It has been suggested that the agonist-induced internalization of G-protein-coupled receptors from the cell surface into intracellular compartments regulates cellular responsiveness. We previously reported that Gq/11 -protein-coupled human histamine H1 receptors internalized via clathrin-dependent mechanisms upon stimulation with histamine. However, the molecular determinants of H1 receptors responsible for agonist-induced internalization remain unclear. In this study, we evaluated the roles of the intracellular C-terminal of human histamine H1 receptors tagged with hemagglutinin (HA) at the N-terminal in histamine-induced internalization in Chinese hamster ovary cells. The histamine-induced internalization was evaluated by the receptor binding assay with [3 H]mepyramine and confocal immunofluorescence microscopy with an anti-HA antibody. We found that histamine-induced internalization was inhibited under hypertonic conditions or by pitstop, a clathrin terminal domain inhibitor, but not by filipin or nystatin, disruptors of the caveolar structure and function. The histamine-induced internalization was also inhibited by truncation of a single amino acid, Ser487, located at the end of the intracellular C-terminal of H1 receptors, but not by its mutation to alanine. In contrast, the receptor-G-protein coupling, which was evaluated by histamine-induced accumulation of [3 H]inositol phosphates, was potentiated by truncation of Ser487, but was lost by its mutation to alanine. These results suggest that the intracellular C-terminal of human H1 receptors, which only comprises 17 amino acids (Cys471-Ser487), plays crucial roles in both clathrin-dependent internalization of H1 receptors and G-protein signaling, in which truncation of Ser487 and its mutation to alanine are revealed to result in biased signaling toward activation of G-proteins and clathrin-mediated internalization, respectively.