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Glycoprotein 120 (gp120) is an HIV envelope glycoprotein. Gp120 can directly stimulate the primary sensory afferent neurons and cause hyperalgesia. The P2X 3 receptor in dorsal root ganglia (DRG) is involved in the transmission of pain. In this study, we aimed to explore the role of the P2X 3 receptor in gp120-induced neuropathic pain. Our data showed that mechanical and thermal hyperalgesia in rats treated with gp120 were increased compared to those in the control group. The expression levels of the P2X 3 mRNA and protein in rats treated with gp120 were higher than those in the control group. The P2X 3 antagonist A317491 decreased mechanical hyperalgesia and thermal hyperalgesia and the up-regulated expression levels of P2X 3 mRNA and protein in rats treated with gp120. A317491 decreased ERK1/2 phosphorylation levels in the gp120-treated rat DRG. In addition, P2X 3 agonist α,β-methylene ATP (α,β-meATP)-activated currents in DRG neurons cultured with gp120 were higher than those in control neurons. The inhibitory effect of A317491 on α,βme-ATP-induced currents in DRG neurons from the gp120-treated neurons was larger than that for control neurons. Molecular docking data showed that A317491 may be acted in the gp120 protein to inhibit the gp120 initiated the P2X 3 activation, decrease the sensitizing DRG primary afferents and reduce the signal transmission of neuropathic pain in gp120-treated rats. Therefore, the inhibition of the P2X 3 receptor in rat DRG neurons relieved gp120-induced mechanical hyperalgesia. [ABSTRACT FROM AUTHOR]