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1.
Brain Commun ; 2(2): fcaa105, 2020.
Article in English | MEDLINE | ID: mdl-32954345

ABSTRACT

Administration of recombinant glial cell line-derived neurotrophic factor into the putamen has been tested in preclinical and clinical studies to evaluate its neuroprotective effects on the progressive dopaminergic neuronal degeneration that characterizes Parkinson's disease. However, intracerebral glial cell line-derived neurotrophic factor infusion is a challenging therapeutic strategy, with numerous potential technical and medical limitations. Most of these limitations could be avoided if the production of endogenous glial cell line-derived neurotrophic factor could be increased. Glial cell line-derived neurotrophic factor is naturally produced in the striatum from where it exerts a trophic action on the nigrostriatal dopaminergic pathway. Most of striatal glial cell line-derived neurotrophic factor is synthesized by a subset of GABAergic interneurons characterized by the expression of parvalbumin. We sought to identify molecular targets specific to those neurons and which are putatively associated with glial cell line-derived neurotrophic factor synthesis. To this end, the transcriptomic differences between glial cell line-derived neurotrophic factor-positive parvalbumin neurons in the striatum and parvalbumin neurons located in the nearby cortex, which do not express glial cell line-derived neurotrophic factor, were analysed. Using mouse reporter models, we have defined the genomic signature of striatal parvalbumin interneurons obtained by fluorescence-activated cell sorting followed by microarray comparison. Short-listed genes were validated by additional histological and molecular analyses. These genes code for membrane receptors (Kit, Gpr83, Tacr1, Tacr3, Mc3r), cytosolic proteins (Pde3a, Crabp1, Rarres2, Moxd1) and a transcription factor (Lhx8). We also found the proto-oncogene cKit to be highly specific of parvalbumin interneurons in the non-human primate striatum, thus highlighting a conserved expression between species and suggesting that specific genes identified in mouse parvalbumin neurons could be putative targets in the human brain. Pharmacological stimulation of four G-protein-coupled receptors enriched in the striatal parvalbumin interneurons inhibited Gdnf expression presumably by decreasing cyclic adenosine monophosphate formation. Additional experiments with pharmacological modulators of adenylyl cyclase and protein kinase A indicated that this pathway is a relevant intracellular route to induce Gdnf gene activation. This preclinical study is an important step in the ongoing development of a specific pro-endo-glial cell line-derived neurotrophic factor pharmacological strategy to treat Parkinson's disease.

2.
Mov Disord ; 35(4): 565-576, 2020 04.
Article in English | MEDLINE | ID: mdl-31930748

ABSTRACT

BACKGROUND: The glial cell line-derived neurotrophic factor has a potent neuroprotective action on mesencephalic dopamine neurons, which are progressively lost in Parkinson's disease. Intrastriatal administration of this factor is a promising therapy for Parkinson's disease. Glial cell line-derived neurotrophic factor is naturally produced in restricted cerebral regions, such as the striatum, septum, and thalamus; however, its effects in the adult brain remain under debate. OBJECTIVES: We sought to clarify the physiologic role of endogenous glial cell line-derived neurotrophic factor in the survival of catecholaminergic neurons of the substantia nigra pars compacta and the locus coeruleus in adult mice. METHODS: We used 2 new Cre recombinase-based mouse models to delete a floxed-glial cell line-derived neurotrophic factor gene. The first model had Cre expression in the parvalbumin expressing interneurons, as these cells represent the major source of striatal glial cell line-derived neurotrophic factor. The second model was an estrogen receptor 2-based inducible Cre triggered by tamoxifen at 2 months of age. RESULTS: We found that the floxed-glial cell line-derived neurotrophic factor gene was resilient to ablation by Cre-induced recombination and that parvalbumin-driven Cre was particularly inefficient to do so. The inducible-Cre model allowed an average 70% to 80% reduction in glial cell line-derived neurotrophic factor messenger ribonucleic acid and protein in striatum and septum with moderate significant loss of catecholamine neurons in the nigrostriatal pathway and, more markedly, in the locus coeruleus. This was accompanied with mild locomotor decline. CONCLUSIONS: Our data support qualitatively the view that brain glial cell line-derived neurotrophic factor is needed for the maintenance of adult central catecholaminergic neurons. © 2020 International Parkinson and Movement Disorder Society.


