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This study provides a general approach to the presence and possible role of orexins and their receptors in the gut (three gastric chambers and intestine) of confined environment bottlenose dolphin. The expression of prepro-orexin, orexin A and B and orexin 1 and 2 receptors were investigated by single immunostaining and western blot analysis. The co-localization of vasoactive intestinal peptide and orexin 1 receptor in the enteric nervous system was examined by double immunostaining. Also, orexin A concentration were measured in plasma samples to assess the possible diurnal variation of the plasma level of peptide in this species. Our results showed that the orexin system is widely distributed in bottlenose dolphin enteric nervous system of the all gastrointestinal tract examined. They are very peculiar and partially differs from that of terrestrial mammals. Orexin peptides and prepro-orexin were expressed in the main stomach, pyloric stomach and proximal intestine; while orexin receptors were expressed in the all examined tracts, with the exception of main stomach where found no evidence of orexin 2 receptor. Co-localization of vasoactive intestinal peptide and orexin 1 receptor were more evident in the pyloric stomach and proximal intestine. These data could suggest a possible role of orexin system on the contractility of bottlenose dolphin gastrointestinal districts. Finally, in agreement with several reports, bottlenose dolphin orexin A plasma level was higher in the morning during fasting. Our results emphasize some common features between bottlenose dolphin and terrestrial mammals. Certainly, further functional investigations may help to better explain the role of the orexin system in the energy balance of bottlenose dolphin and the complex interaction between feeding and digestive physiology.

Calpains are Ca2+-dependent proteases able to cleave a large number of proteins involved in many biological functions. Particularly, in skeletal muscle they are involved in meat tenderizing during post mortem storage. In this report we analyzed the presence and expression of µ- and m-calpains in two skeletal muscles of the Marchigiana cattle soon after slaughter, using immunocytochemical and immunohistochemical techniques, Western blotting analysis and Casein Zymography. Therefore, the presence and the activity of these proteases was investigated until 15th day post-mortem during normal process of meat tenderizing. The results showed m- and µ-calpain immunosignals in the cytoplasm both along the Z disk/I band regions and in the form of intracellular stores. Moreover, the expression level of µ-calpain but not m-calpain decreased after 10 days of storage. Such a decrease in µ-calpain was accompanied by a gradual reduction of activity. On the contrary, m-calpain activity persisted up to 15 days of post-mortem storage. Such data indicate that expression and activity of both µ-calpain and m-calpain analyzed in the Marchigiana cattle persist longer than reported in literature for other bovines and may be related to both the type of muscle and breed examined.

This study provides an immunohistochemical approach to the expression of leptin in the gastrointestinal tract of the monogastric South American sea lion (Otaria flavescens), and the poligastric bottlenose dolphin (Tursiops truncatus). The specific organization of the gastrointestinal tract is examined in relation to the neuroendocrine regulation of the gut exerted by leptin. In the South American sea lion some leptin-like-immunoreactive (ir) cells, and endocrine type cells, were found in the pit of gastric mucosal folds and in the epithelium of duodenum as well as numerous neurons were detected in the submucosal and myenteric plexuses of the stomach. In the bottlenose dolphin, many leptin-like-ir cells, and exocrine type cells, were identified in the mucosal layer of the main stomach as well as several neurons and nervous fibers were detected in nervous plexuses of main stomach, pyloric stomach, proximal, and middle intestine. Our data suggest that the distribution of leptin-like peptides is similar in the two species, notwithstanding the different anatomical organization of the gastrointestinal apparatus of South American sea lion and bottlenose dolphin. These findings "suggest" the presence of a basal plan in the regulation of food intake, body weight, energy balance and of the gastrointestinal functions in general also in marine mammals with different and specific feeding habits.