Distribution of growth hormone-like immunoreactive cells and somatostatin receptors in the nervous system and Hatschek's pit of amphioxus, Branchiostoma belcheri

Using immunohistochemical method and double staining technique, the localization of growth hormone (GH) and somatostatin receptors in the nervous system and Hatschek's pit of amphioxus has been investigated. The results showed that the growth hormone-like nerve cells and endocrine cells as well as three subtypes of somatostatin receptors exist in the nervous system and Hatschek's pit, and GH-like nerve cells and endocrine cells co-exist with three subtypes of somatostatin receptors in the brain vesicle and Hatschek's pit. It is suggested that a primitive control system of inhibitory growth hormone secretion in Hatschek's pit could have been developed in amphioxus, as in vertebrates. The present study provides new evidence for the endocrinology and the evolution of Hatschek's pit.     

Distribution of ghrelin-like immunoreactive cells in amphioxus, Branchiostoma belcheri – A study of immunohistochemistry

The distribution of ghrelin-like immunoreactive cells in amphioxus （Branchiostoma belcheri） was investigated by using immunohistochemical staining with rabbit antiserum against synthetical mammalian ghrelin. The results showed that ghrelin-like immunoreactive cells were distributed widely in the nervous system, Hatschek＇s pit, wheel organ, digestive tract and gonads （ovary and testis）. In nervous system, ghrelin-like immunoreactive neurons and their protrusions were distributed specifically on the dorsal side, ventral side and funnel part of brain vesicle, with a few dispersive immunoreactive nerve cells and their fibers in nerve tube. Ghrelin-like immunoreactivities were also detected in Hatschek＇s pit epithelial cells and wheel organ cells, with positive substance located along cell membrane. In digestive tract, ghrelin-like immunoreactive cells existed in hepatic diverticulum, anterior and posterior region of midgut, and could be classified into two types, closed- and opened-type endocrine cells. The number of positive cells was most in hepatic diverticulum, secondary in posterior region of midgut and least in anterior region of midgut. In gonads, ghrelin-like immunoreactive substance was detected in oogonia, oocytes and follicle cells in ovary at the small and large growth stages and in early spermatogenic cells and Sertoli cells in testis. The extensive distribution of ghrelin-like cells in amphioxus suggested that these kinds of cells are conservative in evolution and diversified in function. At the same time, we found for the first time that ghrelin-like immunoreactive cells existed in brain vesicle and Hatschek＇s pit, which provided new morphological evidence for the existence of an activation pathway between brain vesicle and Hatschek＇s pit for the regulation of growth hormone excretion.     

Distribution of CRH-and ACTH-like immunoreactive cells in amphioxus,Branchiostoma belcheri

Immunocytochemical studies on the nervous system,Hatschek's pit,digestive tract and gonads tissues of an amphioxus(Branchiostoma belcheri)were performed using polyclonal antibodies against human corticotrophin-releasing hormone(CRH)and human adrenocorticotropin(ACTH).The results showed that many CRH-like immunoreactive neurons were distributed specifically on the dorsal side and ventral side of brain vesicle,while a few CRH-like neurons and their fibers in spinal cord.At the same time,the epithelial cells in the basic region of Hatschek's pit were shown immunopositive to CRH antibody.In gonads(ovary and testis),CRH-immunopositive substance was localized in the cytoplasm near oocyte nucleus and in early spermatogenic cells.ACTH-like immunoreactivities were observed specially in the neurons and their protrusions localized on the ventral side of the brain vesicle and in spinal cord,and also in epithelial cells of Hatschek's pit,enteric neurons of digestive tract,oocytes in ovary and in early spermatogenic cells as well.It was found for the first time that CRH-like neurons existed in the middle region of brain vesicle(corresponding to the hypothalamus of vertebrates)and ACTH-like immunopositive cells existed in Hatschek's pit,implying that a control mechanism between brain vesicle and Hatschek's pit maybe had been already built in amphioxus as that in vertebrates.The present study will provide new morphological evidence for the origin and evolution of ACTH.In addition,the immunoreactivities of CRH and ACTH in the digestive tract and gonads suggested other physiological function of CRH and ACTH in amphioxus.     

ULTRASTRUCTURAL STUDY OF THE HATSCHEK''''S PIT EPITHELIAL CELLS OF AMPHIOXUS

The authors have observed, using an electron microscope, that the number of thesecretory granules and the development of various organelles in the apical epithelial cellof the Hatschek's pit showed a positive correlation with the gonadal development andmaturity. At the same time, only the epithelial cells respond to GnRH-A(gonadotropin-releasing hormone analogue). Both cytological and endocrinological experiments demon-strate that the cell may be the primitive gonadotropic cell of amphioxus.     

