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Sara Stokes Patterson, Ralph F Nelson; Spectral properties of a zebrafish transgenic with L-opsin expression in all cone types. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):994.
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© ARVO (1962-2015); The Authors (2016-present)
The cone-rod homeobox gene (crx) is essential for photoreceptor differentiation. Mutations in crx are linked to human photoreceptor degenerations. Thyroxin β2 receptor (trβ2) triggers the later development of red cones. The zebrafish transgenic crx:MYFP-2A-trβ2 (crx:trβ2) causes expression of trβ2 on all cone progenitors. Red cone opsin is expressed in all cone types while the other opsins are suppressed (Suzuki et al, 2013). Retinas with a purely red cone spectral sensitivity are expected.
Larvae (5-12dpf) from an outcross of crx:MYFP-2A-trβ2 and wild type (WT) adults were phenotyped by MYFP pupil fluorescence. Larval eyes were isolated and perfused with oxygenated MEM containing 20mM Na Aspartate to isolate cone electrical signals. Eyes were punctured transcorneally with glass microelectrodes to record these ‘cone PIII’ signals while the eye was stimulated with 280 spectral flashes of 9 wavelengths (330-650nm) and 7 irradiances (0.5 log unit increments) to generate spectral datasets. Dataset amplitudes were fit with an 8 parameter function summing 4 Hill functions, each representing an individual red, green, blue or UV cone contribution (Nelson and Singla, 2009).
At threshold, WT spectral sensitivities peaked in the UV at 360nm while crx:trβ2 spectra peaked at 450-500nm. The crx:trβ2 peak migrated towards the 570nm red cone peak for supra-threshold spectra. In mid and long wavelength spectral regions, crx:trβ2 sensitivity exceeded wild type sensitivity. Red cone sensitivity in crx:trβ2 increased by a factor of 2.7 (P>.999) compared to WTs. Green cone sensitivity increased by a factor of 3.1 (P>.999). For WTs, Hill saturation amplitudes >0 were found for red, green, blue and UV cones (2 SEMs, P>.95, 17 of 17 datasets). In crx:trβ2 Hill saturation amplitudes >0 were found for red cones (32 of 32) and green cones (31 of 32), but rarely blue cones (2 of 32) or UV cones (1 of 32). Results were independent of larval age.
As expected, the crx:trβ2 transgenic was robust in red cone signals, while blue and UV cone signals were nearly absent. Surprisingly green cone signals increased. While green cone saturation amplitude was less than for red cones, a more sensitive half-saturation value allowed domination of threshold spectral sensitivity. The crx:trβ2 zebrafish appear to be dichromats.
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