Dorsal Body Hormone Research Articles

An ultrastructural study of the activation of the endocrine dorsal bodies in the snail Helix aspersa by mating

The ultrastructural organization of the dorsal bodies of Helix aspersa was studied using serial sections. The dorsal body cells, which were found in groups of two to six in the connective tissue surrounding the brain, contacted each other and formed a network. They were innervated by two types of axons. Type I axons conformed with those described in earlier studies as inhibitory and originating from the cerebral green cells. Type II axons contained secretory granules that were morphologically distinct from and smaller than type I granules. Between the dorsal body cells and both axon types, localized modifications of the membrane were evident. Virgin snails, which grew faster (wet weight) than mated snails, did not lay viable eggs. A comparative morphometric analysis of the dorsal bodies of virgin and mated snails showed that in the latter there was an increase in the numbers of mitochondria, secretory granules, and Golgi bodies, with a concomitant decrease in the numbers of lipid droplets. However, no difference was found in the numbers of profiles of exocytotic release of secretory granules with the use of tannic acid methods. Mating, obligatory for oviposition of viable eggs, appeared to cause ultrastructural changes consistent with increased synthesis and release of the dorsal body hormone essential for vitellogenesis, ovulation, and, consequently, egg laying.

Trichobilharzia ocellata: influence of infection on the fecundity of its intermediate snail host Lymnaea stagnalis and cercarial induction of the release of schistosomin, a snail neuropeptide antagonizing female gonadotropic hormones

Subadult and adult specimens of the pond snail Lymnaea stagnalis were infected with the schistosome Trichobilharzia ocellata. Egg production and growth of the snails were monitored over an 8-week period post-infection (p.i.). Snail haemolymph was collected and analysed for the presence of schistosomin, a neuropeptide which antagonizes the action of the snails' female gonadotropic hormones. Snails infected as subadults showed an increase in fecundity during the first 4 weeks p.i. compared with non-infected controls. The possibility is discussed that this increase is caused by an accelerated maturation of the female sex organs due to elevated levels of Dorsal Body Hormone, a female gonadotropic hormone. No difference in fecundity was found between snails infected as adults and control snails during the first 4 weeks p.i. Snails infected as subadults and as adults showed a decrease in fecundity from week 5 p.i. and onwards. This decrease coincided with the appearance of schistosomin in the haemolymph of the snails and with that of differentiating cercariae in the daughter sporocysts. Cercariae are probably involved in the induction of schistosomin release from the snails' CNS into the haemolymph. Snails infected as subadults or as adults grew at approximately the same rate as uninfected snails.

Trichobilharzia ocellata: Influence of infection on the interaction between the dorsal body hormone, a female gonadotropic hormone, and the follicle cells in the gonad of the intermediate snail host Lymnaea stagnalis

Activation of adenylate cyclase (AC)-cAMP system in follicle cells of Lymnaea stagnalis by the gonadotropic dorsal body hormone (DBH) is inhibited by schistosomin, an agent present in hemolymph of snails infected with Trichobilharzia ocellata. AC activation was determined enzyme cytochemically. This conclusion is based on the observation that the percentage of oocytes with AC-positive follicle cells in gonads incubated in the presence of schistosomin, i.e., in serum of infected snails (IS) with DBH, is significantly lower than that in gonads incubated in the absence of schistosomin, i.e., in serum of noninfected snails (NS) with DBH. Follicle cells in gonads preincubated in the absence of schistosomin, i.e., in NS, and subsequently incubated with freshly dissolved DBH showed a considerably lower response to DBH than those in not preincubated gonads. This indicates that the number of receptors for DBH on follicle cells had decreased during preincubation. The response to DBH also appeared to decrease when the hormone was preincubated in NS. This indicates that the activity of DBH decreases during preincubation. These data make it impossible to answer the question of whether or not schistosomin acts as an antagonist of DBH at the receptor level.

Schistosomin: a pronase-sensitive agent in the hemolymph of Trichobilharzia ocellata-infected Lymnaea stagnalis inhibits the activity of albumen glands in vitro.

