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Sperm adhering to glass slides is one of the main problems during fish sperm motility analyses with CASA systems. To mitigate this, albumin is the supplement added most frequently to activating solutions. However, there is no data on the use of supplements other than albumin (in various concentrations) in analyses of European whitefish (Coregonus lavaretus) sperm motility. This issue was investigated in the presented research using three anti-adhesive supplements (albumin, casein, Pluronic F-127) that were added to Billard solution (BS: 20 mM Tris, 1 mM CaCl 2 , 154 mM NaCl, 30 mM glycine at pH 9.0) at different concentrations (0.0; 0.1; 0.2; 0.5; 1.0; 2.0%). It was noted that the addition of the lowest concentration (0.1%) of albumin, casein, or the pluronic to BS had a significant effect on the motility and kinetic parameters of whitefish sperm compared to pure BS. BS supplemented with 0.2-0.5% albumin was the most appropriate variant used for whitefish sperm motility activation in the present experiment. BS supplemented with the pluronic at 1.0-2.0% concentrations resulted in significantly higher values of almost all CASA parameters compared to casein at the same concentrations. Moreover, CASA parameters determined in this variant of the pluronic (1.0-2.0%) were similar to those when BS was supplemented with the same albumin concentrations. This indicated that instead of albumin, the pluronic at higher concentrations in BS might be used to analyze whitefish sperm motility., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The purpose of this study was to compare four activation solutions (AS)-Woynarovich, Lahnsteiner, Kucharczyk, and Perchec-with the addition of 0.5% bovine serum albumin (BSA) for ide ( Leuciscus idus ) sperm activation and analysis with a CASA system. It was found that ide sperm can be activated using each AS within a pH range of 7.4-9.0 and an osmolality range of 160-200 mOsm kg -1 . The effect of Woynarovich and Perchec solutions supplemented with BSA and casein at concentrations of 0.25, 0.5, 1.0, and 2.0% were also analyzed during the experiment. These two AS without protein supplementation (pure solutions) were the controls. Woynarovich and Perchec solutions supplemented with the minimum BSA concentration (i.e., 0.25%) significantly improved sperm motility (89.05% and 86.63%, respectively) compared to the controls (20.39 and 28.48%, respectively). Similar increases were also noted in progressively motile sperm (PRG, %), the curvilinear velocity of sperm (VCL, µm s -1 ), and the amplitude of lateral head displacement (ALH, µm). A similar trend in CASA parameters was also noted when casein was added to Woynarovich and Perchec solutions at a concentration of 0.25%. We concluded that 0.25% doses of each of the proteins were sufficient to prevent sperm adhesion to glass slides, and they can be used in research on ide sperm motility measurements.

The aim of this study was to describe the selected stages of embryonic development in Siberian sturgeon ( Acipenser baerii ) and hybrids of Siberian sturgeons and Russian sturgeons ( Acipenser baerii × Acipenser gueldenstaedtii ). For this purpose, embryos representing nine distinct developmental stages (stage 1-2.0 hpf, stage 2-5.5 hpf, stage 3-13.0 hpf, stage 4-20.0 hpf, stage 5-24.0 hpf, stage 6-26.0 hpf, stage 7-35.0 hpf, stage 8-55.0 hpf, and, stage 9-160.0 hpf; hpf-hours postfertilization) were sampled from each group (group A, group B, and group C) during incubation. Stages of embryogenesis were identified based on a 30-point scale of embryonic development in sturgeons. A total of 13 developmental stages were identified, including early cleavage, blastula formation, early and late gastrulation, onset of neurulation, beginning of organogenesis, and prelarvae. During gastrulation, the survival of hybrid embryos was highest in group B (93.8%) and lowest in group A (86.7%). Embryonic deformation was not observed during experimental incubation. The archived data relating to the embryonic development of Siberian sturgeon × Russian sturgeon hybrids could be applied to identify the individual stages of embryogenesis in hybrid sturgeons during egg incubation.

