Investigations on circumferential microfilament bundles in rat retinal pigment epithelium

The presence of contractile proteins in normal rat retinal pigment epithelium has been studied using fluorescence and electron microscopy. Investigations using the F-actin binding toxin, phallacidin, coupled to the fluorochrome nitrobenzoxadiazole, revealed a band of fluorescence at or near the cell membrane. Immunofluorescent observations with anti-myosin and anti-alpha-actinin antisera gave similar results. Electron microscopy employing glutaraldehyde-8% tannic acid fixation revealed the presence of a circumferential microfilament band beneath the pigment epithelial apical surface that is closely associated with the plasma membrane and junctional complexes. Freeze-fracture studies confirmed the relationship of this band to the junctional complexes. The microfilament band measures approximately 0.5 micron +/- 0.2 micron in width and is composed of numerous 6 to 7 nm filaments. Some microtubules are seen in regions around the band, but no organelles appear to be associated with this structure. In en face sections through the zonula adherens, the circumferential microfilament band is associated with 30-nm electron-dense particles that are bound to the internal side of the membrane. Morphological evidence suggests that these may serve in anchoring the band to the membrane and assist in aligning the microfilament bands of adjoining cells. In the subapical cytoplasm, a microfilament bundle network was detected that interfaced with the circumferential microfilament band. In some cases, pigment epithelium was incubated in media-199 containing 25 to 50 ng/ml phallacidin prior to fixation. Circumferential microfilament bands of tissues treated in this manner exhibited a striated appearance.