Abstract: The presence of binding sites for the β chemokines monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1α (MIP-1α) has recently been identified on human brain microvessels. We extend these findings in this report to reveal that such sites exemplify characteristics of the recognized major receptors for MCP-1 and MIP-1α: CCR2, and CCR1 and CCR5, respectively. Specifically, labeled MCP-1 binding to isolated brain microvessels was inhibited by unlabeled MCP-1 and MCP-3, the latter another CCR2 ligand, but not by MIP-1α. Inhibition of labeled MIP-1α binding was achieved with unlabeled MIP-1α and RANTES, the latter a β chemokine that binds to both CCR1 and CCR5, but not by MCP-1. Labeled MIP-1α binding was also antagonized by unlabeled MCP-3, which is also recognized by CCR1, and MIP-1β, which is a ligand for CCR5. Labeled MCP-1 and MIP-1α were further observed to be internalized within the endothelial cells of brain microvessels, following their binding to the microvascular surface at 37°C. Additionally, exposure of microvessels to unlabeled MCP-1 or MIP-1α was accompanied by the initial loss and subsequent recovery of surface binding sites for these chemokines, which occurred on a time scale consistent with ligand-induced endocytosis and recycling. These collective features bear striking similarity to those that characterize interactions of MCP-1 and MIP-1α with their receptors on leukocytes and underscore the concept of cognate chemokine receptors on brain microvascular endothelium.