Plasma Sex Steroid Levels and Steroidogenesis in the Gonad of the Self-fertilizing Fish Rivulus marmoratus

Synopsis The mangrove killifish, Rivulus marmoratus, is the only known self-fertilizing vertebrate. This species is sexually dimorphic; sexually mature individuals are either hermaphrodite or primary and secondary males. Although the mangrove killifish has a unique reproductive strategy, there has been no study on the reproductive endocrinology of this species. Thus we investigated plasma sex steroid hormone levels and steroidogenesis in the gonads of R. marmoratus by enzyme linked immunosorbent assay (ELISA). Plasma 17β-estradiol (E2) and 11-ketotestosterone (11-KT) were detected both in hermaphrodite and in primary male. Ovarian follicles (follicle-enclosed oocytes) from hermaphrodites, which were categorized into early yolk stage and late yolk stage, and testis tissue of primary males were cultured with different concentrations of 17α-hydroxyprogesterone (OHP) or testosterone (T) for 24 h. Production of T, E2, 11-KT and 17α-20 β-dihydroxy-4-pregnen-3-one (17α,20β-P) in the medium from tissue culture were measured by ELISA. Early and late ovarian follicles of hermaphrodites and testis pieces of primary males synchronously secreted E2, 11-KT, and 17α,20β-P following incubation with OHP or T. We conclude that both hermaphrodite and primary male of the mangrove killifish secrete estrogen, androgen, and progestin synchronously.     

Stimulation of testicular function by exogenous testosterone in male protandrous black porgy, Acanthopagrus schlegeli

When 4 mg of testosterone (T) per kg food was given to 1-year-old protandrous male black porgy Acanthopagrus schlegeli for 7 months, gonadosomatic index was significantly higher than when the dose was 0.5 mg kg⁻¹ food. Both doses of T prolonged the spawning season, and increased the number of spermiating fish and milt volume. Sperm concentrations were similar in spermiating black porgy from the treated and control groups. Low levels of oestradiol-17β were observed during the experimental period while elevated levels of plasma T were observed only in March in both control and T-treated groups. Significantly higher levels of plasma 11-ketotestosterone (11-KT) were observed in the 0.5- and 4.0-mg T-treated groups during the spawning season as compared to the control group. The present data suggest that both 0.5- and 4·0-mg T doses stimulate testicular weight, increase numbers of spermiating males and milt volume without affecting the sperm concentrations. Plasma 11-KT concentrations were elevated during T treatment and closely correlated with testicular development and spermiation.     

Isolation and preliminary characteristics of β-N-acetylglucosaminidase in the sperm of Siberian sturgeon (Acipenser baerii) and rainbow trout (Oncorhynchus mykiss)

The aim of this study was to characterize the enzyme β- N -acetyglucosaminidase (β-NAGase) in the milt and spermatozoa extracts from Siberian sturgeon and rainbow trout. After ion exchange chromatography one protein peak showed β-NAGase activity in sturgeon milt plasma and sperm extracts of both species. Surprisingly, two protein peaks showing β-NAGase activity were found in rainbow trout milt plasma. The molecular mass of β-NAGase was estimated by gel filtration as 127 kDa for rainbow trout spermatozoa, 271 kDa for sturgeon spermatozoa, and 74 kDa for milt plasma from both species. The kinetic parameters were determined for milt plasma and sperm extracts. The optimum pH of the β-NAGases was 3.8 for sturgeon milt plasma, 4.4 for sturgeon sperm extract, and 4.4–4.8 for milt plasma and sperm extract from rainbow trout. K _m value of the β-NAGases was 0.212, 0.563, 0.779 m m for sturgeon milt plasma, sturgeon sperm extract or rainbow trout extract, respectively. The β-NAGase from sperm extracts in both species showed 100% activity even after incubation at 56°C by 20 min, whereas its activity was decreased to 23% in sturgeon milt plasma and to 2% in trout milt plasma.     