In cyprinids, Tyrode's (TLP) and Volckaert's (VRT) solutions are the most frequently used extenders for short-term sperm storage. The effectiveness of TLP and VRT on ide (Leuciscus idus) sperm short-term storage was analyzed using a × 4 (sperm:extender) dilution ratio over 48 h. A × 4 (1:3) dilution ratio was compared to a × 10 (1:9) dilution ratio for ide sperm storage using TLP supplemented with antibiotics and was tested for a 14-day period. Sperm motility (MOT, %), progressively motile sperm (PRG, %), curvilinear velocity of sperm (VCL, µm s -1 ), movement linearity (LIN, %), beat cross frequency (BCF, Hz), and the amplitude of lateral head displacement (ALH, µm) were verified using a CASA system. After 48 h, most CASA parameters were significantly higher in TLP compared to VRT. The dilution ratio also had a significant impact (P < 0.0001) on the efficiency of ide sperm short-term storage over 14 d in comparison to undiluted sperm (control). Ide sperm diluted with TLP supplemented with penicillin/streptomycin and stored short term, regardless of the dilution ratio used, retained motility and fertilization capacity over 14 d at 4 °C. The highest embryo survival rates of 70% and 73% were noted using sperm diluted with TLP at × 4 and × 10 dilution ratios compared to 48% for raw sperm that was not stored short term., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Although ruffe (Gymnocephalus cernua) are widely distributed in Eurasia, in some regions (i.e., North America) ruffe are considered an invasive species. There have been no reports on artificial reproduction of this species. A study, therefore, was conducted to evaluate reproduction of ruffe with results of specimens captured from their natural habitat and cultured (F 1 generation) specimens being compared. Spawning specimens from both stocks were treated with hormonal preparations: carp pituitary homogenate (CPH), Ovopel, Ovaprim and human chorionic gonadotropin (hCG) and results were analyzed by comparing response to the specimens of control groups. Spermiation outcomes and sperm motility of the captured and cultured ruffe were similar and with all hormonal treatments, there was a slightly greater sperm motility (55.6 %-57.1 %) in comparison to specimens of control groups (46.7 %-47.1 %). For captured specimens, there was no asynchronous development of oocytes, whereas in cultured specimens 32 % of females had asynchronous development of oocytes. The ovulation rate in specimens of all treated groups was 100 %, whereas specimens in the control groups did not reproductively mature and have ovulations. The latency time from time of hormonal treatments to initiation of reproductive functions depended on the spawning agent used and oocyte maturation stage and there was the shortest latency after using CPH and the longest with hCG treatment. The embryo survival and hatching rates varied with use of different hormonal preparations to induce reproduction: greatest hatching rates with hCG treatment (86.4 %-88.9 %), followed by Ovaprim (78.2 %-80.2 %) and least hatching rate with Ovopel and CPH treatments (66.0 %-67.1 % and 64.0 %-66.0 %, respectively)., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Optimization of artificial reproduction, by increasing the survival rate of embryos and hatching rate, is of major importance for reducing genetic diversity, especially in fish captured from their natural habitat that subsequently spawn in hatcheries. The artificial reproduction of ide, Leuciscus idus (L.) was conducted in controlled conditions. The spawning agents included: different doses of human chorionic gonadotropin (hCG) that was compared with common carp pituitary homogenate (CPH), Ovopel - a commercial agent for induction of spawning that contains mammalian gonadotropin releasing hormone analogue (mGnRHa) with dopamine antagonists (DA): metoclopramide (MET) and Ovaprim - a commercial agent containing salmon gonadotropin releasing hormone analogue (sGnRHa) and a dopamine antagonists (DA): domperidone (DOM). There were no ovulations in females from control groups. There were no differences between the ovulation rates (90 %) or embryo survival (> 92 %) and hatching rates (> 91 %) when there was administration of hCG doses between 500 and 1000 IU/kg. When there was comparison of different spawning agents, the ovulation rate was 100 % for all treated groups. There were the shortest and longest latency times to the time of ovulation after administration of CPH (26 h) and hCG (79 h), respectively. The greatest embryo survival (> 93 %) and hatching (> 91 %) rates occurred as a result of hCG administration with these values being slightly greater than when there was treatment with Ovaprim. The association between latency time and hatching rate indicated that when there was a slower final oocyte maturation (FOM) there were greater hatching rates., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The effect of different treatment agents, namely, carp pituitary homogenate (CPH), Ovaprim ([D-Arg 6 , Pro 9 NEt]-sGnRH + domperidone) and a dopamine-receptor antagonist (metoclopramide), on the stimulation of northern pike (Esox lucius) spermiation was tested under controlled conditions. To carry out the experiment, males (n = 84) were divided into four groups: control (n = 21); CPH (n = 21); Ovaprim (n = 21); metoclopramide (n = 21). The control group was given 0.9% NaCl but no hormonal treatment. After 24 h, sperm was collected from seven males belonging to control (n = 7), CPH (n = 7), Ovaprim (n = 7) and metoclopramide (n = 7). This procedure was repeated after 48 and 72 h post-treatment. At each time, sperm was collected from seven males from each group only once. After collection, the quantity and quality of sperm were determined. It was confirmed that the treatment agent and latency time (the time between treatment and sperm collection) are two factors affecting the quantity and quality of northern pike sperm collected under controlled conditions. The highest total sperm volume and total sperm production (TSP) were noted in the CPH group compared to the Ovaprim, metoclopramide and control groups. In contrast, the time of sperm collection affected the sperm concentration (SC), TSP and sperm pH. With increasing time, SC and TSP decreased, which indicated the occurrence of sperm hydration being part of the final sperm maturation process. Sperm maturation is in turn a consequence of increases in sperm pH and seminal plasma osmotic pressure between 48 and 72 h post-treatment. Sperm motility and sperm kinetic parameters were affected by treatment agent and the time of sperm collection. This indicates that the sperm's ability to move that is achieved in the optimal environment (in spermatic ducts) is dependent on both factors which determine sperm maturation in northern pike under controlled condition., (Copyright © 2019 Elsevier Inc. All rights reserved.)

