Title: Functional organization of motor neurons during fictive locomotor behavior revealed by large-scale optical imaging
Abstract: The isolated neonatal mouse spinal cord is capable of generating sustained rhythmic network activity, termed fictive locomotion (Kiehn and Kjaerulff 1996, Markin et al. 2012). However, the spatiotemporal pattern of motor neuron activity during fictive locomotion has not been measured at single-cell resolution, nor has the variation across a motor pool been quantified. We have measured the activity of thousands of retrogradely labeled motor neurons using large-scale, cellular resolution calcium imaging. Spike inference methods (Vogelstein et al. 2010) have been used to estimate peak firing phase. This approach was validated in each experiment using antidromic stimulation of ventral roots to generate data where spike timing information is known. Our imaging approach has revealed that neurons within the same motor pool fire synchronously. In contrast, neurons innervating muscles that have slightly different phase tunings during walking also showed slightly offset burst times during fictive locomotion. Neurons innervating antagonist muscles reliably fired 180° out of phase with one another. Finally, groups of motor neurons that fired asynchronously were found at each lumbar spinal segment, suggesting that the recruitment of motor neurons during fictive locomotion is determined by pool identity, rather than by segmental position. These spatiotemporal patterns were each highly reproducible between preparations. Our approach has revealed complexity and specificity in the patterns of motor neuron recruitment during locomotor-like network activity. We are currently analyzing the relationship between the activity of genetically defined pre-motor interneurons and the activity of identified motor neuron pools.