The shift from C-19 to C-21 steroid synthesis in spawning male common carp, Cyprinus carpio, is regulated by the inhibition of androgen production by progestogens produced by spermatozoa

There is a rapid shift in the steroidogenic pathway from androgen to progestogen production in spawning male common carp, Cyprinus carpio. Experiments were conducted to determine the mechanism regulating this shift using in vitro cultures of testicular fragments and isolated sperm of spermiating male carp. The levels of 11-ketotestosterone (11-KT) continually increased for 48 h with or without gonadotropin (GtH) stimulation, suggesting that 11-KT is the principal androgen produced by carp testes. Ovine prolactin (oPRL) enhanced GtH-stimulated 11-KT production, but by itself had no effect. Gonadotropin, carp pituitary extract, and pregnenolone all enhanced the production of 11-KT, testosterone (T), and 17 alpha-hydroxyprogesterone (17-P) in a dose-dependent manner. No 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17,20 beta-P) was detected in response to any of these agents; 17 alpha,20 alpha-dihydroxy-4-pregnen-3-one (17,20 alpha-P) was not measured. Both 17,20 beta-P and 17,20 alpha-P inhibited 11-KT production in a dose-dependent manner in the presence of either GtH, 17-P, or T. Isolated sperm and testicular fragment preparations both produced 17,20 beta-P and approximately tenfold more 17,20 alpha-P when incubated with 17-P. Only testicular fragments, however, produced 11-KT. We conclude that androgen synthesis occurs only within somatic cells of common carp testes. GtH, and perhaps PRL, stimulates the production of steroid precursors that, under normal physiological conditions, are metabolized to androgens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Annual changes in testicular development and plasma sex steroids in the captive male dojo loach <Emphasis Type="Italic">Misgurnus anguillicaudatus</Emphasis>

Testicular development in the captive male dojo loach Misgurnus anguillicaudatus was examined monthly in relation to the levels of plasma sex steroids [testosterone (T), 11-ketotestostrone (11-KT), and 17,20β-dihydroxy-4-pregnen-3-one (DHP)]. On the basis of testicular histology, the annual gonadal cycle was found to be divisible into 3 periods: the recovery and proliferation period, which mainly consists of early spermatogenic testis from August to November (reproductive phase I); the preparation period for the next spawning period, which mainly consists of late spermatogenic testis from December to April (reproductive phase II); and the mature period, characterized by a high proportion of mature testis from May to July (reproductive phase III). Individual variability in testicular development was high, and continuous spermatogenesis was observed throughout the year. High levels of plasma T, 11-KT, and DHP were observed during reproductive phase III. 11-KT began to increase in February, while T was present at low levels in reproductive phase II. These results suggest that the physiologically active season of testis development for breeding in the dojo loach is from May to July, although spermatogenesis occurs throughout the year.     

Reproductive Structure of the Adult Green Sturgeon, Acipenser medirostris, Population in the Rogue River, Oregon

The primary objective of this study was to determine the reproductive structure of the adult green sturgeon population in the Rogue River. Green sturgeon were captured by gillnet in the lower 11.6–68.4 river kilometers in April to July 2000–2003 and September and October 2002–2003. Gonadal tissue, collected by biopsy, was processed histologically, blood was collected from the caudal vasculature, and fork length (FL) and total length (TL) (±0.5 cm) were measured for each individual. Sex steroids, testosterone (T), 11-ketotestosterone (11-KT), and estradiol-17β (E2), were measured by radioimmunoassay. Biological samples were collected from a total of 88 green sturgeon of which 37 females and 41 males were confirmed by histological analysis. Four gravid females, captured in the spring, were visually identified, and oocyte polarization index and ovarian follicle diameter indicated that these females were in spawning condition. Gonadal samples collected from six individuals did not contain gonial cells, hence the sex and stage of maturity in these individuals remains unknown. Of the 20 females captured in the spring, 1 was vitellogenic, 4 were post-vitellogenic, and 15 were post-ovulatory. Twenty-one females were captured in the fall of which 6 were pre-vitellogenic, 7 vitellogenic, and 8 post-ovulatory. Of the 16 males captured in the spring, 2 were pre-meiotic, 8 were ripe or actively spermiating, and 6 were post-spermiation. Twenty-five males were captured in the fall: 11 pre-meiotic males and 14 post-spermiation. The majority of green sturgeon captured in the Rogue River were reproductively active or had recently spawned indicating the importance of this river for the preservation of green sturgeon.     

