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1.
J Neurosci ; 44(22)2024 May 29.
Article in English | MEDLINE | ID: mdl-38604780

ABSTRACT

The autonomic nervous system (ANS) regulates the body's physiology, including cardiovascular function. As the ANS develops during the second to third trimester, fetal heart rate variability (HRV) increases while fetal heart rate (HR) decreases. In this way, fetal HR and HRV provide an index of fetal ANS development and future neurobehavioral regulation. Fetal HR and HRV have been associated with child language ability and psychomotor development behavior in toddlerhood. However, their associations with postbirth autonomic brain systems, such as the brainstem, hypothalamus, and dorsal anterior cingulate cortex (dACC), have yet to be investigated even though brain pathways involved in autonomic regulation are well established in older individuals. We assessed whether fetal HR and HRV were associated with the brainstem, hypothalamic, and dACC functional connectivity in newborns. Data were obtained from 60 pregnant individuals (ages 14-42) at 24-27 and 34-37 weeks of gestation using a fetal actocardiograph to generate fetal HR and HRV. During natural sleep, their infants (38 males and 22 females) underwent a fMRI scan between 40 and 46 weeks of postmenstrual age. Our findings relate fetal heart indices to brainstem, hypothalamic, and dACC connectivity and reveal connections with widespread brain regions that may support behavioral and emotional regulation. We demonstrated the basic physiologic association between fetal HR indices and lower- and higher-order brain regions involved in regulatory processes. This work provides the foundation for future behavioral or physiological regulation research in fetuses and infants.


Subject(s)
Brain Stem , Gyrus Cinguli , Heart Rate, Fetal , Hypothalamus , Magnetic Resonance Imaging , Humans , Female , Male , Gyrus Cinguli/physiology , Gyrus Cinguli/diagnostic imaging , Brain Stem/diagnostic imaging , Brain Stem/physiology , Infant, Newborn , Pregnancy , Heart Rate, Fetal/physiology , Adult , Hypothalamus/physiology , Hypothalamus/diagnostic imaging , Hypothalamus/embryology , Adolescent , Young Adult , Brain Mapping/methods , Neural Pathways/physiology
2.
Cell Rep ; 38(3): 110251, 2022 01 18.
Article in English | MEDLINE | ID: mdl-35045288

ABSTRACT

The hypothalamus regulates many innate behaviors, but its development remains poorly understood. Here, we used single-cell RNA sequencing (RNA-seq) and hybridization chain reaction (HCR) to profile multiple stages of early hypothalamic development in the chick. Hypothalamic neuroepithelial cells are initially induced from prethalamic-like cells. Two distinct hypothalamic progenitor populations then emerge and give rise to tuberal and mammillary/paraventricular hypothalamic cells. At later stages, the regional organization of the chick and mouse hypothalamus is highly similar. We identify selective markers for major subdivisions of the developing chick hypothalamus and many previously uncharacterized candidate regulators of hypothalamic induction, regionalization, and neurogenesis. As proof of concept for the power of the dataset, we demonstrate that prethalamus-derived follistatin inhibits hypothalamic induction. This study clarifies the organization of the nascent hypothalamus and identifies molecular mechanisms that may control its induction and subsequent development.


Subject(s)
Hypothalamus/embryology , Neural Stem Cells/cytology , Neurogenesis/physiology , Animals , Chick Embryo , RNA-Seq , Single-Cell Analysis
3.
Nat Commun ; 12(1): 6749, 2021 11 19.
Article in English | MEDLINE | ID: mdl-34799566

ABSTRACT

The hypothalamus regulates metabolic homeostasis by influencing behavior and endocrine systems. Given its role governing key traits, such as body weight and reproductive timing, understanding the genetic regulation of hypothalamic development and function could yield insights into disease pathogenesis. However, given its inaccessibility, studying human hypothalamic gene regulation has proven challenging. To address this gap, we generate a high-resolution chromatin architecture atlas of an established embryonic stem cell derived hypothalamic-like neuron model across three stages of in vitro differentiation. We profile accessible chromatin and identify physical contacts between gene promoters and putative cis-regulatory elements to characterize global regulatory landscape changes during hypothalamic differentiation. Next, we integrate these data with GWAS loci for various complex traits, identifying multiple candidate effector genes. Our results reveal common target genes for these traits, potentially affecting core developmental pathways. Our atlas will enable future efforts to determine hypothalamic mechanisms influencing disease susceptibility.


