Levels of the native forms of GnRH in the pituitary of the gilthead seabream, Sparus aurata, at several characteristic stages of the gonadal cycle

Brains of the gilthead seabream, Sparus aurata, contain three different forms of gonadotropin-releasing hormone (GnRH): seabream (sb) GnRH, chicken (c) GnRH-II, and salmon (s) GnRH. In the present study, we developed three specific enzyme-linked-immunosorbent assays (ELISA) for sbGnRH, cGnRH-II, and sGnRH and used them to measure the levels of each GnRH form in the pituitary of male and female seabream at different stages of gametogenesis. The sensitivity was 6 pg/well for the sbGnRH assay, 7 pg/well for the cGnRH-II assay, and 2 pg/well for the sGnRH assay. Levels of each of the three GnRH forms were measured in pituitaries from fish sampled at the beginning of gonadal recrudescence and during the spawning season. Of the three forms, only sbGnRH and cGnRH-II were detected in the pituitary, irrespective of reproductive state or sex. Recrudescent fish had similar levels of sbGnRH and cGnRH-II in the pituitary. In sexually mature fish, the levels of sbGnRH were higher than those in recrudescent fish while pituitary cGnRH-II content remained unchanged. Consequently, sbGnRH levels were 3- to 17-fold higher than cGnRH-II levels in mature fish. Positive correlations also existed between pituitary sbGnRH content and pituitary and plasma gonadotropin (GtH) II levels. Surprisingly, mature 1-year-old males had significantly higher levels of sbGnRH in the pituitary than mature 3-year-old males, while pituitary and plasma GtH II levels were similar between these two groups. Although the reason for this difference in sbGnRH levels is unclear, a possible role of sbGnRH in the processes of puberty or sex-inversion is implied. Based on the present results, it can be suggested that in the gilthead seabream, sbGnRH is the most relevant form of GnRH in the control of reproduction.     

The value of a fabric-coated self-expanding stent in iliac arterial occlusions or aneurysms--the primary and long-term results

The present study focused on the role of catecholaminergic neurons and estrogens on the release of gonadotropins I and II in immature and early vitellogenic female rainbow trout. The ovariectomy-induced increase of GtH I blood levels (from about 10 to 15 ng/ml) was prevented in vitellogenic fish by E2 supplementation. E2 implantation of immature fish decreased blood GtH I levels (from about 6 to 1 ng/ml). Blood levels of GtH II were low (about 0.5 ng/ml) and not altered by ovariectomy and E2 treatment. These data demonstrate that estrogens exert a negative feedback on the release of GtH I in trout. A treatment with alpha-methyl-p-tyrosine (MPT), an inhibitor of catecholamine synthesis, increased blood GtH II levels of sham-operated vitellogenic fish and ovariectomized fish implanted with E2, but had no effects in ovariectomized fish. MPT did not modify blood GtH I levels in any experimental group. A treatment of E2-implanted immature or vitellogenic fish with the dopamine antagonist pimozide also increased blood GtH II levels, but did not significantly change blood GtH I levels. These data demonstrate that release of GtH II, but not of GtH I, depends on an E2-activated DA inhibitory tone.     

In vitro action of testosterone in potentiating gonadotropin-releasing hormone-stimulated gonadotropin-II secretion in goldfish pituitary cells: involvement of protein kinase C, calcium, and testosterone metabolites

