Leif A. Havton

Long-Distance Growth and Connectivity of Neural Stem Cells after Severe Spinal Cord Injury

Neural stem cells (NSCs) expressing GFP were embedded into fibrin matrices containing growth factor cocktails and grafted to sites of severe spinal cord injury. Grafted cells differentiated into multiple cellular phenotypes, including neurons, which extended large numbers of axons over remarkable distances. Extending axons formed abundant synapses with host cells. Axonal growth was partially dependent on mammalian target of rapamycin (mTOR), but not Nogo signaling. Grafted neurons supported formation of electrophysiological relays across sites of complete spinal transection, resulting in functional recovery. Two human stem cell lines (566RSC and HUES7) embedded in growth-factor-containing fibrin exhibited similar growth, and 566RSC cells supported functional recovery. Thus, properties intrinsic to early-stage neurons can overcome the inhibitory milieu of the injured adult spinal cord to mount remarkable axonal growth, resulting in formation of new relay circuits that significantly improve function. These therapeutic properties extend across stem cell sources and species.

Diminished Schwann Cell Repair Responses Underlie Age-Associated Impaired Axonal Regeneration

The regenerative capacity of the peripheral nervous system declines with age. Why this occurs, however, is unknown. We demonstrate that 24-month-old mice exhibit an impairment of functional recovery after nerve injury compared to 2-month-old animals. We find no difference in the intrinsic growth capacity between aged and young sensory neurons in vitro or in their ability to activate growth-associated transcriptional programs after injury. Instead, using age-mismatched nerve transplants in vivo, we show that the extent of functional recovery depends on the age of the nerve graft, and not the age of the host. Molecular interrogation of the sciatic nerve reveals that aged Schwann cells (SCs) fail to rapidly activate a transcriptional repair program after injury. Functionally, aged SCs exhibit impaired dedifferentiation, myelin clearance, and macrophage recruitment. These results suggest that the age-associated decline in axonal regeneration results from diminished Schwann cell plasticity, leading to slower myelin clearance.

Pronounced species divergence in corticospinal tract reorganization and functional recovery after lateralized spinal cord injury favors primates

Fundamental differences in the anatomy and function of the corticospinal tract support enhanced recovery of leg and hand function after lateralized spinal cord injury in primates compared to rodents, emphasizing the importance of primate models for spinal cord repair therapies. Species-specific recovery Despite decades of research and success in rodent models, there are no therapies that repair the human spinal cord. Friedli et al. looked at the reorganization and function of the corticospinal tract after spinal cord injury (SCI) in rats, monkeys, and humans. In humans with lateralized SCI (affecting only one side of the spinal cord), there was greater recovery in motor function than those with more symmetric injuries; this recovery was mirrored in monkeys with a similar SCI, but not in rats. The authors looked into why such a species divergence exists, and revealed that monkeys had a greater number of bilateral axonal projections that sprouted into denervated spinal segments below the injury, whereas rats had interrupted projections and near-complete depletion of corticospinal fibers. Thus, monkeys and humans have the potential for synaptic reorganization above and below the lesion, and this corticospinal tract reorganization correlates with functional recovery. The authors suggest that primate models should be considered more frequently for research aimed at SCI repair and therapeutics, but acknowledge the importance of rodent models in the field. Furthermore, because the degree of laterality correlates with a positive outcome, the authors suggest that it be factored into clinical trial design. Experimental and clinical studies suggest that primate species exhibit greater recovery after lateralized compared to symmetrical spinal cord injuries. Although this observation has major implications for designing clinical trials and translational therapies, advantages in recovery of nonhuman primates over other species have not been shown statistically to date, nor have the associated repair mechanisms been identified. We monitored recovery in more than 400 quadriplegic patients and found that functional gains increased with the laterality of spinal cord damage. Electrophysiological analyses suggested that corticospinal tract reorganization contributes to the greater recovery after lateralized compared with symmetrical injuries. To investigate underlying mechanisms, we modeled lateralized injuries in rats and monkeys using a lateral hemisection, and compared anatomical and functional outcomes with patients who suffered similar lesions. Standardized assessments revealed that monkeys and humans showed greater recovery of locomotion and hand function than did rats. Recovery correlated with the formation of corticospinal detour circuits below the injury, which were extensive in monkeys but nearly absent in rats. Our results uncover pronounced interspecies differences in the nature and extent of spinal cord repair mechanisms, likely resulting from fundamental differences in the anatomical and functional characteristics of the motor systems in primates versus rodents. Although rodents remain essential for advancing regenerative therapies, the unique response of the primate corticospinal tract after injury reemphasizes the importance of primate models for designing clinically relevant treatments.