Subject(s)
Gene Expression Regulation, Developmental , Gene Regulatory Networks , Human Embryonic Stem Cells/physiology , Hypothalamus/embryology , Neurons/physiology , Cell Differentiation/genetics , Cell Line , Chromosome Mapping , Genome-Wide Association Study , Humans , Hypothalamus/cytology , Multifactorial Inheritance , RNA-Seq , Regulatory Elements, Transcriptional/genetics
4.
Nutrients ; 13(9)2021 Sep 18.
Article in English | MEDLINE | ID: mdl-34579137

ABSTRACT

We studied whether myo-inositol supplementation throughout lactation, alone and combined with leptin, may reverse detrimental effects on hypothalamic structure and function caused by gestational calorie gestation (CR) in rats. Candidate early transcript-based biomarkers of metabolic health in peripheral blood mononuclear cells (PBMC) were also studied. Offspring of dams exposed to 25% gestational CR and supplemented during lactation with physiological doses of leptin (CR-L), myo-inositol (CR-M), the combination (CR-LM), or the vehicle (CR-V) as well as control rats (CON-V) were followed and sacrificed at postnatal day 25. Myo-inositol and the combination increased the number of neurons in arcuate nucleus (ARC) (only in females) and paraventricular nucleus, and myo-inositol (alone) restored the number of αMSH+ neurons in ARC. Hypothalamic mRNA levels of Lepr in CR-M and Insr in CR-M and CR-LM males were higher than in CR-V and CON-V, respectively. In PBMC, increased expression levels of Lrp11 and Gls in CR-V were partially normalized in all supplemented groups (but only in males for Gls). Therefore, myo-inositol supplementation throughout lactation, alone and combined with leptin, reverts programmed alterations by fetal undernutrition on hypothalamic structure and gene expression of potential early biomarkers of metabolic health in PBMC, which might be attributed, in part, to increased leptin sensitivity.


Subject(s)
Animals, Suckling/physiology , Caloric Restriction/adverse effects , Hypothalamus/embryology , Inositol/administration & dosage , Prenatal Exposure Delayed Effects , Animals , Dietary Supplements , Female , Hypothalamus/chemistry , Hypothalamus/cytology , Lactation/physiology , Leptin , Leukocytes, Mononuclear/chemistry , Male , Pregnancy , RNA, Messenger/analysis , Rats , Rats, Wistar , Receptors, Leptin/genetics
5.
Brain Struct Funct ; 226(3): 759-785, 2021 Apr.
Article in English | MEDLINE | ID: mdl-33544184

ABSTRACT

The prethalamic eminence (PThE), a diencephalic caudal neighbor of the telencephalon and alar hypothalamus, is frequently described in mammals and birds as a transient embryonic structure, undetectable in the adult brain. Based on descriptive developmental analysis of Tbr1 gene brain expression in chick embryos, we previously reported that three migratory cellular streams exit the PThE rostralward, targeting multiple sites in the hypothalamus, subpallium and septocommissural area, where eminential cells form distinct nuclei or disperse populations. These conclusions needed experimental corroboration. In this work, we used the homotopic quail-chick chimeric grafting procedure at stages HH10/HH11 to demonstrate by fate-mapping the three predicted tangential migration streams. Some chimeric brains were processed for Tbr1 in situ hybridization, for correlation with our previous approach. Evidence supporting all three postulated migration streams is presented. The results suggested a slight heterochrony among the juxtapeduncular (first), the peripeduncular (next), and the eminentio-septal (last) streams, each of which followed differential routes. A possible effect of such heterochrony on the differential selection of medial to lateral habenular hodologic targets by the migrated neurons is discussed.


Subject(s)
Hypothalamus/embryology , Neurons/cytology , Quail/embryology , Telencephalon/metabolism , Animals , Cell Differentiation/physiology , Cell Movement/physiology , Chick Embryo , Chickens , Diencephalon/embryology
6.
Neuron ; 109(7): 1150-1167.e6, 2021 04 07.
Article in English | MEDLINE | ID: mdl-33600763

ABSTRACT

The hypothalamus plays crucial roles in regulating endocrine, autonomic, and behavioral functions via its diverse nuclei and neuronal subtypes. The developmental mechanisms underlying ontogenetic establishment of different hypothalamic nuclei and generation of neuronal diversity remain largely unknown. Here, we show that combinatorial T-box 3 (TBX3), orthopedia homeobox (OTP), and distal-less homeobox (DLX) expression delineates all arcuate nucleus (Arc) neurons and defines four distinct subpopulations, whereas combinatorial NKX2.1/SF1 and OTP/DLX expression identifies ventromedial hypothalamus (VMH) and tuberal nucleus (TuN) neuronal subpopulations, respectively. Developmental analysis indicates that all four Arc subpopulations are mosaically and simultaneously generated from embryonic Arc progenitors, whereas glutamatergic VMH neurons and GABAergic TuN neurons are sequentially generated from common embryonic VMH progenitors. Moreover, clonal lineage-tracing analysis reveals that diverse lineages from multipotent radial glia progenitors orchestrate Arc and VMH-TuN establishment. Together, our study reveals cellular mechanisms underlying generation and organization of diverse neuronal subtypes and ontogenetic establishment of individual nuclei in the mammalian hypothalamus.