Overnight preincubation of goldfish pituitary cell culture with testosterone (T) enhanced the gonadotropin (GTH)-II responses to GTH-releasing hormone (GnRH). In this study, the involvement of GnRH signal transduction components and the requirement for T metabolism in mediating this direct, pituitary cell action of T were examined using cultured pituitary cells from both male and female goldfish. Each sets of related experiments were done in at least two different stages of the gonadal reproductive cycle and similar effects were observed. Overnight treatment with 10 nM T increased GTH-II responses to maximal stimulatory doses (100 nM) of either salmon (s)GnRH or chicken (c)GnRH-II, but not the total cellular GTH-II contents measured prior to and after a 2-h GnRH challenge. T increased the efficacy and sensitivity of the GTH-II response to stimulation by a protein kinase C (PKC) activator, tetradecanoyl phorbol acetate (TPA) without altering the ED50 of the dose-response curve. In T-treated cells, addition of a PKC inhibitor attenuated GTH-II responses to 100 nM doses of sGnRH, cGnRH-II, or TPA. T did not affect the GTH-II release stimulated by high concentrations of the Ca2+ ionophore ionomycin (100 microM) and the voltage-sensitive Ca2+ channel (VSCC) agonist Bay K 8644 (10 microM); similarly, the sensitivity of the GTH-II response to ionomycin and Bay K 8644 was also unaltered. Taken together, these data suggest that T potentiates GnRH-stimulated GTH-II release by enhancing the effectiveness of PKC-dependent pathways, but not by increasing the total Ca2+-sensitive GTH-II pool, the sensitivity of the release response to increases in intracellular Ca2+, or the amount of available GTH-II. However, the VSCC agonist nifedipine reduced sGnRH- and cGnRH-II-elicited GTH-II release in T-treated as well as in non-T-treated cells, suggesting that VSCC dependence is still present in the GnRH-induced response following exposure to T. Since total cGnRH-II binding to pituitary cells was not increased by T, increases in GnRH receptor capacity are unlikely following T treatment. The ability of T to increase GnRH-stimulated GTH-II secretion was not mimicked by 11-ketotestosterone or dihydrotestosterone, but was abolished by coincubation with an aromatase inhibitor. When viewed together, these observations suggest that aromatization of T may be required for the pituitary action of T on GnRH-induced GTH-II release.     

In vitro reconstitution of an intermediate assembly stage of vaccinia virus

The effects of chronic exogenous testosterone treatment on the synthesis and/or secretion of two sturgeon gonadotropins (stGTH I and stGTH II) were assessed in 2-year-old juvenile white sturgeon (Acipenser transmontanus) surgically implanted with silastic capsules filled with 75 mg of testosterone and in previtellogenic female white sturgeon females implanted with 150 mg of testosterone. In groups of juvenile white sturgeon sacrificed 30, 60, 90, or 442 days postimplantation, pituitary concentrations of stGTH I were significantly greater in testosterone-treated fish (P < 0.01) when compared to those of controls. Pituitary concentrations of stGTH II were significantly higher (P < 0.01) in juvenile fish treated 60, 90, or 442 days with testosterone when compared to those of controls. Exogenous testosterone had no effect on plasma concentrations of either stGTH. Additional testosterone-treated juvenile sturgeon which were injected intraperitoneally 90 or 442 days postimplantation with 10 microg/kg of the gonadotropin releasing hormone analog d-Ala6-des-Gly10-GnRH ethylamide (GnRHa) also showed no change in plasma concentrations of stGTHs. Similar results were obtained for previtellogenic white sturgeon, as pituitary concentrations of stGTH I and stGTH II were significantly greater (P < 0.01) after 60 days of testosterone treatment compared to those of controls. A second group of 60-day testosterone-treated previtellogenic females also failed to exhibit increases of plasma stGTHs when administered 10 microg/kg of GnRHa. These results indicate that long-term testosterone treatment stimulates the accumulation of pituitary stGTHs in both juvenile and previtellogenic white sturgeon but does not affect basal or GnRHa-induced stGTH secretion.     

Erythromycin modulates IL-8 expression in normal and inflamed human bronchial epithelial cells

Previous studies have shown that blood plasma levels of 17alpha, 20beta-dihydroxy-4-pregnen-3-one (DHP) and 17alpha, 20beta, 21-trihydroxy-4-pregnen-3-one (20beta-S) increase in striped bass (Morone saxatilis) undergoing final oocyte maturation (FOM). Both hormones are produced by ovarian fragments undergoing hCG-induced germinal vesicle breakdown (GVBD) in vitro. In the present study, we investigated binding of DHP and 20beta-S to ovarian membranes from striped bass undergoing FOM. Saturable binding sites for DHP were not detected. Saturation of 20beta-S binding sites with 5 nM [3H]20beta-S occurred within 40 min at 0 degrees C (at 3 min, half of the maximum specific binding of steroid was calculated to have occurred), and the binding was pH-dependent. Scatchard analyses revealed the presence of a single class of high-affinity (dissociation constant [Kd] = 1.4 +/- 0.2 nM), limited-capacity (estimated concentration [Bmax] = 2.7 +/- 0.3 pmol/g ovary) 20beta-S binding sites on membranes from striped bass ovaries undergoing FOM. In contrast, only low levels of specific binding (Bmax < 0.04 pmol/g tissue) were detected on membranes from testes, liver, brain, and muscle. Ovarian membranes prepared from vitellogenic females also had low levels (Bmax < 0.1 pmol/g ovary) of specific 20beta-S binding, less than 5% of that found during FOM. Results of competition assays showed that DHP was approximately 250 times less effective than 20beta-S for displacing 20beta-S from ovarian membranes. In contrast, 20beta, 21-dihydroxy-4-pregnen-3-one was a very effective competitor, although it is only a weak inducer of oocyte GVBD in vitro. Of several other steroids tested, only progesterone showed affinity for the 20beta-S binding site within a physiological range of concentrations. Taken together with previous studies of striped bass FOM, these findings indicate that 20beta-S is the oocyte maturation-inducing steroid hormone in striped bass.     