Subject(s)
Neuroprotective Agents , Parkinson Disease , Animals , Corpus Striatum , Glial Cell Line-Derived Neurotrophic Factor/genetics , Mice , Neostriatum , Neurons , Substantia Nigra
3.
Tissue Barriers ; 6(2): 1-22, 2018.
Article in English | MEDLINE | ID: mdl-29913111

ABSTRACT

There is emerging evidence that glial-derived neurotrophic factor (GDNF) is a potent inducer of restrictive barrier function in tight junction-forming microvascular endothelium and epithelium, including the human blood-nerve barrier (BNB) in vitro. We sought to determine the role of GDNF in restoring BNB function in vivo by evaluating sciatic nerve horseradish peroxidase (HRP) permeability in tamoxifen-inducible GDNF conditional knockout (CKO) adult mice following non-transecting crush injury via electron microscopy, with appropriate wildtype (WT) and heterozygous (HET) littermate controls. A total of 24 age-, genotype- and sex-matched mice >12 weeks of age were injected with 30 mg/kg HRP via tail vein injection 7 or 14 days following unilateral sciatic nerve crush, and both sciatic nerves were harvested 30 minutes later for morphometric assessment by light and electron microscopy. The number and percentage of HRP-permeable endoneurial microvessels were ascertained to determine the effect of GDNF in restoring barrier function in vivo. Following sciatic nerve crush, there was significant upregulation in GDNF protein expression in WT and HET mice that was abrogated in CKO mice. GDNF significantly restored sciatic nerve BNB HRP impermeability to near normal levels by day 7, with complete restoration seen by day 14 in WT and HET mice. A significant recovery lag was observed in CKO mice. This effect was independent on VE-Cadherin or claudin-5 expression on endoneurial microvessels. These results imply an important role of GDNF in restoring restrictive BNB function in vivo, suggesting a potential strategy to re-establish the restrictive endoneurial microenvironment following traumatic peripheral neuropathies.


Subject(s)
Blood-Nerve Barrier/metabolism , Glial Cell Line-Derived Neurotrophic Factor/metabolism , Peripheral Nervous System Diseases/pathology , Animals , Disease Models, Animal , Female , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Nerve Crush , Nerve Regeneration/physiology , Peripheral Nervous System Diseases/metabolism , Permeability , Recovery of Function/physiology
4.
PLoS One ; 13(1): e0192014, 2018.
Article in English | MEDLINE | ID: mdl-29370263

ABSTRACT

[This corrects the article DOI: 10.1371/journal.pone.0176821.].

5.
Neural Regen Res ; 12(11): 1799-1800, 2017 Nov.
Article in English | MEDLINE | ID: mdl-29239320
6.
PLoS One ; 12(5): e0176821, 2017.
Article in English | MEDLINE | ID: mdl-28464043

ABSTRACT

Kisspeptins regulate the mammalian reproductive axis by stimulating release of gonadotrophin releasing hormone (GnRH). Different length kisspeptins (KP) are found of 54, 14, 13 or 10 amino-acids which share a common C-terminal 10-amino acid sequence. KP-54 and KP-10 have been widely used to stimulate the reproductive axis but data suggest that KP-54 and KP-10 are not equally effective at eliciting reproductive hormone secretion after peripheral delivery. To confirm this, we analysed the effect of systemic administration of KP-54 or KP-10 on luteinizing hormone (LH) secretion into the bloodstream of male mice. Plasma LH measurements 10 min or 2 hours after kisspeptin injection showed that KP-54 can sustain LH release far longer than KP-10, suggesting a differential mode of action of the two peptides. To investigate the mechanism for this, we evaluated the pharmacokinetics of the two peptides in vivo and their potential to cross the blood brain barrier (BBB). We found that KP-54 has a half-life of ~32 min in the bloodstream, while KP-10 has a half-life of ~4 min. To compensate for this difference in half-life, we repeated injections of KP-10 every 10 min over 1 hr but failed to reproduce the sustained rise in LH observed after a single KP-54 injection, suggesting that the failure of KP-10 to sustain LH release may not just be related to peptide clearance. We tested the ability of peripherally administered KP-54 and KP-10 to activate c-FOS in GnRH neurons behind the blood brain barrier (BBB) and found that only KP-54 could do this. These data are consistent with KP-54 being able to cross the BBB and suggest that KP10 may be less able to do so.


Subject(s)
Central Nervous System Agents/pharmacology , Kisspeptins/pharmacology , Analysis of Variance , Animals , Blood-Brain Barrier/drug effects , Blood-Brain Barrier/metabolism , Capillary Permeability/drug effects , Capillary Permeability/physiology , Central Nervous System Agents/pharmacokinetics , Dose-Response Relationship, Drug , Enzyme-Linked Immunosorbent Assay , Humans , Hypothalamus/cytology , Hypothalamus/drug effects , Hypothalamus/metabolism , Immunohistochemistry , Kisspeptins/pharmacokinetics , Luteinizing Hormone/blood , Luteinizing Hormone/metabolism , Male , Mice, 129 Strain , Neurons/cytology , Neurons/drug effects , Neurons/metabolism , Proto-Oncogene Proteins c-fos/metabolism
7.
PLoS One ; 11(10): e0164391, 2016.
Article in English | MEDLINE | ID: mdl-27741271