Subject(s)
Hypothalamus/cytology , Hypothalamus/growth & development , Neurons/physiology , Animals , Animals, Genetically Modified , Arcuate Nucleus of Hypothalamus/cytology , Arcuate Nucleus of Hypothalamus/embryology , Cell Lineage , Glutamic Acid/physiology , Homeodomain Proteins/metabolism , Hypothalamus/embryology , Mice , Mice, Inbred C57BL , Nerve Tissue Proteins/metabolism , Neuroglia/physiology , Stem Cells/physiology , T-Box Domain Proteins/metabolism , Transcription Factors/metabolism , Ventromedial Hypothalamic Nucleus/cytology , Ventromedial Hypothalamic Nucleus/embryology , Ventromedial Hypothalamic Nucleus/metabolism , gamma-Aminobutyric Acid/physiology
7.
Nat Commun ; 12(1): 256, 2021 01 11.
Article in English | MEDLINE | ID: mdl-33431871

ABSTRACT

In humans, inactivating mutations in MLL4, which encodes a histone H3-lysine 4-methyltransferase, lead to Kabuki syndrome (KS). While dwarfism is a cardinal feature of KS, the underlying etiology remains unclear. Here we report that Mll4 regulates the development of growth hormone-releasing hormone (GHRH)-producing neurons in the mouse hypothalamus. Our two Mll4 mutant mouse models exhibit dwarfism phenotype and impairment of the developmental programs for GHRH-neurons. Our ChIP-seq analysis reveals that, in the developing mouse hypothalamus, Mll4 interacts with the transcription factor Nrf1 to trigger the expression of GHRH-neuronal genes. Interestingly, the deficiency of Mll4 results in a marked reduction of histone marks of active transcription, while treatment with the histone deacetylase inhibitor AR-42 rescues the histone mark signature and restores GHRH-neuronal production in Mll4 mutant mice. Our results suggest that the developmental dysregulation of Mll4-directed epigenetic control of transcription plays a role in the development of GHRH-neurons and dwarfism phenotype in mice.


Subject(s)
Growth Hormone-Releasing Hormone/biosynthesis , Histone-Lysine N-Methyltransferase/metabolism , Hypothalamus/cytology , Neurons/metabolism , Animals , Base Sequence , Dwarfism/metabolism , Embryo, Mammalian/metabolism , Epigenesis, Genetic , Gene Expression Regulation, Developmental , HEK293 Cells , Humans , Hypothalamus/embryology , Male , Mice, Knockout , Models, Biological , Nuclear Respiratory Factor 1/metabolism , Phenylbutyrates/pharmacology , Transcription Factors/metabolism
8.
Cell Rep ; 34(1): 108587, 2021 01 05.
Article in English | MEDLINE | ID: mdl-33406432

ABSTRACT

Despite a growing appreciation for microglial influences on the developing brain, the responsiveness of microglia to insults during gestation remains less well characterized, especially in the embryo when microglia themselves are still maturing. Here, we asked if fetal microglia could coordinate an innate immune response to an exogenous insult. Using time-lapse imaging, we showed that hypothalamic microglia actively surveyed their environment by near-constant "touching" of radial glia projections. However, following an insult (i.e., IUE or AAV transduction), this seemingly passive touching became more intimate and long lasting, ultimately resulting in the retraction of radial glial projections and degeneration into small pieces. Mechanistically, the TAM receptors MERTK and AXL were upregulated in microglia following the insult, and Annexin V treatment inhibited radial glia breakage and engulfment by microglia. These data demonstrate a remarkable responsiveness of embryonic microglia to insults during gestation, a critical window for neurodevelopment.


Subject(s)
Embryo, Mammalian/metabolism , Ependymoglial Cells/physiology , Hypothalamus/embryology , Hypothalamus/physiology , Microglia/physiology , Proto-Oncogene Proteins/metabolism , Receptor Protein-Tyrosine Kinases/metabolism , c-Mer Tyrosine Kinase/metabolism , Animals , Brain/embryology , Embryonic Development , Gene Expression Regulation, Developmental , Immunity, Innate , Mice , Mice, Transgenic , Optical Imaging/methods , Axl Receptor Tyrosine Kinase
9.
Sci China Life Sci ; 64(3): 419-433, 2021 Mar.
Article in English | MEDLINE | ID: mdl-32803714

ABSTRACT

Cenpj is a centrosomal protein located at the centrosomes and the base of cilia, it plays essential roles in regulating neurogenesis and cerebral cortex development. Although centrosomal and cilium dysfunction are one of the causes of obesity, insulin resistance, and type 2 diabetes, the role that Cenpj plays in the regulation of body weight remains unclear. Here, we deleted Cenpj by crossing Cenpjflox/flox mice with Nkx2.1-Cre mice. Loss of the centrosomal protein Cenpj in Nkx2.1-expressing cells causes morbid obesity in mice at approximately 4 months of age with expended brain ventricles but no change of brain size. We found that hypothalamic cells exhibited reduced proliferation and increased apoptosis upon Cenpj depletion at the embryonic stages, resulting in a dramatic decrease in the number of Proopiomelanocortin (POMC) neurons and electrophysiological dysfunction of NPY neurons in the arcuate nucleus (ARC) in adults. Furthermore, depletion of Cenpj also reduced the neuronal projection from the ARC to the paraventricular nucleus (PVN), with decreased melanocortin-4 receptors (MC4R) expression in PVN neurons. The study defines the roles that Cenpj plays in regulating hypothalamus development and body weight, providing a foundation for further understanding of the pathological mechanisms of related diseases.