Milt characteristics and plasma levels of gonadal steroids in greenback flounder Rhombosolea tapirina following treatment with exogenous hormones

Fish were treated with exogenous hormones, and milt and blood samples were collected for up to 96 h post‐treatment. Blood plasma samples were assayed for the gonadal steroids testosterone (T), 11‐ketotestosterone (11KT) and 17,20ß‐dihydroxy‐4‐pregnen‐3‐one (17.20ßP). Milt volume, spermatocrit and sperm motility were measured from milt samples. Non‐spermiated fish showed increased plasma T and 11KT in response to human chorionic gonadotropin (hCG) but not luteinising hormone releasing hormone analogue (LHRHa). Fish did not become spermiated in response to treatment with hCG, LHRHa, 11KT, 17‐hydroxyprogesterone (17P) or 17,20ßP. Spermiated fish showed an increase in milt volume in response to hCG and LHRHa but not exogenous steroids. Sperm motility declined to zero over 120 s and was not affected by hormone treatment or sampling time. Increased milt volume was accompanied by increased plasma T and 11KT, but not 17.20ßP levels. In a separate experiment, LHRHa delivered by injection or pellet was equally effective at increasing milt volume but had no effect on plasma steroid levels. Spermatocrit declined with stripping but was not affected by hormone treatment, nor was sperm motility. Co‐treatment of fish with 17P plus LHRHa had no additive effect on plasma steroid concentrations or milt volume. The results suggest that as in other teleosts, gonadotropin mediates milt production in greenback flounder.     

Ovarian steroid sulphate functions as priming pheromone in maleBarilius bendelisis (Ham.)

The study reveals that pre-ovulatory females of the fishBarilius bendelisis (Ham.) release sex steroids and their conjugates into the water and that a steroid sulphate of these compounds functions as a potent sex pheromone which stimulates milt production in conspecific males prior to spawning. Since males exposed to the purified sub-fraction III of the steroid sulphate fraction have increased milt volume and more spermatozoa with greater motility, the function of this priming pheromone appears to be to enhance male spawning success. High turbulence and faster water currents render the hillstream ecosystem extremely challenging for chemical communication. Therefore, ovulatory female fish secrete highly water soluble steroid sulphates for rapid pheromonal action in males. Inhibited milt volume in olfactory tract lesioned (OTL) males exposed to the steroid sulphate fraction and 17α,20β-dihydroxy-4-pregnen-3-one supports the concept that the pheromonally induced priming effect in male fish is mediated through olfactory pathways.     

Lack of species-specific primer effects of odours from female Atlantic salmon, Salmo salar, and brown trout, Salmo trutta

We exposed, in two successive spawning seasons, individually placed precocious male Atlantic salmon ( Salmo salar ) and brown trout ( Salmo trutta ) parr to odour stimuli (ovarian fluid and urine mix) from ovulated conspecific or heterospecific anadromous females. Atlantic salmon parr had significantly higher plasma concentrations of the hormones 17α,20β-dihydroxy-4-pregnen-3-one (17,20β-P), 11-ketotestosterone (11-KT) and testosterone (T) after exposure to odours from conspecific females or from brown trout females compared to parr exposed to a control solution (0.9% NaCl). We did not observe any significant differences between the hormone levels in salmon parr exposed to the two female odours. The salmon parr exposed to conspecific odours had significantly higher volumes of strippable milt compared to the controls, but we did not find any significant differences when comparing the effect of the two female odours. Brown trout parr had significantly higher plasma 17,20β-P levels following exposure to heterospecific female odours compared to control males, but there was no significant difference between males exposed to the different female odours. We did not observe any significant differences in plasma levels of T and 11-KT and in milt volumes between exposed and control trout. Taken together, the results from both tested species indicate that the potency of heterospecific stimuli in stimulating increased plasma sex steroid hormone levels in male parr was as strong as stimuli from conspecific females. The results are discussed in connection to observed hybridisation between the two sympatric species.     