ABSTRACT

Gender difference in Parkinson's disease (PD) suggests that female sex steroids may promote dopaminergic neuron survival and protect them from degeneration. The glial cell line-derived neurotrophic factor (GDNF) is believed to be dopaminotrophic; thus it is considered as a potential therapeutic target in PD. Additionally, GDNF is endogenously synthetized in the caudate/putamen of humans and striatum in rodents. A neuroprotective role of estrogens on the nigrostriatal pathway via the stimulation of GDNF has been proposed. Since the GDNF-producing parvalbumin (Parv) interneurons express the estrogen receptor alpha in the mouse striatum, we sought to determine whether ectopic estrogenic compound modulates the GDNF synthesis in mice. Using an ovariectomized-estradiol (E2) replacement regimen, which reliably generates a rise of plasma estradiol, we assessed the effects of different levels of E2 on the activation of striatal neuronal populations, and GDNF production. A strong correlation was found between plasma E2 and the expression of the immediate early gene cFos in the striatum, as well as in other cortical regions. However, moderate and high E2 treatments failed to induce any striatal GDNF mRNA and protein synthesis. High E2 only stimulates cFos induction in a low percentage of striatal Parv neurons whereas the majority of cFos-positive cells are medium spiny neurons. Activation of these projecting neurons by E2 suggests a role of circulating sex steroids in the modulation of striatal neural pathways.


Subject(s)
Estradiol/blood , Glial Cell Line-Derived Neurotrophic Factor/metabolism , Animals , Corpus Striatum/metabolism , Disease Models, Animal , Dopaminergic Neurons/metabolism , Estrogen Receptor alpha/metabolism , Female , Glial Cell Line-Derived Neurotrophic Factor/genetics , Immunoassay , Male , Mice , Mice, Inbred C57BL , Ovariectomy , Parkinson Disease/metabolism , Parkinson Disease/pathology , Proto-Oncogene Proteins c-fos/metabolism , RNA, Messenger/metabolism
8.
Hypoxia (Auckl) ; 3: 15-33, 2015.
Article in English | MEDLINE | ID: mdl-27774479

ABSTRACT

Chronic hypoxemia, as evidenced in de-acclimatized high-altitude residents or in patients with chronic obstructive respiratory disorders, is a common medical condition that can produce serious neurological alterations. However, the pathogenesis of this phenomenon is unknown. We have found that adult rodents exposed for several days/weeks to hypoxia, with an arterial oxygen tension similar to that of chronically hypoxemic patients, manifest a partially irreversible structural disarrangement of the subventricular neurogenic niche (subventricular zone) characterized by displacement of neurons and myelinated axons, flattening of the ependymal cell layer, and thinning of capillary walls. Despite these abnormalities, the number of neuronal and oligodendrocyte progenitors, neuroblasts, and neurosphere-forming cells as well as the proliferative activity in subventricular zone was unchanged. These results suggest that neural stem cells and their undifferentiated progeny are resistant to hypoxia. However, in vivo and in vitro experiments indicate that severe chronic hypoxia decreases the survival of newly generated neurons and oligodendrocytes, with damage of myelin sheaths. These findings help explain the effects of hypoxia on adult neurogenesis and provide new perspectives on brain responsiveness to persistent hypoxemia.

9.
Front Neuroanat ; 9: 10, 2015.
Article in English | MEDLINE | ID: mdl-25762899

ABSTRACT

The glial cell line-derived neurotrophic factor (GDNF) is a well-established trophic agent for dopaminergic (DA) neurons in vitro and in vivo. GDNF is necessary for maintenance of neuronal morphological and neurochemical phenotype and protects DA neurons from toxic damage. Numerous studies on animal models of Parkinson's disease (PD) have reported beneficial effects of GDNF on nigrostriatal DA neuron survival. However, translation of these observations to the clinical setting has been hampered so far by side effects associated with the chronic continuous intra-striatal infusion of recombinant GDNF. In addition, double blind and placebo-controlled clinical trials have not reported any clinically relevant effect of GDNF on PD patients. In the past few years, experiments with conditional Gdnf knockout mice have suggested that GDNF is necessary for maintenance of DA neurons in adulthood. In parallel, new methodologies for exogenous GDNF delivery have been developed. Recently, it has been shown that a small population of scattered, electrically interconnected, parvalbumin positive (PV+) GABAergic interneurons is responsible for most of the GDNF produced in the rodent striatum. In addition, cholinergic striatal interneurons appear to be also involved in the modulation of striatal GDNF. In this review, we summarize current knowledge on brain GDNF delivery, homeostasis, and its effects on nigrostriatal DA neurons. Special attention is paid to the therapeutic potential of endogenous GDNF stimulation in PD.

