The aim of the present study was to investigate the spontaneous motility of spermatozoa and to optimize sperm collection, short-term sperm storage, and fertilization in zebrafish Danio rerio. The movement of spermatozoon in water was propagated along the flagellum at 16 s after sperm activation then damped from the end of the flagellum for 35 s and fully disappeared at 61 s after activation. For artificial fertilization, milt must be added to an immobilizing solution, which stops the movement of sperm and keeps the sperm motionless until fertilization. E400 and Kurokura as isotonic solutions were shown to be suitable extenders to store sperm for fertilization for 6 h. E400 stored sperm for 12 h at 0-2 °C. Sperm motility decreased only to 36% at 12 h post stripping for the E400 extender and to 19% for the Kurokura extender. To achieve an optimal level of fertilization and swim-up larvae rates, a test tube with a well-defined amount of 6,000,000 spermatozoa in E400 extender per 100 eggs and 100 µL of activation solution has proven to be more successful than using a Petri dish. The highest fertilization and swim-up larvae rates reached 80% and 40-60%, respectively, with milt stored for 1.5 h in the E400 extender at 0-2 °C.

College of Life Science Northwest University Xi'an 710069 China

South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses Research Institute of Fish Culture and Hydrobiology Faculty of Fisheries and Protection of Waters University of South Bohemia in Ceske Budejovice Zatisi 728 2 389 25 Vodnany Czech Republic

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Hagedorn M., Carter V.L. Zebrafish reproduction: Revisiting in vitro fertilization to increase sperm cryopreservation success. PLoS ONE. 2011;6:e21059. doi: 10.1371/journal.pone.0021059. PubMed DOI PMC

Franěk R., Tichopad T., Fucikova M., Steinbach C., Psenicka M. Production and use of triploid zebrafish for surrogate reproduction. Theriogenology. 2019;140:33–43. doi: 10.1016/j.theriogenology.2019.08.016. PubMed DOI

Van Eeden F.J.M., Granato M., Odenthal J., Haffter P. Developmental mutant screens in the zebrafish. Methods Cell Biol. 1999;60:21–41. PubMed

Briggs J.P. The zebrafish: A new model organism for integrative physiology. Am. J. Physiol. Regul. Integr. Comp. Physiol. 2002;282:R3–R9. doi: 10.1152/ajpregu.00589.2001. PubMed DOI

Bambino K., Chu J. Zebrafish in toxicology and environmental health. Curr. Top. Dev. Biol. 2017;124:331–367. PubMed PMC

Berghmans S., Jette C., Langenau D., Hsu K., Stewart R., Look T., Kanki J.P. Making waves in cancer research: New models in the zebrafish. Biotechniques. 2005;39:227–237. doi: 10.2144/05392RV02. PubMed DOI

Jiang L., Zhang J., Wang J.J., Wang L., Zhang L., Li G., Yang X., Ma X., Sun X., Cai J., et al. Sperm, but not oocyte, DNA methylome is inherited by zebrafish early embryos. Cell. 2013;153:773–784. doi: 10.1016/j.cell.2013.04.041. PubMed DOI PMC

Potok M.E., Nix D.A., Parnell T.J., Cairns B.R. Reprogramming the maternal zebrafish genome after fertilization to match the paternal methylation pattern. Cell. 2013;153:759–772. doi: 10.1016/j.cell.2013.04.030. PubMed DOI PMC

Cosson J. The ionic and osmotic factors controlling motility of fish spermatozoa. Aquacult. Int. 2004;12:69–85. doi: 10.1023/B:AQUI.0000017189.44263.bc. DOI

Alavi S.M., Cosson J. Sperm motility in fishes. (II) Effects of ions and osmolality: A review. Cell Biol. Int. 2006;30:1–14. doi: 10.1016/j.cellbi.2005.06.004. PubMed DOI

Takai H., Morisawa M. Change in intracellular K+ concentration caused by external osmolality change regulates sperm motility of marine and freshwater teleosts. J. Cell Sci. 1995;108:1175–1181. doi: 10.1242/jcs.108.3.1175. PubMed DOI

Yang H., Carmichael C., Varga Z.M., Tiersch T.R. Development of a simplified and standardized protocol with potential for high-throughput for sperm cryopreservation in zebrafish Danio rerio. Theriogenology. 2007;68:128–136. doi: 10.1016/j.theriogenology.2007.02.015. PubMed DOI PMC

Jing R., Huang C., Bai C., Tanguay R., Dong Q. Optimization of activation, collection, dilution, and storage methods for zebrafish sperm. Aquaculture. 2009;290:165–171. doi: 10.1016/j.aquaculture.2009.02.027. DOI

