The endoplasmic reticulum (ER) in honeybee photoreceptors is organized into structurally distinct subregions. The most prominent of these, the submicrovillar network of ER cisternae, is tightly associated with actin filaments. Electron microscopic techniques have demonstrated that the ER-associated actin filaments are regularly spaced at 60-80 nm and cross-bridged by filamentous structures. A polyclonal antibody against Drosophila alpha-spectrin has been used to examine the distribution of spectrin in the photoreceptors. On Western blots of bee retina, the antibody identifies a 260-kDa protein that exhibits biochemical and immunological properties characteristic of alpha-spectrin. Immunofluorescence microscopy has shown that alpha-spectrin codistributes with the submicrovillar ER but not with other ER subdomains. After cytochalasin-B-induced depolymerization of the ER-associated F-actin system, alpha-spectrin remains colocalized with the ER, indicating that alpha-spectrin is bound to the ER membrane. The F-actin/spectrin system associated with the submicrovillar ER may stabilize the shape of this ER subcompartment and may play a role in maintaining functional ER subregions.