The pharmacological property that most distinguishes rat P2X4 receptors from other P2X receptors is their insensitivity to the purinoceptor antagonists, suramin and pyridoxal-phosphate-6-azophenyl-2',4'-disulphonic acid (PPADS). The molecular basis of this insensitivity is not known. Here, we investigated the possibility that histidine residues in the extracellular loop of P2X4 receptors may be involved in the antagonist sensitivity of these receptors. We found that histidine mutation H241A in the rat P2X4 receptor produced receptors that are sensitive to suramin and PPADS. In contrast, mutation H140A or H286A did not significantly alter antagonist sensitivity. In addition, mutation H241A in the human P2X4 receptor significantly increased antagonist sensitivity. The results suggest that histidine 241of P2X4 receptors is involved in regulating the antagonist sensitivity of these receptors.