Neural Stem Cell Dissemination after Grafting to CNS Injury Sites

Steward and colleagues report that implants of E14 rat spinal-cord-derived multipotent neural progenitor cells are associated with ectopic deposits of cells, occasionally at long distances from a T3 spinal cord lesion and grafting site. Of 20 grafted rats, half showed ectopic cell deposits. One rat had a deposit of cells in the 4th ventricle, from which relatively few axons extended into the tegmentum. Half of animals had cells within six spinal segments of the lesion site, and of these, three had deposits more distantly.

Leveraging biomedical informatics for assessing plasticity and repair in primate spinal cord injury

Recent preclinical advances highlight the therapeutic potential of treatments aimed at boosting regeneration and plasticity of spinal circuitry damaged by spinal cord injury (SCI). With several promising candidates being considered for translation into clinical trials, the SCI community has called for a non-human primate model as a crucial validation step to test efficacy and validity of these therapies prior to human testing. The present paper reviews the previous and ongoing efforts of the California Spinal Cord Consortium (CSCC), a multidisciplinary team of experts from 5 University of California medical and research centers, to develop this crucial translational SCI model. We focus on the growing volumes of high resolution data collected by the CSCC, and our efforts to develop a biomedical informatics framework aimed at leveraging multidimensional data to monitor plasticity and repair targeting recovery of hand and arm function. Although the main focus of many researchers is the restoration of voluntary motor control, we also describe our ongoing efforts to add assessments of sensory function, including pain, vital signs during surgery, and recovery of bladder and bowel function. By pooling our multidimensional data resources and building a unified database infrastructure for this clinically relevant translational model of SCI, we are now in a unique position to test promising therapeutic strategies efficacy on the entire syndrome of SCI. We review analyses highlighting the intersection between motor, sensory, autonomic and pathological contributions to the overall restoration of function. This article is part of a Special Issue entitled SI: Spinal cord injury.

Spinal stimulation of the upper lumbar spinal cord modulates urethral sphincter activity in rats after spinal cord injury

After spinal cord injury (SCI), the neurogenic bladder is observed to develop asynchronous bladder and external urethral sphincter (EUS) contractions in a condition known as detrusor-sphincter dyssnergia (DSD). Activation of the EUS spinal controlling center located at the upper lumbar spinal cord may contribute to reduce EUS dyssynergic contractions and decrease urethral resistance during voiding. However, this mechanism has not been well studied. This study aimed at evaluating the effects of epidural stimulation (EpS) over the spinal EUS controlling center (L3) in combination with a serotonergic receptor agonist on EUS relaxation in naive rats and chronic (6-8 wk) T8 SCI rats. Cystometrogram and EUS electromyography (EMG) were obtained before and after the intravenous administration of 5HT-1A receptor agonist and antagonist. The latency, duration, frequency, amplitude, and area under curve of EpS-evoked EUS EMG responses were analyzed. EpS on L3 evoked an inhibition of EUS tonic contraction and an excitation of EUS intermittent bursting/relaxation correlating with urine expulsion in intact rats. Combined with a 5HT-1A receptor agonist, EpS on L3 evoked a similar effect in chronic T8 SCI rats to reduce urethral contraction (resistance). This study examined the effect of facilitating the EUS spinal controlling center to switch between urine storage and voiding phases by using EpS and a serotonergic receptor agonist. This novel approach of applying EpS on the EUS controlling center modulates EUS contraction and relaxation as well as reduces urethral resistance during voiding in chronic SCI rats with DSD.

PI(3,5)P2 biosynthesis regulates oligodendrocyte differentiation by intrinsic and extrinsic mechanisms

