoid) treatment on performance in a fine motor task (skilled reaching) after cervical hemicontusion. We also asked whether early PGB administration affected PGB responsiveness later on.Methods:Rats received C4/5 cervical hemicontusions. Injury severities ranged from 80 to 150 kdyn. We monitored evidence of skin irritation (peri-incisional and elsewhere) and quantified food pellet retrieval using the Montoya staircase test. Behaviours were assessed in rats receiving early (for 3 weeks from injury induction) and/or late (resuming or beginning at week 8) PGB treatment in animals with 150-kdyn injuries.Results:Contralateral skilled reaching waned in control animals with 150-kdyn injuries. This was prevented in animals, which received early PGB as long as treatment continued. Deterioration of skilled reaching was reversed by later (week 8) PGB only in animals that had received early treatment. Ipsilateral reaching impairment was not improved by PGB. Relief of skin irritation verified early PGB efficacy.Conclusion:Hemicontusive spinal cord injury produces a contralateral motor phenotype evocative of on-going neuropathic pain. Early PGB preserves sensitivity to subsequent PGB treatment, indicating that motor function is impaired by neuropathic pain and can be improved indirectly by early PGB administration. Direct effects of PGB on motor circuitry cannot be excluded but are not supported by our data.

Following unilateral spinal contusion pregabalin has an at‐time effect on pruritus and a protective effect on mechanosensory nociception, but does not improve ipsilateral motor outcomes with early administration in rats

Anticonvulsants like pregabalin (PGB) are the first‐line treatment for neuropathic pain caused by traumatic injury and non‐traumatic diseases of the central nervous system. Recent evidence from a human cohort study suggests that early use of pregabalin after spinal cord injury (SCI) may result in improved motor scores, however, it is unknown to what extent changes in spinal neural circuitry are involved. Using a rat model of unilateral cervical contusion, we examined the effect of pregabalin treatment on both motor and sensory function. For four weeks post‐injury, rats were given daily pregabalin or filtered water via oral gavage. Motor function was scored using the Montoya staircase assessment for food pellet retrieval (MSA). Additionally, pruritus and noxious mechosensation were assessed through behavioural evidence of scratching and the Randall‐Selitto analgesiometre, respectively. Finally, we used immunohistochemistry to characterize the primary afferent axons responsible for conveying signals related to nociception and itch. We found no significant relationship between the early administration of pregabalin and improved motor scores in the affected forelimb following MSA analysis. However, rats receiving pregabalin had improved performance on the MSA using their contralateral paw, suggesting alleviation of ongoing pain. Additionally, we found that self‐injurious scratching, often displayed by animals with this type of injury, was greatly reduced in those treated with PGB. However, following a two‐month sham injury experiment without PGB treatment, it was found that the scratching was not necessarily due to SCI, except for a small number of cases in which scratching included the territory of the greater auricular nerve. Furthermore, results of the Randall‐Selitto analgesiometre indicate a protective effect that lasted after discontinuation of PGB. Finally, we saw no changes in the density of pruritic and nociceptive primary afferents just rostral to the site of injury. Our findings suggest that in rats with a unilateral SCI, PGB treatment has a possible protective effect on mechanosensory nociception, aids in diminishing pain and itch during use, and contrary to the human cohort study, does not improve motor outcomes with early administration.Support or Funding InformationWings for Life; Natural Sciences and Engineering Research Council (NSERC)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Focal Malonate Injection Into the Internal Capsule of Rats as a Model of Lacunar Stroke.

Background: Stroke is the first cause of disability in adults in western countries. Infarct of the internal capsule (IC) may be related to motor impairment and poor prognosis in stroke patients. Functional deficits due to medium-sized infarcts are difficult to predict, except if the specific site of the lesion is taken into account. None of the few pre-clinical models recapitulating this type of stroke has shown clear, reproducible, and long-lasting sensorimotor deficits. Here, we developed a rat model of lacunar infarction within the IC, key structure of the sensorimotor pathways, by precise injection of malonate.Methods: The mitochondrial toxin malonate was injected during stereotactic surgery into the IC of rat brains. Rats were divided in three groups: two groups received malonate solution at 1.5M (n = 12) or at 3M (n = 10) and a sham group (n = 5) received PBS. Three key motor functions usually evaluated following cerebral lesion in the clinic strength, target reaching, and fine dexterity were assessed in rats by a forelimb grip strength test, a skilled reaching task (staircase) for reaching and dexterity, and single pellet retrieval task. Sensorimotor functions were evaluated by a neurological scale. Live brain imaging, using magnetic resonance (MRI), and post-mortem immunohistochemistry in brain slices were performed to characterize the lesion site after malonate injection.Results: Intracerebral injection of malonate produced a 100% success rate in inducing a lesion in the IC. All rats receiving the toxin, regardless the dose injected, had similar deficits in strength and dexterity of the contralateral forepaw, and showed significant neurological impairment. Additionally, only partial recovery was observed with respect to strength, while no recovery was observed for dexterity and neurological deficit. MRI and immunostaining show volume size and precise location of the lesion in the IC, destruction of axonal structures and Wallerian degeneration of fibers in the area above the injection site.Conclusions: This pre-clinical model of lacunar stroke induces a lesion in the IC with measurable and reproducible sensorimotor deficits, and limited recovery with stabilization of performance 2 weeks post-injury. Future therapies in stroke may be successfully tested in this model.

