There are numerous studies of the pruritus-producing effects of histamine, serotonin, tryptase, substance P and interleukin-2 in humans and mice, but very little reported in dogs even though a common reason dogs are presented to veterinarians is pruritus. The aim of this study was to determine whether substances known to cause pruritus in humans also cause pruritus in dogs. Twenty-five clinically healthy research beagle dogs were included in the study. All dogs first received an intradermal injection of 0.05 mL saline as a control substance and were video-recorded for 20 min before and after the injection. Twenty-four hours later the dogs were randomly divided into five groups of five dogs each and randomly assigned to receive histamine, serotonin, tryptase, substance P or interleukin-2 injected intradermally each at the volume of 0.05 mL. On subsequent days, increasing concentrations of each substance were used. Before (baseline) and after the injection of each concentration of the substances, the dogs were video-recorded for 20 min. The frequency and character of pruritus episodes (scratching, licking, chewing, rubbing or rolling) were noted and these data were used for statistical analysis. The number of pruritus episodes was compared among baseline, saline and the different concentrations of each substance. The results showed that dogs did not have a significant increase in pruritic behaviour above baseline or saline after injection of any of the investigated substances (generalized linear model; P = 0.23).