10.
J Clin Invest ; 124(6): 2550-9, 2014 Jun.
Article in English | MEDLINE | ID: mdl-24812663

ABSTRACT

The transition to puberty and adult fertility both require a minimum level of energy availability. The adipocyte-derived hormone leptin signals the long-term status of peripheral energy stores and serves as a key metabolic messenger to the neuroendocrine reproductive axis. Humans and mice lacking leptin or its receptor fail to complete puberty and are infertile. Restoration of leptin levels in these individuals promotes sexual maturation, which requires the pulsatile, coordinated delivery of gonadotropin-releasing hormone to the pituitary and the resulting surge of luteinizing hormone (LH); however, the neural circuits that control the leptin-mediated induction of the reproductive axis are not fully understood. Here, we found that leptin coordinated fertility by acting on neurons in the preoptic region of the hypothalamus and inducing the synthesis of the freely diffusible volume-based transmitter NO, through the activation of neuronal NO synthase (nNOS) in these neurons. The deletion of the gene encoding nNOS or its pharmacological inhibition in the preoptic region blunted the stimulatory action of exogenous leptin on LH secretion and prevented the restoration of fertility in leptin-deficient female mice by leptin treatment. Together, these data indicate that leptin plays a central role in regulating the hypothalamo-pituitary-gonadal axis in vivo through the activation of nNOS in neurons of the preoptic region.


Subject(s)
Leptin/metabolism , Nitric Oxide/metabolism , Preoptic Area/physiology , Reproduction/physiology , Animals , Female , Humans , Kisspeptins/metabolism , Leptin/deficiency , Leptin/genetics , Luteinizing Hormone/metabolism , Male , Mice , Mice, 129 Strain , Mice, Inbred C57BL , Mice, Knockout , Neurons/metabolism , Nitric Oxide Synthase Type I/deficiency , Nitric Oxide Synthase Type I/genetics , Nitric Oxide Synthase Type I/metabolism , Preoptic Area/metabolism , Receptors, G-Protein-Coupled/metabolism , Receptors, Kisspeptin-1 , Receptors, Leptin/deficiency , Receptors, Leptin/genetics , Receptors, Leptin/metabolism , Signal Transduction
11.
J Neurosci ; 32(3): 932-45, 2012 Jan 18.
Article in English | MEDLINE | ID: mdl-22262891

ABSTRACT

Reproduction is controlled in the brain by a neural network that drives the secretion of gonadotropin-releasing hormone (GnRH). Various permissive homeostatic signals must be integrated to achieve ovulation in mammals. However, the neural events controlling the timely activation of GnRH neurons are not completely understood. Here we show that kisspeptin, a potent activator of GnRH neuronal activity, directly communicates with neurons that synthesize the gaseous transmitter nitric oxide (NO) in the preoptic region to coordinate the progression of the ovarian cycle. Using a transgenic Gpr54-null IRES-LacZ knock-in mouse model, we demonstrate that neurons containing neuronal NO synthase (nNOS), which are morphologically associated with kisspeptin fibers, express the kisspeptin receptor GPR54 in the preoptic region, but not in the tuberal region of the hypothalamus. The activation of kisspeptin signaling in preoptic neurons promotes the activation of nNOS through its phosphorylation on serine 1412 via the AKT pathway and mimics the positive feedback effects of estrogens. Finally, we show that while NO release restrains the reproductive axis at stages of the ovarian cycle during which estrogens exert their inhibitory feedback, it is required for the kisspeptin-dependent preovulatory activation of GnRH neurons. Thus, interactions between kisspeptin and nNOS neurons may play a central role in regulating the hypothalamic-pituitary-gonadal axis in vivo.


Subject(s)
Hypothalamus/cytology , Kisspeptins/metabolism , Neurons/metabolism , Nitric Oxide Synthase Type I/metabolism , Ovulation/genetics , Receptors, G-Protein-Coupled/metabolism , Signal Transduction/physiology , Analysis of Variance , Animals , Enzyme Inhibitors/pharmacology , Enzyme-Linked Immunosorbent Assay , Estrous Cycle/drug effects , Estrous Cycle/genetics , Female , Gene Expression Regulation/drug effects , Gene Expression Regulation/genetics , Hypothalamus/drug effects , Kisspeptins/deficiency , Kisspeptins/pharmacology , Luteinizing Hormone/blood , Mice , Mice, Inbred C57BL , Mice, Knockout , Neurons/drug effects , Nitric Oxide Synthase Type I/deficiency , Ovulation/drug effects , Phosphorylation/drug effects , Phosphorylation/genetics , Receptors, G-Protein-Coupled/deficiency , Receptors, Kisspeptin-1 , Signal Transduction/drug effects , Signal Transduction/genetics , Steroids/pharmacology
12.
PLoS One ; 6(11): e27601, 2011.
Article in English | MEDLINE | ID: mdl-22132116

