Glutamatergic neurons of the gigantocellular reticular nucleus shape locomotor pattern and rhythm in the freely behaving mouse.

Because of their intermediate position between supraspinal locomotor centers and spinal circuits, gigantocellular reticular nucleus (GRN) neurons play a key role in motor command. However, the functional contribution of glutamatergic GRN neurons in initiating, maintaining, and stopping locomotion is still unclear. Combining electromyographic recordings with optogenetic manipulations in freely behaving mice, we investigate the functional contribution of glutamatergic brainstem neurons of the GRN to motor and locomotor activity. Short-pulse photostimulation of one side of the glutamatergic GRN did not elicit locomotion but evoked distinct motor responses in flexor and extensor muscles at rest and during locomotion. Glutamatergic GRN outputs to the spinal cord appear to be gated according to the spinal locomotor network state. Increasing the duration of photostimulation increased motor and postural tone at rest and reset locomotor rhythm during ongoing locomotion. In contrast, photoinhibition impaired locomotor pattern and rhythm. We conclude that unilateral activation of glutamatergic GRN neurons triggered motor activity and modified ongoing locomotor pattern and rhythm.

Role of DSCAM in the Development of Neural Control of Movement and Locomotion.

Locomotion results in an alternance of flexor and extensor muscles between left and right limbs generated by motoneurons that are controlled by the spinal interneuronal circuit. This spinal locomotor circuit is modulated by sensory afferents, which relay proprioceptive and cutaneous inputs that inform the spatial position of limbs in space and potential contacts with our environment respectively, but also by supraspinal descending commands of the brain that allow us to navigate in complex environments, avoid obstacles, chase prey, or flee predators. Although signaling pathways are important in the establishment and maintenance of motor circuits, the role of DSCAM, a cell adherence molecule associated with Down syndrome, has only recently been investigated in the context of motor control and locomotion in the rodent. DSCAM is known to be involved in lamination and delamination, synaptic targeting, axonal guidance, dendritic and cell tiling, axonal fasciculation and branching, programmed cell death, and synaptogenesis, all of which can impact the establishment of motor circuits during development, but also their maintenance through adulthood. We discuss herein how DSCAM is important for proper motor coordination, especially for breathing and locomotion.

DSCAM Mutation Impairs Motor Cortex Network Dynamic and Voluntary Motor Functions.

While it is well known that netrin-1 and its receptors UNC5 and UNC40 family members are involved in the normal establishment of the motor cortex and its corticospinal tract, less is known about its other receptor Down syndrome cell adherence molecule (DSCAM). DSCAM is expressed in the developing motor cortex, regulates axonal outgrowth of cortical neurons, and its mutation impairs the dendritic arborization of cortical neurons, thus suggesting that it might be involved in the normal development and functioning of the motor cortex. In comparison to WT littermates, DSCAM2J mutant mice slipped and misplaced their paw while walking on the rungs of a horizontal ladder, and exhibited more difficulties in stepping over an obstacle while walking at slow speed. Anterograde tracing showed a normal pyramidal decussation and corticospinal projection, but a more dorsal distribution of their axonal terminals in the spinal gray matter. Intracortical microstimulations showed a reduced corticospinal and intracortical efficacy, whereas stimulations of the pyramidal tract revealed a normal spinal efficacy and excitability of corticospinal tract axons, thus arguing for a dysfunctional cortical development. Our study reveals impairment of the network dynamics within the motor cortex, reducing corticospinal drive and impairing voluntary locomotor functions upon DSCAM2J mutation.

Distinct dampening effects of progesterone on the activity of nucleus tractus solitarii neurons in rat pups.