The schistosome parasite, Trichobilharzia ocellata, nearly completely inhibits the reproductive activity of its intermediate host, Lymnaea stagnalis. The synthetic activity of albumen glands of infected snails at day 35 postinfection (p.i.) is only 1% of the control value. The parasite acts by humoral means. We tested the hypothesis that (a) specific humoral agent(s) is (are) involved and refer to this (these) agent(s) as schistosomin. The presence of schistosomin in the hemolymph of infected snails was investigated by using galactogen synthesis in albumen glands as an in vitro bioassay. The synthetic activity of albumen glands of noninfected snails decreased by about 50% during a 1-h incubation in the hemolymph of infected snails. This inhibition is attributed to schistosomin. Based on these results, with the present bioassay schistosomin appears in the hemolymph between days 28-36 p.i. onwards. Schistosomin is heat-stable (100 degrees C) and pronase-sensitive, and therefore it might have a peptide nature. Schistosomin suppresses the stimulating action of the female, gonadotrophic dorsal body hormone at relatively low doses, which suggests that it may compete with this hormone for the same receptors. The development of two other bioassays for schistosomin in our laboratory is discussed.

Polysaccharide synthesis stimulating factors from the dorsal bodies and cerebral ganglia of Helisoma duryi (Mollusca: Pulmonata)

Polysaccharide synthesis by the albumen gland of Helisoma duryi is under the endocrine control of two factors, one from the cerebral ganglia and the other from the dorsal bodies. Both factors stimulate albumen synthesis in vitro in a dose-dependent manner. The cerebral ganglia derived factor is probably a peptide or protein, as it is not extracted by acid or base and is sensitive to trypsin. The nature of the dorsal body derived polysaccharide synthesis stimulating factor is less clear. It is heat labile and not extracted by acid or base, but is insensitive to trypsin and general proteases, and is soluble in methanol. These characteristics lend some support to the hypothesis that dorsal body hormone is a steroid.

Ferritin in mantle pore cells and its role in reproduction of Helisoma duryi (mollusca: Pulmonata)

AbstractThis study was undertaken to investigate the ferritin content of pore cells in the mantle of Helisoma duryi and the possibility of these cells being a source of vitellogenic ferritin. Ultrastructurally pore cells in the mantle resemble those previously studied in the heart/kidney region, except that their plasma membrane is less elaborate. The presence of iron is demonstrated in the cells by the Prussian blue reaction, and in the ferritinlike contents of cytoplasmic vesicles by energy dispersive X‐ray analysis. Egg‐laying activity begins 2 days after virgin snails are allowed to mate. By means of radioimmunoassay it is found that mantle (containing pore cells) ferritin content increases rapidly at the onset of egg‐laying, then declines to reproductive levels. As it declines, the blood ferritin content increases to reproductive levels. Ovotestis and digestive gland ferritin contents drop at the onset of egg‐laying, then rapidly return to reproductive levels. The relationships between the ferritin contents of these four tissues at the onset of and subsequent to egg‐laying are discussed. The ferritin contents of various tissues are determined by radioimmunoassay (RIA) after the removal of endocrine dorsal bodies from reproducing snails. The ferritin levels remain unchanged in the blood and digestive gland, increase in the mantle, and decrease in the ovotestis. A possible influence of dorsal body hormone on ferritin uptake by vitellogenic oocytes is discussed.

A neuropeptide (Calfluxin) is involved in the influx of calcium into mitochondria of the albumen gland of the freshwater snail Lymnaea stagnalis

In the course of a study on the role of Ca 2+ in the regulation of the secretion process in a female accessory sex gland, the albumen gland of the freshwater snail Lymnaea stagnalis, it appeared that the percentage of mitochondria containing Ca 2+ deposits varied significantly in the successive stages of the egg-laying process. Ca 2+ was demonstrated at the ultrastructural level with the pyroantimonate precipitation technique. A significant increase of the percentage can be correlated with the onset of synthetic activity. Two gonadotropic (neuro) hormones are known to stimulate synthetic activity, viz. the cerebral caudo-dorsal cell hormone (CDCH) and the dorsal body hormone (DBH). In vitro experiments involving addition of extracts of various parts of the cerebral ganglion-dorsal body (DB) complex to albumen glands showed that neither CDCH nor DBH causes the influx of Ca 2+ into the mitochondria of the gland cells, but that another factor is responsible. The factor is present in the cerebral commissure. It is pronase sensitive and heat stable. The factor is released from isolated cerebral ganglia-DB complexes upon stimulation with 8-chlorophenylthio-cAMP. Probably the factor is a CDC peptide. It is called Calfluxin.

