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PLOS ONE  2013 

Zebrafish Guanylate Cyclase Type 3 Signaling in Cone Photoreceptors

DOI: 10.1371/journal.pone.0069656

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The zebrafish guanylate cyclase type 3 (zGC3) is specifically expressed in cone cells. A specifc antibody directed against zGC3 revealed expression at the protein level at 3.5 dpf in outer and inner retinal layers, which increased in intensity between 3.5 and 7 dpf. This expression pattern differed from sections of the adult retina showing strong immunostaining in outer segments of double cones and short single cones, less intense immunoreactivity in long single cones, but no staining in the inner retina. Although transcription and protein expression levels of zGC3 are similar to that of the cyclase regulator guanylate cyclase-activating protein 3 (zGCAP3), we surprisingly found that zGCAP3 is present in a 28-fold molar excess over zGC3 in zebrafish retinae. Further, zGCAP3 was an efficient regulator of guanylate cyclases activity in native zebrafish retinal membrane preparations. Therefore, we investigated the physiological function of zGCAP3 by two different behavioral assays. Using the morpholino antisense technique, we knocked down expression of zGCAP3 and recorded the optokinetic and optomotor responses of morphants, control morphants, and wild type fish at 5–6 dpf. No significant differences in behavioral responses among wild type, morphants and control morphants were found, indicating that a loss of zGCAP3 has no consequences in primary visual processing in the larval retina despite its prominent expression pattern. Its physiological function is therefore compensated by other zGCAP isoforms.


