Project: Research project

Project Details


The goal of this proposal is to understand the role of second messenger
systems in the regulation of glial fibrillary acidic protein(GFAP) and in
the maturation of bipotential precursor cells from the nervous system in
vitro. GFAP, the astrocyte specific intermediate filament (IF), is an
major component of the astrocyte cytoskeleton. It is developmentally
regulated and is elevated in most neurologic disease and brain traumas.
Defining GFAP regulation in cultured cells will permit a clearer
understanding of how GFAP expression is regulated in normal astrocytes
and in disease states. The effects of agents which modulate cAMP or
protein kinase C on GFAP RNA in primary astrocyte cultures and in the
human astrocytoma cell line HTB-17 are being studies. Northern blot
analysis of steady-state RNA levels, together with nuclear run-off
experiments to assess relative transcription rates, indicate that cAMP,
stimulated by treatment of cells with forskolin plus
isobutylmethylxanthine (IMX) positively regulates GFAP mRNA at the post-
transcriptional level. In contrast, GFAP steady-state mRNA is
dramatically reduced in astrocytes and HTB-17 cultures treated with the
phorbol ester PMA (10nM). This regulation is also post-transcriptional.
Recently serum has been found to inhibit the effect of cAMP-stimulatory
agents. Therefore we plan to refine our model system through the use of
chemically defined (CD) medium, minus fetal calf serum. The exact nature
of post-transcriptional up-regulation and down-regulation of GFAP mRNA
will be determined. To investigate a possible role for second messengers
in the in vitro maturation of bipotential precursor cells from the
nervous system, two types of precursors will be grown in CD medium
containing agents which modulate cAMP, protein kinase C and/ or calcium.
One type of precursor, the AC-36A cell line, has the capacity to express
either glial or neuronal characteristics; while the other, a bipotential
precursor derived from neonatal rat forebrain, can become either
astrocytic or oligodendrocytic, depending on culture conditions. We will
follow the fate of these precursors in the presence of the various agents
using immunocytochemical and DNA markers, as well as morphologic
criteria. These studies will provide insight into the molecular
mechanisms by which extracellular stimuli, such as cell-cell contact and
growth factors, induce commitment of nervous system precursors to
particular lineages.
Effective start/end date12/31/8911/30/92


  • Cell Biology
  • Genetics


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