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In JoVE (1)
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Articles by Chun-Chun Chen in JoVE
Radioativo In situ Hibridação para detecção de padrões de expressão dos genes nos tecidos diversos
Chun-Chun Chen1, Kazuhiro Wada2, Erich D. Jarvis1
1Howard Hughes Medical Institute, Department of Neurobiology, Duke University, 2Department of Biological Sciences, Hokkaido University
Este protocolo é usado com sucesso para detectar quantitativamente os níveis e padrões espaciais de expressão de mRNA em vários tipos de tecido em todas as espécies de vertebrados. O método pode detectar transcritos baixa abundância e permite o tratamento de centenas de lâminas simultaneamente. Apresentamos este protocolo com perfil de expressão de formação cérebro das aves embrionárias como um exemplo.
Other articles by Chun-Chun Chen on PubMed
Distributions of Two Gonadotropin-releasing Hormone Receptor Types in a Cichlid Fish Suggest Functional Specialization
The Journal of Comparative Neurology. Mar, 2006 | Pubmed ID: 16440293
Gonadotropin-releasing hormone 1 (GnRH1) from the brain controls reproduction in vertebrates via a GnRH-specific receptor in the pituitary; however, other forms of GnRH are found in all species, suggesting additional roles for this family of peptides. GnRH action depends critically on the location of its cognate receptors in the brain. To understand the potential roles of additional GnRH forms, we localized two known GnRH receptor types in a cichlid fish, Astatotilapia burtoni, in which GnRH1 is socially regulated. Using in situ hybridization, we describe the mRNA expression pattern of these GnRH receptor (GnRH-R) subtypes in the brain, specifically with respect to GnRH-producing neurons. Our data suggest that following a gene duplication, the two GnRH receptors have evolved to serve different functions. The type 1 receptor (GnRH-R1) is expressed less widely than the type 2 receptor (GnRH-R2). Specifically, GnRH-R1 is expressed in groups of neurons in the telencephalon, preoptic area, ventral hypothalamus, thalamus, and pituitary. In contrast, GnRH-R2 is expressed in many more brain areas, including the olfactory bulb, telencephalon, preoptic area, hypothalamus, thalamus, midbrain, optic tectum, cerebellum, hindbrain, and pituitary. The specific distribution of GnRH-R2 suggests that the GnRH ligands may act via this receptor to influence behavior in A. burtoni. Moreover, only GnRH-R2 mRNA is colocalized in the three known groups of GnRH-containing neurons, suggesting that any direct feedback regulation of GnRH by itself must act through this receptor type. Taken together, these data suggest that the two GnRH receptor types serve different functional roles in A. burtoni.
Expression, Structure, Function, and Evolution of Gonadotropin-releasing Hormone (GnRH) Receptors GnRH-R1SHS and GnRH-R2PEY in the Teleost, Astatotilapia Burtoni
Endocrinology. Oct, 2007 | Pubmed ID: 17595228
Multiple GnRH receptors are known to exist in nonmammalian species, but it is uncertain which receptor type regulates reproduction via the hypothalamic-pituitary-gonadal axis. The teleost fish, Astatotilapia burtoni, is useful for identifying the GnRH receptor responsible for reproduction, because only territorial males reproduce. We have cloned a second GnRH receptor in A. burtoni, GnRH-R1(SHS) (SHS is a peptide motif in extracellular loop 3), which is up-regulated in pituitaries of territorial males. We have shown that GnRH-R1(SHS) is expressed in many tissues and specifically colocalizes with LH in the pituitary. In A. burtoni brain, mRNA levels of both GnRH-R1(SHS) and a previously identified receptor, GnRH-R2(PEY), are highly correlated with mRNA levels of all three GnRH ligands. Despite its likely role in reproduction, we found that GnRH-R1(SHS) has the highest affinity for GnRH2 in vitro and low responsivity to GnRH1. Our phylogenetic analysis shows that GnRH-R1(SHS) is less closely related to mammalian reproductive GnRH receptors than GnRH-R2(PEY). We correlated vertebrate GnRH receptor amino acid sequences with receptor function and tissue distribution in many species and found that GnRH receptor sequences predict ligand responsiveness but not colocalization with pituitary gonadotropes. Based on sequence analysis, tissue localization, and physiological response we propose that the GnRH-R1(SHS) receptor controls reproduction in teleosts, including A. burtoni. We propose a GnRH receptor classification based on gene sequence that correlates with ligand selectivity but not with reproductive control. Our results suggest that different duplicated GnRH receptor genes have been selected to regulate reproduction in different vertebrate lineages.