We describe the technique of pneumatic stripping of whitefish ( Coregonus lavaretus ) eggs with the use of oxygen, nitrogen, and air. Eggs obtained via the traditional method (by pressing the abdominal surfaces) served as a control group. It was established that the gas flow rate during pneumatic stripping should not exceed 0.5 L∙min -1 , since higher air flow resulted in increased post-spawning mortality. The pneumatic stripping method of egg collection was no faster than hand stripping; however, the time required per female was more consistent. It was found that the pH of the ovarian fluid obtained during hand and pneumatic stripping was not related to the success rate of fertilization. Pneumatic stripping resulted in a higher quality of collected eggs and a higher and more consistent hatching rate as compared with the hand-stripped samples, regardless of the gas used. The results presented here lead us to recommend the pneumatic method for obtaining eggs from whitefish, since it is a simple, reproducible method and improves the reproductive performance and developmental success of the fish eggs.

Fish sperm quality assessment is helpful for optimizing production and for monitoring the environmental state. Sperm can be monitored relatively easy and, to date, various analyses have been applied and proven to be helpful in this task. Among them, sperm motility parameters such as sperm speed are one of the main performance traits during assisted fish reproduction. Apart from motility the sperm concentration, volume, and seminal plasma pH and osmolality are also frequently evaluated and are the main sperm quality indicators measured in fish sperm. However, other parameters also determine sperm fertilization potential. Recent knowledge reveals several additional parameters of high importance for sperm function. Among them are DNA integration, membrane stability, mitochondria status and enzymatic activity. Measuring all these parameters in fish sperm provides complex knowledge regarding male fertility and helps to improve broodstock maintenance protocols as well as gamete handling and fertilization processes. This review focuses on the presentation of the sperm quality measures for freshwater and marine species of the fish and provides information regarding recent methods of sperm quality evaluation., (Copyright © 2019 Elsevier Inc. All rights reserved.)

