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Purpose: Postoperative pneumonia remains a common complication of surgery, despite increased attention. The purpose of our study was to determine the effects of routine surgery and post-surgical opioid administration on airway protection risk., Methods: Eight healthy adult cats were evaluated to determine changes in airway protection status and for evidence of dysphagia in two experiments. (1) In four female cats, airway protection status was tracked following routine abdominal surgery (spay surgery) plus low-dose opioid administration (buprenorphine 0.015 mg/kg, IM, q8-12 h; n = 5). (2) Using a cross-over design, four naive cats (2 male, 2 female) were treated with moderate-dose (0.02 mg/kg) or high-dose (0.04 mg/kg) buprenorphine (IM, q8-12 h; n = 5)., Results: Airway protection was significantly affected in both experiments, but the most severe deficits occurred post-surgically as 75% of the animals exhibited silent aspiration., Conclusion: Oropharyngeal swallow is impaired by the partial mu-opioid receptor agonist buprenorphine, most remarkably in the postoperative setting. These findings have implications for the prevention and management of aspiration pneumonia in vulnerable populations., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Gastrointestinal (GI) complications, including motility disorders, metabolic deficiencies, and changes in gut microbiota following spinal cord injury (SCI), are associated with poor outcomes. After SCI, the autonomic nervous system becomes unbalanced below the level of injury and can lead to severe GI dysfunction. The SmartPill™ is a non-invasive capsule that, when ingested, transmits pH, temperature, and pressure readings that can be used to assess effects in GI function post-injury. Our minipig model allows us to assess these post-injury changes to optimize interventions and ultimately improve GI function. The aim of this study was to compare pre-injury to post-injury transit times, pH, and pressures in sections of GI tract by utilizing the SmartPill™ in three pigs after SCI at 2 and 6 weeks. Tributyrin was administered to two pigs to assess the influences on their gut microenvironment. We observed prolonged GET (Gastric Emptying Time) and CTT (Colon Transit Time), decreases in contraction frequencies (Con freq) in the antrum of the stomach, colon, and decreases in duodenal pressures post-injury. We noted increases in Sum amp generated at 2 weeks post-injury in the colon, with corresponding decreases in Con freq. We found transient changes in pH in the colon and small intestine at 2 weeks post-injury, with minimal effect on stomach pH post-injury. Prolonged GETs and CTTs can influence the absorptive profile in the gut and contribute to pathology development. This is the first pilot study to administer the SmartPill™ in minipigs in the context of SCI. Further investigations will elucidate these trends and characterize post-SCI GI function.

The immature central nervous system is recognized as having substantial neuroplastic capacity. In this study, we explored the hypothesis that rehabilitation can exploit that potential and elicit reciprocal walking in nonambulatory children with chronic, severe (i.e., lower extremity motor score < 10/50) spinal cord injuries (SCIs). Seven male subjects (3-12 years of age) who were at least 1-year post-SCI and incapable of discrete leg movements believed to be required for walking, enrolled in activity-based locomotor training (ABLT; clinicaltrials.gov NCT00488280). Six children completed the study. Following a minimum of 49 sessions of ABLT, three of the six children achieved walking with reverse rolling walkers. Stepping development, however, was not accompanied by improvement in discrete leg movements as underscored by the persistence of synergistic movements and little change in lower extremity motor scores. Interestingly, acoustic startle responses exhibited by the three responding children suggested preserved reticulospinal inputs to circuitry below the level of injury capable of mediating leg movements. On the other hand, no indication of corticospinal integrity was obtained with transcranial magnetic stimulation evoked responses in the same individuals. These findings suggest some children who are not predicted to improve motor and locomotor function may have a reserve of adaptive plasticity that can emerge in response to rehabilitative strategies such as ABLT. Further studies are warranted to determine whether a critical need exists to re-examine rehabilitation approaches for pediatric SCI with poor prognosis for any ambulatory recovery., (© 2023 Wiley Periodicals LLC.)

Animal models are necessary to identify pathological changes and help assess therapeutic outcomes following spinal cord injury (SCI). Small animal models offer value in research in terms of their easily managed size, minimal maintenance requirements, lower cost, well-characterized genomes, and ability to power research studies. However, despite these benefits, small animal models have neurologic and anatomical differences that may influence translation of results to humans and thus limiting the success of their use in preclinical studies as a direct pipeline to clinical studies. Large animal models, offer an attractive intermediary translation model that may be more successful in translating to the clinic for SCI research. This is largely due to their greater neurologic and anatomical similarities to humans. The physical characteristics of pig spinal cord, gut microbiome, metabolism, proportions of white to grey matter, bowel anatomy and function, and urinary system are strikingly similar and provide great insight into human SCI conditions. In this review, we address the variety of existing porcine injury models and their translational relevance, benefits, and drawbacks in modeling human systems and functions for neurophysiology, cardiovascular, gastrointestinal and urodynamic functions., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Laryngeal function is vital to airway protection. Although swallow is mediated by the brainstem, the mechanism underlying the increased risk of dysphagia after cervical spinal cord injury (SCI) is unknown. We hypothesized that: 1 ) loss of descending phrenic drive affects swallow and breathing differently, and 2 ) loss of ascending spinal afferent information alters swallow regulation. We recorded electromyograms (EMGs) from upper airway and chest wall muscles in freely breathing pentobarbital-anesthetized cats and rats. Laryngeal abductor activity during inspiration increased about twofold following C2 lateral hemisection. Ipsilateral to the injury, the crural diaphragm EMG amplitude was reduced during breathing (62 ± 25% change postinjury), but no animal had complete termination of activity; 75% of animals had increased contralateral diaphragm recruitment, but this did not reach significance. During swallow, laryngeal adductor and pharyngeal constrictor muscles increased activity, and diaphragm activity was bilaterally suppressed. This was unexpected because of the ipsilateral-specific response during breathing. Swallow-breathing coordination was disrupted by injury, and more swallows occurred during early expiration. Finally, to determine if the chest wall is a major source of feedback for laryngeal regulation, we performed T1 total transections in rats. As in the C2 lateral hemisection, inspiratory laryngeal recruitment was the first feature noted after injury. In contrast to the C2 lateral hemisection, diaphragmatic drive increased after T1 transection. Overall, we found that SCI alters laryngeal drive during swallow and breathing, and alters swallow-related diaphragm activity. Our results show behavior-specific effects, suggesting that swallow is affected more than breathing is by SCI, and emphasizing the need for additional studies on the effect of ascending afferents from the spinal cord on laryngeal function. NEW & NOTEWORTHY This is the first manuscript to determine the impact of cSCI on laryngeal and swallow function, and to describe a possible mechanism for dysphagia and altered airway protection after injury.

Successful propagation throughout the step cycle is contingent on adequate regulation of whole-limb stiffness by proprioceptive feedback. Following spinal cord injury (SCI), there are changes in the strength and organization of proprioceptive feedback that can result in altered joint stiffness. In this study, we measured changes in autogenic feedback of five hindlimb extensor muscles following chronic low thoracic lateral hemisection (LSH) in decerebrate cats. We present three features of the autogenic stretch reflex obtained using a mechanographic method. Stiffness was a measure of the resistance to stretch during the length change. The dynamic index documented the extent of adaptation or increase of the force response during the hold phase, and the impulse measured the integral of the response from initiation of a stretch to the return to the initial length. The changes took the form of variable and transient increases in the stiffness of vastus (VASTI) group, soleus (SOL), and flexor hallucis longus (FHL), and either increased (VASTI) or decreased adaptation (GAS and PLANT). The stiffness of the gastrocnemius group (GAS) was also variable over time but remained elevated at the final time point. An unexpected finding was that these effects were observed bilaterally. Potential reasons for this finding and possible sources of increased excitability to this muscle group are discussed., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature.)

