The differential effects of cell density and NGF on the expression of tyrosine hydroxylase and dopamine β-hydroxylase in PC12 cells

Abstract

Expression of neurotransmitter phenotype during development of the nervous system is determined by several micro-environmental factors including cell aggregation. In order to delineate the role of cell aggregation and nerve growth factor (NGF) in regulating catecholamine expression, tyrosine hydroxylase (TH) and dopamine β-hydroxylase (DBH) mRNA levels were examined in PC12 cells at different cell densities with and without NGF treatment. Upon plating of PC12 cells from low density (0.3–1.0 × 10 5 cells/cm 2) to high density (0.5–2.0 × 10 6 cells/cm 2) TH mRNA levels increased 4-fold within 1 day and remained at this level for several days. In cells replated from high to low density, TH mRNA returned to original levels within 1 day. In addition to TH mRNA, TH protein and dopamine levels were also found to increase in high-density cultures. In contrast to the increase in TH mRNA, DBH mRNA decreased about 40% in cells plated from low to high density. Hence, cell density differentially regulated TH and DBH mRNA levels. Unlike cell density, NGF treatment led to a decrease in both TH and DBH mRNA levels. However, when NGF treated cells were replated from low to high density, TH and dopamine levels increased. Thus NGF did not alter the density dependent regulation of TH. Similarly, TH mRNA levels increased in F4 cells, a mutant PC12 cell line unresponsive to NGF, when plated from low to high density. DBH mRNA decreased to undetectable levels when NGF treated PC12 cells were plated to high density, demonstrating a synergetic effect of cell density and NGF treatment on DBH mRNA levels.