In vitro studies of limb regeneration in adult

Explants of 99 adult newt forelimb blastemata (21to 24-day regenerates) were cultured, with and without implanted dorsal root ganglia, in modified Parker's medium (CMRL-1415) for periods of 72-144 h. Growth and differentiation of the cultured blastemata were compared with ganglionated and non-ganglionated controls fixed at the start of the culture period. The results of these experiments establish that implanted spinal ganglia are able to sustain growth and differentiation of forelimb blastemata in vitro: active proliferation amongst the blastema cells was found to be correlated with the presence of an implanted ganglion. In addition, the blastema cells exhibited a differential growth response which was most pronounced when the ganglion was eccentrically implanted 2-3 days before explantation of the limb regenerate. These results suggest that a causal relationship exists between the position of the implanted ganglion and the localization of growth within the blastema. The nerve influence, believed to be mediated by a chemical factor(s), was localized in the region of the implanted neurons, indicating that a close association between the nerves and the responding blastema cells is essential for normal growth. The importance of the physical presence of nerves for the cultivation of blastemata in vitro is emphasized.