Journal of Endocrinology

Molecular heterogeneity of Crh⁺ neurons in the hypothalamus. (A) Distribution of GFP⁺ neurons outside the PVN in the hypothalamus of Crh-IRES-Cre::GFP mice. (B) Single-cell RNA-seq distinguishes 62 neuronal subclasses along the PVN–arcuate axis of the hypothalamus (Romanov et al. 2015). Expression levels (horizontal axis) were plotted as means of log₂ transformed mRNA copy numbers ± s.e.m. Red and green colors identify GABAergic (#14, #15 and #24) and glutamatergic clusters (#44 and #45) of hypothalamic neurons, which express Crh mRNA at levels exceeding 2× the s.e.m. *q < 0.05 (Wilcoxon rank-sum test corrected for multiple testing). (C) Among both GABA and glutamate neurons, subsets are endowed with Crh mRNAs. (D) CRH co-exists in Lhx6-GFP neurons (asterisks) particularly in the bed nucleus of stria terminalis and preoptic area. (E) Neurotransmitter heterogeneity of Crh⁺ neurons. Venn diagrams demonstrate the proportion of dual and triple neuronal phenotypes in GABA/glutamate (top), GABA/dopamine (middle) and GABA/glutamate/dopamine (bottom) neurons. This analysis was performed using a threshold for mRNA expression at a level of ≥2 mRNA transcripts for each gene. Percentage values indicate the proportion of Crh⁺ neurons falling into groups and intersections (‘dual phenotype’ categories). Scale bars = 250 µm (A), 10 µm (D). Adapted, with permission, from Romanov et al. (2016).