Luteinizing hormone and sexual steroid plasma levels after treatment of European sea bass with sustained-release delivery systems for gonadotropin-releasing hormone analogue

Spermiating male European sea bass Dicentrarchus labrax were treated with gonadotropin-releasing hormone agonists (GnRHa), either a GnRHa injection (IN; 25 μg kg⁻¹ body mass) or one of three types of controlled-release GnRHa-delivery systems: fast release implants (EVAc; 1OO μg kg⁻¹), slow release implants (EVSL; lOO μg kg⁻¹) and slow release microspheres (MC; 50 μg kg⁻¹). Luteinizing hormone (LH) release was highly stimulated by all GnRHa treatments, with elevated plasma levels lasting for 2 days in injected fish (IN) and 2, 4 and 6 weeks in controlled-release-treated fish (EVAc, MC and EVSL, respectively), correlating with a 1, 3, 5 and 5 week period of stimulation of milt production, respectively. Plasma levels of the androgens testosterone (T) and 11-ketotestosterone (11-KT), were not significantly affected by the GnRHa treatments. Plasma T was high at early spermiation and declined sharply near the end of this period. Plasma 11-KT levels declined continuously throughout the experiment. Levels of 17,20 β -dihydroxy-4-pregnen-3-one (17,20 β -P), a proposed maturation-inducing steroid (MIS) in European sea bass, fluctuated around 0.2–1 ng ml⁻¹ and were not greatly affected by the treatments. These results indicated a close correlation between sustained stimulation of LH release, achieved by GnRHa-delivery systems, and long-term enhancement of milt production. They also show an absence of changes in the common sex steroids, associated with elevated LH and enhanced spermiation.     

Physiological effects of the lunar cycle on the spawning of a coral reef fish,Abudefduf Vaigiensis: in vivo and in vitro trait

The aim of the present study was to investigate the lunar cycle effects of the spawning of Audefduf vaigiensis through in vivo and in vitro analysis. For this purpose, the indices of GSI, serum levels of sex steroids, including testosterone (T), 17α-hydroxyprogesterone (OHP), 17α, 20β-dihydroxyprogesterone (DHP), and 11-keto-testosterone (11-KT) as well as the germinal vesicle breakdown (GVBD) were measured. The sampling pattern was weekly, based on the moon cycles as the new moon (NM), the first quarter (FQ), the full moon (FM), and the last quarter (LQ). In females, the highest in vivo values of the GSI index were obtained in FQ and LQ, and in males, this value was significantly higher in LQ than NM. The highest in vivo level of OHP in females was observed in FQ, whereas in males was obtained in FM. In both sexes, the in vivo serum levels of DHP were obtained in LQ. In males, the level of 11-KT were at the peak in NM. In vitro analysis showed the highest rate of GVBD in LQ. Moreover, the in vitro levels of T, OHP, and DHP were significantly higher in LQ compare to NM in both sexes. However, in males, the in vitro levels of 11-KT was significantly higher in NM than LQ. These cyclical changes obtained from in vivo plasma steroid hormones and in vitro data on GVBD suggested that lunar periodicity is a major external regulator that synchronized ovarian and testicular activity of A. vaigiensis with emphasis on spawning phenomenon.

