Project description:The foggy mutant zebrafish show severe deficits in dopaminergic neurons, thus is an important tool to understanding the development of these neurons, which undergo degeneration in Parkinson's disease. Molecular study reveals that the foggy gene encodes a regulator of transcription elongation. Biochemical study suggests that the repressive activity of the Foggy protein is lost in the foggy mutant. The genes whose expression is normally repressed by Foggy are likely to play important roles in dopamine neuron development. However, their identity is unknown. Finding out these target genes of Foggy will provide important insights into dopamine neuron development and regeneration. They will also serve as an entry point to elucidate the role of transcription elongation in controlling neuronal development. We will identify genes whose expression is altered in the foggy mutant. Their expression pattern and functional significance will be examined in zebrafish through in situ hybridization and morpholino antisense knockdowm methods. We hypothesize that the genes whose transcription is regulated by Foggy play a critical role in the development of dopaminergic neurons. The species noted above is Drosophila however we are using ZEBRAFISH. We will examine three stages: 24 hour post fertilization (hpf), 36 hpf, and 48 hpf. During these stages, the dopamine neuron deficits manifest in the foggy mutant. To identify genes whose expression is altered and important for dopamine neuron development, we shall dissect embryonic brain tissues from wildtype and the foggy mutant embryos at the stages mentioned above, and prepare RNA from them. Since the foggy gene is expressed ubiquitously and also required for proper retinal and cardiovascular development, we would like to compare the gene expression between the whole embryos of wildtype and foggy mutant, to identify all possible downstream targets. We are currently applying for a supplement to support the entire microarray experiment. As a pilot experiment, we would like to perform 6 array analysis as soon as possible. Thus, we will examine the 48 hours post fertilzation time point in this pilot experiment and it will be performed in triplicate.

Project description:The foggy mutant zebrafish show severe deficits in dopaminergic neurons, thus is an important tool to understanding the development of these neurons, which undergo degeneration in Parkinson's disease. Molecular study reveals that the foggy gene encodes a regulator of transcription elongation. Biochemical study suggests that the repressive activity of the Foggy protein is lost in the foggy mutant. The genes whose expression is normally repressed by Foggy are likely to play important roles in dopamine neuron development. However, their identity is unknown. Finding out these target genes of Foggy will provide important insights into dopamine neuron development and regeneration. They will also serve as an entry point to elucidate the role of transcription elongation in controlling neuronal development. We will identify genes whose expression is altered in the foggy mutant. Their expression pattern and functional significance will be examined in zebrafish through in situ hybridization and morpholino antisense knockdowm methods. We hypothesize that the genes whose transcription is regulated by Foggy play a critical role in the development of dopaminergic neurons. The species noted above is Drosophila however we are using ZEBRAFISH. We will examine three stages: 24 hour post fertilization (hpf), 36 hpf, and 48 hpf. During these stages, the dopamine neuron deficits manifest in the foggy mutant. To identify genes whose expression is altered and important for dopamine neuron development, we shall dissect embryonic brain tissues from wildtype and the foggy mutant embryos at the stages mentioned above, and prepare RNA from them. Since the foggy gene is expressed ubiquitously and also required for proper retinal and cardiovascular development, we would like to compare the gene expression between the whole embryos of wildtype and foggy mutant, to identify all possible downstream targets. We are currently applying for a supplement to support the entire microarray experiment. As a pilot experiment, we would like to perform 6 array analysis as soon as possible. Thus, we will examine the 48 hours post fertilzation time point in this pilot experiment and it will be performed in triplicate. This dataset is composed of 36 hours post fertilization wild type and mutants and 12-somite stage Foggy knockdown.

Project description:The foggy mutant zebrafish show severe deficits in dopaminergic neurons, thus is an important tool to understanding the development of these neurons, which undergo degeneration in Parkinson's disease. Molecular study reveals that the foggy gene encodes a regulator of transcription elongation. Biochemical study suggests that the repressive activity of the Foggy protein is lost in the foggy mutant. The genes whose expression is normally repressed by Foggy are likely to play important roles in dopamine neuron development. However, their identity is unknown. Finding out these target genes of Foggy will provide important insights into dopamine neuron development and regeneration. They will also serve as an entry point to elucidate the role of transcription elongation in controlling neuronal development. We will identify genes whose expression is altered in the foggy mutant. Their expression pattern and functional significance will be examined in zebrafish through in situ hybridization and morpholino antisense knockdowm methods. We hypothesize that the genes whose transcription is regulated by Foggy play a critical role in the development of dopaminergic neurons. The species noted above is Drosophila however we are using ZEBRAFISH. We will examine three stages: 24 hour post fertilization (hpf), 36 hpf, and 48 hpf. During these stages, the dopamine neuron deficits manifest in the foggy mutant. To identify genes whose expression is altered and important for dopamine neuron development, we shall dissect embryonic brain tissues from wildtype and the foggy mutant embryos at the stages mentioned above, and prepare RNA from them. Since the foggy gene is expressed ubiquitously and also required for proper retinal and cardiovascular development, we would like to compare the gene expression between the whole embryos of wildtype and foggy mutant, to identify all possible downstream targets. We are currently applying for a supplement to support the entire microarray experiment. As a pilot experiment, we would like to perform 6 array analysis as soon as possible. Thus, we will examine the 48 hours post fertilzation time point in this pilot experiment and it will be performed in triplicate. Keywords: other

