EFFECTS OF NERVE GROWTH FACTOR (NGF) ON DIFFERENTIATION OF TRIGEMINAL GANGLIA IN CELL CULTURES

Completely dissociated trigeminal ganglia from 10-day old chick embryos were cultured directly on glass or on collagen-coated coverslips in double-coverslip assemblies. The nutrient fluid medium consisted of human placental serum, chick embryo extract and Tyrode's salt solution. In control cultures without nerve growth factor (NGF) nerve cells were predominant, which regenerated one or two processes and stood out as isolated elements on the background of nonneuronal cells. Their perikarya in course of cultivation increased in diameter (from 10-15 μm to 30-45 μm), but never became surrounded by satellite cells. After 3-4 weeks in vitro they degenerated without attaining complete structural differentiation. To prevent an initial aggregation of neuroblasts in test cultures during the first week of cultivation, the content of NGF in these cultures was increased by successive daily addition of extract of male mouse submaxillary salivary glands to the culture medium. Later the medium with optimal NGF-concentration was renewed every 2 days. This technique stimulates the development of a very rich fibrillar network of nerve fibers among supporting cells as well as two types of sensory nerve cells. The first type consists of rounded cells, resembling both in morphology and time of survival those of the control cultures. A second type, which is the dominant one in NGF-treated cultures is in close contact with the glas or collagen substrate. Only in test cultures after 3 weeks of cultivation these flattened perikarya (15-80 (μm in diameter) recover from chromatolysis to the normal mature Nissl pattern. The somata of these cells are associated with 2-6 satellite cells, their thickened fibers being enveloped by Schwann cells. The nerve processes reach a length of several millimeters. However, there was no evidence for myelin formation. Preliminary studies using media with high glucose content in combination with insulin gave no positive results in this direction. Except for myelinogenesis the neuron-satellite-Schwann cell complexes resemble their in vivo counterparts in some morphological aspects. They can be maintained in good condition for periods up to 2 months in vitro. A limiting factor for the maintenance of the cultures is given by retraction of the collagen film. The patterns of differentiation of neurons in cell and explantate cultures of trigeminal ganglia from chick embryos are compared. © 1969 Springer-Verlag.