Fishing for Drugs from Nature: Zebrafish as a Technology Platform for Natural Product Discovery

Abstract

Emerging challenges within the current drug discovery paradigm are prompting renewed interest in natural products as a source of novel, bioactive small molecules. With the recent validation of zebrafish as a biomedically relevant model for functional genomics and in vivo drug discovery, the zebrafish bioassay-guided identification of natural products may be an attractive strategy to generate new lead compounds in a number of indication areas. Here, we review recent natural product research using zebrafish and evaluate the potential of this vertebrate model as a discovery platform for the systematic identification of bioactive natural products.

Key words

zebrafish - Danio rerio - bioassay-guided fractionation - chemical genetics - in vivo drug discovery

References

• 1 Drews J. Die verspielte Zukunft: Wohin geht die Arzneimittelforschung?. Basel; Birkhäuser Verlag 1998
• 2 Clardy J, Walsh C. Lessons from natural molecules.  Nature. 2004;  432 829-37
• 3 Koehn F E, Carter G T. The evolving role of natural products in drug discovery.  Nat Rev Drug Discov. 2005;  4 206-20
• 4 Greef J van der, McBurney R N. Rescuing drug discovery: in vivo systems pathology and systems pharmacology.  Nat Rev Drug Discov. 2005;  4 961-7
• 5 Zon L I, Peterson R T. In vivo drug discovery in the zebrafish.  Nat Rev Drug Discov. 2005;  4 35-44
• 6 Streisinger G, Walker C, Dower N, Knauber D, Singer F. Production of clones of homozygous diploid zebra fish (Brachydanio rerio).  Nature. 1981;  291 293-6
• 7 Haffter P, Granato M, Brand M, Mullins M C, Hammerschmidt M, Kane D A. et al . The identification of genes with unique and essential functions in the development of the zebrafish, Danio rerio.  Development. 1996;  123 1-36
• 8 Driever W, Solnica-Krezel L, Schier A F, Neuhauss S C, Malicki J, Stemple D L. et al . A genetic screen for mutations affecting embryogenesis in zebrafish.  Development. 1996;  123 37-46
• 9 Amsterdam A, Nissen R M, Sun Z, Swindell E C, Farrington S, Hopkins N. Identification of 315 genes essential for early zebrafish development.  Proc Natl Acad Sci USA. 2004;  101 12 792-7
• 10 Piotrowski T, Schilling T F, Brand M, Jiang Y J, Heisenberg C P, Beuchle D. et al . Jaw and branchial arch mutants in zebrafish II: anterior arches and cartilage differentiation.  Development. 1996;  123 45-56
• 11 Weinstein B M, Schier A F, Abdelilah S, Malicki J, Solnica-Krezel L, Stemple D L. et al . Hematopoietic mutations in the zebrafish.  Development. 1996;  123 303-9
• 12 Stainier D Y, Fouquet B, Chen J N, Warren K S, Weinstein B M, Meiler S E. et al . Mutations affecting the formation and function of the cardiovascular system in the zebrafish embryo.  Development. 1996;  123 285-92
• 13 Farber S A, Pack M, Ho S Y, Johnson I D, Wagner D S, Dosch R. et al . Genetic analysis of digestive physiology using fluorescent phospholipid reporters.  Science. 2001;  292 1385-8
• 14 Jin S W, Herzog W, Santoro M M, Mitchell T S, Frantsve J, Jungblut B. et al . A transgene-assisted genetic screen identifies essential regulators of vascular development in vertebrate embryos.  Dev Biol. 2007;  307 29-42
• 15 Baraban S C, Dinday M T, Castro P A, Chege S, Guyenet S, Taylor M R. A large-scale mutagenesis screen to identify seizure-resistant zebrafish.  Epilepsia. 2007;  48 1151-7