Poupard G.P., Paxion C., Cosson J., Jeulin C., Fierville F., Billard R. Initiation of carp spermatozoa motility and early ATP reduction after milt contamination by urine. Aquaculture. 1998;160:317–328. doi: 10.1016/S0044-8486(97)00301-3. DOI

Rodina M., Cosson J., Gela D., Linhart O. Kurokura solution as immobilizing medium for spermatozoa of tench (Tinca tinca L.) Aquac. Int. 2004;12:119–131. doi: 10.1023/B:AQUI.0000017192.75993.e3. DOI

Linhart O., Benešovský J. Artificial insemination in asp (Aspius aspius L.) Ziv. Viry. 1991;36:973–980.

Tiersch T.R. Cryopreservation in aquarium fishes. Mar. Biotechnol. 2001;3:S212–S223. doi: 10.1007/s10126001-0044-z. PubMed DOI

Matthews J.L., Murphy J.M., Carmichael C., Yang H.P., Tiersch T., Westerfield M., Varga Z.M. Changes to extender, cryoprotective medium, and in vitro fertilization improve zebrafish sperm cryopreservation. Zebrafish. 2018;15:279–290. doi: 10.1089/zeb.2017.1521. PubMed DOI PMC

Horváth L., Tamás G., Seagrave C. Carp and Pond Fish Culture Including Chinese Herbivorous Species, Pike, Tench, Zander, Wels Catfish and Goldfish. Fishing News Books Ltd.; Oxford, UK: 1992. p. 158.

Linhart O., Gela D., Rodina M., Kocour M. Optimization of artificial propagation in European catfish, Silurus glanis L. Aquaculture. 2004;235:619–632. doi: 10.1016/j.aquaculture.2003.11.031. DOI

Linhart O., Cheng Y., Rodina M., Gela D., Tuckova V., Shelton W.L., Tinkir M., Memis D., Xin M. Sperm management of European catfish (Silurus glanis L.) for effective reproduction and genetic conservation. Aquaculture. 2020;529:735620. doi: 10.1016/j.aquaculture.2020.735620. DOI

Nusslein-Volhard C., Dahm R. Zebrafish: A Practical Approach. 7th ed. Oxford University Press; Oxford, UK: 2002. pp. 7–37.

Westerfield M. A Guide for the Laboratory Use of Zebrafish (Danio rerio) 4th ed. University of Oregon Press; Eugene, OR, USA: 2000. The Zebrafish Book.

Sakai N., Burgess S., Hopkins N. Delayed in vitro fertilization of zebrafish eggs in Hank’s saline containing bovine serum albumin. Mol. Mar. Biol. Biotechnol. 1997;6:84–87. PubMed

Cardona-Costa J., Perez-Camps M., Garcia-Ximenez F., Espinos F.J. Effect of gametes aging on their activation and fertilizability in zebrafish (Danio rerio) Zebrafish. 2009;6:93–95. doi: 10.1089/zeb.2008.0578. PubMed DOI

Yang H., Daly J., Carmichael C., Matthews J., Varga Z.M., Tiersch T. A procedure-spanning analysis of plasma membrane integrity for assessment of cell viability in sperm cryopreservation of zebrafish Danio rerio. Zebrafish. 2016;13:144–151. doi: 10.1089/zeb.2015.1176. PubMed DOI PMC

Cosson J., Linhart O., Mims S.D., Shelton W.L., Rodina M. Analysis of motility parameters from paddlefish and shovelnose sturgeon spermatozoa. J. Fish Biol. 2000;56:1348–1367. doi: 10.1111/j.1095-8649.2000.tb02148.x. DOI

Bondarenko V., Prokopchuk G., Cosson J. Fish sperm flagella: Original features and biological implications through the lens of modern technologies. In: Uzbekov R., editor. Flagella and Cilia: Types, Structure and Functions. Nova Publisher; New York, NY, USA: 2018. pp. 49–82.

Bondarenko V., Cosson J. Structure and beating behavior of the sperm motility apparatus in aquatic animals. Theriogenology. 2019;135:152–163. doi: 10.1016/j.theriogenology.2019.06.005. PubMed DOI

R Core Team . R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing; Vienna, Austria: 2018.

