Photoreceptor structure and function is maintained in organotypic cultures of mouse retinas.
OBJECTIVE
Retina organ cultures can be used as a valuable tool to study retina development ex vivo. Comparison between culture methods has revealed that timing the start of the culture and the presence of the retinal pigment epithelium (RPE) are critical for the development of the rods and cones, which are the two types of photoreceptors; rods can develop in the absence of the RPE, cones cannot. One of the necessary compounds produced by the RPE and essential for cone development and survival is the chromophore 11-cis retinal. Here, we further examined rod and cone development, chromophore production by the RPE, and photoreceptor signaling to the inner retina under organ culture conditions.


METHODS
Retina-RPE cultures were prepared from 7-day-old C57BL/6 pups and maintained in culture for 11 days. Rod and cone structure was analyzed by immunohistochemistry, and cell-specific mRNA expression was analyzed by quantitative real-time PCR. We quantified 11-cis retinal spectrophotometrically by measuring rhodopsin. Signal transmission in the rod pathway was studied by analyzing c-fos expression in the inner retina in response to stroboscopic illumination.


RESULTS
In retina-RPE cultures analyzed after 11 days in culture, rod and cone numbers exhibited a similar ratio to those observed in the intact animal. Although photoreceptor outer segments were shorter when grown ex vivo, membrane proteins, such as cone opsin and transducin, were localized appropriately to the outer segment. Relative 11-cis retinal production ex vivo plateaued after 7 days in culture, resulting in approximately 30% of the in vivo level by day 11. The retinas responded to prolonged stroboscopic illumination with the normal nuclear expression of c-fos in cells in the inner retina.


CONCLUSIONS
Mouse retinal structure is maintained in retina-RPE organ cultures. The RPE in organ cultures produces sufficient amounts of 11-cis retinal to promote cone development and support signal transmission in the rod pathway. Organ cultures may be a powerful low-throughput screening tool to identify novel agents to promote photoreceptor cell survival and signaling.
Bandyopadhyay
Mausumi
M
Department of Neurosciences, Division of Research, Medical University of South Carolina, Charleston, SC 29425, USA.
Rohrer
Bärbel
B
D000818:Animals; D005786:Gene Expression Regulation; D055537:Light Signal Transduction; D051379:Mice; D008810:Mice, Inbred C57BL; D009924:Organ Culture Techniques / Q000379:methods*; D009928:Organ Specificity / Q000235:genetics; D020419:Photoreceptor Cells, Vertebrate / Q000166:cytology / Q000378:metabolism*; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000378:metabolism; D012160:Retina / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D012172:Retinaldehyde / Q000378:metabolism
D012172:Retinaldehyde