• 16 Summerton J, Weller D. Morpholino antisense oligomers: design, preparation, and properties.  Antisense Nucleic Acid Drug Dev. 1997;  7 187-95
• 17 Nasevicius A, Ekker S C. Effective targeted gene ‘knockdown’ in zebrafish.  Nat Genet. 2000;  26 216-20
• 18 Heasman J. Morpholino oligos: making sense of antisense?.  Dev Biol. 2002;  243 209-14
• 19 Doitsidou M, Reichman-Fried M, Stebler J, Köprunner M, Dörries J, Meyer D. et al . Guidance of primordial germ cell migration by the chemokine SDF-1.  Cell. 2002;  111 647-59
• 20 Pickart M A, Klee E W, Nielsen A L, Sivasubbu S, Mendenhall E M, Bill B R. et al . Genome-wide reverse genetics framework to identify novel functions of the vertebrate secretome.  PLoS ONE. 2006;  1 e104
• 21 Esguerra C V, Nelles L, Vermeire L, Ibrahimi A, Crawford A D, Derua R. et al . Ttrap is an essential modulator of Smad3-dependent Nodal signaling during zebrafish gastrulation and left-right axis determination.  Development. 2007;  134 4381-93
• 22 Shestopalov I A, Sinha S, Chen J K. Light-controlled gene silencing in zebrafish embryos.  Nat Chem Biol. 2007;  3 650-1
• 23 Dodd A, Chambers S P, Love D R. Short interfering RNA-mediated gene targeting in the zebrafish.  FEBS Lett. 2004;  561 89-93
• 24 Summerton J E. Morpholino, siRNA, and S-DNA compared: impact of structure and mechanism of action on off-target effects and sequence specificity.  Curr Top Med Chem. 2007;  7 651-60
• 25 Wang N, Sun Y H, Liu J, Wu G, Su J G, Wang Y P. et al . Knockdown of gfp and no tail expression in zebrafish embryo by in vivo-transcribed short hairpin RNA with T7 plasmid system.  J Biomed Sci. 2007;  14 767-76
• 26 Amsterdam A, Becker T S. Transgenes as screening tools to probe and manipulate the zebrafish genome.  Dev Dyn. 2005;  234 255-68
• 27 Beis D, Stainier D Y. In vivo cell biology: following the zebrafish trend.  Trends Cell Biol. 2006;  16 105-12
• 28 Lawson N D, Weinstein B M. In vivo imaging of embryonic vascular development using transgenic zebrafish.  Dev Biol. 2002;  248 307-18
• 29 Tran T C, Sneed B, Haider J, Blavo D, White A, Aiyejorun T. et al . Automated, quantitative screening assay for antiangiogenic compounds using transgenic zebrafish.  Cancer Res. 2007;  67 11 386-92
• 30 Pisharath H, Rhee J M, Swanson M A, Leach S D, Parsons M J. Targeted ablation of beta cells in the embryonic zebrafish pancreas using E. coli nitroreductase.  Mech Dev. 2007;  124 218-29
• 31 Esengil H, Chang V, Mich J K, Chen J K. Small-molecule regulation of zebrafish gene expression.  Nat Chem Biol. 2007;  3 154-5
• 32 Chang Y T. Small-molecule switch for zebrafish gene expression.  Nat Chem Biol. 2007;  3 135-6
• 33 Jones R W, Huffman M N. Fish embryos as bio-assay material in testing chemicals for effects on cell division and differentiation.  Trans Am Microsc Soc. 1957;  76 177-83
• 34 Abedi Z H, McKinley W P. Bioassay of captan by zebrafish larvae.  Nature. 1967;  216 1321-2
• 35 Alestrom P, Holter J L, Nourizadeh-Lillabadi R. Zebrafish in functional genomics and aquatic biomedicine.  Trends Biotechnol. 2006;  24 15-21
• 36 Jones R W, Stout M G, Reich H, Huffman M N. Cytotoxic activities of certain flavonoids against zebra-fish embryos.  Cancer Chemother Rep. 1964;  34 19-20
• 37 Thomas R J. The toxicologic and teratologic effects of delta-9-tetrahydrocannabinol in the zebrafish embryo.  Toxicol Appl Pharmacol. 1975;  32 184-90
