Kisspeptin neurons in the mediobasal hypothalamus (MBH) are critical targets of ovarian estrogen feedback regulating mammalian fertility. To reveal molecular mechanisms underlying this signaling, we thoroughly characterized the estrogen-regulated transcriptome of kisspeptin cells from ovariectomized transgenic mice substituted with 17β-estradiol or vehicle. MBH kisspeptin neurons were harvested using laser-capture microdissection, pooled, and subjected to RNA sequencing. Estrogen treatment significantly ( p.adj . < 0.05) up-regulated 1,190 and down-regulated 1,139 transcripts, including transcription factors, neuropeptides, ribosomal and mitochondrial proteins, ion channels, transporters, receptors, and regulatory RNAs. Reduced expression of the excitatory serotonin receptor-4 transcript ( Htr4 ) diminished kisspeptin neuron responsiveness to serotonergic stimulation. Many estrogen-regulated transcripts have been implicated in puberty/fertility disorders. Patients ( n = 337) with congenital hypogonadotropic hypogonadism (CHH) showed enrichment of rare variants in putative CHH-candidate genes (e.g., LRP1B , CACNA1G , FNDC3A ). Comprehensive characterization of the estrogen-dependent kisspeptin neuron transcriptome sheds light on the molecular mechanisms of ovary–brain communication and informs genetic research on human fertility disorders.

Human reproduction is controlled by ~2,000 hypothalamic gonadotropin-releasing hormone (GnRH) neurons. Here we report the discovery and characterization of additional ~150,000-200,000 GnRH-synthesizing cells in the human basal ganglia and basal forebrain. Nearly all extrahypothalamic GnRH neurons expressed the cholinergic marker enzyme choline acetyltransferase. Similarly, hypothalamic GnRH neurons were also cholinergic both in embryonic and adult human brains. Whole-transcriptome analysis of cholinergic interneurons and medium spiny projection neurons laser-microdissected from the human putamen showed selective expression of GNRH1 and GNRHR1 autoreceptors in the cholinergic cell population and uncovered the detailed transcriptome profile and molecular connectome of these two cell types. Higher-order non-reproductive functions regulated by GnRH under physiological conditions in the human basal ganglia and basal forebrain require clarification. The role and changes of GnRH/GnRHR1 signaling in neurodegenerative disorders affecting cholinergic neurocircuitries, including Parkinson's and Alzheimer's diseases, need to be explored.

Kisspeptin neurons residing in the rostral periventricular area of the third ventricle (KPRP3V) and the arcuate nucleus (KPARC) mediate positive and negative estrogen feedback, respectively. Here, we aim to compare transcriptional responses of KPRP3V and KPARC neurons to estrogen. Transgenic mice were ovariectomized and supplemented with either 17β-estradiol (E2) or vehicle. Fluorescently tagged KPRP3V neurons collected by laser-capture microdissection were subjected to RNA-seq. Bioinformatics identified 222 E2-dependent genes. Four genes encoding neuropeptide precursors (Nmb, Kiss1, Nts, Penk) were robustly, and Cartpt was subsignificantly upregulated, suggesting putative contribution of multiple neuropeptides to estrogen feedback mechanisms. Using overrepresentation analysis, the most affected KEGG pathways were neuroactive ligand-receptor interaction and dopaminergic synapse. Next, we re-analyzed our previously obtained KPARC neuron RNA-seq data from the same animals using identical bioinformatic criteria. The identified 1583 E2-induced changes included suppression of many neuropeptide precursors, granins, protein processing enzymes, and other genes related to the secretory pathway. In addition to distinct regulatory responses, KPRP3V and KPARC neurons exhibited sixty-two common changes in genes encoding three hormone receptors (Ghsr, Pgr, Npr2), GAD-65 (Gad2), calmodulin and its regulator (Calm1, Pcp4), among others. Thirty-four oppositely regulated genes (Kiss1, Vgf, Chrna7, Tmem35a) were also identified. The strikingly different transcriptional responses in the two neuron populations prompted us to explore the transcriptional mechanism further. We identified ten E2-dependent transcription factors in KPRP3V and seventy in KPARC neurons. While none of the ten transcription factors interacted with estrogen receptor-α, eight of the seventy did. We propose that an intricate, multi-layered transcriptional mechanism exists in KPARC neurons and a less complex one in KPRP3V neurons. These results shed new light on the complexity of estrogen-dependent regulatory mechanisms acting in the two functionally distinct kisspeptin neuron populations and implicate additional neuropeptides and mechanisms in estrogen feedback.