Proper development of the CNS axon-glia unit requires bi-directional communication between axons and oligodendrocytes (OLs). We show that the signaling lipid phosphatidylinositol-3,5-bisphosphate [PI(3,5)P2] is required in neurons and in OLs for normal CNS myelination. In mice, mutations of Fig4, Pikfyve or Vac14, encoding key components of the PI(3,5)P2 biosynthetic complex, each lead to impaired OL maturation, severe CNS hypomyelination and delayed propagation of compound action potentials. Primary OLs deficient in Fig4 accumulate large LAMP1+ and Rab7+ vesicular structures and exhibit reduced membrane sheet expansion. PI(3,5)P2 deficiency leads to accumulation of myelin-associated glycoprotein (MAG) in LAMP1+perinuclear vesicles that fail to migrate to the nascent myelin sheet. Live-cell imaging of OLs after genetic or pharmacological inhibition of PI(3,5)P2 synthesis revealed impaired trafficking of plasma membrane-derived MAG through the endolysosomal system in primary cells and brain tissue. Collectively, our studies identify PI(3,5)P2 as a key regulator of myelin membrane trafficking and myelinogenesis. DOI: http://dx.doi.org/10.7554/eLife.13023.001

Modulation of the visceromotor reflex by a lumbosacral ventral root avulsion injury and repair in rats.

Increased abdominal muscle wall activity may be part of a visceromotor reflex (VMR) response to noxious stimulation of the bladder. However, information is sparse regarding the effects of cauda equina injuries on the VMR in experimental models. We studied the effects of a unilateral L6-S1 ventral root avulsion (VRA) injury and acute ventral root reimplantation (VRI) into the spinal cord on micturition reflexes and electromyographic activity of the abdominal wall in rats. Cystometrogram (CMG) and electromyography (EMG) of the abdominal external oblique muscle (EOM) were performed. All rats demonstrated EMG activity of the EOM associated with reflex bladder contractions. At 1 wk after VRA and VRI, the duration of the EOM EMG activity associated with reflex voiding was significantly prolonged compared with age-matched sham rats. However, at 3 wk postoperatively, the duration of the EOM responses remained increased in the VRA series but had normalized in the VRI group. The EOM EMG duration was normalized for both VRA and VRI groups at 8-12 wk postoperatively. CMG recordings show increased contraction duration at 1 and 3 wk postoperatively for the VRA series, whereas the contraction duration was only increased at 1 wk postoperatively for the VRI series. Our studies suggest that a unilateral lumbosacral VRA injury results in a prolonged VMR to bladder filling using a physiological saline solution. An acute root replantation decreased the VMR induced by VRA injury and provides earlier sensory recovery.

A Lumbosacral Ventral Root Avulsion Injury and Repair Model for Studies of Neuropathic Pain in Rats

Neuropathic pain may develop after a variety of injuries to peripheral nerves and roots. Most injury models have included a direct injury to primary afferent fibers or neurons. Recently, it has been demonstrated that injury to motor fibers in ventral roots may also result in neuropathic pain. A lumbosacral ventral root avulsion injury results in acute and persistent mechanical allodynia, but not thermal hyperesthesia. Interestingly, an acute replantation of the avulsed ventral roots into the spinal cord results in amelioration of the neuropathic pain. A detailed description of this injury and repair model is provided.

Radiographic and Magnetic Resonance Imaging Identification of Thoracolumbar Spine Variants with Implications for the Positioning of the Conus Medullaris in Rhesus Macaques.

The anatomy of the vertebral column in mammals may differ between species and between subjects of the same species, especially with regards to the composition of the thoracolumbar spine. We investigated, using several noninvasive imaging techniques, the thoracolumbar spine of a total of 44 adult rhesus macaques of both genders. Radiographic examination of the vertebral column showed a predominant spine phenotype with 12 rib‐bearing thoracic vertebrae and 7 lumbar vertebrae without ribs in 82% of subjects, whereas a subset of subjects demonstrated 13 rib‐bearing thoracic vertebrae and 6 lumbar vertebrae without ribs. Computer tomography studies of the thoraco‐lumbar spine in two cases with a pair of supernumerary ribs showed facet joints between the most caudal pair of ribs and the associated vertebra, supporting a thoracic phenotype. Magnetic resonance imaging (MRI) studies were used to determine the relationship between the lumbosacral spinal cord and the vertebral column. The length of the conus medullaris portion of the spinal cord was 1.5u2009±u20090.3 vertebral units, and its rostral and caudal positions in the spinal canal were at 2.0u2009±u20090.3 and 3.6u2009±u20090.4 vertebral units below the thoracolumbar junction, respectively (nu2009=u200944). The presence of a set of supernumerary ribs did not affect the length or craniocaudal position of the conus medullaris, and subjects with13 rib‐bearing vertebrae may from a functional or spine surgical perspective be considered as exhibiting12 thoracic vertebrae and an L1 vertebra with ribs. Anat Rec, 300:300–308, 2017.