Towards cell-type specific neuromodulation for spinal cord injury recovery

Abstract

Spinal cord injury (SCI) causes life-long neurological impairment, with loss of sensory and motor function distal to the point of injury. There are approximately 300,000 patients living with SCI in the United States, and currently no effective treatment, reducing their quality of life. Amongst other things, proprioception, which has been determined essential for normal locomotion, can be lost with SCI. Epidural Electric Stimulation (EES), that is thought to excite large diameter afferent fibers (LDAF), has been found to improve recovery from spinal cord injury in conjunction with movement rehabilitation in animal models and humans. This represents an exciting new approach to help these patients. However, many open questions remain about how and why EES works. Chief among them are 1) which of the afferent fibers are necessary and sufficient to promote better recovery, and 2) what are the mechanisms of plasticity in the spinal cord that underly improvement. Here, we sought to address the first question by using viral and genetic tools to begin to target specific subsets of LDAF. First, we use a viral vector that preferably transduces only in the large diameter afferent fibers (LDAF) in the Dorsal Root Ganglia (DRG), and then specifically only the proprioceptors within the LDAF, by using a transgenic rat line that expresses Cre recombinase in Parvalbumin, a marker for proprioceptive neurons in the DRG. This approach consists of using the chemogenetic modulator of neuronal activity Designer Receptors Exclusively Activated by Designer Drugs (DREADDs), which are activated by a putatively inert drug, clozapine-N-oxide (CNO), that crosses the blood brain barrier. While we were able to specifically target LDAF with excitatory DREADDs in L3-L5 DRGs in wild type rats, we were unsuccessful at specifically targeting proprioceptors by using the Pvalb-iCre rat line. Additionally, we studied the effect of exciting LDAF on rats with a 200KDyn SCI. CNO withdrawal on the week 7 stage of the recovery was associated with worse ladder performance than the previous and following weeks, as well as worse kinematic behavior of the same week on lower speeds in ankle movement. These results suggest that DREADDs activation is necessary for changes in movement at longer times post injury. It does not rule out that plasticity in neural circuitry has occurred but suggests that plasticity may rely on afferent activation. Finally, we sought to develop new methods to overcome skin motion artifact in rat kinematics by tattooing the knee area under the skin and recording infrared high-speed videos of moving rats which would correct joint calculations beyond just triangulation methods, as well as a novel MATLAB application that can accurately and reliably perform automated H-Reflex measurements, test the stimulating electrodes, and carry out typical instantaneous analyses, which in return allows for faster data collection with reduced human error, and subsequently result in higher research quality.