Functional Local Proprioceptive Feedback Circuits Initiate and Maintain Locomotor Recovery after Spinal Cord Injury

Summary: Somatosensory feedback from proprioceptive afferents (PAs) is essential for locomotor recovery after spinal cord injury. To determine where or when proprioception is required for locomotor recovery after injury, we established an intersectional genetic model for PA ablation with spatial and temporal confinement. We found that complete or spatially restricted PA ablation in intact mice differentially affects locomotor performance. Following incomplete spinal cord injury, PA ablation below but not above the lesion severely restricts locomotor recovery and descending circuit reorganization. Furthermore, ablation of PAs after behavioral recovery permanently reverts functional improvements, demonstrating their essential role for maintaining regained locomotor function despite the presence of reorganized descending circuits. In parallel to recovery, PAs undergo reorganization of activity-dependent synaptic connectivity to specific local spinal targets. Our study reveals that PAs interacting with local spinal circuits serve as a continued driving force to initiate and maintain locomotor output after injury. : Takeoka and Arber examined the spatial and temporal requirements of proprioceptive feedback in recovery after spinal cord injury. They reveal an indispensable role for proprioceptive afferents below the injury in the initiation and maintenance of locomotor recovery. The activity of proprioceptive afferents contributes to local and descending circuit rearrangements that parallel recovery. Keywords: spinal cord injury, proprioception, mouse genetics, neuronal circuit reorganization, somatosensory feedback, viral tracing, locomotio

Long-Distance Descending Spinal Neurons Ensure Quadrupedal Locomotor Stability

Locomotion is an essential animal behavior used for translocation. The spinal cord acts as key executing center, but how it coordinates many body parts located across distance remains poorly understood. Here we employed mouse genetic and viral approaches to reveal organizational principles of long-projecting spinal circuits and their role in quadrupedal locomotion. Using neurotransmitter identity, developmental origin, and projection patterns as criteria, we uncover that spinal segments controlling forelimbs and hindlimbs are bidirectionally connected by symmetrically organized direct synaptic pathways that encompass multiple genetically tractable neuronal subpopulations. We demonstrate that selective ablation of descending spinal neurons linking cervical to lumbar segments impairs coherent locomotion, by reducing postural stability and speed during exploratory locomotion, as well as perturbing interlimb coordination during reinforced high-speed stepping. Together, our results implicate a highly organized long-distance projection system of spinal origin in the control of postural body stabilization and reliability during quadrupedal locomotion

Multisensory Signaling Shapes Vestibulo-Motor Circuit Specificity

The ability to continuously adjust posture and balance is necessary for reliable motor behavior. Vestibular and proprioceptive systems influence postural adjustments during movement by signaling functionally complementary sensory information. Using viral tracing and mouse genetics, we reveal two patterns of synaptic specificity between brainstem vestibular neurons and spinal motor neurons, established through distinct mechanisms. First, vestibular input targets preferentially extensor over flexor motor pools, a pattern established by developmental refinement in part controlled by vestibular signaling. Second, vestibular input targets slow-twitch over fast motor neuron subtypes within extensor pools, while proprioceptors exhibit inversely correlated connectivity profiles. Genetic manipulations affecting the functionality of proprioceptive feedback circuits lead to adjustments in vestibular input to motor neuron subtypes counterbalancing the imposed changes, without changing the sparse vestibular input to flexor pools. Thus, two sensory signaling systems interact to establish complementary synaptic input patterns to the final site of motor output processing

Muscle spindle feedback directs locomotor recovery and circuit reorganization after spinal cord injury

Spinal cord injuries alter motor function by disconnecting neural circuits above and below the lesion, rendering sensory inputs a primary source of direct external drive to neuronal networks caudal to the injury. Here, we studied mice lacking functional muscle spindle feedback to determine the role of this sensory channel in gait control and locomotor recovery after spinal cord injury. High-resolution kinematic analysis of intact mutant mice revealed proficient execution in basic locomotor tasks but poor performance in a precision task. After injury, wild-type mice spontaneously recovered basic locomotor function, whereas mice with deficient muscle spindle feedback failed to regain control over the hindlimb on the lesioned side. Virus-mediated tracing demonstrated that mutant mice exhibit defective rearrangements of descending circuits projecting to deprived spinal segments during recovery. Our findings reveal an essential role for muscle spindle feedback in directing basic locomotor recovery and facilitating circuit reorganization after spinal cord injury