Perchec P.G., Cosson J., Billard F.A. La motilite des spermatozoides de truite (Oncorhynchus mykiss) et de carpe (Cyprinus carpio) J. Appl. Ichthyol. 1993;9:129–149. doi: 10.1111/j.1439-0426.1993.tb00389.x. DOI

Rahi D., Dzyuba B., Xin M.M., Cheng Y., Dzyuba V. Energy pathways associated with sustained spermatozoon motility in the endangered Siberian sturgeon Acipenser baerii. J. Fish Biol. 2020;97:435–443. doi: 10.1111/jfb.14382. PubMed DOI

Wilson-Leedy J.G., Kanuga M.K., Ingermann R.L. Influence of osmolality and ions on the activation and characteristics of zebrafish sperm motility. Theriogenology. 2009;71:1054–1062. doi: 10.1016/j.theriogenology.2008.11.006. PubMed DOI

Wasden M.B., Roberts R.L., DeLaurier A. Optimizing sperm collection procedures in zebrafish. J. South Carolina Acad. Sci. 2017;15:7. PubMed PMC

Linhart O., Kouřil J., Hamáčková J. Increased rate of egg fertilization in artificial propagation of sheatfish (Silurus glanis L.) by means of suppressing the movements of spermatozoa with immobilization solution. J. Appl. Ichthyol. 1987;65:353–358. doi: 10.1016/0044-8486(87)90247-X. DOI

Linhart O., Rodina M., Bastl J., Cosson J. Urinary bladder, ionic composition of seminal fluid and urine with characterization of sperm motility in tench (Tinca tinca L.) J. Appl. Ichthyol. 2003;19:177–181. doi: 10.1046/j.1439-0426.2003.00470.x. DOI

Cejko B.I., Żarski D., Targońska K., Krejszeff S., Kucharczyk D., Glogowski J. Osmolality of seminal plasma as an indicator of milt contamination with urine based on the example of the tench Tinca tinca (L.) Pol. J. Nat. Sci. 2010;25:287–298. doi: 10.2478/v10020-010-0026-6. DOI

Linhart O., Walford J., Sivaloganathan B., Lam T.J. Effects of osmolality and ions on the motility of stripped and testicular sperm of freshwater- and seawater-acclimated tilapia, Oreochromis mossambicus. J. Fish Biol. 1999;55:1344–1358.

Gonzalez-Lopez W.A., Ramos-Judez S., Gimenez I., Duncan N.J. Sperm contamination by urine in Senegalese sole (Solea senegalensis) and the use of extender solutions for short-term chilled storage. Aquaculture. 2020;516:734649. doi: 10.1016/j.aquaculture.2019.734649. DOI

Dreanno C., Suquet M., Desbruyeres E., Cosson J., Le Delliou H., Billard R. Effect of urine on semen quality in turbot (Psetta maxima) Aquaculture. 1998;169:247–262. doi: 10.1016/S0044-8486(98)00262-2. DOI

Cosson J., Groison A.L., Suquet M., Fauvel C., Dreanno C., Billard R. Studying sperm motility in marine fish: An overview on the state of the art. J. Appl. Ichthyol. 2008;24:460–486. doi: 10.1111/j.1439-0426.2008.01151.x. DOI

Alavi S.M.H., Linhart O., Coward K., Rodina M. Fish Spermatology: Implications for Aquaculture Management. In: Alavi S.M.H., Cosson J., Coward K., Rafiee G., editors. Fish Spermatology. Alpha Science Ltd.; Oxford, UK: 2008. pp. 397–461.

Alavi S.M.H., Cosson J., Bondarenko O., Linhart O. Sperm motility in fishes: (III) diversity of regulatory signals from membrane to the axoneme. Theriogenology. 2019;136:143–165. doi: 10.1016/j.theriogenology.2019.06.038. PubMed DOI

Morisawa M. Initiation mechanism of sperm motility at spawning in teleosts. Zool. Sci. 1985;2:605–615.

Suquet M., Billard R., Cosson J., Dorange G., Chauvaud L., Mugnier C., Fauvel C. Sperm features in turbot (Scophthalmus maximus): A comparison with other freshwater and marine fish species. Aquat. Living Resour. 1994;7:283–294. doi: 10.1051/alr:1994031. DOI

Koldras M., Loir M., Maisse G., LeGac F. Study of the composition of seminal fluid and of sperm motility along the genital tract, during a spawning season, in the rainbow trout (Oncorhynchus mykiss) Aquat. Living Resour. 1996;9:337–345. doi: 10.1051/alr:1996036. DOI

Billard R., Linhart O., Fierville F., Cosson J. Motility of European catfish Silurus glanis spermatozoa in testes and in milt. Pol. Arch. Hydrobiol. 1997;44:115–122.