     

Testicular development and serum levels of gonadal steroids during the annual reproductive cycle of captive Japanese sardine

Gonad and blood samples were taken throughout the year from captive males of the Japanese sardine,Sardinops melanostictus, and changes in serum levels of gonadal steroids were examined in relation to the annual gonadal cycle. On the basis of testicular histology, the annual gonadal cycle was divisible into four periods: immature (July–September), spermatogenesis (October–December), spermiation (January–April), and post-spawning (May–June). The pattern of seasonal changes in the gonadosomatic index (GSI) was inversely correlated with that of water temperature, and reflected the degree of testicular maturity. The serum testosterone level was relatively low during spermatogenesis (2.2–2.5 ng/ml), rose markedly around the time of spermiation (7.7–24.6 ng/ml), and became low after spawning and during immature periods (0.6–0.7 ng/ml). The serum 17α,20β-dihydroxy-4-pregnen-3-one level was high in males with spermatogenic or spermiating testes (0.6–1.0 ng/ml), but became low (0.2 ng/ ml during the post-spawning period and was undetectable in immature fish. Although 11-ketotestosterone was detectable in some fish, the values obtained were thought to reflect cross-reactivity of the antiserum employed with testosterone. These findings are discussed in relation to male reproduction of the Japanese sardine and steroidal regulation of spermatogenesis and spermiation in other teleosts.     

Effects of constant short and long photoperiod regimes on the spawning performance and sex steroid levels of female and male sea bass

Under constant short photoperiod, the spawning time of 2-year-old sea bass Dicentrarchus labrax was advanced as compared to controls, whereas spawnings were delayed under constant long photoperiod. High plasma levels of 17β-oestradiol (E_2/) and testosterone (T) in females were coincident with the appearance of vitellogenic oocytes in the ovary, while high levels of 11-ketotestosterone (11-KT) and T in males were coincident with the presence of spermiating males. Although plasma levels of E₂ in females and 11-KT in males were low during the remainder of the cycle, levels of T were always >1 ng ml⁻¹ in both sexes, suggesting that T could play an important role during the initial stages of gonadal development. The profiles of E₂ and T in females and 11-KT and T in males exposed to constant short days were similar to those in the control group, but fish which were maintained under constant long photoperiods showed a bimodal pattern of these steroids. The results obtained from fish exposed to constant photoperiod regimes provide further evidence that an endogenous process could be operating to control the reproduction of sea bass.     

Involvement of sex steroid hormones in the early stages of spermatogenesis in Japanese huchen (Hucho perryi )

In higher vertebrates, considerable progress has been made in understanding the endocrine regulation of puberty; however, in teleosts, the regulatory mechanisms of spermatogenesis during the first annual cycle remain unclear. The present study was conducted to understand the regulatory mechanisms of spermatogenesis throughout the different stages of the first spermatogenic cycle and to check the ability of various steroids and hormones to induce in vitro spermatogonial proliferation in Japanese huchen (Hucho perryi ). The results indicate that the serum level of 11-ketotestosterone (11-KT) was positively associated with germ cell type; the level first began to rise with the appearance of late-type B spermatogonia and continued to increase gradually throughout the active spermatogenic stages and spermiogenesis, reaching a peak value 2 wk before spawning, and then declined. During the spermatogenic stages, the serum concentration of 17alpha,20beta-dihydroxy-4-pregnen-3-one (17alpha,20beta-DP) was undetectable. Only a small peak was detected with the appearance of spermatocytes and spermatids, and at the time of spawning, the level increased dramatically, reaching its maximum value with the onset of milt production. Despite the high variation in serum levels of 17beta-estradiol (E2) both between months and among the individuals, E2 was found during the whole reproductive cycle. From these results, we concluded that 1) 11-KT is necessary for the initiation of spermatogenesis and sperm production, and it probably plays a role in spermiation, 2) 17alpha,20beta-DP is essential for the final maturation stage, could play a significant role in the mitosis phase and meiosis process, and probably participates in the regulation of spawning behavior, and 3) estrogen is an indispensable male hormone that plays a physiological role in some aspects of testicular functions, especially during the mitotic phase. The three steroids were also able to induce DNA synthesis, spermatogonial renewal, and/or spermatogonial proliferation in vitro.     