Mutational analysis of two unstructured domains of the 5'' untranslated region of HCV RNA

Substance P (SP) has been suggested to regulate gonadotroph function both directly and indirectly in different species. In pigs, the possible role of hypothalamic and pituitary SP in the control of LH release has not been examined. Here, we investigated SP effects on basal and GnRH-stimulated LH secretion by porcine gonadotrophs in vitro, in both static and dynamic culture systems. SP concentrations of 100 nM or above stimulated LH release from monolayer cultures without affecting intracellular LH content. In the same cultures, SP potentiated GnRH (10 nM)-stimulated LH release and reversed the GnRH-induced decrease of gonadotroph LH stores. The GnRH, but not the SP, effect was completely blocked by the potent GnRH-receptor antagonist antide. In superfused pituitary fragments, three successive pulses of SP or GnRH also stimulated LH release, yet the combined administration of both factors did not result in a synergistic stimulation. These results demonstrate that SP acts directly on porcine gonadotrophs to stimulate LH release, and to maintain the levels of hormonal stores, through a GnRH receptor-independent mechanism. Furthermore, our findings suggest that continuous exposure of gonadotrophs to SP would potentiate GnRH-stimulated LH secretion, thus supporting a possible role of SP as modulator of porcine gonadotroph function.     

The effects of GnRH and adrenergic agents on PRL and beta-endorphin secretion by porcine pituitary cells in vitro

The direct effects of alpha- and beta-adrenergic agents on PRL and beta-endorphin (beta-END) secretion in vitro by porcine pituitary cells have been investigated. Pituitary glands were obtained from mature gilts, which were ovariectomised (OVX) one month before slaughter. Ovariectomised gilts, assigned to four groups, were primed with: (1) vehicle (OVX); (2) and (3) oestradiol benzoate (EB; 2.5 mg/100 kg b.w.) at 30-36 h (OVX+EB I) and 60-66 h (OVX+EB II) before slaughter, respectively; and (4) progesterone (P4; 120 mg/100 kg b.w.) for 5 consecutive days before slaughter (OVX+P4). Isolated anterior pituitary cells were submitted to 3.5 h incubation in the presence of GnRH, alpha- and beta-adrenergic agonists [phenylephrine (PHEN) and isoproterenol (ISOP), respectively], or alpha- and beta-adrenergic blockers [phentolamine (PHENT) and propranolol (PROP), respectively]. The culture media were assayed for PRL (exp. I) and beta-endorphin-like immunoreactivity (beta-END-LI) (experiment II). In experiment I, GnRH did not influence PRL release by pituitary cells in all experimental groups. Some of tested doses of adrenergic agonists, PHEN and ISOP, increased PRL release from pituitary cells of OVX gilts, but not from those of OVX+EB I animals. In the OVX+EB II group, PHEN alone, but ISOP with PROP, potentiated PRL secretion by the cells. In OVX+P4 animals, PHEN alone or in combination with PHENT and also ISOP alone or with PROP enhanced PRL output from the cells. In experiment II, addition of GnRH increased beta-END-LI release from pituitary cells only in the OVX+EB II group. PHEN and PHENT potentiated beta-END-LI secretion by pituitary cells in OVX+EB II and OVX+P4 groups, while ISOP and PROP increased beta-END-LI secretion by the cells of OVX and OVX+EB II animals. In turn, in the OVX+EB I group, effect of PHENT and PROP on PRL secretion by pituitary cells was inhibitory. In conclusion, our results suggest that adrenergic agents can modulate PRL and beta-END secretion by porcine pituitary cells in a manner dependent on the hormonal status of gilts.     

