Visualization of [3H]docosahexaenoic acid trafficking through photoreceptors and retinal pigment epithelium by electron microscopic autoradiography.

PURPOSE

[3H]docosahexaenoic (DHA) acid was followed through the retinal pigment epithelial cells and photoreceptors for up to 5 days after injection to specifically determine which membrane systems of the retinal pigment epithelial cells are used in the handling of [3H]DHA after shedding and phagocytosis of rod tips.

METHODS

Frogs (Rana pipiens) were injected with [3H]DHA in the dorsal lymph sacs, and maintained for up to 5 days. Retinas were processed for electron microscopic autoradiography, stored for various periods of time, and then analyzed by transmission electron microscopy.

RESULTS

After 1 day, [3H]DHA had accumulated within photoreceptor ellipsoids, and had begun to appear as dense label in newly formed discs. By day 5, the basal region of dense label had expanded apically. Newly shed rod outer segment tips were diffusely labeled; but occasionally after several hours, they acquired additional label as they moved near Bruch's membrane. Retinal pigment epithelial cytoplasm maintained a constant level of label, with myeloid bodies sometimes slightly labeled. Oil droplets of the retinal pigment epithelium accumulated dense label throughout this study.

CONCLUSIONS

When [3H]DHA enters the retinal pigment epithelium, some is retained within oil droplets, whereas the rest is passed on to the photoreceptors. [3H]DHA is initially taken up by inner segments and then dispersed to photoreceptor synaptic terminals as well as to ellipsoids where discs are assembled. Phagosomal labeling exactly matches rod outer segment tips, but occasionally increases as degradation occurs near Bruch's membrane. Normally, density of label remains constant throughout the degradation process.