ABSTRACT

Kisspeptins, the ligands of the kisspeptin receptor known for its roles in reproduction and cancer, are also vasoconstrictor peptides in atherosclerosis-prone human aorta and coronary artery. The aim of this study was to further investigate the cardiovascular localisation and function of the kisspeptins and their receptor in human compared to rat and mouse heart. Immunohistochemistry and radioligand binding techniques were employed to investigate kisspeptin receptor localisation, density and pharmacological characteristics in cardiac tissues from all three species. Radioimmunoassay was used to detect kisspeptin peptide levels in human normal heart and to identify any pathological changes in myocardium from patients transplanted for cardiomyopathy or ischaemic heart disease. The cardiac function of kisspeptin receptor was studied in isolated human, rat and mouse paced atria, with a role for the receptor confirmed using mice with targeted disruption of Kiss1r. The data demonstrated that kisspeptin receptor-like immunoreactivity localised to endothelial and smooth muscle cells of intramyocardial blood vessels and to myocytes in human and rodent tissue. [(125)I]KP-14 bound saturably, with subnanomolar affinity to human and rodent myocardium (K(D) = 0.12 nM, human; K(D) = 0.44 nM, rat). Positive inotropic effects of kisspeptin were observed in rat, human and mouse. No response was observed in mice with targeted disruption of Kiss1r. In human heart a decrease in cardiac kisspeptin level was detected in ischaemic heart disease. Kisspeptin and its receptor are expressed in the human, rat and mouse heart and kisspeptins possess potent positive inotropic activity. The cardiovascular actions of the kisspeptins may contribute to the role of these peptides in pregnancy but the consequences of receptor activation must be considered if kisspeptin receptor agonists are developed for use in the treatment of reproductive disorders or cancer.


Subject(s)
Cardiotonic Agents/pharmacology , Cardiovascular System/drug effects , Cardiovascular System/metabolism , Kisspeptins/pharmacology , Puberty/drug effects , Receptors, Cell Surface/metabolism , Adolescent , Adult , Aged , Animals , Aorta/drug effects , Aorta/metabolism , Endothelium, Vascular/drug effects , Endothelium, Vascular/metabolism , Female , Gene Expression Regulation/drug effects , Heart Atria/drug effects , Heart Atria/metabolism , Heart Diseases/metabolism , Heart Diseases/pathology , Humans , In Vitro Techniques , Male , Mice , Middle Aged , Myocardium/metabolism , Myocytes, Cardiac/drug effects , Myocytes, Cardiac/metabolism , Protein Transport/drug effects , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Receptors, Cell Surface/genetics , Vasoconstriction/drug effects , Young Adult
13.
BMC Genomics ; 12: 209, 2011 Apr 28.
Article in English | MEDLINE | ID: mdl-21527035

ABSTRACT

BACKGROUND: Humans and mice with loss of function mutations in GPR54 (KISS1R) or kisspeptin do not progress through puberty, caused by a failure to release GnRH. The transcriptional networks regulated by these proteins in the hypothalamus have yet to be explored by genome-wide methods. RESULTS: We show here, using 1 million exon mouse arrays (Exon 1.0 Affymetrix) and quantitative polymerase chain reaction (QPCR) validation to analyse microdissected hypothalamic tissue from Gpr54 and Kiss1 knockout mice, the extent of transcriptional regulation in the hypothalamus. The sensitivity to detect important transcript differences in microdissected RNA was confirmed by the observation of counter-regulation of Kiss1 expression in Gpr54 knockouts and confirmed by immunohistochemistry (IHC). Since Gpr54 and Kiss1 knockout animals are effectively pre-pubertal with low testosterone (T) levels, we also determined which of the validated transcripts were T-responsive and which varied according to genotype alone. We observed four types of transcriptional regulation (i) genotype only dependent regulation, (ii) T only dependent regulation, (iii) genotype and T-dependent regulation with interaction between these variables, (iv) genotype and T-dependent regulation with no interaction between these variables. The results implicate for the first time several transcription factors (e.g. Npas4, Esr2), proteases (Klk1b22), and the orphan 10-transmembrane transporter TMEM144 in the biology of GPR54/kisspeptin function in the hypothalamus. We show for the neuronal activity regulated transcription factor NPAS4, that distinct protein over-expression is seen in the hypothalamus and hippocampus in Gpr54 knockout mice. This links for the first time the hypothalamic-gonadal axis with this important regulator of inhibitory synapse formation. Similarly we confirm TMEM144 up-regulation in the hypothalamus by RNA in situ hybridization and western blot. CONCLUSIONS: Taken together, global transcriptional profiling shows that loss of GPR54 and kisspeptin are not fully equivalent in the mouse hypothalamus.