• 38 Chang B E, Liao M H, Kuo M Y, Chen C H. Developmental toxicity of arecoline, the major alkaloid in betel nuts, in zebrafish embryos.  Birth Defects Res A: Clin Mol Teratol. 2004;  70 28-36
• 39 Gertsch J, Niomawe , Gertsch-Roost K, Sticher O. Phyllanthus piscatorum: ethnopharmacological studies on a women’s medicinal plant of the Yanomami Amerindians.  J Ethnopharmacol. 2004;  91 181-8
• 40 Gertsch J, Tobler R T, Brun R, Sticher O, Heilmann J. Antifungal, antiprotozoal, cytotoxic and piscicidal properties of justicidin B and a new arylnaphthalide lignan from Phyllanthus piscatorum.  Planta Med. 2003;  69 420-4
• 41 Peterson R T, Link B A, Dowling J E, Schreiber S L. Small molecule developmental screens reveal the logic and timing of vertebrate development.  Proc Natl Acad Sci USA. 2000;  97 12 965-9
• 42 Spring D R, Krishnan S, Blackwell H E, Schreiber S L. Diversity-oriented synthesis of biaryl-containing medium rings using a one bead/one stock solution platform.  J Am Chem Soc. 2002;  124 1354-63
• 43 MacRae C A, Peterson R T. Zebrafish-based small molecule discovery.  Chem Biol. 2003;  10 901-8
• 44 Peterson R T, Shaw S Y, Peterson T A, Milan D J, Zhong T P, Schreiber S L. et al . Chemical suppression of a genetic mutation in a zebrafish model of aortic coarctation.  Nat Biotech. 2004;  22 595-9
• 45 Jung D W, Williams D, Khersonsky S M, Kang T W, Heidary N, Chang Y T. et al . Identification of the F1F0 mitochondrial ATPase as a target for modulating skin pigmentation by screening a tagged triazine library in zebrafish.  Mol Biosyst. 2005;  1 85-92
• 46 Yang C T, Johnson S L. Small molecule-induced ablation and subsequent regeneration of larval zebrafish melanocytes.  Development. 2006;  133 3563-73
• 47 Choi T Y, Kim J H, Ko D H, Kim C H, Hwang J S, Ahn S. et al . Zebrafish as a new model for phenotype-based screening of melanogenic regulatory compounds.  Pigment Cell Res. 2007;  20 120-7
• 48 Murphey R D, Stern H M, Straub C T, Zon L I. A chemical genetic screen for cell cycle inhibitors in zebrafish embryos.  Chem Biol Drug Des. 2006;  68 13-9
• 49 Burns C G, Milan D J, Grande E J, Rottbauer W, MacRae C A, Fishman M C. High-throughput assay for small molecules that modulate zebrafish embryonic heart rate.  Nat Chem Biol. 2005;  1 63-4
• 50 Mathew L K, Sengupta S, Kawakami A, Andreasen E A, Lohr C V, Loynes C A. et al . Unraveling tissue regeneration pathways using chemical genetics.  J Biol Chem. 2007;  282 35 202-10
• 51 Shafizadeh E, Peterson R T, Lin S. Induction of reversible hemolytic anemia in living zebrafish using a novel small molecule.  Comp Biochem Physiol C: Toxicol Pharmacol. 2004;  138 245-9
• 52 Lally B E, Geiger G A, Kridel S, Arcury-Quandt A E, Robbins M E, Kock N D. et al . Identification and biological evaluation of a novel and potent small molecule radiation sensitizer via an unbiased screen of a chemical library.  Cancer Res. 2007;  67 8791-9
• 53 Berger J, Currie P. The role of zebrafish in chemical genetics.  Curr Med Chem. 2007;  14 2413-20
• 54 Lam H W, Lin H C, Lao S C, Gao J L, Hong S J, Leong C WY. et al . The angiogenic effects of Angelica sinensis extract on HUVEC in vitro and zebrafish in vivo.  J Cell Biochem. 2008;  103 195-211
• 55 He M F, But P PH, Shaw P C, Jiang R W, Xu H X. Anti-angiogenic agents from Tripterygium wilfordii. American Society of Pharmacognosy 47th Annual Meeting Arlington, Virginia; 2006