There is an increasing need to develop approaches that will not only improve the clinical management of neurogenic lower urinary tract dysfunction (NLUTD) after spinal cord injury (SCI), but also advance therapeutic interventions aimed at recovering bladder function. Although pre-clinical research frequently employs rodent SCI models, large animals such as the pig may play an important translational role in facilitating the development of devices or treatments. Therefore, the objective of this study was to develop a urodynamics protocol to characterize NLUTD in a porcine model of SCI. An iterative process to develop the protocol to perform urodynamics in female Yucatan minipigs began with a group of spinally intact, anesthetized pigs. Subsequently, urodynamic studies were performed in a group of awake, lightly restrained pigs, before and after a contusion-compression SCI at the T2 or T9-T11 spinal cord level. Bladder tissue was obtained for histological analysis at the end of the study. All anesthetized pigs had bladders that were acontractile, which resulted in overflow incontinence once capacity was reached. Uninjured, conscious pigs demonstrated appropriate relaxation and contraction of the external urethral sphincter during the voiding phase. SCI pigs demonstrated neurogenic detrusor overactivity and a significantly elevated post-void residual volume. Relative to the control, SCI bladders were heavier and thicker. The developed urodynamics protocol allows for repetitive evaluation of lower urinary tract function in pigs at different time points post-SCI. This technique manifests the potential for using the pig as an intermediary, large animal model for translational studies in NLUTD.

Yucatan miniature pigs (YMPs) are similar to humans in spinal cord size as well as physiological and neuroanatomical features, making them a useful model for human spinal cord injury. However, little is known regarding pig gait kinematics, especially on a treadmill. In this study, 12 healthy YMPs were assessed during bipedal and/or quadrupedal stepping on a treadmill at six speeds (1.0, 1.5, 2.0, 2.5, 3.0, and 3.5 km/h). Kinematic parameters, including limb coordination and proximal and distal limb angles, were measured. Findings indicate that YMPs use a lateral sequence footfall pattern across all speeds. Stride and stance durations decreased with increasing speed whereas swing duration showed no significant change. Across all speeds assessed, no significant differences were noted between hindlimb stepping parameters for bipedal or quadrupedal gait with the exception of distal limb angular kinematics. Specifically, significant differences were observed between locomotor tasks during maximum flexion (quadrupedal > bipedal), total excursion (bipedal > quadrupedal), and the phase relationship between the timing of maximum extension between the right and left hindlimbs (bipedal > quadrupedal). Speed also impacted maximum flexion and right-left phase relationships given that significant differences were found between the fastest speed (3.5 km/h) relative to each of the other speeds. This study establishes a methodology for bipedal and quadrupedal treadmill-based kinematic testing in healthy YMPs. The treadmill approach used was effective in recruiting primarily the spinal circuitry responsible for the basic stepping patterns as has been shown in cats. We recommend 2.5 km/h (0.7 m/sec) as a target walking gait for pre-clinical studies using YMPs, which is similar to that used in cats.

Inhibitory pathways from Golgi tendon organs project widely between muscles crossing different joints and axes of rotation. Evidence suggests that the strength and distribution of this intermuscular inhibition is dependent on motor task and corresponding signals from the brainstem. The purpose of the present study was to investigate whether this sensory network is altered after spinal cord hemisection as a potential explanation for motor deficits observed after spinal cord injury (SCI). Force feedback was assessed between the long toe flexor and ankle plantarflexor (flexor hallucis longus), and the three major ankle extensors, (combined gastrocnemius, soleus, and plantaris muscles) in the hind limbs of unanesthetized, decerebrate, female cats. Data were collected from animals with intact spinal cords (control) and lateral spinal hemisections (LSHs) including chronic LSH (4-20 weeks), subchronic LSH (2 weeks), and acute LSH. Muscles were stretched individually and in pairwise combinations to measure intermuscular feedback between the toe flexor and each of the ankle extensors. In control animals, three patterns were observed (balanced inhibition between toe flexor and ankle extensors, stronger inhibition from toe flexor to ankle extensor, and vice versa). Following spinal hemisection, only strong inhibition from toe flexors onto ankle extensors was observed independent of survival time. The results suggest immediate and permanent reorganization of force feedback in the injured spinal cord. The altered strength and distribution of force feedback after SCI may be an important future target for rehabilitation., (Published 2020. This article is a U.S. Government work and is in the public domain in the USA.)

Lung volume is modulated by sensory afferent feedback via vagal and spinal pathways. The purpose of this study was to systematically alter afferent feedback with and without a mechanical challenge (chest compression). We hypothesized that manipulation of afferent feedback by nebulization of lidocaine, extra-thoracic vagotomy, or lidocaine administration to the pleural space would produce differential effects on the motor pattern of breathing during chest compression in sodium pentobarbital anesthetized rats (N = 43). Our results suggest that: 1) pulmonary stretch receptors are not the sole contributor to breathing feedback in adult male and female rats; 2) of our manipulations, chest compression had the largest effect on early expiratory diaphragm activity ("yield"); 3) reduction of spinally-mediated afferent feedback modulates breathing patterns most likely via inhibition; and 4) breathing parameters demonstrate large sex differences. Compared to males, female animals had lower respiratory rates (RR), which were further depressed by vagotomy, while chest compression increased RR in males, and decreased yield in females without changing RR. Collectively, our results suggest that balance between tonic vagal inhibition and spinal afferent feedback maintains breathing characteristics, and that it is important to specifically evaluate sex differences when studying control of breathing., Competing Interests: The authors have declared that no competing interest exist.

Swallow-breathing coordination is influenced by changes in lung volume, which is modulated by feedback from both vagal and spinal sensory afferents. The purpose of this study was to manipulate feedback from these afferents, with and without a simultaneous mechanical challenge (chest compression), in order to assess the influence of each sensory pathway on swallow in rats. We hypothesized that manipulation of afferent feedback would shift the occurrence of swallow toward the inspiratory phase of breathing. Afferent feedback was perturbed by lidocaine nebulization, extra-thoracic vagotomy, or lidocaine administration to the pleural space in sodium pentobarbital anesthetized rats (N = 43). These different afferent perturbations were performed both in control conditions (no chest compression), and with chest compression. Manipulating pulmonary stretch receptor-mediated volume feedback in male animals decreased swallow occurrence. Female rats appear to rely more on spinal afferent feedback, as swallow occurrence shifted to late expiration with chest compression and vagotomy or lidocaine injections. Results suggest that sex-specific mechanisms modulate swallow-breathing coordination, and that vagal feedback is inhibitory to swallow-related muscles, while spinal feedback from pleural afferents has excitatory effects. This study supports the theory that a balance of vagal and spinal afferent feedback is necessary to maintain an optimal swallow pattern and swallow-breathing coordination., Competing Interests: The authors have declared that no competing interests exist.