Subject(s)
Gene Regulatory Networks , Hypothalamus/metabolism , Proteins/genetics , Proteins/metabolism , Receptors, G-Protein-Coupled/deficiency , Receptors, G-Protein-Coupled/genetics , Testosterone/metabolism , Animals , Genotype , Kisspeptins , Male , Mice , Oligonucleotide Array Sequence Analysis , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Kisspeptin-1 , Transcription, Genetic
14.
Neuroendocrinology ; 93(2): 74-89, 2011.
Article in English | MEDLINE | ID: mdl-21335953

ABSTRACT

Nitric oxide (NO) is a peculiar chemical transmitter that freely diffuses through aqueous and lipid environments and plays a role in major aspects of brain function. Within the hypothalamus, NO exerts critical effects upon the gonadotropin-releasing hormone (GnRH) network to maintain fertility. Here, we review recent evidence that NO regulates major aspects of the GnRH neuron physiology. Far more active than once thought, NO powerfully controls GnRH neuronal activity, GnRH release and structural plasticity at the neurohemal junction. In the preoptic region, neuronal nitric oxide synthase (nNOS) activity is tightly regulated by estrogens and is found to be maximal at the proestrus stage. Natural fluctuations of estrogens control both the differential coupling of this Ca²+-activated enzyme to glutamate N-methyl-D-aspartic acid receptor channels and phosphorylation-mediated nNOS activation. Furthermore, NO endogenously produced by neurons expressing nNOS acutely and directly suppresses spontaneous firing in GnRH neurons, which suggests that neuronal NO may serve as a synchronizing switch within the preoptic region. At the median eminence, NO is spontaneously released from an endothelial source and follows a pulsatile and cyclic pattern of secretion. Importantly, GnRH release appears to be causally related to endothelial NO release. NO is also highly involved in mediating the dialogue set in motion between vascular endothelial cells and tanycytes that control the direct access of GnRH neurons to the pituitary portal blood during the estrous cycle. Altogether, these data raise the intriguing possibility that the neuroendocrine brain uses NO to coordinate both GnRH neuronal activity and GnRH release at key stages of reproductive physiology.


Subject(s)
Brain/physiology , Endothelial Cells/physiology , Neuroglia/physiology , Neurons/physiology , Neurosecretory Systems/physiology , Nitric Oxide/physiology , Reproduction/physiology , Signal Transduction/physiology , Animals , Gonadotropin-Releasing Hormone/physiology , Models, Biological , Nitric Oxide/biosynthesis
15.
J Neurosci ; 31(7): 2421-30, 2011 Feb 16.
Article in English | MEDLINE | ID: mdl-21325509

ABSTRACT

The anteroventral periventricular nucleus (AVPV) is thought to play a key role in regulating the excitability of gonadotropin-releasing hormone (GnRH) neurons that control fertility. Using an angled, parahorizontal brain slice preparation we have undertaken a series of electrophysiological experiments to examine how the AVPV controls GnRH neurons in adult male and female mice. More than half (59%) of GnRH neurons located in the rostral preoptic area were found to receive monosynaptic inputs from the AVPV in a sex-dependent manner. AVPV stimulation frequencies <1 Hz generated short-latency action potentials in GnRH neurons with GABA and glutamate mediating >90% of the evoked fast synaptic currents. The AVPV GABA input was dominant and found to excite or inhibit GnRH neurons in a cell-dependent manner. Increasing the AVPV stimulation frequency to 5-10 Hz resulted in the appearance of additional poststimulus inhibitory as well as delayed excitatory responses in GnRH neurons that were independent of ionotropic amino acid receptors. The inhibition observed immediately following the end of the stimulation period was mediated partly by GABA(B) receptors, while the delayed activation was mediated by the neuropeptide kisspeptin. The latter response was essentially absent in Gpr54 knock-out mice and abolished by a Gpr54 antagonist. Together, these studies show that AVPV neurons provide direct amino acid and neuropeptidergic inputs to GnRH neurons. Low-frequency activation generates predominant GABA/glutamate release with higher frequency activation recruiting release of kisspeptin. This frequency-dependent release of amino acid and neuropeptide neurotransmitters greatly expands the range of AVPV control of GnRH neuron excitability.


Subject(s)
Amino Acids/metabolism , Anterior Thalamic Nuclei/cytology , Gonadotropin-Releasing Hormone/metabolism , Neurons/metabolism , Neuropeptides/metabolism , 6-Cyano-7-nitroquinoxaline-2,3-dione/pharmacology , Animals , Biophysics , Electric Stimulation/methods , Excitatory Amino Acid Antagonists/pharmacology , Female , Glutamic Acid/pharmacology , Gonadotropin-Releasing Hormone/genetics , Green Fluorescent Proteins/genetics , In Vitro Techniques , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , N-Methylaspartate/pharmacology , Neurons/drug effects , Neurotransmitter Agents/metabolism , Patch-Clamp Techniques , Reaction Time/physiology , Receptors, G-Protein-Coupled/deficiency , Receptors, Kisspeptin-1 , Statistics, Nonparametric , Synaptic Transmission/drug effects , Synaptic Transmission/physiology , Valine/analogs & derivatives , Valine/pharmacology , alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid/pharmacology , gamma-Aminobutyric Acid/metabolism , gamma-Aminobutyric Acid/pharmacology
16.
Physiology (Bethesda) ; 25(4): 207-17, 2010 Aug.
Article in English | MEDLINE | ID: mdl-20699467