SOX-2 Mediated Conversion of Astrocyte to Neuron

Background: Axonal regeneration following SCI is crucial if individuals are to avoid permanent neurological damage. A plausible solution to circumvent neuronal death/ axonal degeneration may be in the conversion of ubiquitously distributed astrocytes into neurons. Previous research has shown that the neuronal transcription factor SOX-2 has the potential to convert mature glial cells into neuroblasts (iANBs). Furthermore, iANBs possess the capability to differentiate into functional neurons. Our study employs an in vivo mouse model to demonstrate the functional recovery that SOX-2 mediated conversion of astrocytes to neurons may pose following SCI. 
 Methods: Our study utilized two populations of mice. The first population received thoracic T10 contusion injuries while the second population received cervical C5 dorsal hemi-sections. Mice were subsequently blindly categorized into groups A, B, and C (treatment groups) depending on which injection they would be receiving: LV-hNG2-GFP, LV-hNG2-SOX2, or LV-p75-2. Following injection, behavioral studies including Hargreaves, roto-rod, grid walk, BMS (contusion mice) and pellet retrieval (DH mice) were performed to assess functional recovery at 2 week intervals. 
 Results: The astrocyte conversion to neuron project spans more than 16 weeks. Here we present data obtained in the first eight weeks which includes behavioral analyses of contusion injury mice. Currently grid walk, BMS, and Hargreaves testing show similar trends in spontaneous recovery however no significant difference is observed between the independent injection groups. On the contrary roto-rod analyses showed injection group C had a significantly lower latency to fall time in comparison to groups A and B six weeks post injection (wpi). 
 Conclusion: The treatments are hypothesized to not take effect until approximately 8 wpi thus we expect subsequent behavioral testing to reveal significant differences between treatment groups, ultimately taking one step closer towards therapeutic intervention following traumatic SCI.

Cannabinoid receptor-1 signaling contributions to sign-tracking and conditioned reinforcement in rats.

Endocannabinoids (eCBs) are critical gatekeepers of dopaminergic signaling, and disrupting cannabinoid receptor-1 (CB1) signaling alters DA dynamics to attenuate cue-motivated behaviors. Prior studies suggest that dopamine (DA) release plays a critical role in driving sign-tracking. Here, we determine whether systemic injections of rimonabant, a CB1 receptor inverse agonist, during Pavlovian lever autoshaping impair the expression of sign-tracking. We next examine whether rimonabant blocks the reinforcing properties of the Pavlovian lever cue in a conditioned reinforcement test. In Exp. 1, we trained rats in Pavlovian lever autoshaping prior to systemic rimonabant injections (0, 1, 3mg/kg) during early and late Pavlovian lever autoshaping sessions. In Exp. 2, we trained rats in Pavlovian lever autoshaping prior to systemic rimonabant injections (0, 1mg/kg) during a conditioned reinforcement test. Rimonabant dose-dependently decreased lever contact and probability, and increased sign-tracker's latency to approach the lever cue early in Pavlovian training. With extended training, many previously goal-tracking and intermediate rats shifted to lever approach, which remained dose-dependently sensitive to rimonabant. Rimonabant attenuated cue-evoked food cup approach early, but not late, in conditioning, and did not affect pellet retrieval or consumption. The inserted lever cue served as a robust conditioned reinforcer after Pavlovian lever autoshaping, and 1mg/kg rimonabant blocked conditioned reinforcement. Together, our results suggest that CB1 signaling mediates two critical properties of incentive stimuli; their ability to attract (Exp. 1) and their ability to reinforce (Exp. 2) behavior.

Synergistic Effects of Enriched Environment and Task-Specific Reach Training on Poststroke Recovery of Motor Function.

Reach training in concert with environmental enrichment provides functional benefits after experimental stroke in rats. The present study extended these findings by assessing whether intensive task-specific reach training or enrichment initiated alone would provide similar functional benefit. Additionally, we investigated whether the 70% recovery rule, or a combined model of initial poststroke impairment, cortical infarct volume, and rehabilitation intensity, could predict recovery in the single-pellet task, as previously found for the Montoya staircase. Rats were trained on single-pellet reaching before middle cerebral artery occlusion via intracerebral injection of ET-1 (endothelin-1). There were 4 experimental groups: stroke+enrichment, stroke+reaching, stroke+enrichment+reaching, and sham+enrichment+reaching. Reaching rehabilitation utilized a modified Whishaw box that encouraged impaired forelimb reaching for 6 hours per day, 5 days per week, for 4 weeks. All treatment paradigms began 7 days after ischemia with weekly assessment on the single-pellet task during rehabilitation and again 4 weeks after rehabilitation concluded. Rats exposed to the combination of enrichment and reaching showed the greatest improvement in pellet retrieval and comparable performance to shams after 3 weeks of treatment, whereas those groups that received a monotherapy remained significantly impaired at all time points. Initial impairment alone did not significantly predict recovery in single-pellet as the 70% rule would suggest; however, a combined model of cortical infarct volume and rehabilitation intensity predicted change in pellet retrieval on the single-pellet task with the same accuracy as previously shown with the staircase, demonstrating the generalizability of this model across reaching tasks. Task-specific reach training and environmental enrichment have synergistic effects in rats that persist long after rehabilitation ends, and this recovery is predicted by infarct volume and rehabilitation intensity.

