Conditioned medium from activated microglia promotes cholinergic differentiation in the basal forebrain in vitro

Area of Science:

• Neuroscience
• Developmental Biology
• Cell Biology

Background:

• Previous studies showed interferon-gamma (IFN-gamma) significantly increases choline acetyltransferase (ChAT) in cultured septal nuclei with adjacent basal forebrain (SN/BF) cells.
• Younger cultures exhibited a more profound response to IFN-gamma, suggesting a role in neuronal precursor differentiation.

Purpose of the Study:

• To investigate whether IFN-gamma promotes the cholinergic differentiation of neuronal precursors.
• To identify potential mediators of IFN-gamma's action on cholinergic differentiation.

Main Methods:

• Utilized retroviral labeling (beta-galactosidase) to trace neuronal precursor populations in SN/BF cultures of varying ages.
• Quantified ChAT-positive cells within retrovirally labeled clones after IFN-gamma treatment.
• Assessed the response of microglia to IFN-gamma and analyzed conditioned media from activated microglia and macrophage cell lines (RAW 264.7).

Main Results:

• IFN-gamma treatment increased the proportion of ChAT-positive cells in labeled clones by up to 8.8-fold, indicating promotion of cholinergic differentiation.
• Basic fibroblast growth factor (bFGF) increased clone size but not ChAT-positive cell proportion; nerve growth factor (NGF) had no effect.
• Activated microglia responded to IFN-gamma, and conditioned media from activated microglia and RAW 264.7 cells mimicked IFN-gamma's effect on ChAT activity and differentiation.

Conclusions:

• IFN-gamma promotes cholinergic differentiation in neuronal precursors within the SN/BF.
• A microglial-derived, stable, large protein molecule likely mediates IFN-gamma's effects.
• Activated microglia and similar cells produce factors that enhance cholinergic differentiation, suggesting a role in normal neural development.