Subject(s)
Gene Knockdown Techniques , Hypothalamus/physiopathology , Microtubule-Associated Proteins/genetics , Obesity, Morbid/physiopathology , Animals , Apoptosis , Cell Line , Cell Proliferation , Hypothalamus/embryology , Hypothalamus/metabolism , Mice , Neurons/metabolism , Obesity, Morbid/genetics , Pro-Opiomelanocortin/metabolism , Receptor, Melanocortin, Type 4/metabolism , Thyroid Nuclear Factor 1/genetics
10.
Nat Commun ; 11(1): 5772, 2020 11 13.
Article in English | MEDLINE | ID: mdl-33188191

ABSTRACT

Hypothalamic neurons including proopiomelanocortin (POMC)-producing neurons regulate body weights. The non-motile primary cilium is a critical sensory organelle on the cell surface. An association between ciliary defects and obesity has been suggested, but the underlying mechanisms are not fully understood. Here we show that inhibition of ciliogenesis in POMC-expressing developing hypothalamic neurons, by depleting ciliogenic genes IFT88 and KIF3A, leads to adulthood obesity in mice. In contrast, adult-onset ciliary dysgenesis in POMC neurons causes no significant change in adiposity. In developing POMC neurons, abnormal cilia formation disrupts axonal projections through impaired lysosomal protein degradation. Notably, maternal nutrition and postnatal leptin surge have a profound impact on ciliogenesis in the hypothalamus of neonatal mice; through these effects they critically modulate the organization of hypothalamic feeding circuits. Our findings reveal a mechanism of early life programming of adult adiposity, which is mediated by primary cilia in developing hypothalamic neurons.


Subject(s)
Adiposity , Cilia/metabolism , Hypothalamus/embryology , Hypothalamus/metabolism , Lysosomes/metabolism , Animals , Animals, Newborn , Arcuate Nucleus of Hypothalamus/metabolism , Axons/metabolism , Energy Metabolism , Female , Glucose/metabolism , Leptin/metabolism , Malnutrition/pathology , Mice, Inbred C57BL , Microtubule-Associated Proteins/metabolism , Neurogenesis , Obesity/metabolism , Obesity/pathology , Organogenesis , Pro-Opiomelanocortin/metabolism , Proteolysis
11.
Med Sci Monit ; 26: e927334, 2020 Nov 16.
Article in English | MEDLINE | ID: mdl-33191393

ABSTRACT

BACKGROUND This study utilized CRISPR/Cas9 gene editing technology to construct a Mex3c gene-deficient mouse model, and studied C-FOS expression in hypothalamic nuclei. MATERIAL AND METHODS Thirty Mex3c-/+ mice, 30 mice in the normal group, and 30 Mex3c-/+ mice were randomly divided into control, leptin, and ghrelin groups according to different intraperitoneal injections. HE and Nissl staining were performed to observe the morphology of hypothalamic nerve cells. The C-FOS expression in hypothalamic nuclei of each group was analyzed by immunohistochemical techniques. HE staining was used to observe neural tube morphology, and LFB staining was used to observe nerve myelin sheath morphology. TEM was used to observe neuronal ultrastructure and immunohistochemical techniques were utilized to analyze nestin expression. RESULTS C-FOS expression was lower in the normal control group than in the leptin and ghrelin groups. The Mex3c control group and the leptin group had higher C-FOS expression than the ghrelin group. In neural tube studies, no significant differences were found in the neural tube pathological sections of E14.5-day embryos in each group. Nestin results demonstrated lower expression in the normal group and there was little difference between the HD and Mex3c groups. CONCLUSIONS Mex3c appears to participate in the regulation of energy metabolism by inducing C-FOS expression in the hypothalamus. The neural tubes of the offspring of Mex3c-/+ mice had defects during development.


Subject(s)
Cell Nucleus/metabolism , Embryo, Mammalian/anatomy & histology , Hypothalamus/embryology , Neural Tube/anatomy & histology , Neural Tube/embryology , Proto-Oncogene Proteins c-fos/metabolism , RNA-Binding Proteins/genetics , Animals , Genotype , Hypothalamus/anatomy & histology , Mice, Knockout , Models, Animal , RNA-Binding Proteins/metabolism
12.
Alcohol Clin Exp Res ; 44(12): 2519-2535, 2020 12.
Article in English | MEDLINE | ID: mdl-33067812