Localization and Diurnal Expression of Melanopsin, Vertebrate Ancient Opsin, and Pituitary Adenylate Cyclase-activating Peptide MRNA in a Teleost Retina
Journal of Biological Rhythms. Dec, 2007 | Pubmed ID: 18057331
Sequences, Expression Patterns and Regulation of the Corticotropin-releasing Factor System in a Teleost
General and Comparative Endocrinology. Jun, 2008 | Pubmed ID: 18501902
Corticotropin-releasing factor (CRF) is well known for its role in regulating the stress response in vertebrate species by controlling release of glucocorticoids. CRF also influences other physiological processes via two distinct CRF receptors (CRF-Rs) and is co-regulated by a CRF binding protein (CRFBP). Although CRF was first discovered in mammals, it is important for the regulation of the stress response, motor behavior, and appetite in all vertebrates. However, it is unclear how the actions of CRF, CRF-Rs, and CRFBP are coordinated. To approach this problem, we cloned and identified CRF, CRF-Rs, and CRFBP in a teleost fish model system, Astatotilapia burtoni and mapped their expression patterns in the body and brain. We found that CRF, CRFBP, and CRF-Rs gene sequences are highly conserved across vertebrates, suggesting that the CRF system plays an essential role in survival. Members of the CRF system are expressed widely in the brain, retina, gill, spleen, muscle, and kidney, thereby implicating them in a variety of bodily functions, including vision, respiration, digestion, and movement. We also found that following long-term social stress, mRNA expression of CRF in the brain and CRF type 1 receptor in the pituitary decrease whereas CRFBP in the pituitary increases via a homeostatic mechanism.
Heterogeneous Nuclear Ribonucleoprotein A/B and G Inhibits the Transcription of Gonadotropin-releasing-hormone 1
Molecular and Cellular Neurosciences. Jan, 2008 | Pubmed ID: 17920292
Gonadotropin-releasing hormone 1 (GnRH1) causes the release of gonadotropins from the pituitary to control reproduction. Here we report that two heterogeneous nuclear ribonucleoproteins (hnRNP-A/B and hnRNP-G) bind to the GnRH-I upstream promoter region in a cichlid fish Astatotilapia burtoni. We identified these binding proteins using a newly developed homology based method of mass spectrometric peptide mapping. We show that both hnRNP-A/B and hnRNP-G co-localize with GnRH1 in the pre-optic area of the hypothalamus in the brain. We also demonstrated that these ribonucleoproteins exhibit similar binding capacity in vivo, using immortalized mouse GT1-7 cells where overexpression of either hnRNP-A/B or hnRNP-G significantly down-regulates GnRH1 mRNA levels in GT1-7 cells, suggesting that both act as repressors in GnRH1 transcriptional regulation.
Visual Information Alone Changes Behavior and Physiology During Social Interactions in a Cichlid Fish (Astatotilapia Burtoni)
PloS One. 2011 | Pubmed ID: 21633515
Social behavior can influence physiological systems dramatically yet the sensory cues responsible are not well understood. Behavior of male African cichlid fish, Astatotilapia burtoni, in their natural habitat suggests that visual cues from conspecifics contribute significantly to regulation of social behavior. Using a novel paradigm, we asked whether visual cues alone from a larger conspecific male could influence behavior, reproductive physiology and the physiological stress response of a smaller male. Here we show that just seeing a larger, threatening male through a clear barrier can suppress dominant behavior of a smaller male for up to 7 days. Smaller dominant males being "attacked" visually by larger dominant males through a clear barrier also showed physiological changes for up to 3 days, including up-regulation of reproductive- and stress-related gene expression levels and lowered plasma 11-ketotestesterone concentrations as compared to control animals. The smaller males modified their appearance to match that of non-dominant males when exposed to a larger male but they maintained a physiological phenotype similar to that of a dominant male. After 7 days, reproductive- and stress- related gene expression, circulating hormone levels, and gonad size in the smaller males showed no difference from the control group suggesting that the smaller male habituated to the visual intruder. However, the smaller male continued to display subordinate behaviors and assumed the appearance of a subordinate male for a full week despite his dominant male physiology. These data suggest that seeing a larger male alone can regulate the behavior of a smaller male but that ongoing reproductive inhibition depends on additional sensory cues. Perhaps, while experiencing visual social stressors, the smaller male uses an opportunistic strategy, acting like a subordinate male while maintaining the physiology of a dominant male.