Afferent feedback can appreciably alter the pharyngeal phase of swallow. In order to measure the stability of the swallow motor pattern during several types of alterations in afferent feedback, we assessed swallow during a conventional water challenge in four anesthetized cats, and compared that to swallows induced by fixed (20 Hz) and stochastic (1-20Hz) electrical stimulation applied to the superior laryngeal nerve. The swallow motor patterns were evaluated by electromyographic activity (EMG) of eight muscles, based on their functional significance: laryngeal elevators (mylohyoid, geniohyoid, and thyrohyoid); laryngeal adductor (thyroarytenoid); inferior pharyngeal constrictor (thyropharyngeus); upper esophageal sphincter (cricopharyngeus); and inspiratory activity (parasternal and costal diaphragm). Both the fixed and stochastic electrical stimulation paradigms increased activity of the laryngeal elevators, produced short-term facilitation evidenced by increasing swallow durations over the stimulus period, and conversely inhibited swallow-related diaphragm activity. Both the fixed and stochastic stimulus conditions also increased specific EMG amplitudes, which never occurred with the water challenges. Stochastic stimulation increased swallow excitability, as measured by an increase in the number of swallows produced. Consistent with our previous results, changes in the swallow motor pattern for pairs of muscles were only sometimes correlated with each other. We conclude that alterations in afferent feedback produced particular variations of the swallow motor pattern. We hypothesize that specific SLN feedback might modulate the swallow central pattern generator during aberrant feeding conditions (food/liquid entering the airway), which may protect the airway and serve as potentially important clinical diagnostic indicators., (Copyright © 2020 King, Shen, Musselwhite, Huff, Reed, Poliacek, Howland, Dixon, Morris, Bolser, Iceman and Pitts.)

Transfer of information across a spinal lesion is required for many aspects of recovery across diverse motor systems. Our understanding of axonal plasticity and which subpopulations of neurons may contribute to bridging substrates following injury, however, remains relatively incomplete. Most recently, attention has been directed to propriospinal neurons (PSNs), with research suggesting that they are capable of bridging a spinal lesion in rodents. In the current study, subpopulations of both long (C5) and short (T6, T8) PSNs-as well as a supraspinal system, the rubrospinal tract (RST)-were assessed following low thoracic (T9) hemisection in the cat using the retrograde tracer Fluoro-Gold. Acutely, within 2 weeks post-hemisection, the numbers of short and long PSNs, as well as contralateral RST neurons, with axons crossing the lesion were significantly decreased relative to uninjured controls. This decrease persisted bilaterally and was permanent in the long PSNs and the contralateral red nucleus (RN). However, by 16 weeks post-hemisection, the numbers of ipsilesional and contralesional short PSNs bridging the lesion were significantly increased. Further, the number of contralesional contributing short PSNs was significantly greater in injured animals than in uninjured animals. A significant increase over uninjured numbers also was seen in the ipsilateral (non-axotomized) RN. These findings suggest that a novel substrate of undamaged axons, which normally terminates rostral to the lesion, grows past a thoracic lesion after injury. This rostral population represents a major component of the bridging substrate seen and may represent an important anatomical target for evolving rehabilitation approaches as a substrate capable of contributing to functional recovery.

Background: Kinematic and kinetic analysis have been used to gain an understanding of canine movement and joint loading during gait. By non-invasively predicting muscle activation patterns and forces during gait, musculoskeletal models can further our understanding of normal variability and muscle activation patterns and force profiles characteristic of gait., Methods: Pelvic limb kinematics and kinetics were measured for a 2 year old healthy female Dachshund (5.4 kg) during gait using 3-D motion capture and force platforms. A computed tomography scan was conducted to acquire pelvis and pelvic limb morphology. Using the OpenSim modeling platform, a bilateral pelvic limb subject-specific rigid body musculoskeletal computer model was developed. This model predicted muscle activation patterns, muscle forces, and angular kinematics and joint moments during walking., Results: Gait kinematics determined from motion capture matched those predicted by the model, verifying model accuracy. Primary muscles involved in generating joint moments during stance and swing were predicted by the model: at mid-stance the adductor magnus et brevis (peak activation 53.2%, peak force 64.7 N) extended the hip, and stifle flexor muscles (biceps femoris tibial and calcaneal portions) flexed the stifle. Countering vertical ground reaction forces, the iliopsoas (peak activation 37.9%, peak force 68.7 N) stabilized the hip in mid-stance, while the biceps femoris patellar portion stabilized the stifle in mid-stance and the plantar flexors (gastrocnemius and flexor digitorum muscles) stabilized the tarsal joint during early stance. Transitioning to swing, the iliopsoas, rectus femoris and tensor fascia lata flexed the hip, while in late swing the adductor magnus et brevis impeded further flexion as biceps femoris tibial and calcaneal portions stabilized the stifle for ground contact., Conclusion: The musculoskeletal computer model accurately replicated experimental canine angular kinematics associated with gait and was used to predict muscle activation patterns and forces. Thus, musculoskeletal modeling allows for quantification of measures such as muscle forces that are difficult or impossible to measure in vivo ., (Copyright © 2020 Brown, Bertocci, States, Levine, Levine and Howland.)

Anatomical connections are reported between the cerebellum and brainstem nuclei involved in swallow such as the nucleus tractus solitarius, nucleus ambiguus, and Kölliker-fuse nuclei. Despite these connections, a functional role of the cerebellum during swallow has not been elucidated. Therefore, we examined the effects of cerebellectomy on swallow muscle recruitment and swallow-breathing coordination in anesthetized freely breathing cats. Electromyograms were recorded from upper airway, pharyngeal, laryngeal, diaphragm, and chest wall muscles before and after complete cerebellectomy. Removal of the cerebellum reduced the excitability of swallow (i.e., swallow number), and muscle recruitment of the geniohyoid, thyroarytenoid, parasternal (chestwall), and diaphragm muscles, but did not disrupt swallow-breathing coordination. Additionally, diaphragm and parasternal muscle activity during swallow is reduced after cerebellectomy, while no changes were observed during breathing. These findings suggest the cerebellum modulates muscle excitability during recruitment, but not pattern or coordination of swallow with breathing.

Background: Kinematic gait analysis is an important noninvasive technique used for quantitative evaluation and description of locomotion and other movements in healthy and injured populations. Three dimensional (3D) kinematic analysis offers additional outcome measures including internal-external rotation not characterized using sagittal plane (2D) analysis techniques., Methods: The objectives of this study were to 1) develop and evaluate a 3D hind limb multiplane kinematic model for gait analysis in cats using joint coordinate systems, 2) implement and compare two 3D stifle (knee) prediction techniques, and 3) compare flexion-extension determined using the multiplane model to a sagittal plane model. Walking gait was recorded in 3 female adult cats (age = 2.9 years, weight = 3.5 ± 0.2 kg). Kinematic outcomes included flexion-extension, internal-external rotation, and abduction-adduction of the hip, stifle, and tarsal (ankle) joints., Results: Each multiplane stifle prediction technique yielded similar findings. Joint angles determined using markers placed on skin above bony landmarks in vivo were similar to joint angles determined using a feline hind limb skeleton in which markers were placed directly on landmarks ex vivo. Differences in hip, stifle, and tarsal joint flexion-extension were demonstrated when comparing the multiplane model to the sagittal plane model., Conclusions: This multiplane cat kinematic model can predict joint rotational kinematics as a tool that can quantify frontal, transverse, and sagittal plane motion. This model has multiple advantages given its ability to characterize joint internal-external rotation and abduction-adduction. A further, important benefit is greater accuracy in representing joint flexion-extension movements., Competing Interests: The authors have declared that no competing interests exist.

