Transient receptor potential ankyrin 1 (TRPA1) is a nonselective cation channel that is activated by a variety of stimuli and acts as a nociceptor. Mouse and human TRPA1 exhibit different reactivity to some stimuli, including chemicals such as menthol as well as cold stimuli. The cold sensitivity of TRPA1 in mammalian species is controversial. Here, we analyzed the reactivity of heterologously expressed canine TRPA1 as well as the mouse and human orthologs to menthol or cold stimulation in Ca2+-imaging experiments. Canine and human TRPA1 exhibited a similar response to menthol, that is, activation in a concentration-dependent manner, even at the high concentration range in contrast to the mouse ortholog, which did not respond to high concentration of menthol. In addition, the response during the removal of menthol was different; mouse TRPA1-expressing cells exhibited a typical response with a rapid and clear increase in Ca2+i (“off-response”), whereas Ca2+i in human TRPA1-expressing cells was dramatically decreased by the washout of menthol and Ca2+i in canine TRPA1-expressing cells was slightly decreased. Finally, canine TRPA1 as well as mouse and human TRPA1 were activated by cold stimulation (below 19–20°C). The sensitivity to cold stimulation differed between these species, that is, human TRPA1 activated at higher temperatures compared with the canine and mouse orthologs. All of the above responses were suppressed by the selective TRPA1 inhibitor HC-030031. Because the concentration-dependency and “off-response” of menthol as well as the cold sensitivity were not uniform among these species, studies of canine TRPA1 might be useful for understanding the species-specific functional properties of mammalian TRPA1.