Histochemistry of the developing notochord, perichordal sheath and vertebrae in Danforth's short-tail (sd) and normal...

The development of the notochord, perichordal sheath and vertebrae was studied in C57BL/6 and Danforth's short-tail (Sd) mutant mice on days 9-14 of gestation, using histochemical stains to detect possible extracellular matrix (ECM) components or precursors. Stains used were periodic-acid Schiff (PAS) after diastase treatment (glycoproteins, neutral polysaccharides) and alcian blue (glycosaminoglycans). Embryos from C57BL/6Sfd mice were analyzed to establish a normal baseline. In 9-day normal (C57BL/6Sfd; +/+) embryos the notochord is an uninterrupted structure and contains PAS-positive, diastase-resistant granules, whereas in abnormals (Sd/+; Sd/Sd) the notochord is discontinuous and exhibits few, if any, granules. A notochordal sheath is present in normal and abnormal embryos on day 10 and stains with PAS, alcian blue and aniline blue; subsequently, it increases in thickness in normal, but not defective, embryos. In normal embryos, the notochord shows dilatations, and notochordal cells become vacuolated from 13 to 14 days. In contrast, the notochordal fragments of abnormals never develop dilatations, nor do the cells vacuolate. Organization of mesenchymal cells into specific patterns is observed initially in 11-day normal embryos; further mesenchymal organization into vertebral and intervertebral disc analgen occurs during days 13-14. In abnormal embryos, disturbance of mesenchymal cell organization is evident as early as day 11, and by day 12 aberrant patterns of organization have emerged. Mesenchymal cells of abnormal embryos also lack the typical distribution of PAS-positive, diastase-resistant granules that occurs in normal speciments. The possible relationship of these granules in notochordal and mesenchymal cells to ECM materials is discussed.