Introduction: Functional walking requires the ability to modify one's gait pattern to environmental demands and task goals-gait adaptability. Following incomplete spinal cord injury (ISCI), gait rehabilitation such as locomotor training (Basic-LT) emphasizes intense, repetitive stepping practice. Rehabilitation approaches focusing on practice of gait adaptability tasks have not been established for individuals with ISCIs but may promote recovery of higher level walking skills. The primary purpose of this case series was to describe and determine the feasibility of administering a gait adaptability retraining approach-Adapt-LT-by comparing the dose and intensity of Adapt-LT to Basic-LT., Case Presentation: Three individuals with ISCIs (>1 year, AIS C or D) completed three weeks each (15 sessions) of Basic-LT and Adapt-LT. Interventions included practice on a treadmill with body weight support and practice overground (≥30 mins total). Adapt-LT focused on speed changes, obstacle negotiation, and backward walking. Training parameters (step counts, speeds, perceived exertion) were compared and outcomes assessed pre and post interventions. Based on completion of the protocol and similarities in training parameters in the two interventions, it was feasible to administer Adapt-LT with a similar dosage and intensity as Basic-LT. Additionally, the participants demonstrated gains in walking function and balance following each training type., Discussion: Rehabilitation that includes stepping practice with adaptability tasks is feasible for individuals with ISCIs. Further investigation is needed to determine the efficacy of Adapt-LT., Competing Interests: Compliance with ethical standardsThe authors declare that they have no competing interests.The contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH, Department of VA, or the U.S. Government.

Background: Fluorogold (FG) is used by many groups to retrogradely trace nervous system pathways. Fluorogold, while a robust tracer, also is neurotoxic and causes tissue damage at the injection site and leads to motor deficits., New Method: In the current study, we describe a method for enhancing FG-uptake using Triton™ and an overall procedure for reducing FG-related tissue damage while still allowing effective quantification., Results: Triton™ decreases the amount of FG, as well as the time required for long-distance transport from the thoracic spinal cord to the motor cortex by >4 fold when this distance is >10in. Although small FG concentrations and injection volumes are ideal for minimizing associated tissue damage and motor deficits, they result in difficult-to-detect fluorescence. This can be solved using FG antiserum paired with an ABC chromogen reaction. This ABC chromogen reaction product can remain stable for at least 9 years., Comparison With Existing Method(s): This study is the first to collectively address FG-induced tissue damage and describe methods for minimizing this damage., Conclusions: Triton™ enhances the uptake of FG in the nervous system, reduces the FG required, and allows for a substantial decrease in tracing time that limits FG-induced motor deficits. Small FG concentration and volume decreases tissue damage but also decreases FG fluorescent detection. Detection challenges are resolved using FG anti-serum and chromogen reactions., (Published by Elsevier B.V.)

We investigated the hypothesis, motivated in part by a coordinated computational cough network model, that second-order neurons in the nucleus tractus solitarius (NTS) act as a filter and shape afferent input to the respiratory network during the production of cough. In vivo experiments were conducted on anesthetized spontaneously breathing cats. Cough was elicited by mechanical stimulation of the intrathoracic airways. Electromyograms of the parasternal (inspiratory) and rectus abdominis (expiratory) muscles and esophageal pressure were recorded. In vivo data revealed that expiratory motor drive during bouts of repetitive coughs is variable: peak expulsive amplitude increases from the first cough, peaks about the eighth or ninth cough, and then decreases through the remainder of the bout. Model simulations indicated that feed-forward inhibition of a single second-order neuron population is not sufficient to account for this dynamic feature of a repetitive cough bout. When a single second-order population was split into two subpopulations (inspiratory and expiratory), the resultant model produced simulated expiratory motor bursts that were comparable to in vivo data. However, expiratory phase durations during these simulations of repetitive coughing had less variance than those in vivo. Simulations in which reciprocal inhibitory processes between inspiratory-decrementing and expiratory-augmenting-late neurons were introduced exhibited increased variance in the expiratory phase durations. These results support the prediction that serial and parallel processing of airway afferent signals in the NTS play a role in generation of the motor pattern for cough., (Copyright © 2016 the American Physiological Society.)

The effects of 2weeks of intralesional chondroitinase abc (ch'abc) treatment on anatomical plasticity and behavioral recovery are examined in adult cats and compared to results achieved with 4weeks of treatment following tightly controlled lateral hemisection injuries. Analyses also were completed using 35 cats with a range of hemisection magnitudes to assess relationships between treatment duration, lesion size and functional recovery. Results indicate that both 2 and 4weeks of treatment significantly increased the number of rubrospinal tract (RuST) neurons with axons below the lesion, but neither affected the number of corticospinal tract neurons. Similarly, both treatment periods also accelerated recovery of select motor tasks, which carries considerable importance with respect to human health care and rehabilitation. Four weeks of treatment promoted recovery beyond that seen with 2weeks in its significant impact on accuracy of movement critical for placement of the ipsilateral hindlimb onto small support surfaces during the most challenging locomotor tasks. Analyses, which extended to a larger group of cats with a range of lesion magnitudes, indicate that 4weeks of ch'abc treatment promoted earlier recovery as well as significantly greater targeting accuracy even in cats with larger lesions. Together, these results support the potential for ch'abc to promote anatomical and behavioral recovery and suggest that intraspinal treatment with ch'abc continues to enhance motor recovery and performance beyond the subacute injury period and diminishes the impact of lesion size., (Published by Elsevier Inc.)

A module is a functional unit of the nervous system that specifies functionally relevant patterns of muscle activation. In adults, four to five modules account for muscle activation during walking. Neurological injury alters modular control and is associated with walking impairments. The effect of neurological injury on modular control in children is unknown and may differ from adults due to their immature and developing nervous systems. We examined modular control of locomotor tasks in children with incomplete spinal cord injuries (ISCIs) and control children. Five controls (8.6 ± 2.7 yr of age) and five children with ISCIs (8.6 ± 3.7 yr of age performed treadmill walking, overground walking, pedaling, supine lower extremity flexion/extension, stair climbing, and crawling. Electromyograms (EMGs) were recorded in bilateral leg muscles. Nonnegative matrix factorization was applied, and the minimum number of modules required to achieve 90% of the "variance accounted for" (VAF) was calculated. On average, 3.5 modules explained muscle activation in the controls, whereas 2.4 modules were required in the children with ISCIs. To determine if control is similar across tasks, the module weightings identified from treadmill walking were used to reconstruct the EMGs from each of the other tasks. This resulted in VAF values exceeding 86% for each child and each locomotor task. Our results suggest that 1) modularity is constrained in children with ISCIs and 2) for each child, similar neural control mechanisms are used across locomotor tasks. These findings suggest that interventions that activate the neuromuscular system to enhance walking also may influence the control of other locomotor tasks.

Arm and leg coordination naturally emerges during walking, but can be affected by stroke or Parkinson's disease. The purpose of this preliminary study was to characterize arm and leg coordination during treadmill walking at self-selected comfortable walking speeds (CWSs) in individuals using arm swing with motor incomplete spinal cord injury (iSCI). Hip and shoulder angle cycle durations and amplitudes, strength of peak correlations between contralateral hip and shoulder joint angle time series, the time shifts at which these peak correlations occur, and associated variability were quantified. Outcomes in individuals with iSCI selecting fast CWSs (range, 1.0-1.3m/s) and speed-matched individuals without neurological injuries are similar. Differences, however, are detected in individuals with iSCI selecting slow CWSs (range, 0.25-0.65 m/s) and may represent compensatory strategies to improve walking balance or forward propulsion. These individuals elicit a 1:1, arm:leg frequency ratio versus the 2:1 ratio observed in non-injured individuals. Shoulder and hip movement patterns, however, are highly reproducible (coordinated) in participants with iSCI, regardless of CWS. This high degree of inter-extremity coordination could reflect an inability to modify a single movement pattern post-iSCI. Combined, these data suggest inter-extremity walking coordination may be altered, but is present after iSCI, and therefore may be regulated, in part, by neural control., (Published by Elsevier B.V.)