In this study, there was the first attempt to sex immature European river lampreys Lampetra fluviatilis, classified as Agnatha using ultrasonography. This species starts a spawning migration from seas to rivers in the autumn and reproduction is initiated in the late spring. It is recommended to collect breeders soon after the beginning of the spawning migration, however, to date no method has been developed for distinguishing the sex of individuals during this developmental period. The lampreys for the present study were caught in autumn (November) in the Vistula River (northern Poland) during the period of spawning migration and transported to the laboratory. The lampreys were anaesthetised (MS-222, dose: 0.1 g / dm 3 prior to sex determinations (n = 100) using ultrasonography. The images obtained using ultrasonography were verified with post-mortem and histological examinations. The findings with this study confirmed that the sex of the European river lamprey can be effectively ascertained much earlier than can occur with assessment of external secondary sexual characteristics (e.g., sexual papilla, ovipositor, skinfold). The advantages of the method include: 100% effectiveness, survival of the fish after examination, non-invasiveness, rapid verification of the lamprey sex and the possibility of determining extent of gonadal development., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The research reported focuses on reproduction of the river lamprey Lampetra fluviatilis,(Linnaeus, 1758) in controlled conditions. There was specific emphasis on fish harvesting dates (autumn and spring), holding conditions and reproduction in a controlled environment. Attempts were also made to synchronize the time of ovulation among river lampreys, egg and sperm collections. Hormonal stimulation was conducted using carp pituitary homogenate (CPH) at a total dose of 4 mg/kg which allowed for shortening of the egg-laying period from 2 to 3 weeks to a few days while sustaining embryo survival rates and larvae quality. River lamprey males were found to not require hormonal treatment to yield good-quality sperm, as measured using the CASA system. River lamprey broodstocks adapted well to different manipulations in hatchery conditions when harvested in the autumn and spring. The results of the present study may be used to restore endangered natural populations of the river lamprey (egg and sperm collection, fertilization or gamete preservation) because ovulation and spermiation synchronization is very difficult to achieve without hormonal treatment in controlled conditions., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The effect of sodium and potassium concentrations as well as optimal pH on the motility of common carp Cyprinus carpio L. sperm during short-term storage in artificial seminal plasma (ASP) was investigated. Sperm was collected from individual males (n = 5) and each sample diluted tenfold (1:9) in ASP (sperm:extender) containing 2 mM CaCl 2 , 1 mM Mg 2 SO 4 and 20 mM Tris at pH 8.0 and supplemented by the following concentrations of sodium and potassium (mM/mM): 0/150, 20/130, 40/110, 75/75, 110/40, 130/20 and 150/0. The osmolality of all ASP variants was set at 310 mOsm kg -1 . Sperm motility was measured using a CASA system during 72 h of storage. Immediately after dilution, sperm motility was high (90%) both in each variant and in the control group (fresh sperm). After 72-h storage, the highest sperm motility was noted in ASP containing 110 mM NaCl and 40 mM KCl. No differences were found in the motility of samples preserved within the pH range of 7.0-9.0. Our data suggest that for the short-term storage of common carp sperm, whereas the pH of the solution does not play a crucial role, a specific potassium concentration of around 40 mM is required.

Egg collection is one of the most crucial procedures during fish reproduction in salmonid hatcheries. Classic methods involve the use of hand massage on fish abdomens to expel the eggs. An alternative method uses the pressure of gas injected into the body cavity, which causes the subsequent release of the eggs. This method is believed to have less negative effects on both the welfare and egg quality of the broodstocks. Herein, we compare the results of air and hand stripping methods with respect to one-year survival and egg quantity and quality in two salmonid fish, rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta morpha fario). Our results indicate that air stripping yielded a better quality of eggs and higher one-year survival rate in rainbow trout. In addition, air stripping resulted in lower mortality rate than the group subjected to hand stripping (25% vs. 35%). The pH and hatching rate of the hand stripped group was lower than those of the air stripped group. In the case of brown trout, the quality of eggs obtained by both hand and air-stripping methods was similar; however, the one-year losses in fish were higher in air stripped group (15% compared to 0% in hand stripped fish). Although the advantages of air stripping method over hand stripping in terms of egg quality might not be observed in all salmonid species, the air-stripping procedure might be a promising option to be adopted in hatcheries as it ensures a high level of reproducibility and efficiency.