ABSTRACT

Kisspeptins are a group of peptides that stimulate GnRH release and are required for puberty and maintenance of normal reproductive function. This review focuses on our understanding of the way in which kisspeptin signaling regulates mammalian fertility and how they act as central integrators of different hormonal and physiological signals.


Subject(s)
Reproduction/genetics , Reproduction/physiology , Signal Transduction/genetics , Signal Transduction/physiology , Tumor Suppressor Proteins/genetics , Tumor Suppressor Proteins/physiology , Animals , Gonadotropin-Releasing Hormone/genetics , Gonadotropin-Releasing Hormone/physiology , Humans , Kisspeptins , Neurons/physiology , Proteins/genetics , Rats , Tumor Suppressor Proteins/agonists , Tumor Suppressor Proteins/antagonists & inhibitors
17.
J Neurosci ; 30(25): 8581-90, 2010 Jun 23.
Article in English | MEDLINE | ID: mdl-20573904

ABSTRACT

NMDA and kisspeptins can stimulate gonadotropin-releasing hormone (GnRH) release after peripheral or central administration in mice. To determine whether these agonists act independently or through a common pathway, we have examined their ability to stimulate GnRH/luteinizing hormone (LH) release after peripheral or central administration in Kiss1- or Gpr54 (Kiss1r)-null mutant mice. Peripheral injection of NMDA failed to stimulate GnRH/LH release in prepubertal or gonadally intact mutant male mice. Dual-labeling experiments indicated a direct activation of Kiss1-expressing neurons in the arcuate nucleus. In contrast, central injection of NMDA into the lateral ventricle increased plasma LH levels in both Kiss1 and Gpr54 mutant male mice similar to the responses in wild-type mice. Central injection of NMDA stimulated c-Fos expression throughout the hypothalamus but not in GnRH neurons, suggesting an action at the nerve terminals only. In contrast, kisspeptin-10 stimulated LH release after both central and peripheral injection but induced c-Fos expression in GnRH neurons only after central administration. Finally, central injection of NMDA induces c-Fos expression in catecholamine- and nitric oxide-producing neurons in the hypothalamus of mutant mice, indicating a possible kisspeptin-independent GnRH/LH release by NMDA through activation of these neurons. Thus, NMDA may act at both GnRH cell bodies (kisspeptin-independent) and nerve terminals (kisspeptin-dependent) in a dual way to participate in the GnRH/LH secretion in the male mouse.


Subject(s)
Arcuate Nucleus of Hypothalamus/metabolism , Gonadotropin-Releasing Hormone/metabolism , N-Methylaspartate/administration & dosage , Neurons/metabolism , Tumor Suppressor Proteins/metabolism , Analysis of Variance , Animals , Arcuate Nucleus of Hypothalamus/drug effects , Immunohistochemistry , Injections, Intraperitoneal , Injections, Intraventricular , Kisspeptins , Luteinizing Hormone/blood , Male , Mice , Mice, Knockout , Neurons/drug effects , Proto-Oncogene Proteins c-fos/metabolism , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/metabolism , Receptors, Kisspeptin-1 , Tumor Suppressor Proteins/genetics
18.
Endocrinology ; 151(6): 2723-35, 2010 Jun.
Article in English | MEDLINE | ID: mdl-20371700

ABSTRACT

Within the preoptic region, nitric oxide (NO) production varies during the ovarian cycle and has the ability to impact hypothalamic reproductive function. One mechanism for the regulation of NO release mediated by estrogens during the estrous cycle includes physical association of the calcium-activated neuronal NO synthase (nNOS) enzyme with the glutamate N-methyl-d-aspartate (NMDA) receptor channels via the postsynaptic density 95 scaffolding protein. Here we demonstrate that endogenous variations in estrogens levels during the estrous cycle also coincide with corresponding changes in the state of nNOS Ser1412 phosphorylation, the level of association of this isoform with the NMDA receptor/postsynaptic density 95 complex at the plasma membrane, and the activity of NO synthase (NOS). Neuronal NOS Ser1412 phosphorylation is maximal on the afternoon of proestrus when both the levels of estrogens and the physical association of nNOS with NMDA receptors are highest. Estradiol mimicked these effects in ovariectomized (OVX) rats. In addition, the catalytic activity of NOS in membrane protein extracts from the preoptic region, i.e. independent of any functional protein-protein interactions or cell-cell signaling, was significantly increased in estradiol-treated OVX rats compared with OVX rats. Finally, lambda phosphatase-mediated nNOS dephosphorylation dramatically impaired NOS activity in preoptic region protein extracts, thus demonstrating the important role of phosphorylation in the regulation of NO production in the preoptic region. Taken together, these results yield new insights into the regulation of neuron-derived NO production by gonadal steroids within the preoptic region and raise the possibility that changes in nNOS phosphorylation during fluctuating physiological conditions may be involved in the hypothalamic control of key neuroendocrine functions, such as reproduction.