Advances in the neurobiology of spinal cord injury (SCI) have prompted increasing attention to opportunities for moving experimental strategies towards clinical applications. Preclinical studies are the centerpiece of the translational process. A major challenge is to establish strategies for achieving optimal translational progression while minimizing potential repetition of previous disappointments associated with clinical trials. This chapter reviews and expands upon views pertaining to preclinical design reported in recently published opinion surveys. Subsequent discussion addresses other preclinical considerations more specifically related to current and potentially imminent cellular and pharmacological approaches to acute/subacute and chronic SCI. Lastly, a retrospective and prospective analysis examines how guidelines currently under discussion relate to select examples of past, current, and future clinical translations. Although achieving definition of the "perfect" preclinical scenario is difficult to envision, this review identifies therapeutic robustness and independent replication of promising experimental findings as absolutely critical prerequisites for clinical translation. Unfortunately, neither has been fully embraced thus far. Accordingly, this review challenges the notion "everything works in animals and nothing in humans", since more rigor must first be incorporated into the bench-to-bedside translational process by all concerned, whether in academia, clinical medicine, or corporate circles., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Following a lateralized spinal cord injury (SCI) in humans, substantial walking recovery occurs; however, deficits persist in adaptive features of locomotion critical for community ambulation, including obstacle negotiation. Normal obstacle negotiation is accomplished by an increase in flexion during swing. If an object is unanticipated or supraspinal input is absent, obstacle negotiation may involve the spinally organized stumbling corrective response. How these voluntary and reflex components are affected following partial SCI is not well studied. This study is the first to characterize recovery of obstacle negotiation following low-thoracic spinal hemisection in the cat. Cats were trained pre- and post-injury to cross a runway with an obstacle. Assessments focused on the hindlimb ipsilateral to the lesion. Pre-injury, cats efficiently cleared an obstacle by increasing knee flexion during swing. Post-injury, obstacle clearance permanently changed. At 2 weeks, when basic overground walking ability been recovered, the hindlimb was dragged over the obstacle (∼90%). Surprisingly, the stumbling corrective response was not elicited until after 2 weeks. Despite a notable increase, between 4 and 8 weeks, in the ability to modify limb trajectory when approaching an obstacle, limb lift during obstacle approach was insufficient during ∼50% of encounters and continued to evoke the stumbling corrective response even at 16 weeks. A post-injury lead limb bias identified during negotiations with complete clearance, suggests a potential training strategy to increase the number of successful clearances. Therefore, following complete severing of half of the spinal cord, the ability to modify ipsilateral hindlimb trajectory shows significant recovery and by 16 weeks permits effective clearing of an obstacle, without contact, ∼50% of the time. Although this suggests plasticity of supporting circuitry, it is insufficient to support consistent clearance. This inconsistency, even at the most chronic time point assessed (16 weeks), is probably a contributing factor to falls reported for people with SCI.

A number of studies have shown that chondroitinase ABC (Ch'ase ABC) digestion of inhibitory chondroitin sulfate glycosaminoglycans significantly enhances axonal growth and recovery in rodents following spinal cord injury (SCI). Further, our group has shown improved recovery following SCI in the larger cat model. The purpose of the current study was to determine whether intraspinal delivery of Ch'ase ABC, following T10 hemisections in adult cats, enhances adaptive movement features during a skilled locomotor task and/or promotes plasticity of spinal and supraspinal circuitry. Here, we show that Ch'ase ABC enhanced crossing of a peg walkway post-SCI and significantly improved ipsilateral hindlimb trajectories and integration into a functional forelimb-hindlimb coordination pattern. Recovery of these complex movements was associated with significant increases in neurofilament immunoreactivity immediately below the SCI in the ipsilateral white (p = 0.033) and contralateral gray matter (p = 0.003). Further, the rubrospinal tract is critical in the normal cat during skilled movements that require accurate paw placements and trajectories like those seen during peg walkway crossing. Rubrospinal connections were assessed following Fluoro-Gold injections, caudal to the hemisection. Significantly more retrogradely labeled right (axotomized) red nucleus (RN) neurons were seen in Ch'ase ABC-treated (23%) compared with control-treated cats (8%; p = 0.032) indicating that a larger number of RN neurons in Ch'ase ABC-treated cats had axons below the lesion level. Thus, following SCI, Ch'ase ABC may facilitate axonal growth at the spinal level, enhance adaptive features of locomotion, and affect plasticity of rubrospinal circuitry known to support adaptive behaviors in the normal cat.

Study Design: Observational, cross-sectional study from a convenience sample with pretest/posttest data from a sample subset., Objectives: Determine the presence of walking-related arm swing after spinal cord injury (SCI), its associated factors and whether arm swing may change after locomotor training (LT)., Setting: Malcom Randall VAMC and University of Florida, Gainesville, FL., Methods: Arm movement was assessed during treadmill stepping, pre-LT, in 30 individuals with motor incomplete SCI (iSCI, American Spinal Injury Association Impairment Scale grade C/D, as defined by the International Standards for Neurological Classifications of SCI, with neurological level of impairment at or below C4). Partial body weight support and manual-trainer assistance were provided, as needed, to achieve stepping and allow arm swing. Arm swing presence was compared on the basis of cervical versus thoracic neurological levels of impairment and device type. Leg and arm strength and walking independence were compared between individuals with and without arm swing. Arm swing was reevaluated post-LT in the 21 out of 30 individuals who underwent LT., Results: Of 30 individuals with iSCI, 12 demonstrated arm swing during treadmill stepping, pre-LT. Arm movement was associated with device type, lower extremity motor scores and walking independence. Among the 21 individuals who received LT, only 5 demonstrated arm swing pre-LT. Of the 16 individuals lacking arm swing pre-LT, 8 integrated arm swing post-LT., Conclusion: Devices routinely used for walking post-iSCI appeared associated with arm swing. Post-LT, arm swing presence increased. Therefore, arm swing may be experience dependent. Daily neuromuscular experiences provided to the arms may produce training effects, thereby altering arm swing expression.

Background and Purpose: The authors previously reported on walking recovery in a nonambulatory child with chronic, severe, incomplete cervical spinal cord injury (SCI) after 76 sessions of locomotor training (LT). Although clinical measures did not predict his recovery, reciprocal patterned leg movements developed, affording recovery of independent walking with a reverse rolling walker. The long-term functional limitations and secondary complications often associated with pediatric-onset SCI necessitate continued follow-up of children with SCI. Therefore, the purpose of this case report is to describe this child's walking function and musculoskeletal growth and development during the 2 years since his participation in an LT program and subsequent walking recovery., Case Description: Following LT, the child attended elementary school as a full-time ambulator. He was evaluated 1 month (baseline), 1 year, and 2 years after LT. Examination of walking function included measures of walking independence, gait speed and spatiotemporal parameters, gait kinematics, and daily step activity. Growth and development were assessed by tracking his height, weight, incidence of musculoskeletal complications, and gross motor task performance., Outcomes: Over the 2 years, the child continued to ambulate independently with a reverse rolling walker, increasing his fastest gait speed. Spatiotemporal and kinematic features of his walking improved, and daily step activity increased. Height and weight remained on their preinjury trajectory and within age-appropriate norms. The child experienced only minor musculoskeletal complications. Additionally, he gained the ability to use reciprocal patterned leg movements during locomotor tasks such as assisted stair climbing and independent tricycle pedaling., Conclusions: Two years after recovery of walking, this child with incomplete SCI had maintained and improved his walking function and experienced age-appropriate growth and development.

