Crawford, B.J.*; Pang, T.; Campbell, S.S.: Development of the hyaline layer of the asteroid Pisaster ochraceus.

Embryos and larvae of the starfish Pisaster ochraceus are surrounded by a complex ECM layer called the hyaline layer (HL). A similar but less well organized ECM layer lines some regions of the larval gut. In material preserved with conventional fixatives to which anionic dyes have been added, the HL consists of three regions, a outer coarse meshwork (CM), a supporting layer (SL) and an intervillous layer (IV). Examination of material preserved by freeze substitution shows greater details, i.e. the SL is divided into three sub layers H1, H2, and H3 and has a boundary layer (B), consisting of short fibers which are and associated with its outer surface and project into the CM at right angles to the SL. Development of the HL begins at fertilization when exocytosis of the cortical granules releases ECM into the perivitelline space and elevates the fertilization membrane. Shortly after this plaques of dense material with attached fibers are present on the outer surface of the egg plasmalemma. Following this these plaques and fibers are associated with the tips of short microvilli suggesting that they may induce the microvillus formation. Next, the tips of some of the microvilli are joined by short regions of the H1 sub layer. Some of these H1 regions have short segments of boundary layer material associated with their outer surfaces while others are naked. Just prior to hatching, the H1 and B layers completely surround the embryo separating the developing CM and IV layers. Short segments of the H2 and H3 sub layers are also beginning to appear and extend to surround the embryo. Post hatching, the microvilli and all HL layers increase in thickness and density, particularly the H2, boundary and CM layers. The results suggest a sequential organization of HL components from ECM which is secreted into the perivitelline space. Supported by NSERC.