Abstract
BackgroundPrevious studies show that galanin neurons in ventrolateral preoptic nucleus (VLPO-Gal) are essential for sleep regulation. Here, we explored the transcriptional regulation of the VLPO-Gal neurons in sleep by comparing their transcriptional responses between sleeping mice and those kept awake, sacrificed at the same diurnal time.ResultsRNA-sequencing (RNA-seq) analysis was performed on eGFP(+) galanin neurons isolated using laser captured microdissection (LCM) from VLPO. Expression of Gal was assessed in our LCM eGFP(+) neurons via real time qPCR and showed marked enrichment when compared to LCM eGFP(−) cells and to bulk VLPO samples. Gene set enrichment analysis utilizing data from a recent single-cell RNA-seq study of the preoptic area demonstrated that our VLPO-Gal samples were highly enriched with galanin-expressing inhibitory neurons, but not galanin-expressing excitatory neurons. A total of 263 genes were differentially expressed between sleep and wake in VLPO-Gal neurons. When comparing differentially expressed genes in VLPO-Gal neurons to differentially expressed genes in a wake-active neuronal region (the medial prefrontal cortex), evidence indicates that both systemic and cell-specific mechanisms contribute to the transcriptional regulation in VLPO-Gal neurons. In both wake-active and sleep-active neurons, ER stress pathways are activated by wake and cold-inducible RNA-binding proteins are activated by sleep. In contrast, expression of DNA repair genes is increased in VLPO-Gal during wakefulness, but increased in wake-active cells during sleep.ConclusionOur study identified transcriptomic responses of the galanin neurons in the ventrolateral preoptic nucleus during sleep and sleep deprivation. Data indicate that VLPO contains mainly sleep-active inhibitory galaninergic neurons. The VLPO galanin neurons show responses to sleep and wake similar to wake-active regions, indicating these responses, such as ER stress and cold-inducible RNA-binding proteins, are systemic affecting all neuronal populations. Region-specific differences in sleep/wake responses were also identified, in particular DNA repair. Our study expands knowledge about the transcriptional response of a distinct group of neurons essential for sleep.
Highlights
Previous studies show that galanin neurons in ventrolateral preoptic nucleus (VLPO-Gal) are essential for sleep regulation
laser captured microdissection (LCM) enriched galanin expressing neurons from ventrolateral preoptic area (VLPO) Galanin neurons are located in a number of nuclei and sub-structures of VLPO in preoptic area (POA), in addition to the VLPO core
The short post-fix was sufficient for the GFP fluorescent detection in the VLPO core and the medial VLPO extension, but not in the dorsal extension of VLPO and beyond (Supplementary figure 1 in Additional file 1). Enhanced green fluorescent protein (eGFP)(+) cells in VLPO core were dissected using LCM with care taken to avoid picking eGFP(+) cells in the medial VLPO extension, as shown in Supplementary figure 2 (Additional file 1)
Summary
Previous studies show that galanin neurons in ventrolateral preoptic nucleus (VLPO-Gal) are essential for sleep regulation. Studies using c-Fos immunoreactivity to indicate recent neuronal activities demonstrated increased numbers of c-Fos positive neurons in VLPO following consolidated sleep compared to wake, and the number of c-Fos positive neurons increased with recovery sleep after sleep deprivation, and showed a positive correlation with the amount of sleep before sacrifice [17, 18, 50] These sleep-active neurons, from VLPO, project to histaminergic tuberomammillary nucleus (TMN), serotonergic dorsal raphe (DR), and noradrenergic locus coreleus (LC), and the majority are GABAergic inhibitory neurons, suggesting that during sleep they inhibit multiple monoamine arousal systems [49, 51]. We aimed to further characterize the galaninergic neurons in VLPO by examining their behavioral state dependent transcriptional regulation between sleep and wake
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