The effect of carp pituitary homogenate (CPH, n = 7) at a dose of 2.0 mg kg -1 and (D-Arg 6 , Pro 9 NET)-sGnRH + domperidone (Ovaprim, n = 7) at a dose of 0.5 ml kg -1 in northern pike (Esox lucius) sperm maturation under controlled conditions was examined. On the control group, 0.9% NaCl at a dose of 1.0 ml kg -1 (n = 7) was used. Sperm was collected 48 h following injection. Sperm quantity (total sperm volume, total sperm production and sperm concentration), sperm motility using the Computer-assisted sperm analysis (CASA) system and sperm and seminal plasma quality (sperm pH, seminal plasma osmotic pressure and seminal plasma pH) were determined for each male separately in each group. The results of the present study demonstrate that hormonal treatment had a positive effect on sperm maturation in northern pike, regardless of the hormonal preparation used. However, even though no differences were found in total sperm volume and total sperm production between fish injected with either CPH or Ovaprim, it should be highlighted that the highest progressive motile sperm (PRG), straight-linear velocity (VSL) and movement linearity (LIN) was noted in fish treated with Ovaprim. It was also found that it is possible to collect sperm from non-hormonally manipulated fish. However, in such a case, only a small sperm volume (around 0.1 ml) characterised by lowered PRG (below 40%) was noted. Considering the fact that only after Ovaprim application sperm motility with progressive movement of sperm above 50% was observed, the hatchery practice of collection of sperm 48 h after its application (at a dose of 0.5 ml kg -1 ) may be recommended., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Multiple collections of semen during the reproductive period of the common carp Cyprinus carpio L. were used to analyse changes in semen quality. Semen collection was performed on June 1 (first collection), 12 (second collection), and 19 (third collection) from individual males (n=11) by gentle abdominal massage. Semen quantity (semen volume and sperm count), quality (sperm motility and sperm viability), as well as seminal plasma parameters (pH of seminal plasma and seminal plasma osmotic pressure) and its enzymatic activity, e.g., lactate dehydrogenase (LDH) and ß-N-acetylglucosaminidase (ß-NAG) were determined. Moreover, for the first time, the percentage of live, dead, and apoptotic sperm, as well as the proteolytic activity of seminal plasma, were determined using flow cytometry and zymography, respectively, at specific times during the common carp reproductive period. The lowest volumes of semen and sperm concentration were noted during the first semen collection (June 1). Analysis of computer-assisted sperm analysis parameters revealed the greatest sperm motility, sperm velocity, as well as amplitude of lateral head displacement, were evident in the third collection (June 19). There were no differences in progressively motile sperm, movement linearity, wobbling index, and beat cross frequency between the different collection times. The lowest percentage of live sperm was found in the first collection, although with the passage of time values of this parameter increased. Seminal plasma pH and seminal plasma osmotic pressure were at the lowest values in the second collection (June 12), which corresponded with the lowest concentration of sperm. In the first collection, seminal plasma contained the highest values of LDH and ß-NAG activity, whereas there were no differences in the proteolytic activity of seminal plasma determined between the different collections of semen. The results presented here indicate that during the reproductive period, males of common carp produce a large amount of semen of moderate quality. Low sperm motility noted in the second collection might be explained by a significant increase in sperm production during this period, followed by a low seminal plasma pH and high hydration rate. The high LDH and ß-NAG activity noted in the first collection of semen may reflect a reduced stability of the sperm cell membrane and its viability. The significant difference in the percentage of live sperm at June 1 compared to that at June 19 supports this hypothesis., (Copyright © 2017 Elsevier B.V. All rights reserved.)

This study analysed (i) the effect of human chorionic gonadotropin (hCG) and salmon gonadoliberine analogue (sGnRHa) on the effectiveness of induction of spermiation and (ii) the effect of latency time following the application of those spawning agents on the quantity and quality of the sperm of Eurasian perch, Perca fluviatilis, obtained during out-of-season spawning. For this study, pond-reared fish were used which had been acclimated to the controlled conditions. Three groups were distinguished which were treated with either saline (0.9% NaCl; control group), hCG (500 IU kg -1 ) or sGnRHa (100 μg kg -1 ). The fish were kept in a recirculating system at 12 °C throughout the study, during which sperm was collected every two days between the 2nd and 10th day following hormonal treatment. During the study, quantitative (e.g. sperm volume, total sperm production) and qualitative (measured with a computer-assisted sperm analysis system - i.e. CASA) parameters were monitored. The results of the study indicate that the hormonal treatment had a highly beneficial effect on the spermiation rate (100% in experimental groups from day 6 following injection) as well as quantity, which increased 50% in experimental groups (over 2200 × 10 9 of spermatozoa per kg of body weight) by day 4 following injection. For the sperm quality, both spawning agents tested had a rather positive effect, although sperm motility rate (MOT) was seen to be significantly reduced on day 10 following the application of hCG (MOT = 72.8% ± 8.1), which was not observed after the application of sGnRHa (minimum mean MOT 81.7% ± 6.1). The results clearly indicate that hormonal treatment had a positive effect on spermiation in Eurasian perch, most apparent from day 6 following injection, regardless of the hormonal agent used. Though application of sGnRHa allowed a high volume of high quality sperm to be stripped for two days longer (up to day 10 post-injection) compared to the application of hCG., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The application of zymography, with sperm proteins as a substrate, allowed for the first time the visualisation of two serine proteinases with a molecular weight of 76 and 163kDa from common carp Cyprinus carpio L. seminal plasma. Twenty four hours of incubation in a development solution with a pH of 7.5 and incubation at 37°C were the best conditions for the visualisation of serine proteinase; however, proteolysis was also observed at 4°C. Our results indicate that serine proteinase from common carp seminal plasma with a molecular weight of 76 and 163kDa may be involved in the degradative mechanism of sperm proteins. This mechanism may be responsible for the removal of damaged sperms by the digestion of native sperm proteins., (Copyright © 2017 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.)