Enzyme cytochemical evidence for the activation of adenylate cyclase in the follicle cells of vitellogenic oocytes by the dorsal body hormone in the snail Lymnaea stagnalis

It is unknown what cell type(s) in the gonad of the hermaphroditic snail Lymnaea stanalis is the target(s) of the two gonadotropic hormones, dorsal body hormone (DBH) and caudo-dorsal cell neuro-hormone (CDCH). It was considered that the hormones might act by activating the adenylate cyclase (AC)-cAMP system in the targets. AC activation was determined histochemically. The CDCH (ovulation hormone) titer can be manipulated in vivo by inducing egg laying experimentally (“clean water stimulus”). The effect of the oocyte growth-stimulating DBH and also that of CDCH was studied in in vitro experiments. Only the follicle cells appeared to possess and AC-cAMP system (experiments with forskolin). This system can be activated by DBH, but not by CDCH. Activation can only be brought about in certain stages of the egg-laying process. In vivo this activation was apparent from 30–45 min until 2–3 hr after ovulation, i.e., until 0–1 hr after egg mass deposition. In vitro activation was possible 0–30 min after ovulation. Activation was only obvious in certain stages of oogenesis (not in follicle cells of younger oocyte stages). The results indicate that it is the follicle cells that are the targets for DBH. The possible role of the follicle cells is discussed. The question of what are the targets of CDCH remains unsolved. Apparently this hormone does not act via AC-cAMP, but via another second messenger.

Direct effects of the hyperglycemic factor of the freshwater snail Lymnaea stagnalis on isolated glycogen cells

In the freshwater pulmonate snail Lymnaea stagnalis special cells, the glycogen cells (GC) are present for the storage of glycogen reserves. These cells occur in large numbers in the anterior mantle region. In a previous paper in vitro experiments with intact anterior mantle tissue indicated that a hyperglycemic factor is released from the central nervous system (CNS) which stimulates glycogen mobilization in mantle tissue ( M. A. Hemminga, J. J. Maaskant, W. Koomen, and J. Joosse (1985). Neuroendocrine control of glycogen mobilization in the freshwater snail Lymnaea stagnalis. Gen. Comp. Endocrinol. 57, 117–123). In the present study the question of whether this factor affects glycogen metabolism of GC isolated from mantle tissue was investigated. It is reported that in high-K + Ringers the CNS is stimulated to release a factor which, in a dose-dependent way, inhibits glycogen synthesis in isolated GC (measured as a decreased incorporation of [ 14C]glucose into glycogen). Simultaneous with this glycogen synthesis-inhibiting effect, stimulation of glycogen degradation is found (measured as a decreased retention of prelabeled glycogen). It is concluded that (1) the factor released by the CNS having these effects on isolated GC is the same as the hyperglycemic factor which was reported to stimulate glycogen mobilization in intact mantle tissue, and (2) GC after isolation from mantle tissue have retained their ability to respond to this factor. High-K + bathing Ringers of the cerebral/buccal ganglia have a stronger inhibiting effect on glycogen synthesis in isolated GC than bathing Ringers of the remainder part of the CNS, indicating that in this part of the brain the major release sites of the hyperglycemic factor are located. Dorsal body hormone, ovulation hormone, or growth hormone of L. stagnalis probably are not identical with the hyperglycemic factor, as homogenates of parts of the cerebral ganglia which are known to contain these respective hormones do not influence glycogen synthesis in isolated GC.

Effects of elevated haemolymph levels of the female gonadotropic dorsal body hormone on the cercarial production ofTrichobilharzia ocellata inLymnaea stagnalis

Juvenile specimens ofLymnaea stagnalis were exposed to 0 or 4 miracidia ofTrichobilharzia ocellata. The effects of experimentally raised haemolymph levels of the female gonadotropic dorsal body hormone (DBH) on female reproduction of the host were studied, and the effect on the production of cercariae was determined. Operations of snails and implantations of endocrine dorsal bodies (DB) were performed shortly after infection in three successive experiments. DB implants in non-infected snails (+DB snails) advanced the onset of ovipository activity and enhanced oocyte production compared to that in appropriate controls. In this the increase in haemolymph level is proved of DBH produced by the DB grafts. Infected +DB snails did not show any increase in the various aspects of female reproductive activity. DB implants also did not affect the production of cercariae. Thus the blocking action by the parasites on the development of the reproductive system could not be counteracted by elevating the DBH level in the haemolymph. These results are consistent with the hypothesis that the parasites intervene in the reproductive activity of the host at the level of the targets of the gonadotropic hormones.