ABSTRACT

BACKGROUND: Embryonic exposure to ethanol (EtOH) produces marked disturbances in neuronal development and alcohol-related behaviors, with low-moderate EtOH doses stimulating neurogenesis without producing apoptosis and high doses having major cytotoxic effects while causing gross morphological abnormalities. With the pro-inflammatory chemokine system, Cxcl12, and its main receptor Cxcr4, known to promote processes of neurogenesis, we examined here this neuroimmune system in the embryonic hypothalamus to test directly if it mediates the stimulatory effects low-moderate EtOH doses have on neuronal development. METHODS: We used the zebrafish (Danio rerio) model, which develops externally and allows one to investigate the developing brain in vivo with precise control of dose and timing of EtOH delivery in the absence of maternal influence. Zebrafish were exposed to low-moderate EtOH doses (0.1, 0.25, 0.5% v/v), specifically during a period of peak hypothalamic development from 22 to 24 hours postfertilization, and in some tests were pretreated from 2 to 22 hpf with the Cxcr4 receptor antagonist, AMD3100. Measurements in the hypothalamus at 26 hpf were taken of cxcl12a and cxcr4b transcription, signaling, and neuronal density using qRT-PCR, RNAscope, and live imaging of transgenic zebrafish. RESULTS: Embryonic EtOH exposure, particularly at the 0.5% dose, significantly increased levels of cxcl12a and cxcr4b mRNA in whole embryos, number of cxcl12a and cxcr4b transcripts in developing hypothalamus, and internalization of Cxcr4b receptors in hypothalamic cells. Embryonic EtOH also caused an increase in the number of hypothalamic neurons and coexpression of cxcl12a and cxcr4b transcripts within these neurons. Each of these stimulatory effects of EtOH in the embryo was blocked by pretreatment with the Cxcr4 antagonist AMD3100. CONCLUSIONS: These results provide clear evidence that EtOH's stimulatory effects at low-moderate doses on the number of hypothalamic neurons early in development are mediated, in part, by increased transcription and intracellular activation of this chemokine system, likely due to autocrine signaling of Cxcl12a at its Cxcr4b receptor within the neurons.


Subject(s)
Chemokine CXCL12/metabolism , Ethanol/pharmacology , Hypothalamus/drug effects , Neurons/drug effects , Receptors, CXCR4/metabolism , Zebrafish Proteins/metabolism , Animals , Benzylamines/pharmacology , Cell Count , Cyclams/pharmacology , Embryo, Nonmammalian/drug effects , Hypothalamus/cytology , Hypothalamus/embryology , Neurogenesis/drug effects , Zebrafish/embryology
13.
Nat Commun ; 11(1): 4360, 2020 08 31.
Article in English | MEDLINE | ID: mdl-32868762

ABSTRACT

The hypothalamus is a central regulator of many innate behaviors essential for survival, but the molecular mechanisms controlling hypothalamic patterning and cell fate specification are poorly understood. To identify genes that control hypothalamic development, we have used single-cell RNA sequencing (scRNA-Seq) to profile mouse hypothalamic gene expression across 12 developmental time points between embryonic day 10 and postnatal day 45. This identified genes that delineated clear developmental trajectories for all major hypothalamic cell types, and readily distinguished major regional subdivisions of the developing hypothalamus. By using our developmental dataset, we were able to rapidly annotate previously unidentified clusters from existing scRNA-Seq datasets collected during development and to identify the developmental origins of major neuronal populations of the ventromedial hypothalamus. We further show that our approach can rapidly and comprehensively characterize mutants that have altered hypothalamic patterning, identifying Nkx2.1 as a negative regulator of prethalamic identity. These data serve as a resource for further studies of hypothalamic development, physiology, and dysfunction.


Subject(s)
Cell Differentiation , Hypothalamus , Neurons/metabolism , Thyroid Nuclear Factor 1/metabolism , Animals , Base Sequence , Body Patterning , Gene Expression Regulation, Developmental , Hypothalamus/cytology , Hypothalamus/embryology , Hypothalamus/growth & development , Hypothalamus/metabolism , Mice , Mutation , Single-Cell Analysis , Thyroid Nuclear Factor 1/genetics
14.
Mol Metab ; 42: 101079, 2020 12.
Article in English | MEDLINE | ID: mdl-32919096

ABSTRACT

OBJECTIVE: Perinatal exposure to maternal obesity results in predisposition of offspring to develop obesity later in life. Increased weight gain in offspring exposed to maternal obesity is usually associated with hyperphagia, implicating altered central regulation of food intake as a cause. We aimed to define how maternal obesity impacts early development of the hypothalamus to program lasting dysfunction in feeding regulatory pathways. METHODS: Mice offspring of diet-induced obese mothers were compared to the offspring of lean control mothers. We analysed gene expression in the fetal hypothalamus, alongside neurosphere assays to investigate the effects of maternal obesity on neural progenitor cell proliferation in vitro. Western blotting was used to investigate the insulin signalling pathway in the fetal hypothalamus. Characterisation of cell type and neuropeptide profile in adulthood was linked with analyses of feeding behaviour. RESULTS: There was a reduction in the expression of proliferative genes in the fetal hypothalamus of offspring exposed to maternal obesity. This reduction in proliferation was maintained in vitro when hypothalamic neural progenitor cells were grown as neurospheres. Hypothalamic fetal gene expression and neurosphere growth correlated with maternal body weight and insulin levels. Foetuses of obese mothers showed hypothalamic insulin resistance, which may be causative of reduced proliferation. Furthermore, maternal obesity activated the Notch signalling pathway in neonatal offspring hypothalamus, resulting in decreased neurogenesis. Adult offspring of obese mothers displayed an altered ratio of anorexigenic and orexigenic signals in the arcuate nucleus, associated with an inability to maintain energy homeostasis when metabolically challenged. CONCLUSIONS: These findings show that maternal obesity alters the molecular signature in the developing hypothalamus, which is associated with disrupted growth and development of hypothalamic precursor cells and defective feeding regulation in adulthood. This is the first report of fetal hypothalamic insulin resistance in an obese pregnancy and suggests a mechanism by which maternal obesity causes permanent changes to hypothalamic structure and function.