NEW FINDINGS: What is the central question of the study? Progesterone is considered a respiratory stimulant drug, but its effect on medullary respiratory neurons are poorly documented. We investigated whether progesterone alters spontaneous activity of neurons in the nucleus of the solitary tract (NTS). What is the main finding and its importance? In NTS neurons, progesterone decreases the action potential firing frequency in response to current injections and the amplitude of excitatory postsynaptic currents. Based on the established neuroprotective effect of progesterone against excitotoxicity resulting from insults, this inhibitory effect is likely to reflect inhibition of ion fluxes. These results are important because they further our understanding of the mechanisms underlying the diversity of respiratory effects of progesterone. ABSTRACT: Progesterone is known to stimulate breathing, but its actions on the respiratory control system have received limited attention. We addressed this issue at the cellular level by testing the hypothesis that progesterone augments excitatory currents at the level of the nucleus tractus solitarii (NTS). Medullary slices from juvenile male rats (14-17 days of age) containing the commissural region of the NTS (NTScom) were incubated with progesterone (1 µm) or vehicle (0.004% DMSO) for 60 min. We performed whole-cell voltage-clamp recordings of spontaneous excitatory postsynaptic currents (EPSCs) in the NTScom and determined membrane properties by applying depolarizing current steps. In comparison to vehicle-treated cells, progesterone exposure attenuates the firing frequency response to current injection and reduces the EPSC amplitude without modifying the EPSC frequency or the basal membrane properties. These data do not support our hypothesis, because they indicate that incubation with progesterone attenuates intrinsic action potential generation and inhibits excitatory synaptic inputs in the NTS. Given that these results are more in line with the protective effect of progesterone against excitotoxicity resulting from various insults, we propose that progesterone acts via inhibition of ionic flux.

Age- and speed-dependent modulation of gaits in DSCAM2J mutant mice.

Gaits depend on the interplay between distributed spinal neural networks, termed central pattern generators, generating rhythmic and coordinated movements, primary afferents, and descending supraspinal inputs. Recent studies demonstrated that the mouse displays a rich repertoire of gaits. Changes in gaits occur in mutant mice lacking particular neurons or molecular signaling pathways implicated in the normal establishment of these neural networks. Given the role of the Down syndrome cell adherence molecule (DSCAM) to the formation and maintenance of spinal interneuronal circuits and sensorimotor integration, we have investigated its functional contribution to gaits over a wide range of locomotor speeds using freely walking mice. We show in this study that the DSCAM2J mutation, while not precluding any gait, impairs the age- and speed-dependent modulation of gaits. It impairs the ability of mice to maintain their locomotion at high treadmill speeds. DSCAM2J mutation induces the dominance of lateral walk over trot and the emergence of aberrant gaits for mice, such as pace and diagonal walk. Gaits were also more labile in DSCAM2J mutant mice, i.e., less stable, less attractive, and less predictable than in their wild-type littermates. Our results suggest that the DSCAM mutation affects the behavioral repertoire of gaits in an age- and speed-dependent manner. NEW & NOTEWORTHY Gaits evolve throughout development, up to adulthood, and according to the genetic background. Using mutant mice lacking DSCAM (a cell adherence molecule associated with Down syndrome), we show that the DSCAM2J mutation alters the repertoire of gaits according to the mouse's age and speed, and prevents fast gaits. Such an incapacity suggests a reorganization of spinal, propriospinal, and supraspinal neuronal circuits underlying locomotor control in DSCAM2J mutant mice.

Betacellulin regulates schwann cell proliferation and myelin formation in the injured mouse peripheral nerve.

