125I-thrombin binds to clustered receptors on noncoated regions of mouse embryo cell surfaces.
We used electron microscope autoradiography (EMAR) to visualize the interaction of 125I-thrombin with its surface receptors on mouse embryo (ME) cells. Autoradiographic grains were spaced over the surface of cells in a periodic nonrandom pattern, indicating 125I-thrombin association with clusters of thrombin receptors. The grain spacing varied slightly from cell to cell, indicating subpopulations of cells with different numbers of thrombin receptors. The average distance between grains on ME cells after binding 125I-thrombin (125 ng/ml) at 37 degrees C was 1.65 +/- 0.49 microns. The average distance between grains on prefixed cells and cells incubated with 125I-thrombin at 4 degrees C was not significantly different from that observed at 37 degrees C. This indicates that thrombin receptors are clustered before thrombin binding and that the thrombin receptor aggregates do not redistribute into large aggregates on the surface of cells subsequent to thrombin binding. The number of grains per cluster also does not change under these three binding conditions. Thus, the number of occupied receptors in each cluster appears to be constant. On the basis of the average grain number and spacing, we estimate that each cluster is approximately 400 nm in diameter containing approximately 550 thrombin-binding sites. These receptor-clusters are not associated with specialized structures or coated regions of the membrane. Additionally, grains observed within cells were not found associated with coated vesicles. Therefore, neither the clustering patterns nor internalization of 125I-thrombin are characteristic of molecules which bind to receptors and are internalized by receptor-mediated endocytosis.
D000818:Animals; D001345:Autoradiography; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D003034:Coated Pits, Cell-Membrane / Q000032:analysis; D004622:Embryo, Mammalian; D007457:Iodine Radioisotopes; D051379:Mice; D008854:Microscopy, Electron; D011956:Receptors, Cell Surface / Q000032:analysis / Q000378:metabolism*; D018179:Receptors, Thrombin; D013917:Thrombin / Q000378:metabolism*
D007457:Iodine Radioisotopes; D011956:Receptors, Cell Surface; D018179:Receptors, Thrombin; D013917:Thrombin