Evidence for Uptake of Basement Membrane by Differentiating Ameloblasts in the Rat Incisor Enamel Organ

Abstract

Demonstration of type-IV collagen and acid phosphatase (ACPase) was carried out in the rat incisor enamel organ after the animals were fixed by perfusion with periodate-lysine-paraformaldehyde. Their incisors were dissected out, demineralized with EDTA, and prepared into 6-microns-thick frozen sections. The sections, which had been treated by means of antibody incubation for type-IV collagen, were washed with a Trismaleate buffer, incubated in Novikoff's medium for acid phosphatase (ACPase), and then incubated in a 3, 3'-diaminobenzidine solution. After osmification, the sections were embedded in epoxy resin for electron microscopy. The plasma membranes of the distal ends of the inner-enamel-epithelial cells were relatively even and were lined with a basement membrane. Type-IV collagen was localized both in the lamina densa and in the filaments attached to the lamina densa. In differentiating ameloblasts, the remarkably undulating distal plasma membranes formed irregular shallow and deep invaginations, and small cytoplasmic processes that penetrated the basement membrane. Coated pits occurred in various parts of these undulating plasma membranes. Positive reaction to type-IV collagen was observed in the invaginations and coated pits. ACPase-positive granules, present in inner-enamel-epithelial cells, increased in number and sometimes appeared close to both shallow and deep invaginations of differentiating ameloblasts. These results indicate that type-IV collagen in the basement membrane of the enamel organ is removed and degraded by differentiating ameloblasts by means of their engulfing system.