Subject(s)
Hypothalamus/embryology , Insulin Resistance/physiology , Obesity, Maternal/physiopathology , Animals , Arcuate Nucleus of Hypothalamus/metabolism , Body Weight , Brain/metabolism , Diet, High-Fat , Feeding Behavior , Female , Fetus/metabolism , Fetus/physiopathology , Gene Expression/genetics , Gene Expression Regulation, Developmental/genetics , Hypothalamus/metabolism , Insulin/metabolism , Male , Maternal-Fetal Exchange/physiology , Mice , Mice, Inbred C57BL , Neural Stem Cells/metabolism , Neurons/metabolism , Obesity/metabolism , Obesity/physiopathology , Obesity, Maternal/metabolism , Pregnancy , Prenatal Exposure Delayed Effects/metabolism , Weight Gain
15.
Mol Cell Endocrinol ; 518: 111030, 2020 12 01.
Article in English | MEDLINE | ID: mdl-32931849

ABSTRACT

Gonadotropin-releasing hormone (GnRH) neurons control mammalian reproduction and migrate from their birthplace in the nasal placode to the hypothalamus during development. Despite much work on the origin and migration of GnRH neurons, the processes that control GnRH lineage formation are not fully understood. Here, we demonstrate that Nhlh genes control vomeronasal receptor expression in the developing murine olfactory placode associated with the generation of the first GnRH neurons at embryonic days (E)10-12. Inactivation of ß2-microglobulin (ß2-m), which selectively affects surface expression of V2Rs, dramatically decreased the number of GnRH neurons in the Nhlh2 mutant background, preventing rescue of fertility in female Nhlh2 mutant mice by male pheromones. In addition, we show that GnRH neurons generated after E12 fail to establish synaptic connections to the vomeronasal amygdala, suggesting the existence of functionally specialized subpopulations of GnRH neurons, which process pheromonal information.


Subject(s)
Gonadotropin-Releasing Hormone/metabolism , Neurogenesis/genetics , Neurons/metabolism , Receptors, Pheromone/genetics , Animals , Basic Helix-Loop-Helix Transcription Factors/genetics , Cells, Cultured , Chemotactic Factors/genetics , Embryo, Mammalian , Female , Gene Expression Regulation, Developmental , Hypothalamus/embryology , Hypothalamus/metabolism , Male , Mice , Mice, Knockout , Neurons/physiology , Pheromones/metabolism , Pregnancy , Receptors, Pheromone/metabolism
16.
J Neuroinflammation ; 17(1): 207, 2020 Jul 10.
Article in English | MEDLINE | ID: mdl-32650794

ABSTRACT

BACKGROUND: Clinical and animal studies show that alcohol consumption during pregnancy produces lasting behavioral disturbances in offspring, including increased alcohol drinking, which are linked to inflammation in the brain and disturbances in neurochemical systems that promote these behaviors. These include the neuropeptide, melanin-concentrating hormone (MCH), which is mostly expressed in the lateral hypothalamus (LH). Maternal ethanol administration at low-to-moderate doses, while stimulating MCH neurons without affecting apoptosis or gliogenesis, increases in LH the density of neurons expressing the inflammatory chemokine C-C motif ligand 2 (CCL2) and its receptor CCR2 and their colocalization with MCH. These neural effects associated with behavioral changes are reproduced by maternal CCL2 administration, reversed by a CCR2 antagonist, and consistently stronger in females than males. The present study investigates in the embryo the developmental origins of this CCL2/CCR2-mediated stimulatory effect of maternal ethanol exposure on MCH neurons. METHODS: Pregnant rats from embryonic day 10 (E10) to E15 during peak neurogenesis were orally administered ethanol at a moderate dose (2 g/kg/day) or peripherally injected with CCL2 or CCR2 antagonist to test this neuroimmune system's role in ethanol's actions. Using real-time quantitative PCR, immunofluorescence histochemistry, in situ hybridization, and confocal microscopy, we examined in embryos at E19 the CCL2/CCR2 system and MCH neurons in relation to radial glia progenitor cells in the hypothalamic neuroepithelium where neurons are born and radial glia processes projecting laterally through the medial hypothalamus that provide scaffolds for neuronal migration into LH. RESULTS: We demonstrate that maternal ethanol increases radial glia cell density and their processes while stimulating the CCL2/CCR2 system and these effects are mimicked by maternal administration of CCL2 and blocked by a CCR2 antagonist. While stimulating CCL2 colocalization with radial glia and neurons but not microglia, ethanol increases MCH neuronal number near radial glia cells and making contact along their processes projecting into LH. Further tests identify the CCL2/CCR2 system in NEP as a primary source of ethanol's sexually dimorphic actions. CONCLUSIONS: These findings provide new evidence for how an inflammatory chemokine pathway functions within neuroprogenitor cells to mediate ethanol's long-lasting, stimulatory effects on peptide neurons linked to adolescent drinking behavior.