• 56 Maule J, Richardson J, Clements C, Harvey A, Patton E. A pilot screen for natural inhibitors of cancer-relevant signaling pathways. 5th European Zebrafish Genetics and Development Meeting Amsterdam; 2007
• 57 Incardona J P, Gaffield W, Kapur R P, Roelink H. The teratogenic Veratrum alkaloid cyclopamine inhibits sonic hedgehog signal transduction.  Development. 1998;  125 3553-62
• 58 D’Amour K A, Bang A G, Eliazer S, Kelly O G, Aqulnick A D, Smart N G. et al . Production of pancreatic hormone-expressing endocrine cells from human embryonic stem cells.  Nat Biotechnol. 2006;  24 1392-401
• 59 Lauth M, Toftgard R. The Hedgehog pathway as a drug target in cancer therapy.  Curr Opin Invest Drugs. 2007;  8 457-61
• 60 Lipinski R J, Dengler E, Kiehn M, Peterson R E, Bushman W. Identification and characterization of several dietary alkaloids as weak inhibitors of Hedgehog signaling.  Toxicol Sci. 2007;  100 456-63
• 61 Crawford A D, Breyne A, Oosterlynk J, Maes J, Dewaele M, Ruzzene M. et al .Natural product discovery via chemical genetics in zebrafish. 5th International Congress and Annual Meeting of the Society for Medicinal Plant Research Graz; 2007
• 62 Fong T A, Shawver L K, Sun L, Tang C, App H, Powell T J. et al . SU5416 is a potent and selective inhibitor of the vascular endothelial growth factor receptor (Flk-1/KDR) that inhibits tyrosine kinase catalysis, tumor vascularization, and growth of multiple tumor types.  Cancer Res. 1999;  59 99-106
• 63 Yim H, Lee Y H, Lee C H, Lee S K. Emodin, an anthraquinone derivative isolated from the rhizomes of Rheum palmatum, selectively inhibits the activity of casein kinase II as a competitive inhibitor.  Planta Med. 1999;  65 9-13
• 64 Wang X H, Wu S Y, Zhen Y S. Inhibitory effects of emodin on angiogenesis.  Yao Xue Xue Bao (Acta Pharm Sin). 2004;  39 254-8
• 65 Ljubimov A V, Caballero S, Aoki A M, Pinna L A, Grant M B, Castellon R. Involvement of protein kinase CK2 in angiogenesis and retinal neovascularization.  Invest Ophthalmol Visc Sci. 2004;  45 4583-91
• 66 Luesch H, Yoshida W Y, Moore R E, Paul V J, Corbett T H. Total structure determination of apratoxin A, a potent novel cytotoxin from the marine cyanobacterium Lyngbya majuscula. .  J Am Chem Soc. 2001;  123 5418-23
• 67 Luesch H, Chanda S K, Raya R M, DeJesus P D, Orth A P, Walker J R. et al . A functional genomics approach to the mode of action of apratoxin A.  Nat Chem Biol. 2006;  2 158-67
• 68 McCowen M C, Callender M E, Lawlis Jr J F. Fumagillin (H-3), a new antibiotic with amebicidal properties.  Science. 1951;  113 202-3
• 69 Ingber D, Fujita T, Kishimoto S, Sudo K, Kanamaru T, Brem H. et al . Synthetic analogues of fumagillin that inhibit angiogenesis and suppress tumour growth.  Nature. 1990;  348 555-7
• 70 Satchi-Fainaro R, Puder M, Davies J W, Tran H T, Sampson D A, Greene A K. et al . Targeting angiogenesis with a conjugate of HPMA copolymer and TNP-470.  Nat Med. 2004;  10 255-61
• 71 Sin N, Meng L, Wang M Q, Wen J J, Bornmann W G, Crews C M. The anti-angiogenic agent fumagillin covalently binds and inhibits the methionine aminopeptidase, MetAP-2.  Proc Natl Acad Sci USA. 1997;  94 6099-7103
• 72 Zhang Y, Yeh J R, Mara A, Ju R, Hines J F, Cirone P. et al . A chemical and genetic approach to the mode of action of fumagillin.  Chem Biol. 2006;  13 1001-9
• 73 Peterson R T. A noncanonical path to mechanism of action.  Chem Biol. 2006;  13 924-6
