Development And Regulation Of The Gonadotropin Releasing Hormone System

National Institute Of Neurological Disorders And Stroke

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Abstract

GnRH neurons, critical for reproduction, are derived from the nasal placode and migrate into the brain where they become integral members of the hypothalamic-pituitary-gonadal axis. We study mechanism(s) underlying GnRH neuronal differentiation, migration and axonal targeting in normal/transgenic animals, and two in vitro model systems nasal explants and GnRH derived cell lines. Using these animal and in vitro models, our work also addresses the mechanisms regulating (intrinsic and trans-synaptic) GnRH gene expression, peptide synthesis and secretion in GnRH neurons. Multiple approaches are used to identify and understand the multitude of molecules and factors which play a role in directing the GnRH neurons to their final location in the CNS. These include differential screening of libraries obtained from migrating versus non-migrating cells, examination of molecules differentially expressed at key locations along the migratory route, morphological examination of the development of the GnRH system in knockout mice, and perturbation of molecules in vitro and subsequent monitoring of GnRH neuronal movement. As GnRH neurons migrate, they also mature, and the two processes may in fact be linked. To investigate the maturation of GnRH neurons, we use calcium imaging, electrophysiology and biochemical measures to examine GnRH neuronal activity and peptide secretion. In addition, we collaborate with labs performing human genetic screening of patients with reproductive dysfunction. Once a mutation is identified, we analyze the expression pattern in mice and perform biological assays to determine the outcome of the mutated gene on GnRH development. Over the past year, four articles were published, 2 reviews and 2 primary articles. In vertebrates, Gonadotropin releasing hormone-1 (GnRH) neuroendocrine cells originate from two different lineages in the olfactory placode and migrate into the forebrain where they regulate reproduction. Three publications focused on the development of the GnRH system and one focused on regulation of GnRH neuronal activity relevant for reproductive function. Developmental articles: This paper (Welch B, Cho H-J, Ucakturk SA, Farmer, SM, Cetinkaya S, Abaci A, Akkus G, Simsek E, Kotan LD, Turan I, Yuksel B, Wray S, Topaloglu AK. PLXNB1 Mutations in the Etiology of Idiopathic Hypogonadotropic Hypogonadism. J. of Neuroendocrinology, 2022) examines mutations identified in human patients exhibiting delayed or absent puberty. Idiopathic hypogonadotropic hypogonadism (IHH) comprises a group of rare genetic disorders characterized by pubertal failure caused by GnRH deficiency. Genetic factors involved in semaphorin/plexin signaling have been identified in patients with IHH. PlexinB1, a member of the plexin family receptors, serves as the receptor for semaphorin 4D. In mice, perturbations in Sema4D/PlexinB1 signaling leads to improper GnRH development, highlighting the importance of investigating PlexinB1 mutations in IHH families. In total, 336 IHH patients from 290 independent families were included in the present study. Six PLXNB1 rare sequence variants (p.N361S, p.V608A, p.R636C, p.V672A, p.R1031H, and p.C1318R) are described in eight IHH patients from seven independent families. These variants were examined using bioinformatic modeling and compared to mutants reported in PLXNA1. Based on these analyses, the variant p.R1031H was assayed for alterations in cell morphology, PlexinB1 expression, and migration using a GnRH cell line and Boyden chambers. Experiments showed reduced membrane expression and impaired migration in cells expressing this variant compared to the wild-type. Our results provide clinical, genetic, molecular/cellular, and modeling evidence to implicate variants in PLXNB1 in the etiology of IHH. This review (Shan Y and Wray S. Hidden pitfalls in deciphering the gonadotropin releasing hormone neuroendocrine cell lineage. J. Neuroendocrinology, 2021) addresses the lineage of GnRH cells. To this day, the identity of GnRH progenitors remains unclear. However, the visualization of different developmental markers in subsets of GnRH neurons during early embryonic stages raised the possibility of at least two GnRH subpopulations. This observation led directly to a second question. Does visualization of different developmental markers in subsets of GnRH neurons reflect functional heterogeneity? This question remains unanswered, but as we learn more about the GnRH system, functional GnRH subpopulations becomes critically important to understanding GnRH function. This review addresses the development of the neuroendocrine GnRH system, specifically the heterogeneity of the GnRH neuroendocrine population. In this review (Duittoz A, Forni PE, Giacobini P, Golan M, Mollard P, Negrn AL, Radovick S, Wray S. Development of the Gonadotropin releasing hormone system. J. Neuroendocrinology, 2022), I assembled an international group of researchers working on the development of the GnRH system for a special issue celebrating the 50th anniversary of the discovery of GnRH. This review summarizes our current understanding of the development of the GnRH system, including discussion on open questions regarding (1) transcriptional regulation of the Gnrh1 gene; (2) prenatal development of the GnRH1 system in rodents and humans; and (3) paracrine and synaptic communication during migration of the GnRH cells. Regulation of GnRH neuronal activity: This paper (Dairaghi L, Constantin S, Oh A, Shostak D, Wray, S. The Dopamine D4 receptor regulates gonadotropin-releasing hormone neuron excitability in male mice. eNeuro, 2022)clarifies how dopamine regulated GnRH neuronal activity. While it was known that dopamine regulates GnRH neurons, the specific dopamine receptor subtype(s) involved remain unclear. Previous studies in adult rodents have reported juxtaposition of fibers containing tyrosine hydroxylase (TH), a marker of catecholaminergic cells, onto GnRH neurons and that exogenous dopamine inhibits GnRH neurons postsynaptically through dopamine D1-like and/or D2-like receptors. Our microarray data from GnRH neurons revealed a high level of Drd4 transcripts i.e., dopamine D4 receptor(D4R). Single-cell RT-PCR and immunocytochemistry confirmed GnRH cells express the Drd4 transcript and protein, respectively. Calcium imaging identified changes in GnRH neuronal activity during application of subtype-specific dopamine receptor agonists and antagonists when GABAergic and glutamatergic transmission was blocked. Dopamine, dopamine with D1/5R-specific or D2/3R-specific antagonists or D4R-specific agonists decreased the frequency of calcium oscillations. In contrast, D1/5R-specific agonists increased the frequency of calcium oscillations. The D4R-mediated inhibition was dependent on Gai/o protein coupling, while the D1/5R-mediated excitation required Gas protein coupling. Together, these results indicate that D4R plays an important role in the dopaminergic inhibition of GnRH neurons.