| The Ostrich is the biggest, unflyable bird existing in the world, whose significant feeding value has assumed. Because of their incomplete physiological function, low resistance, the stress reaction inducing by the change of internal and external environment and the invasion of pathogen easily occur in ostrich chicks, Thus, the perfect function of hypothalamus-pituitary-adrenal axis (HPAA) can decide the stress ability, growth and development of ostrich chicks. New castle disease (ND) is one of the most serious infection diseases that damage ostrich cultivation. The highest disease incidence of ostrich chicks is 50%. As a stressor, ND could invade the organism and cause a series of pathological changes in body, which induced excessive activation or inactivation of ostrich chicks' HPA axis, and seriously disturbs the organismic internal environment. In the present study, ostrich chicks were divided into normal group (0d, 45d and 90d) and challenge group (45d) at random. Using the methods of HE staining, transmission electron microscopy (TEM), radioactive immunoassay (RIA), immunohistochemistry (SABC), histochemical method and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), the morphological structure, development characteristics of ostrich chicks' HPA axis and the regulation mechanism in the stress reaction after artificially challenged with chicken NDV were illustrated. The main contents and research results are as follows:1. Morphological structure of ostrich chicks' HPA axis1, 45 and 90d healthy ostrich chicks were used in the present study to investigate the morphological structure of ostrich chicks' HPA axis by the methods of HE staining and TEM. The results indicated that the ostrich chicks' HPA axis has its own features comparing with other birds. Differences between the left and right adrenal glands were found in shape, size and location. The left adrenal gland is oblong, and the right gland is often triangle exhibiting wider and weighter than the left gland. The main tissue of adrenal glands of ostrich chicks were interdigitation consisted of the interrenal tissue (adrenal cortex) and chromaffin cell (adrenal medulla). The interrenal tissue was divided into a peripheral zone (PZ) and a central inner zone (CZ), and the PZ could be divided into outer area (sub-capsular zone, SCZ) and inner area (IZ). The arrangement of SCZ cells appeared similar to that of zona glomerulosa in mammalian adrenal cortex and also the arrangement of IZ cells resembled that of zona fasciculata, as well as the arrangement of CZ cells appeared like the zona reticularis, suggesting the significant signs of zonation in African ostrich adrenal cortex. The cytoplasm of interrenal cells located beneath the adrenal capsule (SCZ) were stained lighter than that of interrenal cells located inside the adrenal gland (IZ and CZ) and contained several vacuoles. Additionally, unlike CZ cells, SCZ cells contained more mitochondria and lesser lipid droplets. The medulla cells formed cell lines, contained two types of chromaffin cells, i.e. adrenaline cells and norepinephrine cells. Their ultrastructural features appeared characterist of affluent cellular organ and considerable catecholamine-containing secretory granules, both types of granules were membrane-limited. The type I granules possessed an central core, and a confined halo could be found between limiting membrane and core; however, the type II granules had an eccentric core, which polarized one side of secretory granule and sticked limiting membrane, giving a lager lacouna appearance in another side of secretory granule. These results demonstrate there are obvious differences in the HPA axis between ostrich chicks and other poultries and the histological structure of adrenal glands was similar with that of some mammalian.2. Developmental characteristics of ostrich chicks' HPA axis1, 45 and 90d healthy ostrich chicks were selected from normal group. The development characteristics of ostrich chicks' HPA axis was detailly observed by HE staining, TEM, RIA, SABC and TUNEL. The results showed the HPA axis' histological structure acquired development unceasingly with the ages of ostrich chicks, especially in early postnatal development. The significant signs of zonation, an evident increase of adrenal medullary cells and cell organs were observed in 45d African ostrich adrenal glands. Apoptosis cells in HPA axis could be detected in any developmental period from 1d to 90d ostrich, but the apoptosis degree varying in different organs. In hypothalamus, A large number of apoptosis cells could be observed at 1d (P<0.01), then gradually decreased. A less number and an age-related decline of apoptosis in pituitary could be found. In adrenal gland, a significant apoptosis peak appeared at 45d (P<0.01), then reduced at 90d. The positive signals of GABA could be detected in HPA axis, with the expression increased with age growing of ostrich chicks. However, the average optical density of positive signals presented various regular change in different organs. The expression of GABA positive products in hypothalamus was stronger, and higher average optical density (P<0.01) could be found at 45d. In pituitary, the expression of GABA positive products was relatively weaker, and an age-related drop of average optical density could be detected. The positive signals in adrenal gland gradually increased along with the increasing of age, and the average optical density reached a peak at 90d (P<0.01). The levels of ACTH and Cor in serum changed with ages of ostrich chicks. 1d ostrich showed higher serum ACTH level, gradually increased with age growing which reached maximum levels at 45d (P<0.01), then dropped to lower than that of 1d (P<0.01). While, the levels of Cor in serum ostrich chicks appeared a decreasing trend from 1 d to 45d, come to the lowest point at 45d (P<0.01), then gradually recovered to the levels at 1d. These results demonstrate that the structure and function of HPA axis in ostrich chicks is under continuous perfection and development, especially in early postnatal period.3. The stress-mediated mechanism of ostrich chicks' HPA axis with challenged with NDVThe experimental animals were divided into challenge and control groups at random. The methods of HE staining, TEM, RIA, SABC and TUNEL were used to study the stress-mediated mechanism of ostrich chicks' HPA axis with challenged with NDV. The results are as follows: NDV could cause obvious pathological change of ostrich chicks' HPA axis in which the adrenal gland was relatively serious than that of the hypothalamus and pituitary, and the pathological degree can demonstrate the damaged degree of HPA axis' function. After artificially infected with NDV, the apoptosis in HPA axis of challenge group obviously increased, which indicated that NDV can induce the apoptosis of HPA axis of ostrich chicks. In adrenal gland the number of apoptosis increased significantly at 1 day after challenged, and reached maximum at 5 days (P<0.01), then delined. The dynamic changes of apoptosis in adrenal gland indicate that the adapting and regulating of HPA axis in the pathological stress reaction. The expression of GABA positive cells in HPA axis of ostrich chicks in challenge group reduced significantly compared with control group. The amount of GABA positive cells in adrenal gland decreased sharply at 1 day after challenged (P<0.01), reached minimum at 3 days (P<0.01), then increased again. The changes of GABA positive cells in adrenal gland indicate that GABA participate HPA axis' stress reaction in NDV pathological conditions. The level of ACTH in serum slightly increased at 1 day after challenged, reached its maximum at 5 days (P<0.01), then came back. Whenas, the level of serum Cor always increased until 7 days after challenged, then gradually became normal at 9 days. These results demonstrate that the variation of ACTH and Cor in serum of challenge group correlates closely with the damage and restoration process of HPA axis.
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