When a nerve fiber is cut or crushed, the axon segment that is separated from the soma degenerates distal from the injury in a process termed Wallerian degeneration (WD). C57BL/6OlaHsd-WldS (WldS ) mutant mice exhibit significant delays in WD. This results in considerably delayed Schwann cell and macrophage responses and thus in impaired nerve regenerations. In our previous work, thousands of genes were screened by DNA microarrays and over 700 transcripts were found to be differentially expressed in the injured sciatic nerve of WldS compared with wild-type (WT) mice. One of these transcripts, betacellulin (Btc), was selected for further analysis since it has yet to be characterized in the nervous system, despite being known as a ligand of the ErbB receptor family. We show that Btc mRNA is strongly upregulated in immature and dedifferentiated Sox2+ Schwann cells located in the sciatic nerve distal stump of WT mice, but not WldS mutants. Transgenic mice ubiquitously overexpressing Btc (Tg-Btc) have increased numbers of Schmidt-Lantermann incisures compared with WT mice, as revealed by Coherent anti-Stokes Raman scattering (CARS). Tg-Btc mice also have faster nerve conduction velocity. Finally, we found that deficiency in Btc reduces the proliferation of myelinating Schwann cells after sciatic nerve injury, while Btc overexpression induces Schwann cell proliferation and improves recovery of locomotor function. Taken together, these results suggest a novel regulatory role of Btc in axon-Schwann cell interactions involved in myelin formation and nerve repair. GLIA 2017 GLIA 2017;65:657-669.

Schwann cells generated from neonatal skin-derived precursors or neonatal peripheral nerve improve functional recovery after acute transplantation into the partially injured cervical spinal cord of the rat.

The transplantation of Schwann cells (SCs) holds considerable promise as a therapy for spinal cord injury, but the optimal source of these cells and the best timing for intervention remains debatable. Previously, we demonstrated that delayed transplantation of SCs generated from neonatal mouse skin-derived precursors (SKP-SCs) promoted repair and functional recovery in rats with thoracic contusions. Here, we conducted two experiments using neonatal rat cells and an incomplete cervical injury model to examine the efficacy of acute SKP-SC transplantation versus media control (Experiment 1) and versus nerve-derived SC or dermal fibroblast (Fibro) transplantation (Experiment 2). Despite limited graft survival, by 10 weeks after injury, rats that received SCs from either source showed improved functional recovery compared with media- or fibroblast-treated animals. Compared with media treatment, SKP-SC-transplanted rats showed enhanced rubrospinal tract (RST) sparing/plasticity in the gray matter (GM) rostral to injury, particularly in the absence of immunosuppression. The functional benefits of SC transplantations over fibroblast treatment correlated with the enhanced preservation of host tissue, reduced RST atrophy, and/or increased RST sparing/plasticity in the GM. In summary, our results indicate that: (1) early transplantation of neonatal SCs generated from skin or nerve promotes repair and functional recovery after incomplete cervical crush injury; (2) either of these cell types is preferable to Fibros for these purposes; and (3) age-matched SCs from these two sources do not differ in terms of their reparative effects or functional efficacy after transplantation into the injured cervical spinal cord.

Role of DSCAM in the development of the spinal locomotor and sensorimotor circuits.

Locomotion is controlled by spinal circuits that generate rhythm and coordinate left-right and flexor-extensor motoneuronal activities. The outputs of motoneurons and spinal interneuronal circuits are shaped by sensory feedback, relaying peripheral signals that are critical to the locomotor and postural control. Several studies in invertebrates and vertebrates have argued that the Down syndrome cell adhesion molecule (DSCAM) would play an important role in the normal development of neural circuits through cell spacing and targeting, axonal and dendritic branching, and synapse establishment and maintenance. Although there is evidence that DSCAM is important for the normal development of neural circuits, little is known about its functional contribution to spinal motor circuits. We show here that adult DSCAM(2J) mutant mice, lacking DSCAM, exhibit a higher variability in their locomotor pattern and rhythm during treadmill locomotion. Retrograde tracing studies in neonatal isolated spinal cords show an increased number of spinal commissural interneurons, which likely contributes to reducing the left-right alternation and to increasing the flexor/swing duration during neonatal and adult locomotion. Moreover, our results argue that, by reducing the peripheral excitatory drive onto spinal motoneurons, the DSCAM mutation reduces or abolishes spinal reflexes in both neonatal isolated spinal cords and adult mice, thus likely impairing sensorimotor control. Collectively, our functional, electrophysiological, and anatomical studies suggest that the mammalian DSCAM protein is involved in the normal development of spinal locomotor and sensorimotor circuits.

