Agonist-Antagonist Coactivation Enhances Corticomotor Excitability of Ankle Muscles.

Spinal pathways underlying reciprocal flexion-extension contractions have been well characterized, but the extent to which cortically evoked motor-evoked potentials (MEPs) are influenced by antagonist muscle activation remains unclear. A majority of studies using transcranial magnetic stimulation- (TMS-) evoked MEPs to evaluate the excitability of the corticospinal pathway focus on upper extremity muscles. Due to functional and neural control differences between lower and upper limb muscles, there is a need to evaluate methodological factors influencing TMS-evoked MEPs specifically in lower limb musculature. If and to what extent the activation of the nontargeted muscles, such as antagonists, affects TMS-evoked MEPs is poorly understood, and such gaps in our knowledge may limit the rigor and reproducibility of TMS studies. Here, we evaluated the effect of the activation state of the antagonist muscle on TMS-evoked MEPs obtained from the target (agonist) ankle muscle for both tibialis anterior (TA) and soleus muscles. Fourteen able-bodied participants (11 females, age: 26.1 ± 4.1 years) completed one experimental session; data from 12 individuals were included in the analysis. TMS was delivered during 4 conditions: rest, TA activated, soleus activated, and TA and soleus coactivation. Three pairwise comparisons were made for MEP amplitude and coefficient of variability (CV): rest versus coactivation, rest versus antagonist activation, and agonist activation versus coactivation. We demonstrated that agonist-antagonist coactivation enhanced MEP amplitude and reduced MEP CVs for both TA and soleus muscles. Our results provide methodological considerations for future TMS studies and pave the way for future exploration of coactivation-dependent modulation of corticomotor excitability in pathological cohorts such as stroke or spinal cord injury.

SPINAL V1 INHIBITORY INTERNEURON CLADES DIFFER IN BIRTHDATE, PROJECTIONS TO MOTONEURONS AND HETEROGENEITY.

Spinal cord interneurons play a crucial role in shaping motor output, but their precise identity and circuit connectivity remain unclear. Focusing on the cardinal class of inhibitory V1 interneurons, we define the diversity of four major V1 subsets according to timing of neurogenesis, genetic lineage-tracing, synaptic output to motoneurons, and synaptic inputs from muscle afferents. Birthdating delineates two early-born (Renshaw and Pou6f2) and two late-born V1 clades (Foxp2 and Sp8) suggesting sequential neurogenesis gives rise to different V1 clades. Neurogenesis did not correlate with motoneuron targeting. Early-born Renshaw cells and late-born Foxp2-V1 interneurons both tightly coupled to motoneurons, while early-born Pou6f2-V1 and late-born Sp8-V1 interneurons did not. V1-clades also greatly differ in cell numbers and diversity. Lineage labeling of the Foxp2-V1 clade shows it contains over half of all V1 interneurons and provides the largest inhibitory input to motoneuron cell bodies. Foxp2-V1 subgroups differ in neurogenesis and proprioceptive input. Notably, one subgroup defined by Otp expression and located adjacent to the lateral motor column exhibits substantial input from proprioceptors, consistent with some Foxp2-V1 cells at this location forming part of reciprocal inhibitory pathways. This was confirmed with viral tracing methods for ankle flexors and extensors. The results validate the previous V1 clade classification as representing unique interneuron subtypes that differ in circuit placement with Foxp2-V1s forming the more complex subgroup. We discuss how V1 organizational diversity enables understanding of their roles in motor control, with implications for the ontogenetic and phylogenetic origins of their diversity. SIGNIFICANCE STATEMENT: Spinal interneuron diversity and circuit organization represents a key challenge to understand the neural control of movement in normal adults and also during motor development and in disease. Inhibitory interneurons are a core element of these spinal circuits, acting on motoneurons either directly or via premotor networks. V1 interneurons comprise the largest group of inhibitory interneurons in the ventral horn and their organization remains unclear. Here we present a comprehensive examination of V1 subtypes according to neurogenesis, placement in spinal motor circuits and motoneuron synaptic targeting. V1 diversity increases during evolution from axial-swimming fishes to limb-based mammalian terrestrial locomotion and this is reflected in the size and heterogeneity of the Foxp2-V1 clade which is closely associated to limb motor pools. We show Foxp2-V1 interneurons establish the densest and more direct inhibitory synaptic input to motoneurons, especially on cell bodies. This is of further importance because deficits on motoneuron cell body inhibitory V1 synapses and on Foxp2-V1 interneurons themselves have recently been shown to be affected at early stages of pathology in motor neurodegenerative diseases like amyotrophic lateral sclerosis.

Enabling techniques for in vitro studies on mammalian spinal locomotor mechanisms.

The neonatal rodent spinal cord maintained in vitro is a powerful model system to understand the central properties of spinal circuits generating mammalian locomotion. We describe three enabling approaches that incorporate afferent input and attached hindlimbs. (i) Sacral dorsal column stimulation recruits and strengthens ongoing locomotor-like activity, and implementation of a closed positive-feedback paradigm is shown to support its stimulation as an untapped therapeutic site for locomotor modulation. (ii) The spinal cord hindlimbs-restrained preparation allows suction electrode electromyographic recordings from many muscles. Inducible complex motor patterns resemble natural locomotion, and insights into circuit organization are demonstrated during spontaneous motor burst 'deletions', or following sensory stimuli such as tail and paw pinch. (iii) The spinal cord hindlimbs-pendant preparation produces unrestrained hindlimb stepping. It incorporates mechanical limb perturbations, kinematic analyses, ground reaction force monitoring, and the use of treadmills to study spinal circuit operation with movement-related patterns of sensory feedback while providing for stable whole-cell recordings from spinal neurons. Such techniques promise to provide important additional insights into locomotor circuit organization.