Neural cell adhesion molecule (NCAM) constitutes a group of cell surface gl
ycoproteins that regulate cell-cell interactions in the developing and adul
t brain. Endocytosis is a mechanism which dynamically controls the amount o
f cell surface NCAM expression and may involve the rapid changes occurring
in NCAM expression under certain physiological or pathological conditions.
However, the endocytic pathway of NCAM is presently unknown. Using astrocyt
es in culture and immunofluorescence we show that NCAM is internalized and
that the immunolabelling presents a high degree of colocalization with clat
hrin, alpha -adaptin and transferrin, suggesting that NCAM is endocytosed b
y a clathrin-dependent pathway. Potassium depletion which disrupts clathrin
-mediated endocytosis, inhibited internalization of NCAM. Electron microsco
py and immunogold studies also demonstrate that the surface of clathrin-coa
ted vesicles are also immunolabelled for both alpha -adaptin and PSA-NCAM,
the highly sialylated isoform of NCAM. Furthermore, immunoprecipation studi
es demonstrate that NCAM is associated with both clathrin and alpha -adapti
n, a component of adaptor complex AP-2, in brain, neurons and astrocytes. T
hese findings indicate that NCAM is mainly endocytosed via clathrin-coated
vesicles, suggesting a possible mechanism that may contribute to the rapid
changes in NCAM expression at the cell surface.