Motor hypertonia and lack of locomotor coordination in mutant mice lacking DSCAM.

Down syndrome cell adherence molecule (DSCAM) contributes to the normal establishment and maintenance of neural circuits. Whereas there is abundant literature regarding the role of DSCAM in the neural patterning of the mammalian retina, less is known about motor circuits. Recently, DSCAM mutation has been shown to impair bilateral motor coordination during respiration, thus causing death at birth. DSCAM mutants that survive through adulthood display a lack of locomotor endurance and coordination in the rotarod test, thus suggesting that the DSCAM mutation impairs motor control. We investigated the motor and locomotor functions of DSCAM(2J) mutant mice through a combination of anatomical, kinematic, force, and electromyographic recordings. With respect to wild-type mice, DSCAM(2J) mice displayed a longer swing phase with a limb hyperflexion at the expense of a shorter stance phase during locomotion. Furthermore, electromyographic activity in the flexor and extensor muscles was increased and coactivated over 20% of the step cycle over a wide range of walking speeds. In contrast to wild-type mice, which used lateral walk and trot at walking speed, DSCAM(2J) mice used preferentially less coordinated gaits, such as out-of-phase walk and pace. The neuromuscular junction and the contractile properties of muscles, as well as their muscle spindles, were normal, and no signs of motor rigidity or spasticity were observed during passive limb movements. Our study demonstrates that the DSCAM mutation induces dystonic hypertonia and a disruption of locomotor gaits.

Lhx3-Chx10 reticulospinal neurons in locomotor circuits.

Motor behaviors result from the interplay between the brain and the spinal cord. Reticulospinal neurons, situated between the supraspinal structures that initiate motor movements and the spinal cord that executes them, play key integrative roles in these behaviors. However, the molecular identities of mammalian reticular formation neurons that mediate motor behaviors have not yet been determined, thus limiting their study in health and disease. In the medullary reticular formation of the mouse, we identified neurons that express the transcription factors Lhx3 and/or Chx10, and demonstrate that these neurons form a significant component of glutamatergic reticulospinal pathways. Lhx3-positive medullary reticular formation neurons express Fos following a locomotor task in the adult, indicating that they are active during walking. Furthermore, they receive functional inputs from the mesencephalic locomotor region and have electrophysiological properties to support tonic repetitive firing, both of which are necessary for neurons that mediate the descending command for locomotion. Together, these results suggest that Lhx3/Chx10 medullary reticular formation neurons are involved in locomotion.

Functional plasticity of glutamatergic neurons of medullary reticular nuclei after spinal cord injury in mice.

Spinal cord injury disrupts the descending command from the brain and causes a range of motor deficits. Here, we use optogenetic tools to investigate the functional plasticity of the glutamatergic reticulospinal drive of the medullary reticular formation after a lateral thoracic hemisection in female mice. Sites evoking stronger excitatory descending drive in intact conditions are the most impaired after injury, whereas those associated with a weaker drive are potentiated. After lesion, pro- and anti-locomotor activities (that is, initiation/acceleration versus stop/deceleration) are overall preserved. Activating the descending reticulospinal drive improves stepping ability on a flat surface of chronically impaired injured mice, and its priming enhances recovery of skilled locomotion on a horizontal ladder. This study highlights the resilience and capacity for reorganization of the glutamatergic reticulospinal command after injury, along with its suitability as a therapeutical target to promote functional recovery.

Towards a mini-endoscope design with spatially selective excitation and imaging.

We describe a mini-endoscope design that uses a new type of electrically tunable liquid crystal lens array enabling the dynamic increase of spatial resolution by adjusting the working distance in various zones of interest over a relatively large field of view (FoV) without mechanical movement. The characterization of the system is performed by using uniform fluorescent films, fluorescent micro spheres and a tissue sample expressing the fluorescent calcium indicator GCaMP6s. Lateral resolution of up to 2 µm over the FoV between 300 µm - 400 µm is experimentally demonstrated.

