Fluorescent photoreceptors of transgenic Xenopus laevis imaged in vivo by two microscopy techniques.
OBJECTIVE
To develop a method for imaging individual photoreceptors in an intact transgenic Xenopus eye, thus allowing in vivo observation of the effects of various transgenes on photoreceptor development, degeneration, or both.


METHODS
Albino and pigmented transgenic Xenopus laevis that express enhanced green fluorescent protein (GFP) in the major ("red") rods were generated. The distribution of GFP throughout the retina and within the rods was evaluated by confocal microscopy of frozen sections and immunoelectron microscopy. In vivo images of photoreceptors were obtained using conventional fluorescence microscopes to image through the lens of the eye or a laser scanning confocal microscope to image through the hypopigmented iris of albino eyes.


RESULTS
Confocal and immunoelectron microscopy of tissue sections showed that GFP was predominantly localized to the inner segments of the major rods; a smaller amount was in the outer segments. In a number of animals, not all the major rods expressed GFP. It was possible to identify these animals by obtaining fluorescence images of the retinas of intact, living tadpoles with conventional fluorescence microscopes, using the lens of the tadpole as part of the optical path. Confocal images of living animals could be used to visualize the distribution of GFP within the photoreceptors.


CONCLUSIONS
The ability to observe individual photoreceptors noninvasively allows in vivo longitudinal microscopic analysis of photoreceptor development in transgenic Xenopus tadpoles.
Moritz
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Department of Pharmacology, University of Connecticut Health Center, Farmington 06030, USA. omoritz@neuron.uchc.edu
Tam
B M
BM
Knox
B E
BE
Papermaster
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DS
D000818:Animals; D030801:Animals, Genetically Modified; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000032:analysis* / Q000235:genetics; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D015345:Oligonucleotide Probes / Q000737:chemistry; D020419:Photoreceptor Cells, Vertebrate / Q000737:chemistry / Q000166:cytology*; D017299:Rod Opsins / Q000235:genetics; D014162:Transfection; D014982:Xenopus laevis / Q000033:anatomy & histology*
D008164:Luminescent Proteins; D015345:Oligonucleotide Probes; D017299:Rod Opsins; D049452:Green Fluorescent Proteins