The Chinese alligator (Alligator sinensis), a freshwater crocodilian endemic to China, is one of the most endangered crocodilian species; up to this date, very little is known about the endocrine regulation of its metabolic activities during different physiological states. In this study, we characterized the structure of the prepro-vasoactive intestinal peptide in Chinese alligator (prepro-caVIP) for the first time and examined its expression profiles in various tissues during the active and hibernating periods. The prepro-caVIP cDNA consists of a 221-bp 5'-untranslated region (UTR), a 606-bp complete coding region (CDS), and a 312-bp 3'-UTR, which encodes the 201-amino acid prepro-caVIP containing a 28-amino acid vasoactive intestinal peptide (VIP) and a 27-amino acid PHI (peptide histidine isoleucine). Multiple alignment analysis showed that VIP shares 100% identity with the given birds, reptiles, and African clawed frog, and 89% identity with mammals, 96% with fishes. Real-time quantitative PCR showed that the prepro-caVIP is widely expressed in all the examined tissues, and the expression level is significantly higher in small intestine, stomach, pancreas, lung, and skeletal muscle, whereas lower in heart, liver, spleen, kidney, ovary, and oviduct. During hibernation, the expression level of caVIP was significantly decreased in small intestine (P < 0.01), pancreas, and skeletal muscle (P < 0.05), whereas significantly increased in liver, spleen, and lung (P < 0.01). The wide distribution of caVIP and its differential expression changes in various tissues during hibernation implicated that it might play multiple effects in Chinese alligator and participate in the physiological adaptation of various organs in a paracrine and/or neurocrine manner.