Perchec G., Jeulin C., Cosson J., Andre F., Billard R. Relationship between sperm ATP content and motility of carp spermatozoa. J. Cell Sci. 1995;108:747–753. doi: 10.1242/jcs.108.2.747. PubMed DOI

Boryshpolets S., Dzyuba B., Stejskal V., Linhart O. Dynamics of ATP and movement in Eurasian perch (Perca fluviatilis L.) sperm in conditions of decreasing osmolality. Theriogenology. 2009;72:851–859. doi: 10.1016/j.theriogenology.2009.06.005. PubMed DOI

Linhart O., Alavi S.M.H., Rodina M., Gela D., Cosson J. Comparison of sperm velocity, motility and fertilizing ability between firstly and secondly activated spermatozoa of common carp (Cyprinus carpio) J. Appl. Ichthyol. 2008;24:386–392. doi: 10.1111/j.1439-0426.2008.01138.x. DOI

Xin M., Cheng Y., Rodina M., Tučková V., Shelton W.L., Linhart O. Improving motility and fertilization capacity of low-quality sperm of sterlet Acipenser ruthenus during storage. Theriogenology. 2020;156:90–96. doi: 10.1016/j.theriogenology.2020.07.004. PubMed DOI

Poupard G.P., Gatti J.L., Cosson J., Jeulin C., Fierville F., Billard R. Effects of extracellular environment on the osmotic signal transduction involved in activation of motility of carp spermatozoa. J. Reprod. Fertil. 1997;110:315–327. doi: 10.1530/jrf.0.1100315. PubMed DOI

Billard R., Cosson J., Perchec G., Linhart O. Biology of sperm and artificial reproduction in carp. Aquaculture. 1995;129:95–112. doi: 10.1016/0044-8486(94)00231-C. DOI

Aoki K., Okamoto M., Tatsumi K., Ishikawa Y. Cryopreservation of medaka spermatozoa. Zool. Sci. 1997;14:641–644. doi: 10.2108/zsj.14.641. DOI

Huang C., Dong Q., Tiersch T.R. Sperm cryopreservation of a live-bearing fish, the platyfish Xiphophorus couchianus. Theriogenology. 2004;62:971–989. doi: 10.1016/j.theriogenology.2003.12.022. PubMed DOI PMC

Huang C., Dong Q., Walter R.B., Tiersch T.R. Initial studies on sperm cryopreservation of a live-bearing fish, the green swordtail Xiphophorus helleri. Theriogenology. 2004;62:179–194. doi: 10.1016/j.theriogenology.2003.09.019. PubMed DOI PMC

Aurich J.E., Kuhne A., Hoppe H., Aurich C. Seminal plasma affects membrane integrity and motility of equine spermatozoa after cryopreservation. Theriogenology. 1996;46:791–797. doi: 10.1016/S0093-691X(96)00237-3. PubMed DOI

Linhart O., Rodina M., Flajshans M., Gela D., Kocour M. Cryopreservation of European catfish Silurus glanis sperm: Sperm motility, viability, and hatching success of embryos. Cryobiology. 2005;51:250–261. doi: 10.1016/j.cryobiol.2005.07.005. PubMed DOI

Contreras P., Dumorné K., Ulloa-Rodríguez P., Merino O., Figueroa E., Farías J.G., Valdebenito I., Risopatrón J. Effects of short-term storage on sperm function in fish semen: A review. Rev. Aquacult. 2020;12:1373–1389. doi: 10.1111/raq.12387. DOI

Beirao J., Boulais M., Gallego V., O’Brien J.K., Peixoto S., Robeck T.R., Cabrita E. Sperm handling in aquatic animals for artificial reproduction. Theriogenology. 2019;133:161–178. doi: 10.1016/j.theriogenology.2019.05.004. PubMed DOI

Linhart O., Cheng Y., Xin M.M., Rodina M., Tuckova V., Shelton W.L., Kaspar V. Standardization of egg activation and fertilization in sterlet (Acipenser ruthenus) Aquac Rep. 2020;17:100381. doi: 10.1016/j.aqrep.2020.100381. PubMed DOI

Ginsburg A.S. In: Fertilization in Fishes and the Problem of Polyspermy. Dettlaff T.A., editor. Volume 71 U.S. Department of Commerce, National Technical Information Service; Springfield, IL, USA: 1972. Israel Program for Scientific Translations.

In vivo and in vitro aging of common carp Cyprinus carpio sperm after multiple hormonal application and stripping of males

A Review on Environmental Contaminants-Related Fertility Threat in Male Fishes: Effects and Possible Mechanisms of Action Learned from Wildlife and Laboratory Studies