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Virpi Alarautalahti, Symantas Ragauskas, Jenni J. Hakkarainen, Hannele Uusitalo-Järvinen, Hannu Uusitalo, Jari Hyttinen, Giedrius Kalesnykas, Soile Nymark; Viability of Mouse Retinal Explant Cultures Assessed by Preservation of Functionality and Morphology. Invest. Ophthalmol. Vis. Sci. 2019;60(6):1914-1927. doi: 10.1167/iovs.18-25156.
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Retinal explant cultures provide simplified systems where the functions of the retina and the effects of ocular therapies can be studied in an isolated environment. The purpose of this study was to provide insight into long-term preservation of retinal tissue in culture conditions, enable a deeper understanding of the interdependence of retinal morphology and function, and ensure the reliability of the explant technique for prolonged experiments.
Retinal explants from adult mice were cultured as organotypic culture at the air-medium interface for 14 days in vitro (DIV). Retinal functionality was assessed by multielectrode array technique and morphology by immunohistochemical methods at several time points during culture.
Retinal explants retained viability for 14 DIV, although with diminishing neuronal activity, progressing neuronal loss, and increasing reactive gliosis. We recorded spontaneous retinal ganglion cell (RGC) activity up to 14 DIV with temporally changing distribution of RGC firing rates. Light responsiveness was measurable from RGCs for 7 DIV and from photoreceptors for 2 DIV. Apoptotic cells were detected beginning at 3 DIV with their density peaking at 7 DIV. The number of RGCs gradually decreased by 70% during 14 DIV. The change was accompanied by the loss of RGC functionality, resulting in 84% loss of electrically active RGCs.
Retinal explants provide a valuable tool for studies of retinal functions and development of ocular therapies. However, critical for long-term use, retinal functionality was lost before structural loss, emphasizing a need for both functional and morphologic readouts to determine the overall state of the cultured retina.
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