Yasuo Katagiri

Demonstration of a rhodopsin-retinochrome system in the stalk eye of a marine gastropod, Onchidium, by immunohistochemistry.

The stalk eye of Onchidium sp. (Gastropoda, Mollusca) is the principal photoreceptor in a multiple photoreceptive system that consists of the stalk and dorsal eyes, dermal photoreceptor cells, and photosensitive neurons. To examine the localization of photopigments, the stalk eyes were immunostained with specific antibodies to rhodopsin, retinochrome, and retinal‐binding protein (RALBP), which had been generated against squid retinal proteins. The retina of the stalk eye was divided into villous, pigmented, somatic, and neural layers. It was comprised mainly of two types of visual and pigmented supportive cells. The type 1 visual (VC1) cell was characterized by well‐developed microvilli on its apical protrusion and photic vesicles in the cytoplasm. The photic vesicles were specifically blackened by prolonged osmification. The type 2 visual (VC2) cell had less numerous, shorter microvilli on its concave apical surface and lacked photic vesicles. The anti‐squid rhodopsin antiserum was localized specifically to the villous layer that corresponded to the VC1 microvilli. With the anti‐retinochrome peptide antibody, the somatic layer showed specific but patchy, positive staining that corresponded to the cytoplasm of the VC1 cells. Because the photic vesicles are known to contain retinochrome, these results indicate that this retinochrome is localized in the VC1 cytoplasm. Anti‐RALBP antibody stained the supranuclear cytoplasm to the distal cytoplasm of VC1 cells. This is the first demonstration of the localization of RALBP in the Gastropoda Onchidium stalk eye. In squid retina that were immunostained as positive controls, the anti‐rhodopsin antibody stained rhabdomeric microvilli, the anti‐retinochrome antibody stained the inner segment and the basal region of the outer segment, and the anti‐RALBP antibody stained the outer and inner segments, respectively. These results suggest that the rhodopsin‐retinochrome system that has been established in cephalopod eyes is present in the Onchidium stalk eye. J. Comp. Neurol. 433:380–389, 2001.

Localization of Retinal Proteins in the Stalk and Dorsal Eyes of the Marine Gastropod, Onchidium

Abstract Onchidium possesses stalk eye (SE) and dorsal eye (DE) which comprise part of a unique multiple photoreceptive system. The retina of SE consists of rhabdomeric-type visual cells, whereas the DE contains two types of photoreceptor cells; ciliary-type cells in the retina and rhabdomeric-type cells in the lens. High-performance liquid chromatography (HPLC) analyses revealed the presence of 11-cis-retinal as well as all-trans-retinal in both eyes. The amount of retinal of one DE (0.17 pmol) is far less than that in one SE (0.41 pmol) in the dark-adapted Onchidium. In the dark-adapted SE, the amount of all-trans-retinal was higher than that of 11-cis-retinal. This finding is consistent with the presence of photic vesicles, including retinochrome, in rhabdomeric-type visual cells. In contrast, a higher amount of 11-cis-retinal than all-trans-retinal was present in dark-adapted DE, although this was decreased in light-adapted DE. Upon UV irradiation following treatment with sodium borohydride (NaBH4), the fluorescence (derived from retino-chrome) was observed in the somatic layer of SE. Additional fluorescence (due to rhodopsin) was observed in the villous layer upon treatment with NaBH4 after denaturation. However, only weak, obscure fluorescence of retinyl proteins was observed in the DE, not in a specific but an indefinite area on treatment with NaBH4 with or without denaturation. With fluorescence histochemistry, the localization of rhodopsin and retinochrome was confirmed at specific regions in the retina of the SE, whereas no distinct localization of these photopigments in DE was demonstrated. The amount of retinal to detect the fluorescence may be too low in the DE, or photopigments of DE may differ in chemical nature from those of SE.

Extraocular Photoreception of a Marine Gastropoda, Onchidium: Three-Dimensional Analysis of the Axons of Dermal Photoreceptor Cells in the Dorsal Mantle Examined with a High-Voltage Electron Microscope

Extraocular photoreception exists in a variety of forms among invertebrates [1], and cannot be identified visually from a surface view. Messenger [2] defined extraocular photoreception as, “a response to light that is not mediated by an eye.” Dermal light sense is one type of extraocular photoreception. In molluscs, extraocular photoreceptors are widespread and exhibit a surprising range of complexity. The shadow responses, categorized as dermal sensitivity by Messenger [2], are widespread in both gastropods and bivalves and are mediated by cells or sensory endings that remain unidentified. The marine gastropod Onchidium is thought to have extraocular photoreception based on behavioral responses to light and shadow [1–3]. Onchidium possesses paired stalk eyes (SEs) and dorsal eyes (DEs), demonstrating that both dermal and ocular systems can coexist. Yoshida [1] has described the dominant role the dermal system plays in Onchidium. However, the dermal light sensitivity of Onchidium should be reconsidered, as a photoreceptor cell has been found in the dorsal mantle [3, 4].

Three-Dimensional Reconstruction of the Axon Extending from the Dermal Photoreceptor Cell in the Extraocular Photoreception System of a Marine Gastropod, Onchidium

The marine gastropod Onchidium has a multiple photoreceptive system consisting of stalk eyes, dorsal eyes, photosensitive neurons, and extraocular dermal photoreceptor cells (DPCs). The DPCs were widespread all over the dorsal mantle and distributed singly or in groups in the dermis, but were not discernible by the naked eye. The DPC was oval in shape and large in size, and characterized by features specific to gastropod photoreceptor cells such as massive microvilli, photic vesicles, and a depolarized response. DPC-17, one of a group of 19 DPCs, was examined on serial semi-thin sections of 0.4 µm in thickness with a high-voltage transmission electron microscope (HVTEM). The axon emerged specifically from the lateral side between the distal microvillous portion and proximal cytoplasm, travelled through the connective tissue, and joined a small nerve bundle (NB). Two types of supportive cells were found along the length of the axon. The first type was a covering cell (CC) surrounding the surface of the DPC body and continuing onward to the axon sheath. DPC-17 was covered by 11 CCs, while the larger DPC-6 was only covered by four CCs. The second type was a sheath cell (ShC) wrapping the surface of the small NB where the axon of the DPC merged with undefined nerve fibers. The axon extending directly from DPC-17 was reconstructed three-dimensionally (3D) using DeltaViewer software. The 3D-reconstructed image of the sheath of the axon and the CC demonstrated the continuity between the two structures, especially when the image was rotated using DeltaViewer.