ImmunohistochemistryVibratome and cryosections were blocked in 5% BSA/1% Tween (SigmaAldrich) in PBS (pH 7.4) for 1 hour at room temperature. Mouse monoclonal anti-RC2 (IgM) antibody (Developmental Hybridoma Bank) , which are composed of formamide or formamide/polyethylene glycol, respectively, embryos, whole mounts and thick brain sections can be rapidly cleared with minimal volume changes. Unlike other available clearing techniques, these methods do not use detergents or solvents, and thus preserve lipophilic dyes, fluorescent tracers and immunohistochemical labeling, as well as fluorescent-protein labeling.

Summary Transcription factors control cell identity by regulating diverse developmental steps such as differentiation and axon guidance. The mammalian binocular visual circuit is comprised of projections of retinal ganglion cells (RGCs) to ipsilateral and contralateral targets in the brain. A transcriptional code for ipsilateral RGC identity has been identified, but less is known about the transcriptional regulation of contralateral RGC development. Here we demonstrate that SoxC genes (Sox4, 11, and 12) act on the progenitor-to-postmitotic transition to implement contralateral, but not ipsilateral, RGC differentiation, by binding to Hes5 and thus repressing Notch signaling. When SoxC genes are deleted in postmitotic RGCs, contralateral RGC axons grow poorly on chiasm cells in vitro, project ipsilaterally at the chiasm midline in vivo, and Plexin-A1 and Nr-CAM expression in RGCs is downregulated. These data implicate SoxC transcription factors in the regulation of contralateral RGC differentiation and axon guidance.

Brainstem olivocochlear neurons (OCNs) modulate the earliest stages of auditory processing through feedback projections to the cochlea and have been shown to influence hearing and protect the ear from sound-induced damage. Here, we used single-nucleus sequencing, anatomical reconstructions, and electrophysiology to characterize murine OCNs during postnatal development, in mature animals, and after sound exposure. We identified markers for known medial (MOC) and lateral (LOC) OCN subtypes, and show that they express distinct cohorts of physiologically relevant genes that change over development. In addition, we discovered a neuropeptide-enriched LOC subtype that produces Neuropeptide Y along with other neurotransmitters. Throughout the cochlea, both LOC subtypes extend arborizations over wide frequency domains. Moreover, LOC neuropeptide expression is strongly upregulated days after acoustic trauma, potentially providing a sustained protective signal to the cochlea. OCNs are therefore poised to have diffuse, dynamic effects on early auditory processing over timescales ranging from milliseconds to days.

Background In animal stroke models, peri-infarct cortical stimulation (CS) combined with rehabilitative reach training (RT) enhances motor functional outcome and cortical reorganization, compared with RT alone. It was unknown whether the effects of CS+RT: 1) persist long after treatment, 2) can be enhanced by forcing greater use of the paretic limb and 3) vary with treatment onset time. Objective To test the endurance, time-sensitivity, and the potential for augmentation by forced forelimb use of CS+RT treatment effects following ischemic stroke. Methods Adult rats that were proficient in skilled reaching received unilateral ischemic motor cortical lesions. RT was delivered for 3 weeks alone or concurrently with 100Hz cathodal epidural CS, delivered at 50% of movement thresholds. In study 1, this treatment was initiated at 14 days postinfarct, with some subgroups receiving an overlapping period of continuous constraint of the nonparetic forelimb to force use of the paretic limb. The function of the paretic limb was assessed weekly for 9–10 mo post-treatment. In study 2, rats underwent CS, RT and the combination during the chronic post-infarct period. Results Early onset CS+RT resulted in greater functional improvements than RT alone. The CS-related gains persisted for 9–10 mo post-treatment and were not significantly influenced by forced-use of the paretic limb. When treatment onset was delayed until 3 mo post-infarct, RT alone improved function, but CS+RT was no more effective than RT alone. Conclusion CS can enhance the persistence, as well as the magnitude of RT-driven functional improvements, but its effectiveness in doing so may vary with time post-infarct.

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