Effects of gonadotropin-releasing hormone agonist administered in microparticles on sperm quality and quantity,and plasma sex steroid levels in northern pike

Artificial reproduction of northern pike Esox lucius is impeded by the likelihood of obtaining only a small volume of sperm of inconsistent quality. A controlled-release hormone delivery system has the potential to enhance sperm production while avoiding multiple injections The objective of this study was to investigate the effects of mammalian gonadotropin-releasing hormone agonist (mGnRHa) incorporated into poly(lactic-co-glycolic acid) (PLGA) microparticles on milt production, spermatozoon characteristics, and secretion of 17β-estradiol (E2), 11-keto testosterone (11-KT), and testosterone in northern pike. Fish were divided into four groups and injected with 2 mg/kg BW carp pituitary extract (CPE), 20 µg/kg BW mGnRHa in PLGA microparticles, or 20 µg/kg BW mGnRHa plus 20 mg/kg BW metoclopramide (MET) in PLGA microparticles (PLGA + MET), along with a control group injected with 1 ml/kg 0.9% NaCl. At 48 h postinjection, the volume of milt produced was significantly greater in groups treated with CPE and PLGA + MET than in other groups. At 96 h postinjection, all hormone-treated groups exhibited significantly higher spermatozoon average velocity than recorded in the control group. Spermatozoon motility was significantly increased (P < 0.05) in the CPE and PLGA groups compared to baseline values. All treated groups showed significantly lower levels of 11-KT after the hormone injection compared to baseline values and to controls. Plasma testosterone levels increased in all hormone-treated groups. The use of PLGA microparticles, with or without metoclopramide, is suitable for use as a carrier of hormone treatments to regulate spermiation in mature northern pike.     

Female rainbow trout urine contains a pheromone which causes a rapid rise in plasma 17,20β-dihydroxy-4-pregnen-3-one levels and milt amounts in males

Previous studies have shown that spermiating male rainbow trout respond to the presence of female urine in the water with significant increases in plasma levels of gonadotrophin II, 17,20β-dihydroxy-4-pregnen-3-one (17,20β-P) and testosterone. The present results show that males only need a single brief exposure to female urine in order to respond; levels of 17,20β-P rise significantly within 1 h of exposure, and peak between 3 and 4 h. Also, milt amounts increase significantly following exposure of males to female urine. Levels of 17,20β-P are also related positively to the amount of female urine to which the males are exposed. Furthermore, when live females are placed, out of physical and visual contact, in the same tank as males, levels of 17,20β-P rise in the same way as in males which are exposed to female urine. However, if females are fitted with urinary catheters (which drain the urine outside the tank), males respond more slowly. These results indicate that urine is the main source of the male 'priming' pheromone.     

Induced sex change in black sea bass

Experiments were conducted to identify factors involved in sex change in the protogynous black sea bass Centropristis striata . Black sea bass maintained in the ratio of 8 females (F):0 males (M) for 9 months reversed sex while those kept at the ratios of 6F:2M or 4F:4M did not. Female black sea bass implanted with 1·0 mg 11-ketotestosterone (11-KT) or 10 mg fadrozole (FAD) changed sex and began spermiating while those implanted with 0·1 mg 11-KT or 1·0 mg FAD underwent incomplete sex reversal. One fish implanted with 1·0 mg FAD initiated sex change but was not spermiating at the end of the study. One fish in the control group, the largest fish in the study, initiated sex change. These results suggest that the presence of males may restrict sex reversal in black sea bass and that high 17β-oestradiol:11-KT is required for maintaining ovarian function.     

