Evidence of a new transitory extracellular structure within the developing rhombencephalic cavity

The first rudiment of the central nervous system is a simple tube, the neural tube, and its cavities become the cerebro-ventricular system. The elements located within this system, their composition and precise morphogenetic role are poorly understood. This study used transmission (TEM) and scanning (SEM) electron microscopy, and immunoelectron microscopy, and describes in the chick the development, ultrastructure, composition, and regression of a previously undescribed extracellular structure located in close relationship with the luminal pole of the developing rhombencephalic tectoria lamina. We have called it the rhombencephalic roof network (RRN). The RRN was first observed in stage 12, closely related to a cluster of apoptotic cells. Between stages 15 and 18, the RRN attained its greatest development; it was rhomboid in shape and SEM revealed a network of fibers. Between stages 19 and 22, the RRN underwent a process of fragmentation and regression, and was not observed after stage 23. With TEM, the RRN appeared formed by amorphous ruthenium-red-positive material and sets of tubes between 4 and 25 nm in diameter. Each tube was formed by the superposition of annular units. Immunolabelling showed the presence of laminin and heparan sulfate proteoglycan in both the amorphous material and fibers; the former also contained tenascin. In terms of ultrastructure and composition, the fibers were similar to one the basic components of the lamina densa of basement membranes. The developing tectoria lamina exhibited openings as early as stage 12+, showing that the neural cavity is not a closed system and that the neural tube fluid (NTF) could be a circulating liquid. The presence in the RRN of three molecules of the extracellular materials actively involved in several developmental processes and the very early appearance of the RRN suggest that this structure plays a developmental role in rhombencephalic morphogenesis.