Circadian disruption as a result of shift work is associated with adverse metabolic consequences. Internal desynchrony between the phase of the suprachiasmatic nuclei (SCN) and peripheral clocks is widely believed to be a major factor contributing to these adverse consequences, but this hypothesis has never been tested directly. A GABAergic Cre driver combined with conditional casein kinase mutations ( ) was used to lengthen the endogenous circadian period in GABAergic neurons, including the SCN, but not in peripheral tissues, to create a Discordant mouse model. These mice had a long (27.4 h) behavioral period to which peripheral clocks entrained in vivo albeit with an advanced phase (∼6 h). Thus, in the absence of environmental timing cues, these mice had internal desynchrony between the SCN and peripheral clocks. Surprisingly, internal desynchrony did not result in obesity in this model. Instead, Discordant mice had reduced body mass compared with Cre-negative controls on regular chow and even when challenged with a high-fat diet. Similarly, internal desynchrony failed to induce glucose intolerance or disrupt body temperature and energy expenditure rhythms. Subsequently, a lighting cycle of 2-h light/23.5-h dark was used to create a similar internal desynchrony state in both genotypes. Under these conditions, Discordant mice maintained their lower body mass relative to controls, suggesting that internal desynchrony did not cause the lowered body mass. Overall, our results indicate that internal desynchrony does not necessarily lead to metabolic derangements and suggest that additional mechanisms contribute to the adverse metabolic consequences observed in circadian disruption protocols.

Mice with targeted gene disruption have provided important information about the molecular mechanisms of circadian clock function. A full understanding of the roles of circadian-relevant genes requires manipulation of their expression in a tissue-specific manner, ideally including manipulation with high efficiency within the suprachiasmatic nuclei (SCN). To date, conditional manipulation of genes within the SCN has been difficult. In a previously developed mouse line, Cre recombinase was inserted into the vesicular GABA transporter (Vgat) locus. Since virtually all SCN neurons are GABAergic, this Vgat-Cre line seemed likely to have high efficiency at disrupting conditional alleles in SCN. To test this premise, the efficacy of Vgat-Cre in excising conditional (fl, for flanked by LoxP) alleles in the SCN was examined. Vgat-Cre-mediated excision of conditional alleles of Clock or Bmal1 led to loss of immunostaining for products of the targeted genes in the SCN. Vgat-Cre; Clock; Npas2 mice and Vgat-Cre; Bmal1 mice became arrhythmic immediately upon exposure to constant darkness, as expected based on the phenotype of mice in which these genes are disrupted throughout the body. The phenotype of mice with other combinations of Vgat-Cre, conditional Clock, and mutant Npas2 alleles also resembled the corresponding whole-body knockout mice. These data indicate that the Vgat-Cre line is useful for Cre-mediated recombination within the SCN, making it useful for Cre-enabled technologies including gene disruption, gene replacement, and opto- and chemogenetic manipulation of the SCN circadian clock.

The functional state of denervated muscle is a critical factor in the ability to restore movement after injury- or disease-related paralysis. Here we used peripheral optogenetic stimulation and transcriptome profiling in the mouse whisker system to investigate the time course of changes in neuromuscular function following complete unilateral facial nerve transection. While most skeletal muscles rapidly lose functionality after lower motor neuron denervation, optogenetic muscle stimulation of the paralyzed whisker pad revealed sustained increases in the sensitivity, velocity, and amplitude of whisker movements, and reduced fatigability, starting 48 h after denervation. RNA-seq analysis showed distinct regulation of multiple gene families in denervated whisker pad muscles compared with the atrophy-prone soleus, including prominent changes in ion channels and contractile fibers. Together, our results define the unique functional and transcriptomic landscape of denervated facial muscles and have general implications for restoring movement after neuromuscular injury or disease. NEW & NOTEWORTHY Optogenetic activation of muscle can be used to noninvasively induce movements and probe muscle function. We used this technique in mice to investigate changes in whisker movements following facial nerve transection. We found unexpectedly enhanced functional properties of whisker pad muscle following denervation, accompanied by unique transcriptomic changes. Our findings highlight the utility of the mouse whisker pad for investigating the restoration of movement after paralysis.

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14The functional state of denervated muscle is a critical factor in the ability to restore movement 15 after injury-or disease-related paralysis. Here we used peripheral optogenetic stimulation and 16 transcriptome profiling in the mouse whisker system to investigate the time course of changes in 17 neuromuscular function following complete unilateral facial nerve transection. While most 18 skeletal muscles rapidly lose functionality after lower motor neuron denervation, optogenetic 19 muscle stimulation of the paralyzed whisker pad revealed sustained increases in the sensitivity, 20 velocity, and amplitude of whisker movements, and reduced fatigability, starting 48 h after 21 denervation. RNA-seq analysis showed distinct regulation of multiple gene families in 22 denervated whisker pad muscles compared to the atrophy-prone soleus, including prominent 23 changes in ion channels and contractile fibers. Together, our results define the unique functional 24 and transcriptomic landscape of denervated facial muscles, and have general implications for 25 restoring movement after neuromuscular injury or disease. 26 27New & Noteworthy: Optogenetic activation of muscle can be used to non-invasively induce 31 movements and probe muscle function. We used this technique in mice to investigate changes 32 in whisker movements following facial nerve transection. We found unexpectedly enhanced 33 functional properties of whisker pad muscle following denervation, accompanied by unique 34 transcriptomic changes. Our findings highlight the utility of the mouse whisker pad for 35 investigating the restoration of movement after paralysis. 36 37 Haidarliu et al 2010, Park et al 2016), collectively referred to here as "whisker pad muscles". In 61 the present study, we used ChAT-ChR2 and Emx1-ChR2 mice to evoke whisker movements via 62 stimulation of the facial motor nerve (cranial nerve VII) or the whisker pad muscles, respectively. 63 This allowed us to investigate the functional changes that occur in nerve and muscle after the 64 paralysis of whisker movements caused by facial nerve transection. 65 66 One recent study used optogenetic muscle stimulation in the hindlimb triceps surae after sciatic 67 nerve lesion to demonstrate dramatic atrophy and loss of function (Magown et al 2015), 68consistent with classic studies in this system (Nelson 1969), that could be attenuated by daily 69 optogenetic activation. We considered it possible that whisker pad muscles undergo distinct 70 denervation-induced changes compared to other muscle types, in part because whisker pad 71 position of the nerve in ChAT-ChR2 mice, the buccal branch of the facial nerve (cranial nerve 126 VII) was targeted with the light spot at a position between the stylomastoid foramen, ventral to 127 the ear canal and caudal to the whisker pad. Different illumination positions in this region were 128 tested for each subject to optimize the evoked whisker protraction. To test effects on 129 fasciculations, dantrolene (1 mM) was applied subcutaneously to the lesioned (right-...