Endocrine therapy of transsexualism and potential complications of long-term treatment

Physiological principles of the interrelationship of sex hormones and their regulation are the foundation of understanding appropriate treatment of the transsexual patient. While both genetic males and females have estrogens and androgens, the quantitative sex hormone production is genetically predetermined by sex hormone production both in the gonads and via peripheral conversion of hormone precursors to sex steroids. Sex hormones exert a negative feedback on the hypothalamus and pituitary gland whereby gonadotropin-releasing hormone (GnRH), pituitary luteinizing hormone (LH), and follicle-stimulating hormone (FSH) are regulated or suppressed by the endogenous levels of these hormones. Sex hormonal therapy induces attenuated GnRH stimulation of LH and FSH causing a reduction of serum sex hormone levels. It is clear that estrogen as well as androgen therapy have a dual role: (i) induction of feminization or virilization and (ii) suppression of the hypothalamic-pituitary-gonadal axis leading to a reduction of endogenous estradiol or testosterone secretion. Cross-sex hormonal treatment may have substantial medical side effects. The smallest dosage of hormonal therapy compatible with the above clinical aims should be used.     

Short-term GnRH-analogue administration in patients with PCOS and its impact on hemodynamics and hormonal profile

PURPOSE: The ovarian response to 14 days of GnRH-analogue (GnRHa) application in patients with polycystic ovary syndrome (PCOS) was evaluated by measuring hemodynamic parameters and hormonal profile. METHODS: Twenty patients with PCOS represented the study group, and 22 patients with normal menstrual cycle (NMC) the reference group. The resistance index (RI) of uteroovarian arterial blood flow and serum level of LH and androstendione were measured on days 4, 13, and 22 of the cycle and after 14 days of GnRHa administration. RESULTS: On day 22 of the cycle the RI in the uterine artery was 0.86 in the PCOS and 0.82 in the NMC group. Throughout the cycle the ovarian arterial blood flow in both ovaries in the PCOS group was similar to that in inactive ovaries in the NMC group. After the 14-day GnRHa administration the RI of uteroovarian blood flow and the serum level of androstenedione remained unchanged in the PCOS group. CONCLUSIONS: RI in the uteroovarian blood flow and serum androstendione level in the PCOS group are not affected by 14-day GnRHa administration.     

Revisiting gonadotrophin-releasing hormone agonist protocols and management of poor ovarian responses to gonadotrophins

Within the past decade, gonadotrophin-releasing hormone (GnRH) agonists have contributed greatly to the success of cycles programmed for in-vitro fertilization and embryo transfer. However, apart from a preventive effect on the luteinizing hormone (LH) surge, most of the beneficial effects of these molecules are still only partly known. A precise analysis of regimens using GnRH agonists for ovarian stimulation shows that many parameters may interfere with the outcome of long-term and short-term protocols. The great variability between these protocols hampers our comprehension of the mechanisms involved in the overall clinical improvement seen with this therapy. The hypophyseal desensitization induced by GnRH agonists is greatly dependent on the dose and duration of their administration, but the residual gonadotrophin secretion is imperfectly estimated by hormonal measurements using radioimmunometric assays. Moreover, the specific role of GnRH agonist-induced ovarian quiescence on subsequent ovarian responsiveness to gonadotrophins and on endometrial receptivity deserves further investigation. Finally, a direct ovarian action of GnRH agonists on steroidogenesis, folliculogenesis and embryo quality is still controversial in humans. These putative deleterious effects of GnRH agonists have led some authors to recommend a reduction of both dose and duration of GnRH agonist administration for women identified by a poor response to gonadotrophins. Using this approach, a few reports have recently shown some clinical advantages for ovarian responsiveness but no convincing evidence for any improvement in pregnancy rate. It thus appears that the overall impact of GnRH agonists on reproductive function is still partly misunderstood.     

Staging and treatment for patients with pancreatic cancer. How small is an early pancreatic cancer?

