The ectomesenchymal-endodermal interaction system of Triturus alpestris in tissue culture: morphological and histochemical characterization of developing neural derivatives.

In the ectomesenchymal-endodermal interaction system (EEIS), a hanging drop culture in which an explant of neural fold from behind the prospective ear region of a Triturus alpestris neurula together with a piece of ventrolateral pharynx endoderm is cultivated, the differentiation of a variety of neural crest-derived cells (nerve cells, pigment cells, cartilage cells, perichondrial cells, Schwann cells) can be followed. While cartilage cells develop only in contact to pharynx endoderm, the differentiation of neural crest-derived neuroblasts, rhombencephalic brain derivatives and epidermis which segregate from the explant of neural fold, does not depend on the presence of pharynx endoderm. The development of neural derivatives was studied with respect to morphology and histochemistry. Outgrowing axons may be observed 2-3 days after the beginning of cultivation. The fibres increase in thickness and length during further development and frequently make contact with other cells. Cytophotometric measurements of Feulgen-stained neural crest cells in the EEIS revealed that 72% of cells had a 2c and 9% a 4c DNA content while 18% were in S phase. Nerve cells had a 2c DNA content and prospective chondroblasts were in S phase. About 60-70% of S phase and 4c cells were observed at the margin of the culture. Neural crest-derived neuroblasts and melanoblasts obviously reacted catecholamine-positive upon formaldehyde-induced fluorescence (FIF), while axons from the brain derivative were catecholamine-negative but acetylcholinesterase (AChE)-positive. Neural crest cells prior to visible differentiation and neural crest-derived neuroblasts were AChE-negative. Catecholamine-positive (= fluorescing) cells can be recognized as either neuro- or melanoblasts prior to displaying phenotypic features. In cultures incubated with 3H-Dopa, freeze-dried, treated with the FIF-procedure, and subjected to autoradiography, only melanoblasts specifically accumulated the label.