In an attempt to model occupational and environmental Mn exposures and their possible interaction, young male Wistar rats were exposed to Mn by oral administration in dissolved form (MnCl2 ·4H2O, 14.84 and 59.36 mg/kg b.w.) and by intratracheal application of MnO2 nanoparticles (2.63 mg/kg b.w.). After 3 and 6 weeks oral, or 3 weeks oral plus 3 weeks intratracheal, exposure, general toxicological, and electrophysiological tests were done. Body weight gain was significantly reduced after 6 and 3 plus 3 weeks exposure, but the effect of the latter on the pace of weight gain was stronger. Organ weights signalized systemic stress and effect on lungs. Changes in evoked electrophysiological responses (cortical sensory evoked potential and nerve action potential) indicated that the 3 plus 3 weeks combined exposure caused equal or higher changes in the latency of these responses than 6 weeks of exposure, although the calculated summed Mn dose in the former case was lower. The results showed the importance of the physicochemical form of Mn in determining the toxic outcome, and suggested that neurofunctional markers of Mn action may indicate the human health effect better than conventional blood Mn measurement.

Kisspeptin (KP) synthesizing neurons of the hypothalamic infundibular region are critically involved in the central regulation of fertility; these cells regulate pulsatile gonadotropin-releasing hormone (GnRH) secretion and mediate sex steroid feedback signals to GnRH neurons. Fine structural analysis of the human KP system is complicated by the use of post mortem tissues. To gain better insight into the neuroanatomy of the somato-dendritic cellular compartment, we introduced the diolistic labeling of immunohistochemically identified KP neurons using a gene gun loaded with the lipophilic dye, DiI. Confocal microscopic studies of primary dendrites in 100-µm-thick tissue sections established that 79.3% of KP cells were bipolar, 14.1% were tripolar, and 6.6% were unipolar. Primary dendrites branched sparsely, contained numerous appendages (9.1 ± 1.1 spines/100 µm dendrite), and received rich innervation from GABAergic, glutamatergic, and KP-containing terminals. KP neuron synaptology was analyzed with immunoelectron microscopy on perfusion-fixed specimens. KP axons established frequent contacts and classical synapses on unlabeled, and on KP-immunoreactive somata, dendrites, and spines. Synapses were asymmetric and the presynaptic structures contained round and regular synaptic vesicles, in addition to dense-core granules. Although immunofluorescent studies failed to detect vesicular glutamate transporter isoforms in KP axons, ultrastructural characteristics of synaptic terminals suggested use of glutamatergic, in addition to peptidergic, neurotransmission. In summary, immunofluorescent and DiI labeling of KP neurons in thick hypothalamic sections and immunoelectron microscopic studies of KP-immunoreactive neurons in brains perfusion-fixed shortly post mortem allowed us to identify previously unexplored fine structural features of KP neurons in the mediobasal hypothalamus of humans.

