NGF and Anti-NGF: Evidence against Effects on Fiber Growth in Locus coeruleus from Cultures of Perinatal CNS Tissues

Abstract

The present study examines whether the developing noradrenergic neurons of locus coeruleus depend on endogenous nerve growth factor (NGF) for nerve fiber production and if exogenous NGF stimulates fiber growth in this nucleus, using a collagen gel tissue culture technique. Lucus coeruleus from perinatal rat brain was used in three culture experiments: (1) lucus coeruleus, parietal cerebral cortex, and the superior cervical ganglion, prepared from newborn rats and cultured in different sectors of the same dishes; (2) locus coeruleus and parietal cerebral cortex from 17-day-old rat fetuses cultured in the same manner, and (3) locus coeruleus from 17-day-old rat fetuses co-cultured with spinal, sympathetic and ciliary ganglia from 8-day chick embryos. Experiments 1 and 2 were run with and without addition of NGF and anti-NGF, experiment 3 with and without anti-NGF. Total fiber production in all cultured tissues was evaluated daily by dark field and phase contrast microscopy during 4 days. Adrenergic nerve fiber production was then studied in the same locus coeruleus and superior cervical ganglia from the rats by Falck-Hillarp fluorescence histochemistry. Locus coeruleus and cortex cerebri from fetal rats produced dense fiber halos in culture. Locus coeruleus from newborn rats produced considerably less fibers, newborn cortex only few fibers. Superior cervical ganglia from the same newborn animals produced no or almost no fibers. Addition of NGF was not able to stimulate fiber growth in locus coeruleus nor in cortex cerebri as observed both in the living cultures and by fluorescence microscopy. Likewise, addition of anti-NGF did not affect fiber production in the CNS areas. The negative results with NGF on newborn locus coeruleus and cortex cerebri were in sharp contrast to the strong, highly significant fiber growth response demonstrated by the superior cervical ganglion from the same animals cultured in the same dishes. The third experiment tested whether locus coeruleus in tissue culture contained or produced nerve growth factor or any one of the three chick embryo ganglia. No response whatsoever in these three ganglia was observed. It is concluded that the developing locus coeruleus area does not contain or produce NGF, does not depend on NGF for fiber production, and is not stimulated by exogenous NGF.