Subject(s)
Chemokine CCL2/metabolism , Ethanol/toxicity , Hypothalamus/metabolism , Neuroepithelial Cells/metabolism , Receptors, CCR2/metabolism , Sex Characteristics , Animals , Embryonic Development/drug effects , Embryonic Development/physiology , Ethanol/administration & dosage , Female , Hypothalamus/drug effects , Hypothalamus/embryology , Male , Neuroepithelial Cells/drug effects , Neuroglia/drug effects , Neuroglia/metabolism , Neurons/drug effects , Neurons/metabolism , Pregnancy , Rats , Rats, Sprague-Dawley , Stem Cells/drug effects , Stem Cells/metabolism
17.
Nature ; 582(7811): 246-252, 2020 06.
Article in English | MEDLINE | ID: mdl-32499648

ABSTRACT

A wealth of specialized neuroendocrine command systems intercalated within the hypothalamus control the most fundamental physiological needs in vertebrates1,2. Nevertheless, we lack a developmental blueprint that integrates the molecular determinants of neuronal and glial diversity along temporal and spatial scales of hypothalamus development3. Here we combine single-cell RNA sequencing of 51,199 mouse cells of ectodermal origin, gene regulatory network (GRN) screens in conjunction with genome-wide association study-based disease phenotyping, and genetic lineage reconstruction to show that nine glial and thirty-three neuronal subtypes are generated by mid-gestation under the control of distinct GRNs. Combinatorial molecular codes that arise from neurotransmitters, neuropeptides and transcription factors are minimally required to decode the taxonomical hierarchy of hypothalamic neurons. The differentiation of γ-aminobutyric acid (GABA) and dopamine neurons, but not glutamate neurons, relies on quasi-stable intermediate states, with a pool of GABA progenitors giving rise to dopamine cells4. We found an unexpected abundance of chemotropic proliferation and guidance cues that are commonly implicated in dorsal (cortical) patterning5 in the hypothalamus. In particular, loss of SLIT-ROBO signalling impaired both the production and positioning of periventricular dopamine neurons. Overall, we identify molecular principles that shape the developmental architecture of the hypothalamus and show how neuronal heterogeneity is transformed into a multimodal neural unit to provide virtually infinite adaptive potential throughout life.


Subject(s)
Gene Expression Regulation, Developmental , Hypothalamus/cytology , Hypothalamus/embryology , Morphogenesis , Animals , Cell Differentiation , Cell Lineage , Dopamine/metabolism , Dopaminergic Neurons/cytology , Dopaminergic Neurons/metabolism , Ectoderm/cytology , Ectoderm/metabolism , Female , GABAergic Neurons/cytology , GABAergic Neurons/metabolism , Gene Regulatory Networks , Genome-Wide Association Study , Glutamic Acid/metabolism , Hypothalamus/metabolism , Male , Mice , Morphogenesis/genetics , Nerve Tissue Proteins/metabolism , Neuroglia/cytology , Neuroglia/metabolism , Neuropeptides/metabolism , Neurotransmitter Agents/metabolism , Receptors, Immunologic/metabolism , Regulon/genetics , Signal Transduction , Transcription Factors/metabolism , gamma-Aminobutyric Acid/metabolism , Roundabout Proteins
18.
Gen Comp Endocrinol ; 295: 113524, 2020 09 01.
Article in English | MEDLINE | ID: mdl-32526331

ABSTRACT

Early life stages are sensitive to environmental insults and changes during critical developmental periods; this can often result in altered adult behaviour and physiology. Examining the development of the hypothalamus-pituitary-interrenal (HPI) axis and its responsiveness, or lack thereof, during development are important for understanding the short- and long-term impacts of stressors on embryonic and larval fish. We examined the ontogeny of the HPI axis in embryonic (21, 38, 63, 83 and 103 days post-fertilisation (dpf)) and larval (1, 2, 3 and 4 weeks post-hatch (wph)) lake whitefish (Coregonus clupeaformis) by quantifying changes in mRNA levels of several genes associated with HPI axis functioning and whole animal cortisol levels throughout development and in response to a severe or mild hypoxic stress. Cortisol, and crh, crhbp1, pomc and star transcripts were detected from the earliest embryonic age studied. Cortisol levels in control embryos decreased between 21 and 63 dpf, suggesting the utilisation of maternal cortisol deposits. However, by 83 dpf (70% developed) endogenous de novo synthesis had generated a 4.5-fold increase in whole embryo cortisol. Importantly, we provide novel data showing that the HPI axis can be activated even earlier. Whole body cortisol increased in eyed lake whitefish embryos (38 dpf; ~32% developed) in response to hypoxia stress. Coincident with this hypoxia-induced increase in cortisol in 38 dpf embryos were corresponding increases in crh, crhbp1, pomc and star transcript levels. Beyond 38 dpf, the HPI axis in lake whitefish embryos was hyporesponsive to hypoxia stress at all embryonic ages examined (63, 83 and 103 dpf; 54, 72 and 85% developed, respectively). Post-hatch, larvae responded to hypoxia with an increase in cortisol levels and HPI axis genes at 1 wph, but this response was lost and larvae appeared hyporesponsive at subsequent ages (2, 3 and 4 wph). Collectively our work demonstrates that during fish embryogenesis and the larval stage there are windows where the HPI axis is responsive and windows where it is truly hyporesponsive; both could be beneficial in ensuring undisrupted development particularly in the face of increasing environmental changes.