A function of the abdominal expiratory muscles is the generation of cough, a critical respiratory defense mechanism that is often disrupted following spinal cord injury. We assessed the effects of a lateral T9/10 hemisection on cough production at 4, 13 and 21 weeks post-injury in cats receiving extensive locomotor training. The magnitudes of esophageal pressure as well as of bilateral rectus abdominis electromyogram activity during cough were not significantly different from pre-injury values at all time points evaluated. The results show that despite considerable interruption of the descending pre-motor drive from the brainstem to the expiratory motoneuron pools, the cough motor system shows a significant function by 4 weeks following incomplete thoracic injury., (Published by Elsevier Inc.)

Pulmonary morbidity is high following spinal cord injury and is due, in part, to impairment of airway protective behaviors. These airway protective behaviors include augmented breaths, the cough reflex, and expiration reflexes. Functional recovery of these behaviors has been reported after spinal cord injury. In humans, evidence for functional recovery is restricted to alterations in motor strategy and changes in the frequency of occurrence of these behaviors. In animal models, compensatory alterations in motor strategy have been identified. Crossed descending respiratory motor pathways at the thoracic spinal cord levels exist that are composed of crossed premotor axons, local circuit interneurons, and propriospinal neurons. These pathways can collectively form a substrate that supports maintenance and/or recovery of function, especially after asymmetric spinal cord injury. Local sprouting of premotor axons in the thoracic spinal cord also can occur following chronic spinal cord injury. These mechanisms may contribute to functional resiliency of the cough reflex that has been observed following chronic spinal cord injury in the cat.

Background: Poststroke reaching is characterized by excessive trunk motion and abnormal shoulder-elbow coordination. Little attention is typically given to arm-trunk kinematics during task practice. Preventing compensatory trunk motion during short-term practice immediately improves kinematics, but effects of longer-term practice are unknown., Objective: This study compared the effects of intensive task practice with and without trunk restraint on poststroke reaching kinematics and function., Methods: A total of 11 individuals with chronic stroke, baseline Fugl-Meyer Upper Extremity Assessment scores 26 to 54, were randomized to 2 constraint-therapy intervention groups. All participants wore a mitt on the unaffected hand for 90% of waking hours over 14 days and participated in 10 days/6 hours/day of supervised progressive task practice. During supervised sessions, one group trained with a trunk restraint (preventing anterior trunk motion) and one group did not. Tasks for the trunk-restraint group were located to afford repeated use of a shoulder flexion-elbow extension reaching pattern. Outcome measures included kinematics of unrestrained targeted reaching and tests of functional arm ability., Results: Posttraining, the trunk-restraint group demonstrated straighter reach trajectories (P=.000) and less trunk displacement (P=.001). The trunk-restraint group gained shoulder flexion (P=.006) and elbow extension (P=.022) voluntary ranges of motion, the nonrestraint group did not. Posttraining angle-angle plots illustrated that individuals from the trunk-restraint group transitioned from elbow flexion to elbow extension during mid-reach; individuals in the nonrestraint group retained pretraining movement strategies. Both groups gained functional arm ability (P<.05 all tests)., Conclusion: Intensive task practice structured to prevent compensatory trunk movements and promote shoulder flexion-elbow extension coordination may reinforce development of "normal" reaching kinematics.

Background and Purpose: Locomotor training (LT) enhances walking in adult experimental animals and humans with mild-to-moderate spinal cord injuries (SCIs). The animal literature suggests that the effects of LT may be greater on an immature nervous system than on a mature nervous system. The purpose of this study was to evaluate the effects of LT in a child with chronic, incomplete SCI., Subject: The subject was a nonambulatory 4 1/2-year-old boy with an American Spinal Injury Association Impairment Scale (AIS) C Lower Extremity Motor Score (LEMS) of 4/50 who was deemed permanently wheelchair-dependent and was enrolled in an LT program 16 months after a severe cervical SCI., Methods: A pretest-posttest design was used in the study. Over 16 weeks, the child received 76 LT sessions using both treadmill and over-ground settings in which graded sensory cues were provided. The outcome measures were ASIA Impairment Scale score, gait speed, walking independence, and number of steps., Result: One month into LT, voluntary stepping began, and the child progressed from having no ability to use his legs to community ambulation with a rolling walker. By the end of LT, his walking independence score had increased from 0 to 13/20, despite no change in LEMS. The child's final self-selected gait speed was 0.29 m/s, with an average of 2,488 community-based steps per day and a maximum speed of 0.48 m/s. He then attended kindergarten using a walker full-time., Discussion and Conclusion: A simple, context-dependent stepping pattern sufficient for community ambulation was recovered in the absence of substantial voluntary isolated lower-extremity movement in a child with chronic, severe SCI. These novel data suggest that some children with severe, incomplete SCI may recover community ambulation after undergoing LT and that the LEMS cannot identify this subpopulation.

Chondroitin sulfate proteoglycans (CSPGs) are upregulated in the central nervous system following injury. Chondroitin sulfate glycosaminoglycan (CS GAG) side chains substituted on this family of molecules contribute to the limited functional recovery following injury by restricting axonal growth and synaptic plasticity. In the current study, the effects of degrading CS GAGs with Chondroitinase ABC (Ch'ase ABC) in the injured spinal cords of adult cats were assessed. Three groups were evaluated for 5 months following T10 hemisections: lesion-only, lesion+control, and lesion+Ch'ase ABC. Intraspinal control and Ch'ase ABC treatments to the lesion site began immediately after injury and continued every other day, for a total of 15 treatments, using an injectable port system. Delivery and in vivo cleavage were verified anatomically in a subset of cats across the treatment period. Recovery of skilled locomotion (ladder, peg, and beam) was significantly accelerated, on average, by >3 weeks in Ch'ase ABC-treated cats compared to controls. Ch'ase ABC-treated cats also showed greater recovery of specific skilled locomotor features including intralimb movement patterns and significantly greater paw placement onto pegs. Although recovery of basic locomotion (bipedal treadmill and overground) was not accelerated, intralimb movement patterns were more normal in the Ch'ase ABC-treated cats. Qualitative assessment of serotonergic immunoreactivity also suggested that Ch'ase ABC treatment enhanced plasticity. Finally, analyses using fluorophore-assisted carbohydrate electrophoresis (FACE) indicate CS GAG content is similar in cat and human. These findings show, for the first time, that intraspinal cleavage of CS GAGs can enhance recovery of function following spinal cord injury in large animals with sophisticated motor behaviors and axonal growth requirements similar to those encountered in humans.

Chondroitinase ABC (Ch'ase ABC) is a bacterial lyase that degrades chondroitin sulfate (CS), dermatan sulfate, and hyaluronan glycosaminoglycans (GAGs). This enzyme has received significant attention as a potential therapy for promoting central nervous system and peripheral nervous system repair based on its degradation of CS GAGs. Determination of the stability of Ch'ase ABC activity at temperatures equivalent to normal (37 degrees C) and elevated (39 degrees C) body temperatures is important for optimizing its clinical usage. We report here data obtained from examining enzymatic activity at these temperatures across nine lots of commercially available protease-free Ch'ase ABC. CS GAG degrading activity was assayed by using 1) immunohistochemical detection of unsaturated disaccharide stubs generated by digestion of proteoglycans in tissue sections and 2) fluorophore-assisted carbohydrate electrophoresis (FACE) and/or high-performance liquid chromatography (HPLC) to separate and quantify unsaturated disaccharide digestion products. Our results indicate that there is a significant effect of lot and time on enzymatic thermostability. Average enzymatic activity is significantly decreased at 1 and 3 days at 39 degrees C and 37 degrees C, respectively. Furthermore, the average activity seen after 1 day was significantly different between the two temperatures. Addition of bovine serum albumin as a stabilizer significantly preserved enzymatic activity at 1 day, but not 3 days, at 39 degrees C. These results show that the CS GAG degrading activity of Ch'ase ABC is significantly decreased with incubation at body temperature over time and that all lots do not show equal thermostability. These findings are important for the design and interpretation of experimental and potential clinical studies involving Ch'ase ABC., ((c) 2007 Wiley-Liss, Inc.)