We report a case of a 66-yr-old woman with progressive hair balding, hirsutism and virilization. Gonadotropins and estradiol levels were in the postmenopausal range; total testosterone (TT), free testosterone (FT) and 17-hydroxyprogesterone (17-OHP) were elevated with dehydroepiandrosterone sulphate, androstendione and cortisol serum levels in the normal range, as 24-hr free urinary cortisol. TT, FT and 17-OHP were normalized, and FSH and LH fell to premenopausal levels on 18th day after a single i.m. injection of the GnRH analogue (GnRHa), triptorelin. Then, a diagnosis of hyperandrogenism of ovarian origin was made and bilateral ovariectomy was performed. Histological study of gonadal tissue revealed diffuse stromal hyperplasia of both ovaries with occasional nests of luteinized cells. With immunoperoxidase techniques these cells stained positively for testosterone and progesterone. One month after surgery, androgen levels were normalized together with regression of most of the clinical signs of virilization. In conclusion, our patient showed a severe virilization developed after menopause; hormonal investigations suggested a gonadotropin dependent ovarian hyperandrogenism, confirmed by histological examination; the presence of luteinized cells in the ovarian stroma was responsible for hyperandrogenism, as confirmed by the immunoperoxidase technique.     

Rhythmicity of luteinizing hormone secretion expressed in vitro

In the present study we explored the possibility that the pituitary functions as an autonomous clock and is capable of generating rhythms of luteinizing hormone (LH) release independently of hypothalamic control. Pituitaries from estrous or diestrous day 1 female mice were perifused separately with Medium-199. Effluent samples were collected at 10-min intervals and assayed for LH levels. Fourier analysis and curve-fit analysis served to elucidate the presence of prominent periods whose significance was then determined by best-fit cosinor. The latter method was used to determine additional parameters for the significant rhythm. All perifused pituitaries exhibited LH release patterns that were composed of significantly long ultradian rhythms (approximately 16 and 8 h, p < 0.001). Continuous stimulation with gonadotropin-releasing hormone (GnRH) or estradiol did not alter the periods of the observed rhythms but affected other rhythm parameters. Gonadotropin-releasing hormone increased the mesor of the rhythm and estradiol increased the amplitude. The results indicate that pituitary gonadotropes are capable of producing rhythms of LH release for a long duration in vitro, in the absence of hypothalamic control. Both GnRH and estradiol affect different rhythm parameters but do not change the periods of these rhythms.     

Ovarian hyperstimulation following the sole administration of agonistic analogues of gonadotrophin releasing hormone

Ovarian hyperstimulation following the sole administration of gonadotrophin-releasing hormone agonists (GnRHa) is exceedingly rare. We hereby report on two infertile patients undergoing in-vitro fertilization-embryo transfer who developed ovarian hyperstimulation under such circumstances. In both patients, GnRHa were administered using the 'long protocol' regimen. The first patient developed ovarian hyperstimulation on two occasions, with mid-luteal depot administration of triptorelin and with early follicular triptorelin, administered as daily subcutaneous injections. In both cycles, within 2 weeks of triptorelin therapy, massive ovarian multifollicular enlargement occurred, concomitant with high serum oestradiol concentrations, which resolved spontaneously following expectant management. The second patient developed ovarian hyperstimulation following daily injections of leuprolide acetate starting at the mid-luteal phase. The final stage of ovulation was triggered by human chorionic gonadotrophin (HCG) and 11 oocytes were retrieved. In-vitro fertilization resulted in embryo formation, but failed to result in pregnancy. The same phenomenon recurred in a subsequent cycle despite preventive pretreatment with an oral contraceptive. A negative GnRH test, performed just before HCG administration, suggested than an ongoing 'flare-up effect' was unlikely to cause ovarian stimulation. Ovarian hyperstimulation can occur following the sole administration of GnRHa irrespective of the preparation used and the administration protocol. Although spontaneous resolution is the rule, once this condition has developed, HCG administration and oocyte retrieval are feasible. This rare entity probably represents an exaggerated form of ovarian cyst formation following GnRHa administration, the underlying pathophysiology of which remains unresolved.     

