Ultrastructural cytochemistry of monoamines in cultured human fetal sympathetic neurons

Abstract

The presence and storage of adrenergic neurotransmitter (monoamines) in cultured human fetal sympathetic neurons was investigated by chromate-dichromate cytochemistry, formaldehyde-induced fluorescence and potassium permanganate fixation. Monoamines were specifically identified in the neurons by the presence of an electron dense precipitate following cytochemical treatment. Reaction product was found in cell somas and processes in all chromate—dichromate treated cultures. The size range and morphology of the precipitate indicated a vesicular storage site within large dense core vesicles. Neurons fluoresced after treatment with formaldehyde vapors, further confirming the presence of monoamines. When potassium permanganate was employed as the fixative, occasional positive dense core vesicles were found but their frequency was greatly reduced from that seen in the chromate-dichromate treated cultures. These findings show that cultured human fetal sympathetic neurons retain an adrenergic phenotype during long-term serum-free culture. In addition, the storage site for the adrenergic neurotransmitter in the developing neuron is within large dense core vesicles. The lack of dense core vesicles in potassium permanganate fixed material is believed to be due to the depletion of monoamines during fixation.