Distribution of histamine receptors in the intestines
Distribution of histamine receptors in the intestines varies among species, but our findings are similar to those previously reported in humans and guinea pigs (Thurmond, 2010, Thurmond, 2015). H1 receptors were located mainly in the gastric mucosa, smooth muscle, and lymphoid tissue, corresponding to the known function of this receptor in smooth muscle contraction and humoral immunity (Shahid et al., 2009). H2 receptors were most abundant in the gastric mucosa, supporting the major role of this receptor in controlling gastric Alisol B 23-acetate sale secretion. The H3 receptor predominantly mediates neurotransmission, and findings of intense staining of the H3 receptor antibody in all ganglia of the canine GI tract are consistent with this function (Breunig et al., 2007). Similar to other species, the H4 receptor was not as widely distributed in the canine GI tract as the other histamine receptors (Coruzzi et al., 2012). H4 receptors were, however, located in the gastric and colonic mucosa, as well as in lymphoid tissue. In humans, the H4 receptor appears to have a major influence on many aspects of the immune response, including chemotaxis of neutrophils and eosinophils, and T cell responses (Deiteren et al., 2015). Therefore, presence of the H4 receptor in lymphoid tissue was expected. To date, canine-specific histamine receptor antibodies are not commercially available. In our study, we used antibodies predicted by the manufacturers to cross react with canine histamine receptors. The specificity of the H4 receptor antibody was confirmed with western blotting, which was consistent with the limited amount of non-specific staining observed on immunohistochemistry. Non-specific staining of nuclei and collagen was noted with the H1 and H3 receptor antibodies on immunohistochemistry and, although western blotting demonstrated bands at the expected molecular weights of these histamine receptors, additional bands were also present. In addition to non-specific binding of collagen and nuclear material, the additional bands could be due to the presence of isoforms of the H1 and H3 antibodies (Xie and He, 2005). Despite some degree of non-specific staining, immunohistochemistry results with the H1 and H3 antibodies were typically consistent and repeatable among all dogs, and were also consistent with previously published findings in other species, including humans. The H2 receptor antibody yielded excellent immunohistochemical staining with no non-specific background staining, and results were consistent and repeatable. Immunohistochemical findings in the dogs in our study were also very comparable with previously documented H2 receptor distribution in other species (Thurmond, 2010). Despite multiple attempts, a band at the correct molecular weight of the H2 receptor could not be obtained with western blotting. The authors suspect that the rabbit H2 receptor antibody used in our study recognizes a native conformational protein instead of a linear sequence produced as a result of necessary reducing conditions for western blot, and is, therefore, not compatible with western blot methodology. To confirm the specificity of the H2 antibody, an additional method, such as immunoprecipitation followed by mass spectrophotometry, would need to be performed. Whether histamine receptor distribution is altered in canine GI diseases remains unknown, but human research has documented altered histamine receptor density in certain GI diseases (Sander et al., 2006, Deiteren et al., 2015, Von Rahden et al., 2011). Additionally, a novel H4 receptor antagonist, JNJ7777120, has been shown to reduce histological inflammation and clinical signs in experimentally-induced colitis (Schirmer et al., 2015). Interestingly, the dog in our study with histologic evidence of mild gastritis had stronger mucosal staining for the H1 receptor and stronger lymphoid tissue staining for the H3 receptor compared to the dogs with normal gastric biopsies.