Single-cell multiplex qPCR evidence for sex-dimorphic glutamate decarboxylase, estrogen receptor, and 5'-AMP-activated protein kinase alpha subunit mRNA expression by ventromedial hypothalamic nucleus GABAergic neurons

Abstract

The inhibitory amino acid transmitter γ-aminobutryic acid (GABA) acts within the ventromedial hypothalamus to regulate systemic glucose homeostasis, but the issue of whether this neurochemical signal originates locally or is supplied by afferent innervation remains controversial. Here, combinatory in situ immunocytochemistry/laser-catapult microdissection/single-cell multiplex qPCR techniques were used to investigate the premise that ventromedial hypothalamic nucleus ventrolateral (VMNvl) and/or dorsomedial (VMNdm) division neurons contain mRNAs that encode glutamate decarboxylase (GAD)₆₅ or GAD₆₇ and metabolic-sensory biomarkers, and that expression of these genes is sex-dimorphic. In male and female rats, GAD₆₅ mRNA was elevated in VMNvl versus VMNdm GAD_65/67-immunopositive (-ir) neurons, yet the female exhibited higher GAD₆₇ transcript content in VMNdm versus VMNvl GABAergic nerve cells. Estrogen receptor (ER)-alpha transcripts were lower in female versus male GABA neurons from either VMN division; ER-beta and G-protein-coupled ER-1 mRNA expression profiles were also comparatively reduced in cells from female versus male VMNvl. VMNvl and VMNdm GAD_65/67-ir-positive neurons showed equivalent levels of glucokinase and sulfonylurea receptor-1 mRNA between sexes. 5'-AMP-activated protein kinase-alpha 1 (AMPK_α1) and -alpha 2 (AMPK_α2) transcripts were lower in female versus male VMNdm GABAergic neurons, yet AMPK_α2 mRNA levels were higher in cells acquired from female versus male VMNvl. Current studies document GAD₆₅ and -₆₇ gene expression in VMNvl and VMNdm GAD_65/67-ir-positive neurons in each sex. Results infer that GABAergic neurons in each division may exhibit sex differences in receptiveness to estradiol. Outcomes also support the prospect that energy sensory function by this neurotransmitter cell type may predominate in the VMNvl in female versus VMNdm in the male.

Keywords: AMPK; Estrogen receptor-alpha; G-protein-coupled estrogen receptor-1; Glucokinase regulatory protein; Glutamate decarboxylase; Ventromedial hypothalamic nucleus.