Functional contribution of mesencephalic locomotor region nuclei to locomotor recovery after spinal cord injury.

Spinal cord injury (SCI) results in a disruption of information between the brain and the spinal circuit. Electrical stimulation of the mesencephalic locomotor region (MLR) can promote locomotor recovery in acute and chronic SCI rodent models. Although clinical trials are currently under way, there is still debate about the organization of this supraspinal center and which anatomic correlate of the MLR should be targeted to promote recovery. Combining kinematics, electromyographic recordings, anatomic analysis, and mouse genetics, our study reveals that glutamatergic neurons of the cuneiform nucleus contribute to locomotor recovery by enhancing motor efficacy in hindlimb muscles, and by increasing locomotor rhythm and speed on a treadmill, over ground, and during swimming in chronic SCI mice. In contrast, glutamatergic neurons of the pedunculopontine nucleus slow down locomotion. Therefore, our study identifies the cuneiform nucleus and its glutamatergic neurons as a therapeutical target to improve locomotor recovery in patients living with SCI.

Heterozygous Dcc Mutant Mice Have a Subtle Locomotor Phenotype.

Axon guidance receptors such as deleted in colorectal cancer (DCC) contribute to the normal formation of neural circuits, and their mutations can be associated with neural defects. In humans, heterozygous mutations in DCC have been linked to congenital mirror movements, which are involuntary movements on one side of the body that mirror voluntary movements of the opposite side. In mice, obvious hopping phenotypes have been reported for bi-allelic Dcc mutations, while heterozygous mutants have not been closely examined. We hypothesized that a detailed characterization of Dcc heterozygous mice may reveal impaired corticospinal and spinal functions. Anterograde tracing of the Dcc+/- motor cortex revealed a normally projecting corticospinal tract, intracortical microstimulation (ICMS) evoked normal contralateral motor responses, and behavioral tests showed normal skilled forelimb coordination. Gait analyses also showed a normal locomotor pattern and rhythm in adult Dcc+/- mice during treadmill locomotion, except for a decreased occurrence of out-of-phase walk and an increased duty cycle of the stance phase at slow walking speed. Neonatal isolated Dcc+/- spinal cords had normal left-right and flexor-extensor coupling, along with normal locomotor pattern and rhythm, except for an increase in the flexor-related motoneuronal output. Although Dcc+/- mice do not exhibit any obvious bilateral impairments like those in humans, they exhibit subtle motor deficits during neonatal and adult locomotion.

Impact of ovariectomy and CO2 inhalation on microglia morphology in select brainstem and hypothalamic areas regulating breathing in female rats.

The neural network that regulates breathing shows a significant sexual dimorphism. Ovarian hormones contribute to this distinction as, in rats, ovariectomy reduces the ventilatory response to CO2. Microglia are neuroimmune cells that are sensitive to neuroendocrine changes in their environment. When reacting to challenging conditions, these cells show changes in their morphology that reflect an augmented capacity for producing pro- and anti-inflammatory cytokines. Based on evidence suggesting that microglia contribute to sex-based differences in reflexive responses to hypercapnia, we hypothesized that ovariectomy and hypercapnia promote microglial reactivity in selected brain areas that regulate breathing. We used ionized calcium-binding-adapter molecule-1 (Iba1) immunolabeling to compare the density and morphology of microglia in the locus coeruleus (LC), the caudal medullary raphe, the caudal part of the nucleus of the tractus solitarius (cNTS), and the paraventricular nucleus of the hypothalamus (PVN). Tissue was obtained from SHAM (metaestrus) female rats or following ovariectomy. Rats were exposed to normocapnia or hypercapnia (5% CO2, 20 min). Ovariectomy and hypercapnia did not affect microglial density in any of the structures studied. Ovariectomy promoted a reactive phenotype in the cNTS and LC, as indicated by a larger morphological index. In these structures, hypercapnia had a relatively modest opposing effect; the medullary raphe or the PVN were not affected. We conclude that ovarian hormones attenuate microglial reactivity in CO2/H+ sensing structures. These data suggest that microglia may contribute to neurological diseases in which anomalies of respiratory control are associated with cyclic fluctuations of ovarian hormones or menopause.