The diploid-polyploid populations of Cobitis distributed in Poland are usually composed of the spined loach Cobitis taenia or, less often, the Danubian loach C. elongatoides and their triploid (females) and tetraploid hybrids (females and males). The aim of this study was to determine whether tetraploid males participate in the reproduction process by analyzing their testis ultrastructure and the process of spermatogenesis in comparison with diploid males of both parental species. Tetraploid loaches were obtained from three different diploid-polyploid populations distributed in Poland. The structure of Cobitis testes are typical for most Teleostei fish with cystic-type spermatogenesis. The successive stages of developing germ cells are enclosed within cysts formed by the Sertoli cells. This paper morphologically describes the different germ cell stages of spermatogenesis (spermatogonia, spermatocytes, spermatids, and spermatozoa) of C. taenia and C. elongatoides and provides a pioneering ultrastructural analysis of tetraploid Cobitis testes which reveals their unusual structure for the first time. Thus, cysts with normal spermatogonia and spermatocytes (pachyten or leptoten stages) containing synaptonemal complexes were present and no spermatids or spermatozoa were observed. Moreover, in contrast to previously analyzed diploid species, single cells or all of the cells within the cysts displayed chromatin condensation and/or chromatin fragmentation. The obtained results clearly demonstrated that tetraploid males are sterile and diploids are fertile and are the only sperm donors in the reproduction processes of diploid-polyploid Cobitis populations., (© 2017 Wiley Periodicals, Inc.)

Reproductive performance (ovulation/spermation rate and relative fecundity of females) of adult river lamprey (Lampetrafluviatilis) was compared among adults held under three different controlled thermal regimes (7, 10 and 14°C). The quantity of semen (volume of semen, sperm concentration, total sperm production and total number of sperm) and the weight of the eggs as well as the semen quality (sperm motility, seminal plasma osmolality and pH, sperm pH and total protein content) were determined. Housing temperature had no apparent effect on quality or quantity of eggs produced, but did influence time of ovulation. On the other hand, temperature had a significant effect on the quantity and quality of sperm produced; 70% of males held at 10°C and 14°C did not spermiate. The best ambient temperature for river lamprey adults held under controlled conditions was 7°C. All males kept in this temperature yielded mature semen with the highest sperm motility parameters. It was also found that among the six tested solutions, the most suitable artificial medium for river lamprey sperm activation was 20mM Tris buffer containing 40mM NaHCO 3 at pH 8.5 and 100mOsmkg -1 ., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Substrate specificity in the seminal plasma and testes fluids of the common carp Cyprinus carpio was determined using gelatin, casein, albumin and haemoglobin. Proteolytic profiles of the testes and seminal plasma were compared. Different ranges of pH (5·5-9·5) and temperature (4-37° C) were used during incubations of seminal plasma proteinases. Differences in proteolytic activity between testes and seminal plasma may reflect specific functions of the testes and sperm ducts in semen production. Seminal plasma metalloproteinases were characterized by higher substrate specificity than were serine proteinases. Zymography optimization for seminal plasma indicated that pH 7·5 and 22° C were the optimal conditions for gel incubations., (© 2016 The Fisheries Society of the British Isles.)