[1]  Luo DG, Xue T, Yau KW (2008) How vision begins: an odyssey. Proc Natl Acad Sci USA 105: 9855–9862.
[2]  Bilotta J, Saszik S (2001) The zebrafish as a model visual system. Int J Dev Neurosci 19: 621–629.
[3]  Ciluffo MC, Matthews HR, Brockerhoff SE, Fain GL (2004) Light-induced Ca2+ release in the visible cones of the zebrafish. Vis Neurosci 21: 599–609.
[4]  Leung YT, Fain GL, Matthews HR (2007) Simultaneous measurements of current and calcium in the ultraviolet-sensitive cones of zebrafish. J Physiol 579(1): 15–27.
[5]  Summerton JE (2007) Morpholino, siRNA, and S-DNA compared: impact of structure and mechanism of action on off-target effects and sequence specificity. Curr Top Med Chem 7: 651–660.
[6]  Baier H (2000) zebrafish on the move: towards a behavior-genetic analysis of vertebrate vision. Curr Opin Neurobiol 10: 451–455.
[7]  Mueller KP, Neuhauss SCF (2010) Quantitative measurements of the optokinetic response in adult fish. J Neurosci Meth 186: 29–34.
[8]  R?tscho N, Scholten A, Koch KW (2009) Expression profiles of three novel sensory guanylate cyclases and guanylate cyclase-activating proteins in the zebrafish retina. Biochim Biopyhs Acta 1793: 1110–1114.
[9]  Fries R, Scholten A, S?ftel W, Koch KW (2012) Operation profile of zebrafish guanylate cyclase-activating protein 3. J Neurochem 121: 54–65.
[10]  Rinner O, Makhankov YV, Biehlmaier O, Neuhauss SCF (2005) Knockdown of cone-specific kinase GRK7 in larval zebrafish leads to impaired cone response recovery and delayed dark adaptation. Neuron 47: 231–42.
[11]  Vogalis F, Shiraki T, Kojima D, Wada Y, Nishiwaki Y, Jarvinen JLP, Sugiyama J, Kawakami K, Masai I, Kawamura S, Fukada Y, Lamb TD (2011) Ectopic expression of cone-specific G-protein-coupled receptor kinase GRK7 in zebrafish rods leads to lower photosensitivity and altered responses. J. Physiol. 589: 2321–2348.
[12]  Stearns G, Evangelista M, Fadool JM, Brockerhoff SE (2007) A mutation in the cone-specific pde6 gene causes rapid cone photoreceptor degeneration in zebrafish. J Neurosci 27: 13866–13874.
[13]  Palczewski K, Polans A, Baehr W, Ames JB (2000) Ca2+-binding proteins in the retina: structure, function, and the etiology of human visual diseases. BioEssays 22: 337–350.
[14]  Koch KW, Duda T, Sharma RK (2010) Ca2+-modulated vision-linked ROS-GC guanylate cyclase transduction machinery. Mol. Cell. Biochem. 334: 105–115.
[15]  Koch KW, Dell'Orco D. (2013) A Calcium –Relay Mechanism in Vertebrate Phototransduction. ACS Chem Neurosci, in press. DOI: 10.1021/cn400027z.
[16]  Imanishi Y, Yang L, Sokal I, Filipek S, Palczewski K, Baehr W (2004) Diversity of guanylate cyclase-activating proteins (GCAPS) in teleost fish: characterization of three novel GCAPs (GCAP4, GCAP5, GCAP7) from zebrafish (Danio rerio) and prediction of eight GCAPs (GCAP1-8) in pufferfish (Fugu rubripes). J Mol Evol 59: 2204–217.
[17]  Scholten A, Koch KW (2011) Differential calcium signaling by cone specific guanylate cyclase-activating proteins from the zebrafish retina. PLoS One 6(8): e23117.
[18]  Stiebel-Kalish H, Reich E, Rainy N, Vaatine G, Nisgav Y, Tovar A (2012) Gucy2f zebrafish knockdown – a model for Gucy2d-related leber congenital amaurosis. Eur J Hum Genet (20): 884–889.
[19]  Muto A, Orger MB, Wehman AM, Smear MC, Kay JN, Page-McCaw PS (2005) PLoS Genet. 1(5): e66.
[20]  Behnen P, Scholten A, R?tscho N, Koch KW (2009) The cone-specific calcium sensor guanylate cyclase activating protein 4 from the zebrafish retina. J Biol Inorg Chem 14: 89–99.
[21]  Kretschmer F, Hein A, Kretzberg J (2011) Virtual experimental arena for behavioral experiments on small vertebrates. IEEE Proceedings of the 4th International Congress on Image and Signal Processing (CISP) vol 1: 441–445.
[22]  Brockerhoff SE, Hurley JB, Niemi GA, Dowling JE (1997) A new form of inherited red-blindness identified in zebrafish. J Neurosci 17: 4236–4242.
[23]  Haug MF, Biehlmaier O, Mueller KP, Neuhauss SCF (2010) Visual acuity in larval zebrafish: behavior and histology. Front. Zool. 7: 8.
[24]  Orger MB, Baier H (2005) Channeling of red and green cone inputs to the zebrafish optomotor response. Vis Neurosci 22: 275–281.
[25]  Reed R, Holmes D, Weyers J, Jones A. (2007) Practical skills in Biomolecular Sciences (third Edition). Pearson Education Limited, Edinburgh Gate, Harlow, Essex.
[26]  Hwang JY, Lange C, Helten A, H?ppner-Heitmann D, Duda T, Sharma RK, Koch KW (2003) Regulatory modes of rod outer segment membrane guanylate cyclase differ in catalytic efficiency and Ca2+-sensitivity. Eur J Biochem 270: 3814–3821.
[27]  Yang RB, Garbers DL (1997) The two eye guanylyl cyclases are expressed in the same photoreceptor cells and form homomers in preference to heteromers. J Biol Chem 272: 13738–13742.
[28]  Goridis C, Virmaux N, Urban PF, Mandel P (1973) Guanyl cyclase in a mammalian photoreceptor. FEBS Lett 30: 163–166.
[29]  R?tscho N, Scholten A, Koch KW (2010) Diversity of sensory guanylate cyclases in teleost fishes. Mol Cell Biochem 334: 207–214.
[30]  Takemoto N, Tachibanaki S, Kawamura S (2009) High cGMP synthetic activity in carp cones. Proc Natl Acad Sci USA 106: 11788–93.
[31]  Kitiratschky VB, Behnen P, Kellner U, Heckenlively JR, Zrenner E, J?gle H, Kohl S, Wissinger B, Koch KW (2009) Hum Mutat. 30: E782–796.
[32]  Dell'Orco D, Behnen P, Linse S, Koch KW (2010) Calcium binding, structural stability and guanylate cyclase activation in GCAP1 variants associated with human cone dystrophy. Cell Mol Life Sci 67: 973–984.
[33]  Brockerhoff SE, Rieke F, Matthews HR, Taylor MR, Kennedy B, Ankoudinova I, Niemi GA, Tucker CL, Xiao M, Cilluffo MC, Fain GL, Hurley JB (2003) Light stimulates a transducin-independent increase of cytoplasmic Ca2+ and suppression of current in cones from the zebrafish mutant nof. J Neurosci 23: 470–480.
[34]  Thompson D, Larson G (1992) Western blots using stained protein gels. Biotechniques 5: 656–658.


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