The human infundibular nucleus (corresponding to the rodent arcuate nucleus) serves as an important integration center for neuronal signals and hormones released by peripheral endocrine organs. Kisspeptin (KP) producing neurons of this anatomical site many of which also synthesize neurokinin B (NKB) are critically involved in sex hormone signaling to gonadotropin-releasing homone (GnRH) neurons. In recent years, the basic topography, morphology, neuropeptide content and connectivity of human KP neurons have been investigated with in situ hybridization and immunohistochemistry on post mortem tissues. These studies revealed that human KP neurons differ neurochemically from their rodent counterparts and show robust aging-related plasticity. Earlier immunohistochemical experiments also provided evidence for temporal changes in the hypothalamus of aging men whose NKB and KP neurons undergo hypertrophy, increase in number, exhibit increased neuropeptide mRNA expression and immunoreactivity and give rise to higher numbers of immunoreactive fibers and afferent contacts onto GnRH neurons. Increasing percentages of KP-expressing NKB perikarya, NKB axons and NKB inputs to GnRH neurons raise the intriguing possibility that a significant subset of NKB neurons begins to co-synthesize KP as aging proceeds. Although use of post mortem tissues is technically challenging, recently-available single-cell anatomical and molecular approaches discussed in this review article provide promising new tools to investigate the aging-related anatomical and functional plasticity of the human KP neuronal system.

Approximately one month after a red sludge industrial accident, a high concentration of atmospheric aerosol was reported in the affected area. This increased aerosol load originated mainly from the fugitive dust emitted from the red sludge sediment, representing a serious threat to public health. In a study presented here, a recently developed four-wavelength photoacoustic spectrometer (4λ-PAS), which simultaneously measures optical aerosol absorption in the near infrared, visible and ultra-violet spectral ranges with high temporal resolution, was operated in the centre of the sludge hit area. Based on the results of the photoacoustic absorption measurements of temporal changes in light absorption and their correlations with other aerosol variables, including size distribution, were carried out in special atmospheric conditions (including with the presence of "atypical" red mud particles). Correlations between segregated absorption Ångström exponents (AAEs) and specific size ranges were tabulated, and the resulting data were used to draw conclusions about the reasons for the temporal changes to the measured quantities. The absorption behavior of red mud particles was determined in UV, VIS, and near IR spectral regions under laboratory conditions. The absorption spectrum of red mud aerosol was found to be atypical (i.e., it did not follow a power law), and this was used as a selective parameter for its identification in ambient bulk. Each measurement day, at around noon, dust from the red sludge sediment was identified; while during the rest of the day atmospheric aerosol composition was dominated by carbonaceous aerosols, originating most probably from home heating and/or from traffic. In-situ number concentration and size distribution measurements confirm this interpretation of the measurements obtained with 4λ-PAS.

Background: Kisspeptin (KP) neurons in the rostral periventricular region of the 3rd ventricle (RP3V) of female rodents mediate positive estrogen feedback to gonadotropin-releasing hormone neurons and, thus, play a fundamental role in the mid-cycle luteinizing hormone (LH) surge. The RP3V is sexually dimorphic, and male rodents with lower KP cell numbers are unable to mount estrogen-induced LH surges. Objective: To find and characterize the homologous KP neurons in the human brain, we studied formalin-fixed post­mortem hypothalami. Methods: Immunohistochemical techniques were used. Results: The distribution of KP neurons in the rostral hypothalamus overlapped with distinct subdivisions of the paraventricular nucleus. The cell numbers decreased after menopause, indicating that estrogens positively regulate KP gene expression in the rostral hypothalamus in humans, similarly to several other species. Young adult women and men had similar cell numbers, as opposed to rodents reported to have more KP neurons in the RP3V of females. Human KP neurons differed from the homologous rodent cells as well, in that they were devoid of enkephalins, galanin and tyrosine hydroxylase. Further, they did not contain known KP neuron markers of the human infundibular nucleus, neurokinin B, substance P and cocaine- and amphetamine-regulated transcript, while they received afferent input from these KP neurons. Conclusions: The identification and positive estrogenic regulation of KP neurons in the human rostral hypothalamus challenge the long-held view that positive estrogen feedback may be restricted to the mediobasal part of the hypothalamus in primates and point to the need of further anatomical, molecular and functional studies of rostral hypothalamic KP neurons.