The effectiveness of Ovopel (two pellets/kg body weight), a complex of [D-Ala(6) Pro(9)NEt-mGnRH] and dopamine antagonist (DA e.g. Metoclopramide, MET), was compared with the application MET alone at the dose of 20mg/kg body weight in spawning crucian carp Carassius carassius (L.) in the present study. Ovulation and spermation rates were greater after MET application (90% and 100%, respectively) in comparison to the Ovopel (80% and 87.5%, respectively) when used as a broodstock reproductive stimulation agent. Sperm quality (CASA parameters) after MET treatment was very acceptable without a significant decrease in comparison to the samples from Ovopel-treated crucian carp when spermation stimulation treatments were employed. Though there were no differences in egg and sperm production between Ovopel and MET treatments, hatching rates were significantly greater after MET (84.4%) compared with Ovopel (76.2%) treatment. The usefulness of MET application without GnRHa addition for stimulation of reproduction was ascertained in the present study for the first time in crucian carp., (Copyright © 2015 Elsevier B.V. All rights reserved.)

β-N-Acetylglucosaminidase (β-NAGase) is an enzyme found in the sperm acrosome of numerous animal species including fish. Fish spermatozoa differ in their morphology including acrosome or acrosomeless aquasperm in chondrostean (e.g., sturgeon) and teleostean (e.g., rainbow trout). It has been shown that β-NAGase exists with high activity in both eggs and sperm of these species. The present study shows the potency of β-NAGase in fertilization. In rainbow trout, increase in sperm motility parameters (VAP and MOT) were observed in the presence of acetamide, an inhibitor for β-NAGase. In contrast, sperm motility parameters (VCL, VSL, VAP, MOT, and PRG) were reduced on the Siberian sturgeon in the presence of acetamide. The inhibition of the activity of β-NAGase in rainbow trout spermatozoa was led to a reduction in the number of fertilized eggs from 79% to 40%, whereas in sturgeon no change was observed in fertilization. Moreover, inhibition of β-NAGase in both spermatozoa and eggs of trout and sturgeon resulted in significant decrease in fertilization rate from 79% to 1% in rainbow trout and from 84% to 12% in Siberian sturgeon. Our research proves that β-NAGase can play a significant role in the fertilization process in teleosteans., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The effect of Ovaprim (salmon GnRH analogue and a dopamine antagonist) treatment on the quantity and quality of smelt (Osmerus eperlanus L.) sperm was studied in relation to time after hormonal stimulation. Sperm was obtained at 0, 24, 48 and 72 h after treatment (n=13/each time point). Computer Assisted Sperm Analysis (CASA) was used to evaluate sperm motility parameters and histological analysis was used to examine the testis morphology. Only a small volume of semen (1-5 µl) was collected at the beginning of the experiment (time 0) but it dramatically increased 24 h after hormonal treatment. A further increase in semen volume was recorded 48 h after hormonal stimulation. CASA parameters, such as percentage of motile cells, progressive motility, curvilinear velocity, straightlinear velocity, straightness and amplitude of lateral head displacement of stripped sperm increased 48 h after hormonal treatment, which indicates high quality of sperm. No further increase in sperm quality was recorded at 72 h. Compared to stripped semen, testicular semen was characterized by a slightly lower quality. In addition, histological analysis indicated that 24 h after hormonal treatment, a high number of spermatozoa was released from the testis. Ovaprim-stimulated smelt became clearly darker than the control fish. In conclusion, our results suggest that smelt semen should be collected 48 h after hormonal stimulation to ensure high quality of semen. This time may vary depending on maturation status of testis.

The aim of this work was to compare the effects of controlled reproduction of cultured and wild common barbel, Barbus barbus (L.). Preparations containing different GnRH analogues and dopamine receptor antagonists (Ovopel, Ovaprim) as well as human chorionic gonadotropin (hCG) (in the case of cultured fish) were applied and their influence on ovulation, spermiation and quality of gametes obtained was determined. No differences in the qualitative or quantitative parameters of semen were found between fish stimulated with different hormonal preparations and those not receiving hormonal stimulation. The high suitability of Ovaprim for ovulation induction in (cultured and wild) barbel was confirmed. The highest synchronisation of ovulation was obtained after the application of Ovopel (18 ± 3 h), but the best results of controlled reproduction (expressed as the percentage of ovulations and survival of embryos) were obtained by applying Ovaprim (83.2 ± 4.1). A significantly higher percentage of ovulation was obtained in cultured fish (80-90%) than in wild fish (< 25%).