Histamine H3 receptor ligands are usually tested in guinea-pig intestine preparations. A possible desensitization of agonist-induced twitch inhibition was studied in longitudinal muscle-myenteric plexus from ileal segments. A cumulative concentration-response curve for R-alpha-methylhistamine was made; when a second curve was made 30 min afterwards, a marked decrease of pD2 and a more modest decrease of Emax were observed without changes in tissue sensitivity to electrical stimulation or morphine inhibition. At 120 min, pD2 and Emax were not different from those for the first curve. Receptor desensitization seems homologous and reversible and could interfere with repetitive testing of histamine H3 receptor ligands.