Project description:Best2009 - Homeostatic mechanisms in dopamine synthesis and release Encoded non-curated model. Issues: - Initial concentrations for hva, tyrpool, NADPH and NADP are missing - Reaction VMAT requires two substrates to match the formula proposed in the article This model is described in the article: Homeostatic mechanisms in dopamine synthesis and release: a mathematical model. Best JA, Nijhout HF, Reed MC. Theor Biol Med Model 2009; 6: 21 Abstract: BACKGROUND: Dopamine is a catecholamine that is used as a neurotransmitter both in the periphery and in the central nervous system. Dysfunction in various dopaminergic systems is known to be associated with various disorders, including schizophrenia, Parkinson's disease, and Tourette's syndrome. Furthermore, microdialysis studies have shown that addictive drugs increase extracellular dopamine and brain imaging has shown a correlation between euphoria and psycho-stimulant-induced increases in extracellular dopamine 1. These consequences of dopamine dysfunction indicate the importance of maintaining dopamine functionality through homeostatic mechanisms that have been attributed to the delicate balance between synthesis, storage, release, metabolism, and reuptake. METHODS: We construct a mathematical model of dopamine synthesis, release, and reuptake and use it to study homeostasis in single dopaminergic neuron terminals. We investigate the substrate inhibition of tyrosine hydroxylase by tyrosine, the consequences of the rapid uptake of extracellular dopamine by the dopamine transporters, and the effects of the autoreceoptors on dopaminergic function. The main focus is to understand the regulation and control of synthesis and release and to explicate and interpret experimental findings. RESULTS: We show that the substrate inhibition of tyrosine hydroxylase by tyrosine stabilizes cytosolic and vesicular dopamine against changes in tyrosine availability due to meals. We find that the autoreceptors dampen the fluctuations in extracellular dopamine caused by changes in tyrosine hydroxylase expression and changes in the rate of firing. We show that short bursts of action potentials create significant dopamine signals against the background of tonic firing. We explain the observed time courses of extracellular dopamine responses to stimulation in wild type mice and mice that have genetically altered dopamine transporter densities and the observed half-lives of extracellular dopamine under various treatment protocols. CONCLUSION: Dopaminergic systems must respond robustly to important biological signals such as bursts, while at the same time maintaining homeostasis in the face of normal biological fluctuations in inputs, expression levels, and firing rates. This is accomplished through the cooperative effect of many different homeostatic mechanisms including special properties of tyrosine hydroxylase, the dopamine transporters, and the dopamine autoreceptors. This model is hosted on BioModels Database and identified by: MODEL1502230000. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

Project description:This project aimed at identifying developmental stage specific transcript profiles for catecholaminergic neurons in embryos and early larvae of zebrafish (Danio rerio). Catecholaminergic neurons were labeled using transgenic zebrafish strains to drive expression of GFP. At stages 24, 36, 72 and 96 hrs post fertilization, embryos were dissociated and GFP expressing cells sorted by FACS. Isolated RNAs were processed using either polyA selection and libray generation or NanoCAGE. This is the first effort to determine stage specific mRNA profiles of catecholaminergic neurons in zebrafish. Catecholaminergic neurons were labeled by four different strategies: (1) 24 hrs old embryos: we used the ETvmat2:GFP transgenic line (Wen et al. 2007). Visualization of monoaminergic neurons and neurotoxicity of MPTP in live transgenic zebrafish. Dev Biol. 2008 Vol 314 p84-92) which at this early stage labels catecholaminergic neurons in posterior tuberculum and locus coeruleus; (2) 24 hrs old embryos: we used Tg(otpb.A:egfp)zc48 transgenic line (Fujimoto et al. Identification of a dopaminergic enhancer indicates complexity in vertebrate dopamine neuron phenotype specification. Dev Biol 2011, Vol 352, p393–404) which at this stage label ventral diencephalic dopaminergic neurons and some preoptic neurons. (3) For 72 and 96 hrs old zebrafish larvae we used a th:GFP BAC transgenic lines that labels catecholaminergic neurons (Tay et al., Comprehensive catecholaminergic projectome analysis reveals single-neuron integration of zebrafish ascending and descending dopaminergic systems. Nat Comms 2011 Vol 2, 171; also: T. Leng and W. Driever, unpublished). (4) for the 36 and 48 hrs old zebrafish larvae we used a th:Gal4VP16 driver and UAS:EGFP responder transgenic line system to label catecholaminergic cells (Fernandes et al., Deep brain photoreceptors control light-seeking behavior in zebrafish larvae. Curr Biol. 2012 Vol 22 DOI 10.1016/j.cub.2012.08.016). We used the different transgenic lines, because lines (3) and (4) do not efficiently label catecholaminergic neurons at early stages, while lines (1) and (2) also have GFP expression in several other non-catecholaminergic populations at later stages of development. Embryos were dissociated and catecholaminergic neurons were FACS sorted from GFP-tagged zebrafish (Manoli and Driever, 2012, Cold Spring Harbor Protoc. DOI 10.1101/pdb.prot069633). RNA was either processed for NanoCAGE, or mRNA was isolated and amplified. cDNA was sequenced by Illumina technique. This data submission is a series of data files consisting of three independent experiments with diffrent RNA-Seq depth: Samples 1-4 (NanoCage): Samples 5-8 (RNA-Seq high read numbers), and SAmples 9-12 (RNA-Seq low read numbers).

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Foggy mutant: brain (Guo-1R01NS042626-01A2)

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