Immunohistochemical localization of gonad stimulating substance in the seastar Pycnopodia helianthoides
Date
1984
Authors
Caine, Gary David
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Abstract
Spawning in seastars is initiated by gonad stimulating substance (GSS), a peptide found primarily 1n the radial nerve cords. GSS is thought to stimulate the ovarian follicle cells to produce 1-methyl adenine, which initiates oocyte maturation and spawning. Although much is known about the biochemical and physiological properties of GSS, little experimentation has been done to localize the site of production of GSS or the secretory pathway it takes to the ovary. It has been suggested that GSS is produced in the supporting cells of the radial nerve and moves to the ovary by diffusion through the coelomic fluid; however, most of the evidence for this has come from biochemical inference and histological tests, which may not be specific for GSS alone. The objective of this study was to develop a histochemical test specific for GSS so that it could be localized in the tissues of the seastar Pycnopodia helianthoides.
Polyclonal antibodies were developed that were specific for GSS purified by gel filtration and ion-exchange chromatography. The antibodies were used on formalin fixed sections of radial nerve cords, axial organ, tube feet, ovarian wall and pyloric caeca. Binding sites of GSS antibodies were visualized using the indirect immunoperoxidase technique. Immunoreactivity, indicating the presence of GSS, was observed as discrete, brown granules that averaged 2 um in diameter.
In the radial nerve cords, GSS was found in the lower portion of the perihemal epithelium, which encloses the radial hemal sinus. GSS was not found in either the ectoneural or hyponeural nerves. Ultrastructural examination of the radial perihemal epithelium revealed an axonal plexus in the region where immunoreactivity was observed. The axons varied 1n size, but in general there were large (2 um diameter) and small (less than 1 um diameter) axons. The axons contained clusters of 100 nm diameter, electron-dense vesicles. The location of these axons, their size and distribution appeared to correspond to the size and distribution of the immunoreactive granules observed by light microscopy. It is possible that GSS is localized to the axon plexus of the radial perihemal epithelium.
The axial organ, which is an integral part of the hemal system, also showed immunoreactivity to GSS antibodies. Immunoreactive granules similar to those observed in the radial perihemal epithelium were found in the axial perihemal epithelium. Ultrastructural examination of the axial organ revealed cells and axons that contained 100 nm diameter, electron-dense vesicles in the axial perihemal epithelium. The abundance and size of the vesicular clusters appeared similar to that of the immunoreactive granules observed by light microscopy; therefore, GSS in the axial organ may be localized to the vesicles in the axial perihemal epithelium.
Immunoreactive granules were found in the coelomic epithelium of the tube feet, mostly in the region that apposes the podial connective tissue. Ultrastructural correlates of the granules were not found in this study; however, previous works report 100 to 200 um diameter, electron-dense vesicles found in granulocytes. GSS may be localized to these cells in the coelomic epithelium of the tube feet.
Sections of ovarian wall and pyloric caecum were used as immunohistochemical controls because they do not contain GSS. As expected, no immunoreactivity was observed in either ovarian wall or pyloric caeca. The ultrastructure of the ovarian wall was examined to determine if there were differences between the ovarian perihemal epithelium, and the perihemal epithelia of the radial nerve cord and axial organ. The ovarian perihemal epithelium was much reduced and did not contain the vesiculated axons or cells that may contain to GSS in the other tissues. Lack of vesicles in this region of the ovary may explain why no GSS is found in the ovarian wall.
Since GSS is not found in the ovary, except during spawning, it must be transported from its site of synthesis to the ovary. The proximity of GSS to the radial hemal sinus, the structural link between the radial and ovarian hemal sinus, and recent evidence for the tranlocative ability of the hemal system implicates the hemal system as being involved in GSS transport. I propose that GSS is secreted into the radial hemal sinus and axial hemal sinus, and is tranported to the ovary through the hemal sinus system.
That GSS is found in other tissues not directly connected with the ovaries (ie. the tube feet) suggests that GSS may have other physiological roles besides initiating spawning. One role could be the initiation of the characteristic spawning posture that seastars assume prior to spawning. Further study of GSS is needed to better define the physiological role of this hormone in asteroid neuroendocrine regulation.