GnRH Neurons Provide Direct Input to Hypothalamic Tyrosine Hydroxylase Immunoreactive Neurons Which Is Maintained During Lactation

Abstract

Gonadotropin releasing hormone (GnRH) neurons provide neuronal input to the preoptic area (POA) and the arcuate nucleus (Arc), two regions involved critically in the regulation of neuroendocrine functions and associated behaviors. These areas contain tyrosine hydroxylase immunoreactive (TH-IR) neurons, which play location-specific roles in the neuroendocrine control of both the luteinizing hormone and prolactin secretion, as well as, sexually motivated behaviors. Concerning changes in the activity of GnRH neurons and the secretion pattern of GnRH seen under the influence of rising serum estrogen levels and during lactation, we tested the hypothesis that the functional state of GnRH neurons is mediated via direct synaptic connections to TH-IR neurons in the POA and Arc. In addition, we examined putative changes of these inputs in lactating mice and in mothers separated from their pups. Confocal microscopic and pre-embedding immunohistochemical studies on ovariectomized mice treated with 17β-estradiol (OVX+E2) provided evidence for direct appositions and asymmetric synapses between GnRH-IR fiber varicosities and TH-IR neurons in the POA and the Arc. As TH co-localizes with kisspeptin (KP) in the POA, confocal microscopic analysis was continued on sections additionally labeled for KP. The TH-IR neurons showed a lower level of co-labeling for KP in lactating mice compared to OVX+E2 mice (16.1 ± 5% vs. 57.8 ± 4.3%). Removing the pups for 24 h did not alter significantly the KP production in TH-IR neurons (17.3 ± 4.6%). The mean number of GnRH-IR varicosities on preoptic and arcuate TH cells did not differ in the three animal models investigated. This study shows evidence that GnRH neurons provide direct synaptic inputs to POA and Arc dopaminergic neurons. The scale of anatomical connectivity with these target cells was unaltered during lactation indicating a maintained GnRH input, inspite of the altered hormonal condition.

Development of a versatile LCM-Seq method for spatial transcriptomics of fluorescently tagged cholinergic neuron populations

“…Cholinergic cells were recognized based on their ZsGreen fluorescence. Dopaminergic neuronal elements were detected with chicken anti-TH antibodies (AB_10013440, Aves Laboratories; 1:1000; 48 h; 4 °C) ( 53 ) and TRH neurons with rabbit proTRH antibodies (PA5-57331, Invitrogen; 1:1000; 48 h; 4 °C) ( 54 ). Primary antibodies were reacted with Cy3-conjugated secondary antibodies (Jackson ImmunoResearch; 1:500; 2 h; room temperature).…”

Development of a versatile LCM-Seq method for spatial transcriptomics of fluorescently-tagged cholinergic neuron populations

“…Cholinergic cells were recognized based on their ZsGreen fluorescence. Dopaminergic neuronal elements were detected with chicken anti-TH antibodies (AB_10013440, Aves Laboratories, Davis, CA, USA; 1:1,000; 48h; 4 °C) (49) and TRH neurons with rabbit proTRH antibodies (PA5-57331, Invitrogen, Waltham, MA, USA; 1:1,000; 48h; 4 °C) (50). Primary antibodies were reacted with Cy3-conjugated secondary antibodies (Jackson ImmunoResearch; 1:500; 2h; RT).…”

Expression of type one cannabinoid receptor in different subpopulation of kisspeptin neurons and kisspeptin afferents to GnRH neurons in female mice

“…Besides producing various neuropeptides (neurokinin B, dynorphin, galanin) (Goodman et al 2007 ; Murakawa et al 2016 ; Kallo et al 2012 ; Porteous et al 2011 ) or biogenic amines (dopamine) (Clarkson and Herbison 2011 ; Skrapits et al 2015 ; Bardoczi et al 2018 ), subpopulations of kisspeptin neurons use classical neurotransmitters like glutamate and GABA, which contribute to the excitatory and/or inhibitory innervation of the neuroendocrine cells. Contrasting the peptide-type transmitters, these classical neurotransmitters are released spontaneously, as well as at an evoked manner, which raises a potential role of endocannabinoid signaling to discretely influence the release of neurotransmitters/neuromodulators from the different kisspeptin subpopulations.…”