• 74 Braida D, Limonta V, Pegorini S, Zani A, Guerini-Rocco C, Gori E. et al . Hallucinatory and rewarding effect of salvinorin A in zebrafish: kappa-opioid and CB1-cannabinoid receptor involvement.  Psychopharmacology. 2007;  190 441-8
• 75 Ninkovic J, Bally-Cuif L. The zebrafish as a model system for assessing the reinforcing properties of drugs of abuse.  Methods. 2006;  39 262-74
• 76 Yu P B, Hong C C, Sachidanandan C, Babitt J L, Deng D Y, Hoyng S A. et al . Dorsomorphin inhibits BMP signals required for embryogenesis and iron metabolism.  Nat Chem Biol. 2008;  4 33-41
• 77 Sar A M van der, Appelmelk B J, Vandenbroucke-Grauls C M, Bitter W. A star with stripes: zebrafish as an infection model.  Trends Microbiol. 2004;  12 51-7
• 78 Mukhopadhyay A, Peterson R T. Fishing for new antimicrobials.  Curr Opin Chem Biol. 2006;  10 327-33
• 79 Pressley M E, Phelan PE 3 rd, Witten P E, Mellon M T, Kim C H. Pathogenesis and inflammatory response to Edwardsiella tarda infection in the zebrafish.  Dev Comp Immunol. 2005;  29 501-13
• 80 Watzke J, Schirmer K, Scholz S. Bacterial lipopolysaccharides induce genes involved in the innate immune response in embryos of the zebrafish (Danio rerio).  Fish Shellfish Immunol. 2007;  23 901-5
• 81 Lieschke G J, Oates A C, Crowhurst M O, Ward A C, Layton J E. Morphologic and functional characterization of granulocytes and macrophages in embryonic and adult zebrafish.  Blood. 2001;  98 3087-96
• 82 Renshaw S A, Loynes C A, Trushell D M, Elworthy S, Ingham P W, Whyte M K. A transgenic zebrafish model of neutrophilic inflammation.  Blood. 2006;  108 3976-8
• 83 Mathias J R, Perrin B J, Liu T X, Kanki J, Look A T, Huttenlocher A. Resolution of inflammation by retrograde chemotaxis of neutrophils in transgenic zebrafish.  J Leukoc Biol. 2006;  80 1281-8
• 84 Redd M J, Kelly G, Dunn G, Way M, Martin P. Imaging macrophage chemotaxis in vivo: studies of microtubule function in zebrafish wound inflammation.  Cell Motil Cytoskeleton. 2006;  63 415-22
• 85 Baraban S C. Emerging epilepsy models: insights from mice, flies, worms and fish.  Curr Opin Neurol. 2007;  20 164-8
• 86 Baraban S C, Taylor M R, Castro P A, Baier H. Pentylenetetrazole induced changes in zebrafish behavior, neural activity and c-fos expression.  Neuroscience. 2005;  131 759-68
• 87 Berghmans S, Hunt J, Roach A, Goldsmith P. Zebrafish offer the potential for a primary screen to identify a wide variety of potential anticonvulsants.  Epilepsy Res. 2007;  75 18-28
• 88 Tiedeken J A, Ramsdell J S. Embryonic exposure to domoic acid increases the susceptibility of zebrafish larvae to the chemical convulsant pentylenetetrazole.  Environ Health Perspect. 2007;  115 1547-52
• 89 Milan D J, Peterson T A, Ruskin J N, Peterson R T, MacRae C A. Drugs that induce repolarization abnormalities cause bradycardia in zebrafish.  Circulation. 2003;  107 1355-8
• 90 Langheinrich U, Vacun G, Wagner T. Zebrafish embryos express an orthologue of HERG and are sensitive toward a range of QT-prolonging drugs inducing severe arrhythmia.  Toxicol Appl Pharmacol. 2003;  193 370-82
• 91 Heideman W, Antkiewicz D S, Carney S A, Peterson R E. Zebrafish and cardiac toxicology.  Cardiovasc Toxicol. 2005;  5 203-14
• 92 Arnaout R, Ferrer T, Huisken J, Spitzer K, Stainier D Y, Tristani-Firouzi M. et al . Zebrafish model for human long QT syndrome.  Proc Natl Acad Sci USA. 2007;  104 11 316-21
