Purchase this article with an account.
K. Homma, M. Mandai, M. Takahashi; Measurement of Neuronal Activities of Transplanted Retinal Cells in Retinal Degeneration Models. Invest. Ophthalmol. Vis. Sci. 2010;51(13):5246.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
In order to restore or retain the visual functions in the RP or AMD patients, retinal cell transplantation is a hopeful technique. However, whether the transplanted cells could function as photoreceptors is unclear. In this study, for the detection of neuronal activities, we utilized multi-electrode array (MEA) system in the transplanted retinal areas, and Ca2+ imaging technique in the transplanted cells.
The dark-adopted retinas of mice were taken out from eye cups, and used for the ex-vivo study. Total of 8x8 electrodes in 1.2 mm square (Alpha med science, Japan) were positioned against each retina, and electrical recording was obtained with each probe after light stimulation. For Ca2+ imaging, the retinas were dissociated or sliced and cultured. When the experiment started, these cells or tissues were stained with Fura-2, Ca2+ indicator. Fluorescent images were obtained by using Aquacosmos imaging system (Hamamatsu Photonics, Japan).
By using MEA system, ERG-like "a-waves" and "b-waves" were detected. The light-induced "a-waves" were observed in the retinas of P12 and older, with increase in amplitude until P15, by which time the response level reached equivalent to that of adult retina to light stimulation. Next, we transplanted juvenile retinal neurons into sub-retinal spaces in C3H/Hej (rd1) mice, retinal degeneration models. These cells located in outer nuclear layers and showed photoreceptor-like morphologies, and synaptic marker, Bassoon, in the outer plexiform layer. Despite these successes in cell integration into outer nuclear layers, MEA measurement could not detect obvious improvement of "a-waves" in the transplanted regions. These results could be accounted for by the disruption of outer segment layer where the "a-wave" originates. We also investigated neuronal activities of transplanted cells by using Ca2+ imaging technique. In primary culture, we detected Ca2+ responses in photoreceptors by the application of glutamate or 80mM KCl, though these responses were weaker than that of non-photoreceptor retinal neurons. Similar Ca2+ responses were obtained from transplanted cells in retinal slice culture. These responses were comparable with those of endogenous photoreceptors.
Multi-electrode array system could detect subtle light responses of photoreceptors, though could not detected those of transplanted cells in rd1 mice. However, transplanted cells showed Ca2+ responses by application of glutamate or membrane depolarization.
This PDF is available to Subscribers Only