Netrin-1 regulates axon extension during embryonic development and is expressed by neurons and myelinating oligodendrocytes in the adult CNS. To investigate the potential role of netrin-1 after spinal cord injury, we examined the expression of netrin-1 and netrin receptors after sagittal myelotomy in adult rats. This lesion targets spinal commissural projections, which respond to netrin-1 during development. Netrin-1 mRNA and protein levels were dramatically reduced at the site of injury and reduced expression persisted for at least 7 months. Neither netrin-1 protein nor mRNA was associated with the glial scar, but netrin-1 was expressed by neurons and oligodendrocytes immediately adjacent to the lesion. The post-injury distribution detected is similar to that reported for myelin-associated inhibitors of axon regeneration, such as Nogo, and is distinct from the distribution of inhibitors associated with a glial scar. DCC and UNC-5 homologue (UNC5H) expression also was reduced after injury. Although UNC5H levels recovered, DCC expression at the site of injury remained approximately 50% of pre-injury values at 7 months. Increased UNC5H immunoreactivity was associated with fibers in the superficial layers of the dorsal horn and in fibers located in white matter adjacent to the lesion. The dominant expression of UNC5H on axons and neurons in the spinal cord after injury and the persistent expression of netrin-1 by oligodendrocytes surrounding the lesion are consistent with the hypothesis that netrin-1 is a myelin-associated inhibitor of axonal regeneration after spinal cord injury.

Several lines of reasoning suggest that the phosphorylated axonal form of the neurofilament subunit NF-H is likely to be released from damaged and diseased neurons in significant amounts. Detection of this protein in serum or CSF might therefore provide information about the presence and degree of neuronal loss. We therefore developed a sensitive NF-H ELISA capable of detecting picogram quantities of phosphorylated NF-H (pNF-H). This assay showed that soluble pNF-H immunoreactivity is readily detectable in the sera of adult rats following various types of experimental spinal cord injury (SCI) and traumatic brain injury (TBI), but is undetectable in the sera of normal animals. Here we describe details of the time course and extent of serum pNF-H expression following experimental SCI and TBI. Following SCI, serum pNF-H showed an initial peak of expression at 16h and a second, usually larger, peak at 3 days. Following TBI, lower levels of serum pNF-H were detected with a peak at 2 days post-injury. We also show that the higher levels of pNF-H released from injured spinal cord as compared to brain are in line with the approximately 20-fold higher levels of pNF-H present in spinal cord. These findings suggest that serum levels of pNF-H immunoreactivity may be used to conveniently monitor neuronal damage and degeneration in experimental and presumably clinical situations.

Aggrecan is a chondroitin sulfate (CS)/keratan sulfate (KS)-substituted proteoglycan (PG) abundant in cartilage which is also present within the mammalian embryonic, adult, and injured adult central nervous system (CNS). Although its role within the CNS is not clear, cell culture studies show that when substituted with CS, aggrecan inhibits neurite extension. To better understand the inhibitory effect of aggrecan on injured adult axons in vivo, we developed a model to independently test intact aggrecan and CS-depleted aggrecan core glycoprotein. Acute rat spinal cord hemisection cavities were filled with a growth-promoting matrix, Matrigel, and severed dorsal rootlets were placed into this matrix. This created an assay in which axons readily grew. The extent of ingrowth in this baseline assay was compared to the ingrowth in Matrigel loaded with intact aggrecan or the purified core glycoprotein of aggrecan. Our results show that both intact aggrecan and equivalent concentrations of the core glycoprotein component significantly inhibit axonal growth in this model system. These results confirm that aggrecan can inhibit the growth of adult axons in vivo and suggest that the inhibitory effects of aggrecan may be mediated, at least in part, by structures located on the core glycoprotein in the absence of the bulk of the CS chains.

Aggrecan is a large proteoglycan (PG) that has been grouped with different PG families on the basis of its physical characteristics. These families include the chondroitin sulfate PGs, which appear to inhibit the migration of cells and axons during development. Although aggrecan has been studied primarily in cartilage, in the present study, tissue samples from developing, mature, and injured-adult rat spinal cords were used to determine whether aggrecan is present in the mammalian spinal cord. By the use of Western blot analysis, tissues were probed with aggrecan-specific antibodies (ATEGQV, TYKHRL, and LEC-7) and aggrecan-specific neoepitope antibodies (NITEGE, FVDIPEN, and TFKEEE) to identify full-length aggrecan and several fragments. Unlike many other aggrecan gene family members, aggrecan species were similar in embryonic day 14, postnatal day 1, and adult spinal cords. Spinal cord injury caused significant decreases in aggrecan. Partial recovery in some aggrecan species was evident by 2 weeks after injury. The presence of specific aggrecan neoepitopes suggested that aggrecan is cleaved in the spinal cord by both a disintegrin and metalloproteinase thrombospondin (also known as aggrecanase) and metalloproteinase-like activities. Many aggrecan species found in the spinal cord were similar to species in cartilage. Additional antibodies were used to identify two other aggrecan gene family members, neurocan and brevican, in the adult spinal cord. These studies present novel information on the aggrecan core protein species and enzymes involved in aggrecan cleavage in vivo in the rat spinal cord throughout development and after injury. They also provide the basis for investigating the function of aggrecan in the spinal cord.

Extrinsic factors appear to contribute to the lack of regeneration in the injured adult spinal cord. It is likely that these extrinsic factors include a group of putative growth inhibitory molecules known as chondroitin sulfate proteoglycans (CSPGs). The aims of this study were to determine: (1) the consequences of spinal cord contusion injury on CSPG expression, (2) if CSPGs can be degraded in vivo by exogenous enzyme application, and (3) the effects of intraspinal transplantation on the expression of CSPGs. Chondroitin 6-sulfate proteoglycan immunoreactivity (CSPG-IR) dramatically increased following spinal cord contusion injury both at and adjacent to the injury site compared to normal controls (no surgical procedure) and laminectomy-only controls by 4 days postinjury. The dramatic increase in CSPG-IR persisted around the lesion and in the dorsal one-half to two-thirds of the spinal cord for at least 40 days postinjury. Glial fibrillary acidic protein (GFAP)-IR patterns were similarly intensified and spatially restricted as CSPG-IR patterns. These results suggest that: (1) CSPGs may contribute to the lack of regeneration following spinal cord injury and (2) astrocytes may contribute to the production of CSPGs. In addition, our results show that CSPGs could be cleaved in vivo with exogenous chondroitinase ABC application. This demonstration of cleavage may the basis for a model to directly assess CSPGs' role in growth inhibition in vivo (studies in progress) and hold potential as a therapeutic approach to enhance growth. Interestingly, the robust, injury-induced CSPG-IR patterns were not altered by intraspinal grafts of fetal spinal cord. The CSPG expression profile in the host spinal cord was similar to time-matched contusion-only animals. This was also true of GFAP-IR patterns. Furthermore, the fetal spinal cord tissue, which was generally CSPG negative at the time of transplantation, developed robust CSPG expression by 30 days posttransplantation. This increase in CSPG expression in the graft was paired with a moderate increase in GFAP-IR. CSPG-IR patterns suggest that these molecules may contribute to the limited regeneration seen following intraspinal transplantation. In addition, it suggests that the growth permissiveness of the graft may change overtime as CSPG expression develops within the graft. These correlations in the injured and transplanted spinal cord support CSPGs' putative growth inhibitory effect in the adult spinal cord.

