DISTRIBUTION AND MORPHOLOGY OF NEUROPEPTIDE Y AND β-ENDORPHIN ENDOCRINE CELLS IN THE GUT OF GREY MULLET,MUGIL CEPHALUS L.

Neuropeptide Y (NPY), an amidated 36 amino acid peptide and a member of the pancreatic polypeptide family, was first isolated from porcine brain.β-endorphin(β-EP), a member of opioid peptide family, was isolated from the pituitary gland as a fragment of the carboxyl terminal sequence of β-eipotropin consisting of 61 91 amino acids. Biochemical and immunochemical studies have shown that NPY and β-EP are widely distributed in the nervous system (brain and spinal cord),pituitary and retina of vertebrates from fishes to mammals,and involved in a wide array of physiological effects, such as regulation of the secretion of piturtary hormones, control of circadian rhythms, memory processing, food and water intake, regulation of feeding, energy balance and plasma insulin levels. Available information on their distribution in the gastroenteritic tract is much restricted. Chiba (1998) reported the coexistence of sertonin and neuropeptide Y in the gut epithelium of the cloudy dogfish, Scyliorhinus torazame. Zhang et al.(1994) investigated the distribution of β-EP in the pancreas of rabbit. As to the teleost, little information was available. In the present study, the identification and localization of endocrine cells in the various parts of intestinal tract in grey mullet, Mugil cephalus were studied by immunocytochemical method of Strept Avidin Biotin Complex (SABC)and two kinds of rabbit antisera raised against mammalian neuropeptide Y and β-endorphin. The grey mullet Mugil cephalus, used in the present study were collected from Dajing Mariculture Farm in Ronghai, Fujian Province. The fish were 21.5-24.7 cm and weighted 90-225g. They were kept deprived of food in aquarium for 2 days. The fish were vivisected, guts were detached and divided into four blocks induding anterior part of anterior intestine, posterior part of anterior intestine, midgut and posterior intestine. The tissue blocks were fixed in Bouins solution without acetic acid for 8-12h, then dehydrated in a series of alcohol and embeded in paraffin. Sections at a thickness of 6 um in transverse plane were immunocytochemically stained with a commercial kit using the Strept Avidin Biotin Complex (SABC)method. The sections were de waxed, hydrated and incubated in 3% H 2O 2 for 20 min to remove endogenous peroxidase activity. Incubation was performed in the primary antibodies, rabbit anti NPY antibody (1∶1000) and rabbit anti β-EP antibody (1∶400),at 4℃ for 24 h. After three washes in PBS, sections were incubated in the secondary antibody (biotinylated goat anti rabbit immunoglobulin G, 1∶100)for 30 min at room temperature, then rinsed in PBS three times and incubated for 30 min at room temperature with strep avidin biotin peroxidase complex(ABC) The tissue bound peroxidase was visualized for 10-30 min with diaminobenzidine (DAB)method. Immunocytochemistry showed NPY and β-EP immunoreactive cells were localized at varying degrees in the anterior part and posterior part of anterior intestine,midgut and posterior intestine. Immunoreactive cells brown, on a unstained background, were easily identified. NPY immunoreactive cells, majority of which were open type with cytoplasmic protrusion and minority of which were close type, had polymorphi and were mainly localized in the middle and apical part of intestinal fold and minorly in the basic part of intestinal fold. Conversly,β-endorphin immunoreactive cells were almost close type and were mainly localized in the basic part of intestinal fold. NPY were distributed mainly in the anterior part and posterior part of anterior intestine at a density of 18.7 cells/mm2 and 26.3 cells /mm2, respectively. But only a few of NPY cells were found in the midgut and posterior intestine, at a density of 5.1 cells/mm2 and 5.5 cells/mm2, respectively. The highest distribution density of β-endorphin, about 31.5cells/mm, was observed in the posterior intestine, and then, the density decreased sequentially from anterior intestine to midgut, at a density of 26.9, 11.6and 9.1 cells/mm respectively.