Characterization of striped bass growth hormone receptors by disulfide-bond reduction and cross-linking studies

Growth hormone (GH) receptors were analyzed in striped bass (Morone saxatilis) by addition of disulfide-bond reducing agents to radioreceptor assays and by cross-linking both striped bass and coho salmon (Oncorhynchus kisutch) crude membrane preparations to radiolabeled hormone. Dithiothreitol (DTT) caused a dose-dependent increase in specific binding of 125I-tilapia (Oreochromis mossambicus) GH to striped bass membrane preparations. Maximal enhancement of 3.4-fold was obtained with 1 mM DTT and 0.03 trypsin inhibitor units/ml of aprotinin. Addition of N-ethylmaleimide (NEM), which binds covalently to free sulfhydryl groups, decreased specific binding. Scatchard analysis of striped bass membrane preparations indicated a single class of GH receptors. Addition of DTT with aprotinin increased GH-binding site concentration from 278 to 507 fmol/mg, while the dissociation constant of 0.56 nM remained unchanged. Cross-linking 125I-tilapia GH to striped bass hepatic membrane preparations and 125I-salmon GH to coho salmon membrane preparations yielded two to three specifically labeled proteins on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Endoglycosidase H treatment was without effect on specifically labeled proteins from either species. Following digestion with N-glycosidase F, relative molecular weights of specifically labeled 125I-GH complexes were reduced, suggesting that hepatic GH-binding proteins in striped bass and salmon are N-linked glycoproteins.     

Immune responses to intestinal parasites: protection, pathology and prophylaxis

It has been suggested that adjunctive growth hormone (GH) therapy improves ovarian response and in vitro fertilization (IVF) outcome in specific groups of patients. The correlation between insulin-like growth factor (IGF) and GH is well established. The aim of this study was to determine whether changes in plasma GH correlate with IGF blood levels in patients during IVF treatment. Thirty-six women undergoing IVF and embryo transfer (ET) were examined. Ovarian stimulation was carried out by gonadotropin-releasing hormone agonists (GnRHa) and gonadotropins. Blood was drawn at the early and late follicular phase, on the day of human chorionic gonadotropin (hCG) injection and at the mid- and the late luteal phases. The samples were assayed for IGF-I, IGF-II, IGF-binding protein-3 (IGF BP-3), GH and estradiol. According to the IGF-I and GH plasma levels, patients were divided into three major groups: Group I consisted of patients in whom peak levels of GH reached more than 4 ng/ml and IGF-I decreased significantly. In this group, estradiol levels were 1863 +/- 149 pg/ml. Group II consisted of patients in whom peak blood GH levels did not exceed 2.5 ng/ml and the IGF-I level remained unchanged. In this group estradiol levels were 630 +/- 57 pg/ml. Group III consisted of patients in whom blood GH levels were low and remained unchanged while estradiol levels were 1600 +/- 420 pg/ml. In this group no significant increase in IGF-levels were observed. There was no significant change in the levels of either IGF-II or IGF BP-3 in any of the groups. We can conclude that (1) there is a negative correlation between GH and IGF-I plasma levels in patients undergoing controlled ovarian hyperstimulation (COH)-IVF, when levels of estradiol and GH are elevated; (2) plasma levels of IGF-I under ovarian hyperstimulation are probably regulated by a multifactorial system; and (3) no correlation was found between the plasma levels of IGF-I and those of IGF-II and IGF BP-3 in all patient groups.     

Gonadotropin-releasing hormone neuronal system of the freshwater snails Helisoma trivolvis and Lymnaea stagnalis: possible involvement in reproduction

Peptides of the gonadotropin-releasing hormone (GnRH) family are present in neural and nonneural tissues throughout the chordate phylum. Although GnRH peptides have been implicated in nonreproductive functions, their primary function is to control reproduction by regulating sexual behaviors and inducing gonadotropin hormone release from the pituitary. Evidence suggesting the presence of a similar peptide in the central nervous system (CNS) of the gastropod mollusc Helisoma trivolvis has recently been provided. In the present study, we examined the tissue distribution of the peptide and found that it is likely restricted to the nervous system. The neuronal system containing the endogenous GnRH-like peptide is described further and is shown, in part, to innervate the male reproductive tract. Immunostaining in the closely related snail, Lymnaea stagnalis, showed a conservation in the locations of some immunoreactive neurons. Notably, staining occurred in and adjacent to the lateral lobes of both snails. Because these lobes contain neurons involved in the stimulation of egg laying and GnRH staining occurred in additional areas in the Helisoma CNS that are involved in reproduction, we suggest that the endogenous GnRH-like peptide plays a role in regulating reproduction in freshwater snails.