The contribution of the stimulatory guanine nucleotide regulatory protein (Gs) to prostaglandin E2 (PGE2)-induced hyperalgesia was investigated in the hairy skin of the rat hindpaw using the Randall-Selitto paw-withdrawal test. Although without effect alone, guanosine-5'-[gamma-thio]triphosphate (GTP gamma S) and cholera toxin--which activate Gs--both increased, while guanosine-5'-[beta-thio] diphosphate (GDP beta S)--which prevents the activation of Gs--decreased the hyperalgesia induced by PGE2. These data support the hypothesis that the action of PGE2 on primary afferent nociceptors leading to decreases in paw-withdrawal threshold is Gs-mediated.