Development of the observation of membrane fusion with label-free liposomes by calcium imaging

Liposomes are artificial vesicles composed of lipid bilayers that have enabled drugs to be encapsulated and delivered to tumor tissue. Membrane-fusogenic liposomes fuse with the plasma membranes of cells to deliver encapsulated drugs directly to the cytosol, which makes it a promising method for rapid and highly efficient drug delivery. In a previous study, liposomal lipid bilayers were labeled with fluorescent probes, and colocalization of labeled lipids with plasma membrane was observed under a microscope. However, there was concern that fluorescent labeling would affect lipid dynamics and cause liposomes to acquire membrane fusogenic ability. In addition, encapsulation of hydrophilic fluorescent substances in the inner aqueous phase sometimes requires an additional step of removing unencapsulated substances after preparation, and there is a risk of leakage. Herein, we propose a new method to observe cell interaction with liposomes without labeling. Our laboratory has developed two types of liposomes with different cellular internalization pathways, i.e., endocytosis and membrane fusion. We found that cytosolic calcium influx would be triggered following the internalization of cationic liposomes, and different cell entry routes led to different calcium responses. Thus, the correlation between cell entry routes and calcium responses could be utilized to study liposome-cell interactions without fluorescent labeling lipids. Briefly, liposomes were added to phorbol 12-myristate 13-acetate (PMA)-primed THP-1 cells, and calcium influx was measured by time-lapse imaging using a fluorescent indicator (Fura 2-AM). Liposomes with high membrane fusogenic ability elicited a strong transient calcium response immediately after adding liposomes, whereas those taken up mainly by endocytosis elicited multiple weak calcium responses. In order to verify the cell entry routes, we also tracked the intracellular distribution of fluorescent-labeled liposomes in PMA-primed THP-1 cells using a confocal laser scanning microscope. It was shown that for fusogenic liposomes, colocalization with plasma membrane occurred at the same time as calcium elevation, whereas for liposomes with a high endocytosis potential, fluorescent dots were observed in the cytoplasm, suggesting the cell internalization by endocytosis. These results suggested that the calcium response patterns correspond to cell entry routes, and membrane fusion can be observed by calcium imaging

Further evidence of olfactory ensheathing glia facilitating axonal regeneration after a complete spinal cord transection

11 pages, 11 figures, 1 table, PMID:21272578[PubMed] PMCID: PMC3085566Spinal Wistar Hannover rats injected with olfactory ensheathing glia (OEG) have been shown to recover some bipedal stepping and climbing abilities. Given the intrinsic ability of the spinal cord to regain stepping with pharmacological agents or epidural stimulation after a complete mid-thoracic transection, we asked if functional recovery after OEG injections is due to changes in the caudal stump or facilitation of functional regeneration of axons across the transection site. OEG were injected rostral and caudal to the transection site immediately after transection. Robotically assisted step training in the presence of intrathecal injections of a 5-HT(2A) receptor agonist (quipazine) was used to facilitate recovery of stepping. Bipedal stepping as well as climbing abilities were tested over a 6-month period post-transection to determine any improvement in hindlimb functional due to OEG injections and/or step training. The ability for OEG to facilitate regeneration was analyzed electrophysiologically by transcranially stimulating the brainstem and recording motor evoked potentials (MEP) with chronically implanted intramuscular EMG electrodes in the soleus and tibalis anterior with and without intrathecal injections of noradrenergic, serotonergic, and glycinergic receptor antagonists. Analyses confirmed that along with improved stepping ability and increased use of the hindlimbs during climbing, only OEG rats showed recovery of MEP. In addition the MEP signals were eliminated after a re-transection of the spinal cord rostral to the original transection and were modified in the presence of receptor antagonists. These data indicate that improved hindlimb function after a complete transection was coupled with OEG-facilitated functional regeneration of axons. This article is part of a Special Issue entitled: Understanding olfactory ensheathing glia and their prospect for nervous system repair.This work was supported by the National Institute of Neurological Disorders and Stroke Grant NS054159.Peer reviewe