The effect of infection ofLymnaea stagnalis withTrichobilharzia ocellata on the presence of, and reactivity to the ovulation hormone of the host

Oviposition does not start in the hermaphrodite gastropodLymnaea stagnalis if it is infected withTrichobilharzia ocellata when juvenile. Ovulation and oviposition are induced by the gonadotropic hromone CDCH, produced by paired groups of neuroendocrine caudodorsal cells (CDC) located in the cerebral ganglia of the snail. CDCH can be extracted from the neurohaemal area of the CDC, the cerebral commissure (COM), and assayed by a rapid in vivo procedure. In this bioassay CDCH activity was found after injection of COM extracts prepared from infected donor snails, but these COMs contained a smaller amount of CDCH. Parasitized recipient animals in the assay did not respond to CDCH injections, indicating that the mechanism involved in ovulation was unable to react to CDCH. The absence of reactivity of the targets to CDCH and the smaller amount of CDCH in COMs of infected snails might both be caused by incomplete maturity of the CDC and the targets of CDCH in infected snails. Since the maturity of the female reproductive organs is controlled by the female gonadotropic dorsal body hormone (DBH), it is suggested that the parasites intervene at the level of the reactivity of the targets to DBH.

Effects of two gonadotropic hormones on polysaccharide synthesis in the albumen gland of Lymnaea stagnalis, studied with the organ culture technique

A medium for culturing organs of the basommatophoran freshwater snail Lymnaea stagnalis is described. One of the accessory sex glands, the albumen gland, which produces proteins and galactogen as nutritive substances for the embryos, was cultured along with parts of the central nervous system (CNS), including the female gonadotropic hormone-producing dorsal bodies (DB). Culture time was 4 days. The cerebral ganglia and the DB appeared to have a strong stimulating influence on polysaccharide synthesis in the albumen gland. The remaining part of the CNS has a weak stimulating effect. The use of extracts (culture time 6–26 hr) of parts of the cerebral ganglia and of the DB showed that not only the dorsal body hormone (DBH), but also the ovulation hormone (CDCH)—a neurohormone produced by the caudodorsal cells located in the cerebral ganglia—stimulates the synthetic activity of the albumen gland. The action of these hormones is direct; i.e., it is not exerted via the gonad. Dose- and time-response relations for the DBH and the CDCH are very similar. No additional rise in stimulation occurred when the two hormones were administered together. Experiments with albumen glands of adult snails demonstrated that the presence of large quantities of secretory material in the gland inhibits the response to DBH and CDCH.

The stimulating effect of the Dorsal-Body Hormone on cell differentiation in the female accessory sex organs of the hermaphrodite freshwater snail, Lymnaea stagnalis

An experimental study has been made on the role of the Dorsal Body Hormone (DBH) of the Dorsal Bodies (DB), of the growth hormone of the neurosecretory Light Green Cells (LGC) of the cerebral ganglia, and of the differentiation of the various secretory cell types in the accessory sex organs (ASO) of the hermaphrodite freshwater pulmonate snail L. stagnalis. To study this problem, undifferentiated rudiments of the ASO of juvenile snails were transplanted into the haemocoel of the head sinus of adult acceptors subjected to various types of operations. After 6 weeks the male and female ASO of sham-operated control acceptor snails showed clear signs of differentiation. The female ASO, with the exception of the ciliated groove, of DB or of DB + LGC cauterized acceptors had not differentiated. Reimplantation of DB or of DB + LGC stimulated the normal differentiation process. It is concluded that the cellular differentiation of the female ASO is controlled by the DBH. This hormone has no effect on the differentiation of the male ASO. The growth hormone of the LGC has no effect on the differentiation of either the female or the male ASO. Whether the differentiation of the male ASO is controlled by other endocrine centers remains to be studied.

