Microtubule-associated proteins and in vitro astrocyte differentiation.

Abstract

Primary cultures of mouse brain astrocytes have been used to identify the microtubule-associated proteins (MAPs) present in this cell type at different stages of in vitro differentiation. The MAPs of the astrocyte have been identified by polyacrylamide gel electrophoresis and immunological detection. Two antisera were raised against two brain MAPs, tau and MAP-2. These antisera were also used to label the microtubular network in the intact astrocytes at different stages of the culture. The mature astrocyte contains a variety of MAP-like proteins. Anti-MAP-2 serum detected several proteins of high molecular weight (380,000, 260,000, 205,000 and 165,000 mol wt) and one microheterogeneous peak of 83,000 mol wt. Anti-tau also detected high molecular weight components (380,000 to approximately 200,000 mol wt) but not the 165,000-mol-wt peak; in addition two microheterogeneous peaks of 83,000 and 62,000 mol wt were detected by the anti-tau serum. The 62,000-mol-wt peak was therefore detected only by the anti-tau serum whereas the 83,000-mol-wt component cross-reacted with both antisera. At early stages of the culture the immature cell contained about two times less immunoreactive material than at mature stages. Qualitative changes of the high molecular weight components were also observed. In the intact cell both antisera revealed a dense fibrous network. At early stages of the culture the astroblasts were stained by the antisera but the reaction was very diffuse in the cytoplasm; few fibrous cells were intensively stained. Morphological differentiation, which began after serum deprivation and which was accelerated by forskolin (a drug that induces cyclic AMP accumulation), led to high labeling of both the cell body and the cellular processes. In the presence of colchicine the staining regressed, the processes shortened, and the cell returned to a less-apparently differentiated state.