• 93 Le X, Langenau D M, Keefe M D, Kutok J L, Neuberg D S, Zon L I. Heat shock-inducible Cre/Lox approaches to induce diverse types of tumors and hyperplasia in transgenic zebrafish.  Proc Natl Acad Sci USA. 2007;  104 9410-5
• 94 Feng H, Langenau D M, Madge J A, Quinkertz A, Gutierrez A, Neuberg D S. et al . Heat-shock induction of T-cell lymphoma/leukaemia in conditional Cre/lox-regulated transgenic zebrafish.  Br J Haematol. 2007;  138 169-75
• 95 Langenau D M, Feng H, Berghmans S, Kanki J P, Kutok J L, Look A T. Cre/lox-regulated transgenic zebrafish model with conditional myc-induced T cell acute lymphoblastic leukemia.  Proc Natl Acad Sci USA. 2005;  102 6068-73
• 96 Berghmans S, Murphey R D, Wienholds E, Neuberg D, Kutok J L, Fletcher C D. et al . tp53 mutant zebrafish develop malignant peripheral nerve sheath tumors.  Proc Natl Acad Sci USA. 2005;  102 407-12
• 97 Mizgireuv I V, Revskoy S Y. Transplantable tumor lines generated in clonal zebrafish.  Cancer Res. 2006;  66 3120-5
• 98 Goessling W, North T E, Zon L I. Ultrasound biomicroscopy permits in vivo characterization of zebrafish liver tumors.  Nat Methods. 2007;  4 551-3
• 99 Topczewska J M, Postovit L M, Margaryan N V, Sam A, Hess A R, Wheaton W W. et al . Embryonic and tumorigenic pathways converge via Nodal signaling: role in melanoma aggressiveness.  Nat Med. 2006;  12 925-32
• 100 Haldi M, Ton C, Seng W L, McGrath P. Human melanoma cells transplanted into zebrafish proliferate, migrate, produce melanin, form masses and stimulate angiogenesis in zebrafish.  Angiogenesis. 2006;  9 139-51
• 101 Nicoli S, Ribatti D, Cotelli F, Presta M. Mammalian tumor xenografts induce neovascularization in zebrafish embryos.  Cancer Res. 2007;  67 2927-31
• 102 Hendrix M J, Seftor E A, Seftor R E, Kasemeier-Kulesa J, Kulesa P M, Postovit L M. Reprogramming metastatic tumour cells with embryonic microenvironments.  Nat Rev Cancer. 2007;  7 246-55
• 103 Geiger G A, Parker S E, Beothy A P, Tucker J A, Mullins M C, Kao G D. Zebrafish as a ”biosensor”? Effects of ionizing radiation and amifostine on embryonic viability and development.  Cancer Res. 2006;  66 8172-81
• 104 Moore J L, Rush L M, Breneman C, Mohideen M A, Cheng K C. Zebrafish genomic instability mutants and cancer susceptibility.  Genetics. 2006;  174 585-600
• 105 Shepard J L, Amatruda J F, Stern H M, Subramanian A, Finkelstein D, Ziai J. et al . A zebrafish bmyb mutation causes genome instability and increased cancer susceptibility.  Proc Natl Acad Sci USA. 2005;  102 13 194-9
• 106 Stern H M, Murphey R D, Shepard J L, Amatruda J F, Straub C T, Pfaff K L. et al . Small molecules that delay S phase suppress a zebrafish bmyb mutant.  Nat Chem Biol. 2005;  1 366-70
• 107 Arbiser J L, Kau T, Konar M, Narra K, Ramchandran R, Summers S A. et al . Solenopsin, the alkaloidal component of the fire ant (Solenopsis invicta), is a naturally occurring inhibitor of phosphatidylinositol-3-kinase signaling and angiogenesis.  Blood. 2007;  109 60-5
• 108 Langenau D M, Traver D, Ferrando A A, Kutok J L, Aster J C, Kanki J P. et al . Myc-induced T cell leukemia in transgenic zebrafish.  Science. 2003;  299 887-90
• 109 McAleer M F, Davidson C, Davidson W R, Yentzer B, Farber S A, Rodeck U. et al . Novel use of zebrafish as a vertebrate model to screen radiation protectors and sensitizers.  Int J Radiat Oncol Biol Phys. 2005;  61 10-3
• 110 Fleming A, Sato M, Goldsmith P. High-throughput in vivo screening for bone anabolic compounds with zebrafish.  J Biomol Screen. 2005;  10 823-31