Control of vitellogenesis and of growth of female accessory sex organs by the dorsal body hormone (DBH) in the hermaphroditic freshwater snail Lymnaea stagnalis

A study has been made of the role of the dorsal bodies (DB) and the adjacent neurosecretory light green cells (LGC) of the cerebral ganglia in reproductive activity of the hermaphroditic pulmonate snail Lymnaea stagnalis. The experiment included one series of juvenile snails and one of adult snails. Each series included the following groups: controls, sham-operated controls, and snails with cauterized DB, LGC, or DB+LGC. Implantations were carried out on some of the operated snails on postoperative Day 60. The final observations on all animals were performed on Day 90. Removal of the DB in juvenile as well as in adult snails resulted in a complete absence of ovipository activity. Removal of the LGC resulted only in a temporary decrease of ovipository activity. Ninety Days after the DB were removed from juvenile snails, the percentages of vitellogenic and degenerating oocytes in the ovotestes had decreased, whereas the percentage of previtellogenic oocytes had increased. This indicates that in the absence of the DB the production of previtellogenic oocytes continues, but that vitellogenesis ceases. Reimplantation of the DB restored the normal proportional frequencies and was followed by the onset of ovipository activity within 30 days. Removal of the DB from adult snails had no effect on the proportional frequencies of the female cell types, and also did not result in degeneration of the developing and ripe oocytes. Subsequent reimplantation of the DB was followed by the restart of oviposition. Cauterization and subsequent implantation of the LGC in juvenile and adult snails had no effect on the proportional frequencies of the various types of female cells. Growth of the female accessory sex organs is entirely dependent on the presence of the DB. Again, this was most clearly demonstrated by extirpation and implantation of DB in juvenile snails. No clear effects of DB or LGC on spermatogenesis and growth of the male accessory sex organs were found. It is concluded that the DB produce an endocrine factor, the dorsal body hormone (DBH), that stimulates vitellogenesis and growth of the female accessory sex organs. The stimulation of oviposition by the DBH is thought to be due to the stimulation of vitellogenesis. It is suggested that the effect of the DBH on the female organs will be mediated by the ovotestis.

Effects of Starvation, Low Temperature and the Dorsal Body Hormone On the in Vitro Synthesis of Galactogen and Glycogen in the Albumen Gland and the Mantle of the Pond Snail Lymnaea Stagnalis

Previous experiments showed that the synthesis of polysaccharides (mainly galactogen) in the albumen glands of starving pond snails is severely reduced. The present experiments deal with the in vitro synthesis of galactogen in albumen glands and of glycogen in mantle tissue, taken from snails kept under various experimental conditions. The syntheses were studied by the addition of 14 C-glucose to the snail-Ringer medium. Within 3 days starvation causes a decrease in the synthetic activity of the albumen gland of about 80%. During long periods of starvation (up to 6 weeks) it remained at this low level. The wet weights of the glands decreased gradually by about 65%. Also the total polysaccharide content of the albumen glands decreased considerably during starvation. At low temperature (6°C) the rate of incorporation of label in the polysaccharides of the albumen glands declined similar as in starving snails, but the wet weights and the polysaccharide contents of the glands did not differ significantly from those of glands of snails kept at 19°C. In snails from which the Dorsal Bodies (DB) had been extirpated-these animals feed normally but produce no egg masses-the synthetic activity of the albumen glands had decreased with about 80% after 18 days, i.e. to the same level as found after 2 weeks of starvation or of low temperature. The weights of the organs decreased considerably, but the polysaccharide content only a little. When immediately after the operation DB are implanted, the glands remained normal in all respects. From these results it is concluded that the Dorsal Body Hormone (DBH) stimulates the synthesis of galactogen in the albumen gland, and that in starved snails the release of the DBH will be inhibited. The inactivation of the DB is most likely caused by the ceased influx of dietary glucose in the haemolymph of starving snails. It is suggested that in the absence of the DBH, the galactogen in the albumen gland is no longer protected against catabolism, which becomes manifest during starvation. A decrease in the galactogen content was not observed in the low temperature experiments. However, DBH is not absent from the haemolymph of these snails. The influence of the DBH on the albumen gland may be either directly, or indirectly via a gonadal hormone. The glycogen synthetizing system of the mantle is not markedly affected by starvation, low temperature or the absence or presence of DBH, when measured with the in vitro technique. In view of the literature it is suggested that the relatively high glucose concentration of the incubation medium had a direct and rapid effect on the glycogen phosphorylase-synthetase system, resulting in an activation of glycogen synthetase in all mantle pieces, taken from the snails of the various experimental groups. This means that the synthesis of glycogen in the mantle is independent of the DBH and is influenced directly by the glucose levels of the haemolymph. A tentative scheme of the factors involved in the control of galactogen and glycogen metabolism is presented.