Combination of olfactory ensheathing cells with local versus systemic cAMP treatment after a cervical rubrospinal tract injury.

The failure of CNS axons to regenerate following traumatic injury is due in part to a growth-inhibitory environment in CNS as well as a weak intrinsic neuronal growth response. Olfactory ensheathing cell (OECs) transplants have been reported to create a favorable environment promoting axonal regeneration, remyelination, and functional recovery after spinal cord injury. However, in our previous experiments, OEC transplants failed to promote regeneration of rubrospinal axons through and beyond the site of a dorsolateral funiculus crush in rats. Rubrospinal neurons undergo massive cell atrophy and limited expression of regeneration-associated genes after axotomy. Using the same injury model, we tested the hypothesis that treatment of the red nucleus with cAMP, known to stimulate the intrinsic growth response in other neurons, will promote rubrospinal regeneration in combination with OEC transplants. In addition, we assessed a systemic increase of cAMP using the phosphodiesterase inhibitor rolipram. OECs prevented cavity formation, attenuated astrocytic hypertrophy and the retraction of the axotomized rubrospinal axons, and tended to reduce the overall lesion size. OEC transplantation lowered the thresholds for thermal sensitivity of both forepaws. None of our treatments, alone or in combination, promoted rubrospinal regeneration through the lesion site. However, the systemic elevation of cAMP with rolipram resulted in greater numbers of OECs and axonal density within the graft and improved motor performance in a cylinder test in conjunction with enhanced rubrospinal branching and attenuated astrocytic hypertrophy.

Functional characterization of cardiac progenitor cells and their derivatives in the embryonic heart post-chamber formation.

There is scant information on the fate of cardiac progenitor cells (CPC) in the embryonic heart after chamber specification. Here we simultaneously tracked three lineage-specific markers (Nkx2.5, MLC2v, and ANF) and confirmed that CPCs with an Nkx2.5+MLC2v-ANF- phenotype are present in the embryonic (E) day 11.5 mouse ventricular myocardium. We demonstrated that these CPCs could give rise to working cardiomyocytes and conduction system cells. Using a two-photon imaging analysis, we found that the majority of CPCs are not capable of developing Ca2+ transients in response to beta-adrenergic receptor stimulation. In contrast, Nkx2.5+ cells expressing MLC2v but not ANF are capable of developing functional Ca2+ transients. We showed that Ca2+ transients could be invoked in Nkx2.5+MLC2v+ANF+ cells only upon inhibition of Gi, muscarinic receptors, or nitric oxide synthase (NOS) signaling pathways. Our data suggest that these inhibitory pathways may delay functional specification in a subset of developing ventricular cells.

Disruption of estradiol regulation of orexin neurons: a novel mechanism in excessive ventilatory response to CO2 inhalation in a female rat model of panic disorder.

Panic disorder (PD) is ~2 times more frequent in women. An excessive ventilatory response to CO2 inhalation is more likely during the premenstrual phase. While ovarian hormones appear important in the pathophysiology of PD, their role remains poorly understood as female animals are rarely used in pre-clinical studies. Using neonatal maternal separation (NMS) to induce a "PD-like" respiratory phenotype, we tested the hypothesis that NMS disrupts hormonal regulation of the ventilatory response to CO2 in female rats. We then determined whether NMS attenuates the inhibitory actions of 17-ß estradiol (E2) on orexin neurons (ORX). Pups were exposed to NMS (3 h/day; postnatal day 3-12). The ventilatory response to CO2-inhalation was tested before puberty, across the estrus cycle, and following ovariectomy. Plasma E2 and hypothalamic ORXA were measured. The effect of an ORX1 antagonist (SB334867; 15 mg/kg) on the CO2 response was tested. Excitatory postsynaptic currents (EPSCs) were recorded from ORX neurons using whole-cell patch-clamp. NMS-related increase in the CO2 response was observed only when ovaries were functional; the largest ventilation was observed during proestrus. SB334867 blocked this effect. NMS augmented levels of ORXA in hypothalamus extracts. EPSC frequency varied according to basal plasma E2 levels across the estrus cycle in controls but not NMS. NMS reproduces developmental and cyclic changes of respiratory manifestations of PD. NMS disrupts the inhibitory actions of E2 on the respiratory network. Impaired E2-related inhibition of ORX neurons during proestrus is a novel mechanism in respiratory manifestations of PD in females.

