Date of Award
Purpose: To examine retinal horizontal and bipolar cells cultured from goldfish (Carassius auratus), black crappie (Pomoxis nigromaculatus) and bluegill (Lepomis macrochirus) on the light and electron microscope levels; and to compare them with cells from a more established model, the wild white bass (Roccus chrysops). Methods: Retinas from all species were dissociated by methods developed with wild white bass (Dowling et al., 1985). Cell type was identified within the first 3 hr and cells were cultured for up to 2 wk in L-15 medium. At various intervals, cells were fixed in glutaraldehyde, photographed with phase LM and processed for SEM by either the ethanol dehydration method or the OTOTO method (Thompson et al., 1992 and modified Gabriel, 1982). Specimens were viewed with an Hitachi SEM. Each technique was evaluated for safety, simplicity, and image quality. Results: Bluegill and goldfish retina yielded few cells of inferior quality. Neurite outgrowth was limited. In contrast, crappie retina yielded adequate number of recognizable cell morphology, including: rods, cones, horizontal cells (H1s, H2s, H3s, H4s), and bipolar cells (BPs). These cells closely resembled the same cell types obtained from white bass (Vaughan & Lasater, 1990). The most numerous cells were H2s and H4s. Neurite outgrowth was acceptable and long-term survival of HCs and BPs was routine. H1s were observed still in contact with one another and presumably still connected by gap junctions. Neither of the two SEM techniques applied yielded superior results; rather both resulted in distracting artifacts. Conclusion: Crappie are easier to capture and maintain in the laboratory than wild white bass and their retinal cells perform in culture more like those from wild white bass than the other species tested. SEM observations confirm some axon-like and dendrite-like process growth of the bipolar cells and marginal growth in the H1s. The ethanol dehydration method for preparing cells for SEM is quick, simple and cost effective although it lacks the contrast and high resolution provided by the OTOTO method.
Steinfeldt, Lori Ann, "A New Model Species for Cultured Teleost Retinal Cell Studies: Light and Scanning Electron Microscopy" (1994). Undergraduate Honors Capstone Projects. 321.
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Dana K. Vaughan