• 111 Barrett R, Chappell C, Quick M, Fleming A. A rapid, high content, in vivo model of glucocorticoid-induced osteoporosis.  Biotechnol J. 2006;  1 651-5
• 112 Jagadeeswaran P, Paris R, Rao P. Laser-induced thrombosis in zebrafish larvae: a novel genetic screening method for thrombosis.  Methods Mol Med. 2006;  129 187-95
• 113 Lockwood B, Bjerke S, Kobayashi K, Guo S. Acute effects of alcohol on larval zebrafish: a genetic system for large-scale screening.  Pharmacol Biochem Behav. 2004;  77 647-54
• 114 Darland T, Dowling J E. Behavioral screening for cocaine sensitivity in mutagenized zebrafish.  Proc Natl Acad Sci USA. 2001;  98 11 691-6
• 115 Saint-Amant L, Sprague S M, Hirata H, Li Q, Cui W W, Zhou W. et al . The zebrafish ennui behavioral mutation disrupts acetylcholine receptor localization and motor axon stability.  Dev Neurobiol. 2007;  68 45-61
• 116 Best J D, Berghmans S, Hunt J J, Clarke S C, Fleming A, Goldsmith P. et al .Non-associative learning in larval zebrafish. Neuropsychopharmacology June 20 2007, DOI: 10.1038/sj.npp.1301489
• 117 Hicks C, Sorocco D, Levin M. Automated analysis of behavior: a computer-controlled system for drug screening and the investigation of learning.  J Neurobiol. 2006;  66 977-90
• 118 Kuhlman J, Eisen J S. Genetic screen for mutations affecting development and function of the enteric nervous system.  Dev Dyn. 2007;  236 118-27
• 119 Pogoda H M, Sternheim N, Lyons D A, Diamond B, Hawkins T A, Woods I G. et al . A genetic screen identifies genes essential for development of myelinated axons in zebrafish.  Dev Biol. 2006;  298 118-31
• 120 Kazakova N, Li H, Mora A, Jessen K R, Mirsky R, Richardson W D. et al . A screen for mutations in zebrafish that affect myelin gene expression in Schwann cells and oligodendrocytes.  Dev Biol. 2006;  297 1-13
• 121 Tomasiewicz H G, Flaherty D B, Soria J P, Wood J G. Transgenic zebrafish model of neurodegeneration.  J Neurosci Res. 2002;  70 734-45
• 122 Lemmens R, Van Hoecke A, Hersmus N, Geelen V, D’Hollander I, Thijs V. et al . Overexpression of mutant superoxide dismutase 1 causes a motor axonopathy in the zebrafish.  Hum Mol Genet. 2007;  16 2359-65
• 123 Kim H J, Sumanas S, Palencia-Desai S, Dong Y, Chen J N, Lin S. Genetic analysis of early endocrine pancreas formation in zebrafish.  Mol Endocrinol. 2006;  20 194-203
• 124 Ober E A, Verkade H, Field H A, Stainier D Y. Mesodermal Wnt2b signalling positively regulates liver specification.  Nature. 2006;  442 688-91
• 125 Sadler K C, Amsterdam A, Soroka C, Boyer J, Hopkins N. A genetic screen in zebrafish identifies the mutants vps18, nf2 and foie gras as models of liver disease.  Development. 2005;  132 3561-72
• 126 Ton C, Parng C. The use of zebrafish for assessing ototoxic and otoprotective agents.  Hear Res. 2005;  208 79-88
• 127 Ou H C, Raible D W, Rubel E W. Cisplatin-induced hair cell loss in zebrafish (Danio rerio) lateral line.  Hear Res. 2007;  233 46-53
• 128 DeBruyne J, Hurd M W, Gutiérrez L, Kaneko M, Tan Y, Wells D E. et al . Isolation and phenogenetics of a novel circadian rhythm mutant in zebrafish.  J Neurogenet. 2004;  18 403-28
• 129 Kim J H, Baek S H, Kim D H, Choi T Y, Yoon T Y, Hwang J S. et al .Downregulation of melanin synthesis by haginin A and its application to in vivo lightening model. J Invest Dermatol 2007; DOI: 10.1038/sj.jid. 5701177