Annual reproductive and spawning cycles of femaleSebastiscus marmoratus

Synopsis Changes in serum steroid hormones were studied during the reproductive cycle of a viviparous rockfish,Sebastiscus marmoratus. Serum levels of estradiol-17β (E₂) and testosterone (T) were moderately high throughout the spawning period from December until February (E₂), and until post-spawning in April (T). Serum progesterone (prog) fluctuated but remained low throughout the annual reproductive cycle; 17α,20β-dihydroxy-4-pregnen-3-one (17α, 20β-diOHprog), on the other hand, was relatively high during the spawning period. During the spawning period, 7 of 12 females reared under laboratory conditions spawned twice at 10-to 16-day intervals. Histological observations indicated that oocytes developed gradually during gestation of the preceding brood and; after parturition, developed more quickly towards the end of vitellogenesis and subsequent fertilization. In repeat spawners, E₂ and female-specific serum proteins remained high several days after the first parturition, then gradually decreased. Prog showed no significant changes over the period. The 17α, 20β-diOHprog, however, was low immediately after parturition, then rapidly increased, remained elevated during the middle of the period and then decreased. These results indicate that E₂ is involved in vitellogenesis, and 17α, 20β-diOHprog may have some important roles in gestation in the multiple spawnerS. marmoratus.     

Polar steroids from <Emphasis Type="Italic">Solaster endeca</Emphasis> starfish and the physiological activity of polar steroids from three starfish species

Four polyhydroxylated steroids, new (20R)-5α-cholestan-3β,6α,8,15α,24,26-hexaol (I) and known (20R,25S)-5α-cholestan-3β,6α,8,15β,16β,26-hexaol, (20R,25S)-5α-cholestan-3β,6α,15β,16β,26-pentaol, and marthasterone sulfate were isolated from the Solaster endeca starfish inhabiting the Sea of Okhotsk and characterized. Steroid (I) contains a 24,26-dihydroxylated side chain, which is unique for starfish polyols. The isolated steroids and related metabolites from two starfish species of the Evasterias genus (in total, 15 compounds) were weakly cytotoxic in a human HeLa cell culture and some of them were inhibitors of non-specific esterase from mouse Ehrlich carcinoma. The effects of these compounds on the p53 protein activity were studied in a yeast two-hybrid test system and both inhibitors and stimulators of this activity were found among them.     

Responses of round goby (<Emphasis Type="Italic">Neogobius melanostomus</Emphasis>) olfactory epithelium to steroids released by reproductive males

The wild perciform teleost Neogobius melanostomus (the round goby) originated from the Ponto-Caspian region and is now a highly successful invasive species in the Laurentian Great Lakes. Males may attract females into their nests for spawning by releasing reproductive pheromones, and it has been previously shown that reproductive males synthesize and release the 5β-reduced and 3α-hydroxyl steroids 3α-hydroxy-5β-androstane-11,17-dione (11-oxo-etiocholanolone; 11-O-ETIO) and 3α-hydroxy-5β-androstane-11,17-dione 3-sulfate (11-oxo-etiocholanolone-3-sulfate; 11-O-ETIO-3-s) and 3α,17β-dihydroxy-5β-androstan-11-one 17-sulfate. In this study, we investigated properties of these released steroids by recording field potential responses from the olfactory epithelium (electro-olfactogram, EOG). The steroid 3α,17β-dihydroxy-5β-androstan-11-one 17-sulfate did not elicit olfactory responses while both 11-O-ETIO and 11-O-ETIO-3-s stimulated olfactory field potentials in the round goby, but not in the goldfish. Cross-adaptation analysis demonstrated that round gobies discriminated between11-O-ETIO and 11-O-ETIO-3-s (as well as etiocholanolone, ETIO) at the sensory level. Second messenger cascades depending on both cAMP and IP₃ were inferred for steroids from pharmacological inhibition studies, while the canonical teleost odors taurocholic acid (a bile acid) and l-alanine (an amino acid) used only cAMP and IP₃, respectively. The round goby presents itself as an excellent species for the study of olfactory function of fish in the wild, given its possible use of these released steroids as pheromones.