BioProject

The immense molecular diversity of neurons challenges our ability to deconvolve the relationship between the genetic and the cellular underpinnings of neuropsychiatric disorders. More...

The immense molecular diversity of neurons challenges our ability to deconvolve the relationship between the genetic and the cellular underpinnings of neuropsychiatric disorders. Hypocretin (orexin) containing neurons of the lateral hypothalamus are clearly essential for the normal regulation of sleep and wake behaviors, and have been implicated in feeding, anxiety, depression and reward. However, little is known about the molecular phenotypes of these cells, or the mechanism of their specification. We have generated a Hcrt bacTRAP line for comprehensive translational profiling of these neurons in vivo. From this profile, we have identified 188 transcripts, as enriched in these neurons, in additions to thousands more moderately enriched or nominally expressed. We validated many of these at the RNA and protein level, including the transcription factor Lhx9. Lhx9 protein is found in a subset of these neurons, and ablation of these gene results in a 30% loss of Hcrt neuron number, and a profound hypersomnolence in mice. This data suggests that Lhx9 may be important for specification of some Hcrt neurons, and the subsets of these neurons may contribute to discrete sleep phenotypes. Overall design: Four independent TRAP replicates were collected, and total RNA from both the immunoprecipitate(IP) and unbound(total) fractions were seperately amplified and hybridized. Unbound fractions are provided to serve as total RNA controls for the tissue. Biological replicates are GCRMA normalized within groups. Following averaging of replicates, we recommend further global normalization between groups, using affymetrix biotinylated controls, to correct for any broad biases in scanning and hybridization. Finally for many analyses, we also recommend filtering to remove those probesets with low IP/Total fold change values from each cell type(see PMID:20962086). Researchers can contact us for spreadsheets where these additional steps have been completed. Less...