An Atoh1 CRE Knock-In Mouse Labels Motor Neurons Involved in Fine Motor Control

Abstract

Motor neurons (MNs) innervating the digit muscles of the intrinsic hand (IH) and intrinsic foot (IF) control fine motor movements. The ability to reproducibly label specifically IH and IF MNs in mice would be a beneficial tool for studies focused on fine motor control. To this end, we find that a CRE knock-in mouse line of Atoh1 , a developmentally expressed basic helix-loop-helix (bHLH) transcription factor, reliably expresses CRE-dependent reporter genes in ∼60% of the IH and IF MNs. We determine that CRE-dependent expression in IH and IF MNs is ectopic because an Atoh1 mouse line driving FLPo recombinase does not label these MNs although other Atoh1 -lineage neurons in the intermediate spinal cord are reliably identified. Furthermore, the CRE-dependent reporter expression is enriched in the IH and IF MN pools with much sparser labeling of other limb-innervating MN pools such as the tibialis anterior (TA), gastrocnemius (GS), quadricep (Q), and adductor (Ad). Lastly, we find that ectopic reporter expression begins postnatally and labels a mixture of α and γ-MNs. Altogether, the Atoh1 CRE knock-in mouse strain might be a useful tool to explore the function and connectivity of MNs involved in fine motor control when combined with other genetic or viral strategies that can restrict labeling specifically to the IH and IF MNs. Accordingly, we provide an example of sparse labeling of IH and IF MNs using an intersectional genetic approach.

Structure of long-range direct and indirect spinocerebellar pathways as well as local spinal circuits mediating proprioception

“…First, the sensory neurons misexpress tdTomato and their axons can be seen terminating in the dorsal horn of the cleared spinal cord (Movie 3). Atoh1 is not known to be expressed in sensory neurons and we do not see sensory neurons labeled in Atoh1 P2A-FLPo mice, whose expression overlaps quite well with the Atoh1 Cre knockin mouse for spinal cord interneurons (data not shown and Ogujiofor et al, 2021). Therefore, the sensory neuron labeling is due to misexpression of the CRE recombinase in sensory neurons.…”

“…The following mouse strains were used: Gdnf IRES2-CreERT2 (Cebrian et al, 2014)(abbreviated Gdnf CreER , JAX #024948), Neurog1BAC-Cre (Quinones et al, 2010)(JAX #012859), Atoh1 P2A-FLPo (Ogujiofor et al, 2021), Atoh1 Cre knock-in (Yang et al, 2010), R26 LSL-LacZ (Soriano, 1999)(JAX #003474), R26 LSL-tdTom (Ai14)(JAX #007914) (Madisen et al, 2010), R26 LSL-FSF-tdTom (Ai65)(JAX #032864) (Madisen et al, 2015), Cdx2::FLPo (Bourane et al, 2015), R26 LSL-FSF-TeTx (Kim et al, 2009). All mice were outbred and thus, are mixed strains (including C57Bl/6J, C57Bl/6N, and ICR).…”

Structure of Long-Range Direct and Indirect Spinocerebellar Pathways as Well as Local Spinal Circuits Mediating Proprioception

“…First, the sensory neurons misexpress tdTomato, and their axons can be seen terminating in the dorsal horn of the cleared spinal cord (Movie 6). Atoh1 is not known to be expressed in sensory neurons and we do not see sensory neurons labeled in Atoh1 P2A-FLPo mice, whose expression overlaps quite well with the Atoh1 Cre knockin mouse for spinal cord interneurons (data not shown; Ogujiofor et al, 2021). Therefore, the sensory neuron labeling is because of misexpression of the CRE recombinase in sensory neurons.…”

“…We generated the Atoh1 P2A-FLPo knock-in mouse using the Easi-CRISPR method (Quadros et al, 2017;Miura et al, 2018). The model design and its initial characterization are described in our recent work (Ogujiofor et al, 2021). All mice were outbred and are thus mixed strains (including C57Bl/6J, C57Bl/6N, and ICR).…”

Knock-in mouse models for studying somatostatin and cholecystokinin expressing cells

“…Knock-in mice were generated using the Easi-CRISPR method 35,36 . Knock-in models containing new sequences of up to 2kb can be created using Easi-CRISPR with high e ciency [57][58][59][60] . The Easi-CRISPR approach utilizes three components: guide RNA, Cas9 protein to create a dsDNA break in the genome, and a long ssDNA donor to serve as a template to repair the dsDNA break.…”