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Ultrastructural studies on the intracellular fate of 125 I‐nerve growth factor in cultured rat sympathetic neurons
Author(s) -
Claude Philippa,
Hawrot Edward,
Parada Isabel
Publication year - 1982
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.240200102
Subject(s) - neurite , nerve growth factor , intracellular , microbiology and biotechnology , ultrastructure , chemistry , axoplasmic transport , biology , organelle , cell bodies , biophysics , anatomy , endocrinology , central nervous system , biochemistry , in vitro , receptor
Primary cell cultures of sympathetic neurons from rat were exposed to 125 I‐nerve growth factor (NGF) and the fate of the NGF in the cell was followed using electron microscopic autoradiography. The intracellular localization of NGF was determined in the cell bodies and in the proximal neurites of neurons that had been grown in three‐chamber dishes, following 5 or 24 hr of retrograde transport of NGF from the distal portions of the neurites. Label in the proximal neurites was predominantly associated with lysosomes and multivesicular bodies (MVBs), and at 5 hr elongated tubular elements were especially heavily labeled. Most of the label in the cell bodies was concentrated in lysosomes and MVBs. Lysosomes accounted for the largest fraction (45–60%) of the grains in the cell body, with a labeling density (LD = % grains/% area) of 3–5, while MVBs accounted for 5–10% of the grains with an LD of 5–20. We observed no evidence of nuclear labeling after 5 or 24 hr of retrograde transport. Mass cultures of neurons were incubated for 22 hr with NGF in the presence of the lysosomal inhibitors chloroquine (CQ, 0.05 mM) or methylamine (MA, 10 mM). In both agents the lysosomes were swollen with membranous material but still sequestered NGF, especially in CQ where the lysosomes were associated with almost 65% of the grains and had an LD of 6. CQ and MA had different effects on the morphology of the MVBs: in CQ they were few in number and compact while in MA they were numerous and appeared swollen and vacuolated. We observed no evidence for the nuclear accumulation of NGF even in the presence of the lysosomotropic agents.