Biological and immunological characterization of multiple GnRH in an opisthobranch mollusk, Aplysia californica

Gonadotropin-releasing hormone (GnRH) is a neurohormone central to the regu lation of reproductive functions in vertebrates. Recently, several studies have reported the presence of GnRH immunoreactivity (IR) in a number of mol lusks, suggesting that the distribution of GnRH may not be restricted to Ph ylum Chordata. In the present study we extend our investigations to an opis thobranch mollusk, Aplysia californica, to characterize the source, chemica l nature, and biological activity of molluscan GnRH-related molecules. Spec ific radioimmunoassays (RIAs) of various tissue extracts of Aplysia reveale d that only ovotestis, hemocytes, and hemolymph contained significant amoun ts of GnRH that crossreacts with antisera raised against tunicate-I (tI) an d mammalian (m) GnRH. Further RIAs and extractions revealed that the GnRH-I R in the hemolymph is biochemically and immunologically distinct from the G nRH-IR in the hemocytes and ovotestis. Using reverse-phase high-performance liquid chromatography coupled with RIAs, the GnRH-IR in the hemolymph was resolved into two major peaks. The first peak eluted earlier than most know n forms of vertebrate GnRH, and the later peak coeluted with m, lamprey I, chicken II, and tI-GnRH. However, both peaks were broad and may contain a h eterogeneous mixture of GnRH-IR. Immunocytochemical study showed that tI-Gn RH-IR was present in the connective sheath surrounding the central nervous system, with a strong presence in what appeared to be vascular space, again suggesting the close association between Aplysia GnRH-IR and circulation. Finally, treatment of the neuroendocrine bag cells with chicken II GnRH sig nificantly decreased the duration of the afterdischarge (AD, a characterist ic pattern of electrical firing in bag cell neurons) and the number of acti on potentials fired during an AD, indicating the presence of a correspondin g GnRH receptor in the Aplysia central nervous system. Overall, the results demonstrated the presence of multiple forms of GnRH-IR that crossreact wit h tI- and mGnRH antisera in A. californica and the ability of a vertebrate GnRH to alter Aplysia neural activity. Together, these data suggest that Gn RH may be a factor released by the ovotestis and hemocytes into the circula tion to alter neural functions. GnRH-IR produced by the latter may serve as a novel mediator of the neural and immune functions in Aplysia. (C) 2000 A cademic Press.