Distribution of growth hormone-like immunoreactive cells and somatostatin receptors in the nervous system and Hatschek''s pit of amphioxus, Branchiostoma belcheri

Using immunohistochemical method and double staining technique, the localization of growth hormone (GH) and somatostatin receptors in the nervous system and Hatschek's pit of amphioxus has been investigated. The results showed that the growth hormone-like nerve cells and endocrine cells as well as three subtypes of somatostatin receptors exist in the nervous system and Hatschek's pit, and GH-like nerve cells and endocrine cells co-exist with three subtypes of somatostatin receptors in the brain vesicle and Hatschek's pit. It is suggested that a primitive control system of inhibitory growth hormone secretion in Hatschek's pit could have been developed in amphioxus, as in vertebrates. The present study provides new evidence for the endocrinology and the evolution of Hatschek's pit.     

Distribution of growth hormone-like immunoreactive cells and somatostatin receptors in the nervous system and Hatschek''''s pit of amphioxus, Branchiostoma belcheri

Using immunohistochemical method and double staining technique, the localization of growth hormone (GH) and somatostatin receptors in the nervous system and Hatschek's pit of amphioxus has been investigated. The results showed that the growth hormone-like nerve cells and endocrine cells as well as three subtypes of somatostatin receptors exist in the nervous system and Hatschek s pit, and GH-like nerve cells and endocrine cells co-exist with three subtypes of somatostatin receptors in the brain vesicle and Hatschek s pit. It is suggested that a primitive control system of inhibitory growth hormone secretion in Hatschek s pit could have been developed in amphioxus, as in vertebrates. The present study provides new evidence for the endocrinology and the evolution of Hatschek's pit.     

Localization and expression of LHβmRNA in the brain and Hatschek's pit of amphioxus, Branchiostoma belcheri

To determine whether gonadotropin-like substance in the brain and Hatschek' s pit of amphioxus issynthesized by the tissue in situ or transported from other tissue, a histochemical study was carried out by in situ hy- bridization using digoxigenin (DIG)-labelled LHβRNAprobes. The results showed that LHβmRNA expressed in the nerve cells of brain and the epithelial cells of Hatschek' s pit, thus providing new evidence for the homology of pituitary of ver- tebrates with Hatschek' s pit and the functional evolution of gonadotropin.     

Distribution of α-MSH-like immunoreactive cells in the nervous system, Hatschek''''s pit and other tissues of amphioxus, Branchiostoma belcheri

Immunohistochemical localization of a-melanocyte-stimulating hormone (α-MSH) in the nervous system, Hatschek's pit and other tissues of amphioxus (Branchiostoma belcheri) was performed using the antibody against synthetic α-MSH. The results revealed that α-MSH-like immunoreactive cells were distributed at the dorsal side and ventral side of brain vesicle, the dorsal side and the surrounding of nerve tube, and in the epithelial cells of Hatschek's pit, the zone 1, 3, and 6 of endostyle and gut. The immunoreactive substance was also found in the primary oocytes of the small and large growth stage of ovary and early stage spermatogenic cells in testis. These findings indicate that α-MSH is an ancient and highly conserved hormone and it is extensively distributed in amphioxus. Although Hatschek's pit in amphioxus does not have a structure of the intermediate lobe of vertebrate adenohypophysis, it has already hosted α-MSH-like endocrine cells, implying that the functional differentiation of α-MSH-like cells occurred earlier than the differentiation of the tissue structure. The results of the present study provided a new evidence for the endocrinology of Hatschek's pit and for the origin and evolution of vertebrate adenohypophysis.     

mmunohistochemical localization of gonadotropin-releasing hormone receptors (GnRHR) in the nervous system, Hatschek's pit and gonads of amphioxus, Branchiostoma belcheri

Using gonadotropin-releasing hormone (GnRH) anti-idiotypic antibodies and APA im-munohistochemical method, the immunoreactivity of GnRHR in the nervous system, Hatschek's pit and gonads of amphioxus has been located. It is found for the first time that the immunoreactivity of GnRHR exists in the nerve cells and fibers in the amphioxus's brain and nerve tube and the epithelial cells of Hatschek's pit at the different stages of gonadal development. At the same time, it is also found that GnRHR also exists in the ovary and testis of different developed stages. These findings provide morphological new proof for the informative transfer and regulation between brain and Hatschek's pit mediation by GnRHR, and for the understanding of the mechanism of action on the reproductive endocrine control axis among brain-Hatschek's pit-go-nads.