We have studied the locomotor development of kittens that received complete low thoracic spinal cord transections and embryonic spinal cord transplants as newborns. Embryonic spinal cord (E21-E26) transplanted into the site of a transection integrated well with the host spinal cord and promoted the development of overground locomotion. Spinalized kittens with transplants were first distinguished from spinalized kittens during the 2nd and 3rd postnatal weeks when kittens with transplants positioned their hindlimbs underneath their bodies which promoted support of the hindquarters. By postnatal Week 6, kittens with transplants exhibited overground locomotion characterized by full weight support and moderate balance control. By 20 weeks of age, as many as 96% of the step cycles showed full weight support and as few as 2% of the step cycles were interrupted by a fall. Most kittens also showed coordination between the forelimbs and the hindlimbs. They differed from normal in the precocious onset of reflex stepping and in the less precise interlimb coordination and more precarious balance during overground locomotion. The overground locomotor performance of kittens with transplants greatly exceeded that of spinal kittens without transplants since few spinalized kittens showed any full-weight-supported step cycles and none showed coordination between the forelimbs and the hindlimbs. In the absence of a transplant, no fibers could grow across the lesion site. In the presence of a transplant, fibers grew across the lesion site and established anatomical connectivity with the host. Host segmental systems identified by the presence of calcitonin gene-related peptide- and substance P-immunoreactive fibers were found throughout the transplants. Descending host systems of supraspinal origin were identified by serotonin- and dopamine beta-hydroxylase-immunoreactive fibers throughout the transplants. The growth of supraspinal axons into the transplant, and in one case into the caudal host spinal cord, provided a possible anatomical basis for the development of coordinated overground locomotion.

The development of bipedal treadmill locomotion and overground locomotion has previously been studied in the kitten; the development of quadrupedal treadmill locomotion has not. We evaluated and compared all three forms of locomotion in the normal kitten and present quantitative data comparing the development of quadrupedal treadmill and overground locomotion. Overground locomotion was studied from the day of birth to 5 months of age and quadrupedal treadmill locomotion was studied in the same animals from 9 weeks to 5 months of age. Treadmill locomotion was initiated postweaning, since it could not be reliably elicited without a food reward. Three locomotor characteristics (weight support, balance, and coordination between the forelimbs and the hindlimbs) were evaluated quantitatively. Kittens first consistently demonstrated overground steps with the ventral surface of their bodies supported above the walking surface throughout the entire step cycle during the second and third postnatal weeks. By 4 weeks of age, overground locomotion consistently showed full weight support and midline positioning of the hindquarters. Coordination between the forelimbs and the hindlimbs developed differently in the two forms of quadrupedal locomotion evaluated. During overground locomotion, the kittens initially used a single pattern in which only one limb was in swing at any time. As the kittens' weight support and trunk control improved, additional swing phase coordination patterns emerged and these patterns were correlated with the animals' ability to change speeds during locomotion. The consistency with which a dominant interlimb swing phase pattern was used at a particular speed increased with age and, by 6 weeks, the frequency of each speed-related dominant pattern approached 100% during overground locomotion. At 6 weeks, interlimb coordination also was evident in the nearly consistent interlimb phase interval present between the forelimb's initiation of the first extension subphase and the ipsilateral hindlimb's initiation of the flexion phase. The consistent patterns appeared to be fostered by maturation of weight support and balance. In contrast, the interlimb phase interval was inconsistent during quadrupedal treadmill locomotion until 20 weeks of age. Moreover, the interlimb swing phase patterns used during quadrupedal treadmill locomotion differed from those used during overground locomotion. The differences in the developmental time course and patterns of interlimb coordination between overground and quadrupedal treadmill locomotion suggest that different mechanisms regulate the control of interlimb coordination during these two different forms of quadrupedal locomotion.

This study was undertaken to determine the locomotor capability of kittens whose spinal cords were transected at birth. The postnatal development of reflex and goal-directed locomotion was examined during the first 5 postnatal months in kittens that received low thoracic spinal cord transections as newborns. Some spinal kittens developed aberrant quadrupedal forms of locomotion. The onset of quadrupedal locomotion, however, was delayed by 2-3 months compared to the normal kitten (42) and deteriorated by 5 months of age. Qualitative and quantitative analyses demonstrated that the quadrupedal locomotion was abnormal. Although some step cycles were characterized by full weight support, the typical hindlimb step cycle of the best performing cat showed inadequate weight support and balance. No spinal cat was able to coordinate the hindlimbs with the forelimbs during overground locomotion on a runaway or during quadrupedal locomotion on a treadmill. Neuroanatomical tracing with WGA-HRP and immunocytochemical techniques showed no axonal regeneration or growth into or across the lesion sites. The aberrant form of quadrupedal locomotion developed without descending input to the caudal spinal cord. The variability in performance among animals suggested that compensatory strategies were important factors in the spinal kitten's achievement of quadrupedal locomotion. Hindlimb weight-supported stepping during quadrupedal locomotion in some animals underscored the capacity of the isolated caudal spinal cord to generate both rhythmical stepping movements and weight support. The maintenance of developmentally immature, but functional, hindlimb postures suggested that the development of the isolated caudal spinal cord was arrested in the absence of descending input.

Solid or suspension grafts of fetal spinal cord (FSC), caudal brainstem (FBSt), neocortex (FNCx) or a combination of either FSC/FNCx or FSC/FBSt were placed into cavities produced by static loading (i.e., compression) of the spinal cord of adult cats two to 30 weeks after injury. Extensively vascularized, viable graft tissue was found in all animals with the exception of two cats which showed active rejection of their transplants. Surviving grafts showed many immature characteristics 6-9 weeks after transplantation. However, by 20-30 weeks, FSC and FBSt grafts were more mature. Grafts integrated with the host gray and white matter and neuritic processes from both host and graft were seen crossing the host-graft interface. Host calcitonin gene related peptide (CGRP)-like immunoreactive axons could be traced into FSC and FBSt grafts. A more restricted ingrowth of host serotonin (5-HT)-like immunoreactive fibers was seen in FSC grafts. Our results suggest that the capacity of homotypic transplants to promote recovery of function is greater than heterotypic transplants. Additionally, it appears that the functional capacity of the graft depends upon graft survival, the time interval between injury and transplantation, and whether or not the lesion cavity was debrided prior to grafting.

There has been much research studying the effect of homotypic embryonic neural tissue transplantation in the central nervous system of rats and cats, with promising results. The benefits of transplantation are dependent upon the appropriate aged donor. Therefore, it is critical that the exact age of the embryonic tissue be known. For this reason we have developed a method of determining, indirectly, the pregnancy status and embryonic age in the domestic cat (Felis catus) using serum progesterone levels. Serum progesterone levels were monitored in 16 domestic cats during 26 breeding trials. Blood samples were taken prior to and 6 days after natural breeding to determine if ovulation had occurred, indirectly indicating pregnancy status. By knowing the exact date bred and if it resulted in pregnancy, an accurate embryonic age was calculated. A day 6 post-breeding serum progesterone concentration of less than 5.0 ng/ml was considered a negative indication of pregnancy, greater than 5.0 ng/ml a positive indication of pregnancy. Pregnancy was confirmed by abdominal palpation 21 to 26 days after breeding. Serum progesterone levels taken on the sixth day after observed breeding provide an accurate, indirect evaluation of pregnancy in the timed pregnant domestic cat (81%, p = 0.003).