Project Summary
Mammalian locomotion is an essential behavior for mobility and survival. Locomotion involves
coordinated and alternating rhythmic activity between opposing limbs, as well as between antagonistic muscles
of the same limb. The locomotor central pattern generator (CPG), a network of spinal interneurons, is thought to
produce the locomotor pattern and rhythm and directly activate motor neurons, which in turn activate peripheral
muscles resulting in movement. We have recently reported that a set of spinal interneurons, known as ventral
spinocerebellar tract (VSCT) neurons, are both necessary and sufficient for locomotor behavior in neonatal and
adult mice. VSCT neurons possess two rhythmogenic properties, namely the exhibit an Ih current and they are
electrically coupled. Additionally, they possess spinal axon collaterals within the same spinal segment as well as
extra-segmentally. VSCTs are also contacted by motor neuron axon collaterals and they are electrically coupled
to them, at least during early postnatal development. VSCTs in turn, connect with other spinal interneurons
involved in the locomotor central pattern generator, via their axon collaterals. Our published observations provide
a strong foundation to dissect further their cellular characteristics and their neuronal circuits that they form with
other spinal neurons, including motor neurons. We hypothesize that both cellular characteristics in VSCTs and
the spinal neuronal targets contacted by VSCTs’ axon collaterals will be key determinants for locomotor behavior
in mice. In Aim 1, we will study the contribution of the HCN channels responsible for the Ih current in VSCTs and
determine the role of several connexins that we have identified through our preliminary results in their role as
key neurons in locomotor rhythmogenesis. To do so, we will utilize conditional knock out mice, whole-cell patch
clamp protocols along with pharmacological studies, immunohistochemistry, and virally-mediated channel
knockdown. In Aim 2, we will study the neuronal connectivity of VSCTs with Chx10+, Sim1+ and motor neurons.
We will investigate their functional role in these connections utilizing mouse genetics, behavioral assays,
physiological approaches and viral-mediated approaches to map out some of the key neuronal circuits involved
in locomotor behavior. Furthermore, we will employ a novel “clearing” technique together with 2-photon laser
confocal microscopy to image the entire spinal cord of neonatal and adult mice to uncover the relationship
between VSCTs and their potential neuronal targets within the spinal cord and across many different spinal
segments. In Aim 3, we will establish whether a novel set of transcription factors marking this class of spinal
interneurons can be manipulated by chemogenetic and optogenetic approaches and determine their involvement
in locomotor-like behavior. In summary, this comprehensive set of experiments will provide a solid foundation to
further our understanding of spinal locomotor networks.