The toxin MPTP generates similar cognitive and locomotor deficits in hTau and tau knock-out mice.

Parkinson's disease (PD) is characterized by motor deficits, although cognitive disturbances are frequent and have been noted early in the disease. The main pathological characteristics of PD are the loss of dopaminergic neurons and the presence of aggregated α-synuclein in Lewy bodies of surviving cells. Studies have also documented the presence of other proteins within Lewy bodies, particularly tau, a microtubule-associated protein implicated in a wide range of neurodegenerative diseases, including Alzheimer's disease (AD). In AD, tau pathology correlates with cognitive dysfunction, and tau mutations have been reported to lead to dementia associated with parkinsonism. However, the role of tau in PD pathogenesis remains unclear. To address this question, we induced parkinsonism by injecting the toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in hTau mice, a mouse model of tauopathy expressing human tau, and a mouse model knock-out for tau (TKO). We found that although MPTP impaired locomotion (gait analysis) and cognition (Barnes maze), there were no discernable differences between hTau and TKO mice. MPTP also induced a slight but significant increase in tau phosphorylation (Thr205) in the hippocampus of hTau mice, as well as a significant decrease in the soluble and insoluble tau fractions that correlated with the loss of dopaminergic neurons in the brainstem. Overall, our findings suggest that, although MPTP can induce an increase in tau phosphorylation at specific epitopes, tau does not seem to causally contribute to cognitive and locomotor deficits induced by this toxin.

Undesired effects of a combinatorial treatment for spinal cord injury--transplantation of olfactory ensheathing cells and BDNF infusion to the red nucleus.

Transplantations of olfactory ensheathing cells (OECs) have been reported to promote axonal regeneration and functional recovery after spinal cord injury, but have demonstrated limited growth promotion of rat rubrospinal axons after a cervical dorsolateral funiculus crush. Rubrospinal neurons undergo massive atrophy after cervical axotomy and show only transient expression of regeneration-associated genes. Cell body treatment with brain-derived neurotrophic factor (BDNF) prevents this atrophy, stimulates regeneration-associated gene expression and promotes regeneration of rubrospinal axons into peripheral nerve transplants. Here, we hypothesized that the failure of rubrospinal axons to regenerate through a bridge of OEC transplants was due to this weak intrinsic cell body response. Hence, we combined BDNF treatment of rubrospinal neurons with transplantation of highly enriched OECs derived from the nasal mucosa and assessed axonal regeneration as well as behavioral changes after a cervical dorsolateral funiculus crush. Each treatment alone as well as their combination prevented the dieback of the rubrospinal axons, but none of them promoted rubrospinal regeneration beyond the lesion/transplantation site. Motor performance in a food-pellet reaching test and forelimb usage during vertical exploration (cylinder test) were more impaired after combining transplantation of OECs with BDNF treatment. This impaired motor performance correlated with lowered sensory thresholds in animals receiving the combinatorial therapy - which were not seen with each treatment alone. Only this combinatorial treatment group showed enhanced sprouting of calcitonin gene-related peptide-positive axons rostral to the lesion site. Hence, some combinatorial treatments, such as OECs with BDNF, may have undesired effects in the injured spinal cord.