Mathematical Modeling of Neurons and Endocrine Cells

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Together with the Stojilkovic lab and the Genomics core of NICHD, we have initiated a new direction in these studies, single-cell RNAseq analysis of gene expression in the anterior pituitary. This gland is composed of five hormone producing cell types, glia-like folliculostellate cells, and endothelial and blood cells comprising the pituitary sinusoidal capillary network. Some key lines of inquiry in pituitary physiology include three-dimensional organization tissue organization and intercellular communication among cells, development and regeneration of pituitary cells, and the heterogeneity and function of folliculostellate cells. Great efforts have been dedicated towards these aims, but many of the molecular underpinnings remain unknown. We have carried out the first single-cell transcriptome study of anterior pituitary cells. We profiled over 6700 freshly dispersed cells from postpubertal male and female rats. In addition to confirming known markers, our transcriptome analysis highlights many novel genetic markers contributing to pituitary cell type identity and sexual dimorphism. We provide for the first time an estimate of the percentage of folliculostellate cells, and we identify at least two main subtypes of this heterogeneous cell population. Our data support the hypothesis that folliculostellate cells provide structural support for the hormone producing cells, as they express many genes encoding detoxification enzymes. Our analysis of expression of developmental, stem cell/progenitor, and neuroendocrine markers suggests that folliculostellate and hormone producing cells are sister cells with a common origin. We also provide a detailed view of cell type-dependent expression of genes encoding extracellular matrix, cell adhesion, and endogenous ligand proteins, critical for the tridimensional organization of the anterior pituitary and the cross-talk between its constituent cells. We expect the marker genes identified for pituitary cell types will serve as a solid base for future investigations into pituitary structure, function, and pathophysiology, as well as for the study of postnatal pituitary development, regeneration, and reorganization. The results were reported in ref. #1. Continuous, as opposed to pulsatile, delivery of hypothalamic gonadotropin-releasing hormone (GnRH) leads to a marked decrease in secretion of pituitary gonadotropins LH and FSH and impairment of reproductive function. Here we studied the expression profile of gonadotropin subunit and GnRH receptor genes in rat pituitary in vitro and in vivo to clarify their expression profiles in the absence and continuous presence of GnRH. Culturing of pituitary cells in GnRH-free conditions downregulated Fshb, Cga, and Gnrhr expression, whereas continuous treatment with GnRH agonists upregulated Cga expression progressively and Gnrhr and Fshb expression transiently, accompanied by a prolonged blockade of Fshb but not Gnrhr expression. In contrast, Lhb expression was relatively insensitive to loss of endogenous GnRH and continuous treatment with GnRH, probably reflecting the status of Egr1 and Nr5a1 expression. Similar patterns of responses were observed in vivo after administration of a GnRH agonist. However, continuous treatment with GnRH stimulated LH secretion in vitro and in vivo, leading to decrease in LH cell content despite high basal Lhb expression. These data suggest that blockade of Fshb expression and depletion of the LH secretory pool are two major factors accounting for weakening of the gonadotroph secretory function during continuous GnRH treatment. The results were published in ref. #2.