5-Hydroxytryptamine and Dopamine in Biomphalaria glabrata

Abstract

5-hydroxytryptamine (5-HT) and dopamine (DA) have been isolated and identified in the tissues of the snail, Biomphalaria glabrata. The brain contained approximately 15 tAg, the cephalopedal region 2 ,ug, and the mantle region 1 jug of 5-HT per gram of fresh weight. The concentrations of DA in the brain and the cephalopedal region were of the same order of magnitude as those of 5-HT, and 5 times lower in the mantle region. Neither 5-HT nor DA were detectable in the hepatopancreas-ovotestis region. The distribution of these 2 biogenic amines was also studied by the histochemical fluorescence technique of Falck and Owman (1965). Only green (catecholamine) fluorescent neurons were observed in the buccal ganglia; they are tentatively considered as DA neurons. Neuropile areas probably contain a mixture of 5-HT and DA. The nerve fibers emanating from the ganglia predominantly contain DA. With the exception of the buccal ganglia, 5-HT neurons were more numerous and comparatively large in the other ganglia. Green catecholamine fluorescence was detected in areas nearest to the epithelium of the foot. The presence of two biogenic amines, 5hydroxytryptamine (5-HT) and dopamine (DA), has been demonstrated in a variety of gastropods (Carpenter et al., 1971; Cottrell and Maser, 1967; Dahl et al., 1966; Gerschenfeld, 1971; Sweeney, 1963; Twarog, 1968; Walker et al., 1968; Kerkut and Cottrell, 1963; Kerkut et al., 1966). These amines are localized predominantly in the nervous system of this class of animals, and evidence for their function as neural transmitters has been reported (Carpenter et al., 1971; Dahl et al., 1966; Gerschenfeld, 1971; Gerschenfeld and Stefani, 1968; Sweeney, 1963; Twarog, 1968). The gastropod Biomphalaria glabrata is an intermediate host of the parasitic trematode, Schistosoma mansoni. 5-HT and norepinephrine have been identified as constituents of the adult form of this parasite (Bennett et al., 1969; Bennett and Bueding, 1971). Furthermore, recent evidence indicates the presence of dopamine receptors in the longitudinal muscle of adult S. mansoni, a function of 5-HT as an excitatory neuromuscular transmitter in the parasite. (T. Tomosky et al., 1974), and a high affinity uptake Received for publication 2 November 1973. * This investigation was supported by NIH training grant AI-149, and research grants AI-03515 and AI-08022. t Present address: National Institute of Child Health and Human Development, Gerontology Research Center, Baltimore City Hospitals, Baltimore, Maryland 21224. mechanism for 5-HT providing a neurotransmitter which the adult worm cannot synthesize de novo. The question arises whether biogenic amines play a role in the development of the larval stages of schistosomes in their intermediate host; this, in turn, requires information about the distribution of these amines in B. glabrata. The present study is concerned with this problem. MATERIALS AND METHODS Biomphalaria glabrata with shell diameters of 9 to 14 mm were used. Determination of biogenic amines The tissues were dissected, quickly frozen in liquid nitrogen, and homogenized in a solution containing perchloric acid (0.4 M), ascorbic acid (10 mM), and EDTA (5.4 mM) (Anden and Magnusson, 1967) in a motor-driven all-glass homogenizer at 4 C. One ml of solution was used per 100 mg tissue. The mixture was transferred into a centrifuge tube and, after washing of the container with a measured volume of the same solution, was centrifuged at 10,000 g for 15 min at 4 C. The pH of the supernatant solution was adjusted to 5 to 6 by addition of 0.5 ml of 2 M potassium phosphate buffer (pH 6.5) and a measured volume of 2 N KOH. After centrifugation (10,000 g, 15 min, 4 C) the supernatant solution was used for the determinations of 5-HT and of DA.