Subject(s)
Estrogens/metabolism , Hypothalamus/metabolism , Menstrual Cycle/metabolism , Nitric Oxide Synthase Type I/metabolism , Nitric Oxide/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Animals , Blotting, Western , Cell Membrane/metabolism , Estradiol/pharmacology , Female , Hypothalamus/drug effects , Immunoprecipitation , Menstrual Cycle/physiology , Ovariectomy , Phosphorylation/drug effects , Protein Binding/physiology , Rats , Rats, Sprague-Dawley
19.
Endocrinology ; 151(4): 1760-72, 2010 Apr.
Article in English | MEDLINE | ID: mdl-20133455

ABSTRACT

In the ever-changing physiological context of the neuroendocrine brain, the mechanisms by which cellular events involving neurons, astroglia, and vascular cells are coordinated to bring forth the appropriate neuronal signaling is not yet known but is amenable to examination. In the median eminence of the hypothalamus, endothelial cells are key players in the plasticity of tanycytes (specialized astroglia) and neuroendocrine synapse efficacy. Here we report that estradiol acts on both purified endothelial cells and isolated tanycytes to trigger endothelial-to-glial communication that leads to a sudden and massive retraction of tanycyte processes. The blockade of endothelial nitric oxide synthase by in vitro adenoviral-mediated gene transfer of a dominant-negative form of endothelial nitric oxide synthase abrogates the estradiol-induced tanycyte plasticity mediated by endothelial cells. In parallel, increases in prostaglandin-E(2) (PGE(2)) due to changes in cyclooxygenase (COX)-1 and COX-2 expression induced by the exposure of tanycytes to estradiol promote acute tanycyte plasticity. We also demonstrate by electron microscopy that the administration of PGE(2) to median eminence explants induces rapid neuroglial plasticity at the neurovascular junction of neurons that release GnRH (the neuropeptide controlling reproduction). Conversely, preventing local PGE(2) synthesis in the median eminence of adult female rats with the COX inhibitor indomethacin impairs the ovarian cycle, a process that requires a pulsatile, coordinated delivery of GnRH into the hypothalamo-hypophyseal portal system. Taken together, our findings show that estradiol controls the dialog between endothelial cells and astroglia to regulate neuroglial plasticity in the neuroendocrine brain.


Subject(s)
Cell Shape/physiology , Endothelial Cells/physiology , Ependyma/physiology , Estradiol/physiology , Median Eminence/physiology , Neuroglia/physiology , Analysis of Variance , Animals , Blotting, Western , Cell Communication/drug effects , Cell Communication/physiology , Cell Culture Techniques , Cell Shape/drug effects , Cells, Cultured , Dinoprostone/pharmacology , Endothelial Cells/drug effects , Ependyma/drug effects , Estradiol/pharmacology , Hypothalamo-Hypophyseal System/physiology , Neuroglia/drug effects , Nitric Oxide Synthase Type III/physiology , Prostaglandin-Endoperoxide Synthases/physiology , Rats , Rats, Sprague-Dawley
20.
Mol Cell Endocrinol ; 324(1-2): 12-20, 2010 Aug 05.
Article in English | MEDLINE | ID: mdl-20083157

ABSTRACT

At puberty, the mammalian reproductive axis is activated by neuroendocrine events within the hypothalamus that initiate pulsatile secretion of gonadotropin releasing hormone (GnRH) to activate the pituitary/gonadal axis. Thus, puberty is critically dependent on the integrity of GnRH neuronal activity. Defects in the migration of GnRH neurons into the forebrain during development or in GnRH synthesis or release prevent pubertal maturation of the reproductive axis. Both naturally occurring and genetically modified mutant mice have provided valuable information about the cellular and molecular events required for normal pubertal development. This review focuses specifically on the molecules that have been identified from studies in mutant mice that act centrally to control entry into puberty.


Subject(s)
Models, Animal , Puberty/physiology , Sexual Maturation/physiology , Animals , Gonadotropin-Releasing Hormone/biosynthesis , Gonadotropin-Releasing Hormone/metabolism , Humans , Mice , Neurons/cytology , Neurons/metabolism , Neurotransmitter Agents/metabolism
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