Does Stroke Rehabilitation Really Matter? Part B: An Algorithm for Prescribing an Effective Intensity of Rehabilitation

Background. The proportional recovery rule suggests that current rehabilitation practices may have limited ability to influence stroke recovery. However, the appropriate intensity of rehabilitation needed to achieve recovery remains unknown. Similarities between rodent and human recovery biomarkers may allow determination of rehabilitation thresholds necessary to activate endogenous biological recovery processes. Objective. We determined the relative influence that clinically relevant biomarkers of stroke recovery exert on functional outcome. These biomarkers were then used to generate an algorithm that prescribes individualized intensities of rehabilitation necessary for recovery of function. Methods. A retrospective cohort of 593 male Sprague-Dawley rats was used to identify biomarkers that best predicted poststroke change in pellet retrieval in the Montoya staircase-reaching task using multiple linear regression. Prospective manipulation of these factors using endothelin-1-induced stroke (n = 49) was used to validate the model. Results. Rehabilitation was necessary to reliably predict recovery across the continuum of stroke severity. As infarct volume and initial impairment increased, more intensive rehabilitation was required to engage recovery. In this model, we prescribed the specific dose of daily rehabilitation required for rats to achieve significant motor recovery using the biomarkers of initial poststroke impairment and infarct volume. Conclusions. Our algorithm demonstrates an individualized approach to stroke rehabilitation, wherein imaging and functional performance measures can be used to develop an optimized rehabilitation paradigm for rats, particularly those with severe impairments. Exploring this approach in human patients could lead to an increase in the proportion of individuals experiencing recovery of lost motor function poststroke.

Does Stroke Rehabilitation Really Matter? Part A: Proportional Stroke Recovery in the Rat

Background. In human upper-limb stroke, initial level of functional impairment or corticospinal tract injury can accurately predict the degree of poststroke recovery, independent of rehabilitation practices. This proportional recovery rule implies that current rehabilitation practices may play little or no role in brain repair, with recovery largely a result of spontaneous biological recovery processes. Objective. The present study sought to determine if similar biomarkers predict recovery of poststroke function in rats, indicating that an endogenous biological recovery process might be preserved across mammalian species. Methods. Using a cohort of 593 male Sprague-Dawley rats, we predicted poststroke change in pellet retrieval in the Montoya staircase-reaching task based on initial impairment alone. Stratification of the sample into “fitters” and “nonfitters” of the proportional recovery rule using hierarchical cluster analysis allowed identification of distinguishing characteristics of these subgroups. Results. Approximately 30% of subjects were identified as fitters of the rule. These rats showed recovery in proportion to their initial level of impairment of 66% (95% CI = 62%-70%). This interval overlaps with those of multiple human clinical trials. A number of variables, including less severe infarct volumes and initial poststroke impairments distinguished fitters of the rule from nonfitters. Conclusions. These findings suggest that proportional recovery is a cross-species phenomenon that can be used to uncover biological mechanisms contributing to stroke recovery.

Axotomized Corticospinal Neurons Increase Supra-Lesional Innervation and Remain Crucial for Skilled Reaching after Bilateral Pyramidotomy

Skilled upper limb function heavily depends on the corticospinal tract. After bilateral lesions to this tract, motor control is disrupted but can be partially substituted by other motor systems to allow functional recovery. However, the remaining roles of motor cortex and especially of axotomized corticospinal neurons(CSNs) are not well understood. Using the single pellet retrieval task in adult rats, we induced significant recovery of skilled reaching after bilateral pyramidotomy by rehabilitative reaching training, and show that reach-related motor cortex activity, recorded in layer V, topographically reappeared shortly after axotomy. Using a chemogenetic neuronal silencing technique, we found that axotomized CSNs retained a crucial role for the recovered pellet retrieval success. The axotomized CSNs sprouted extensively in the red nucleus supplying new innervation to its magnocellular and parvocellular parts. Specific silencing of the rubrospinal tract(RST) also strongly abolished the recovered pellet retrieval success, suggesting a role of this cervically projecting nucleus in relaying cortical motor control. In summary, our results show that after bilateral corticospinal axotomy, motor cortex still actively engages in forelimb motor control and axotomized CSNs are crucially involved in the recovered reaching movement, potentially by relaying motor control via the RST.