Subject(s)
Hypothalamus/embryology , Hypoxia/embryology , Lakes , Pituitary Gland/embryology , Salmonidae/embryology , Animals , Embryo, Nonmammalian/metabolism , Embryonic Development , Hydrocortisone/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Salmonidae/genetics
19.
Nutrients ; 12(5)2020 May 19.
Article in English | MEDLINE | ID: mdl-32438566

ABSTRACT

Fetal brain development is closely dependent on maternal nutrition and metabolic status. Maternal protein restriction (PR) is known to be associated with alterations in the structure and function of the hypothalamus, leading to impaired control of energy homeostasis and food intake. The objective of this study was to identify the cellular and molecular systems underlying these effects during fetal development. We combined a global transcriptomic analysis on the fetal hypothalamus from a rat model of maternal PR with in vitro neurosphere culture and cellular analyses. Several genes encoding proteins from the mitochondrial respiratory chain complexes were overexpressed in the PR group and mitochondrial metabolic activity in the fetal hypothalamus was altered. The level of the N6-methyladenosine epitranscriptomic mark was reduced in the PR fetuses, and the expression of several genes involved in the writing/erasing/reading of this mark was indeed altered, as well as genes encoding several RNA-binding proteins. Additionally, we observed a higher number of neuronal-committed progenitors at embryonic day 17 (E17) in the PR fetuses. Together, these data strongly suggest a metabolic adaptation to the amino acid shortage, combined with the post-transcriptional control of protein expression, which might reflect alterations in the control of the timing of neuronal progenitor differentiation.


Subject(s)
Diet, Protein-Restricted/adverse effects , Fetus/metabolism , Hypothalamus/embryology , Maternal Nutritional Physiological Phenomena/genetics , Mitochondria/genetics , Animals , Female , Fetal Development/genetics , Hypothalamus/metabolism , Pregnancy , Rats
20.
J Neuroinflammation ; 17(1): 146, 2020 May 06.
Article in English | MEDLINE | ID: mdl-32375817

ABSTRACT

BACKGROUND: Although historically microglia were thought to be immature in the fetal brain, evidence of purposeful interactions between these immune cells and nearby neural progenitors is becoming established. Here, we examined the influence of embryonic microglia on gliogenesis within the developing tuberal hypothalamus, a region later important for energy balance, reproduction, and thermoregulation. METHODS: We used immunohistochemistry to quantify the location and numbers of glial cells in the embryonic brain (E13.5-E17.5), as well as a pharmacological approach (i.e., PLX5622) to knock down fetal microglia. We also conducted cytokine and chemokine analyses on embryonic brains in the presence or absence of microglia, and a neurosphere assay to test the effects of the altered cytokines on hypothalamic progenitor behaviors. RESULTS: We identified a subpopulation of activated microglia that congregated adjacent to the third ventricle alongside embryonic Olig2+ neural progenitor cells (NPCs) that are destined to give rise to oligodendrocyte and astrocyte populations. In the absence of microglia, we observed an increase in Olig2+ glial progenitor cells that remained at the ventricle by E17.5 and a concomitant decrease of these Olig2+ cells in the mantle zone, indicative of a delay in migration of these precursor cells. A further examination of maturing oligodendrocytes in the hypothalamic grey and white matter area in the absence of microglia revealed migrating oligodendrocyte progenitor cells (OPCs) within the grey matter at E17.5, a time point when OPCs begin to slow their migration. Finally, quantification of cytokine and chemokine signaling in ex vivo E15.5 hypothalamic cultures +/- microglia revealed decreases in the protein levels of several cytokines in the absence of microglia. We assayed the influence of two downregulated cytokines (CCL2 and CXCL10) on neurosphere-forming capacity and lineage commitment of hypothalamic NPCs in culture and showed an increase in NPC proliferation as well as neuronal and oligodendrocyte differentiation. CONCLUSION: These data demonstrate that microglia influence gliogenesis in the developing tuberal hypothalamus.


Subject(s)
Astrocytes/cytology , Hypothalamus/cytology , Hypothalamus/embryology , Microglia/cytology , Oligodendroglia/cytology , Animals , Cell Differentiation/physiology , Mice , Neural Stem Cells/cytology
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