Influence of solutions of different osmolarity on morphometric parameters of 15-32-cells bovine embryos

DOI: 10.32900/2312-8402-2022-128-104-118

Lisin Vadim,
Senior Researcher,,
Khmelkov Viacheslav,
Senior Researcher,,
Bugrov O. D,
Doctor of Biological Sci., Professor,
Dibirov M. K.,
Doctor of Biological Sci., Senior Researcher,
Institute of animal science of NAAS of Ukraine

Keywords: bovine; embryo morphometric parameters; pre-compacted 15-17-cells embryos; early morulae; intact, denuded and deblastomerised states; isotonic, hypertonic and hypotonic conditions


The illustrative material was obtained that allows to observe clearly the effect of different tonicity solutions on the morphology of bovine 5-days pre-compacted 15-17-cells embryos and 6-days early morulae obtained in vivo, depending on their technological state.
The average values and rates of variation of the following morphometric parameters were determined in embryos at the stages of the developmental study: outer and inner zona pellucida (ZP) diameters, ZP thickness, embryonic cell mass (CM) diameter and individual embryonic cell diameters. The mentioned parameters were determined in each of the embryos consistently in intact, denuded and deblastomerised states in isotonic (0.29 Osm), hypertonic (1.5 Osm) and hypotonic (0.2 Osm) conditions.
Manipulations with embryos were carried out in such a way that during consistent transfer through solutions of different osmolarity, it was possible to identify the dynamics of changes in the morphology and parameters of each of the embryos (in the intact state and after denudation) and of each of the individual embryonic cells (after deblastomerisation of the embryo).
A decrease or increase in the size of the CM of embryos of the studied developmental stages during their successive transfer through solutions with different osmolarity occurred with the preservation of the approximate proportionality of their shape. After partial dehydration in hypertonia, the denuded embryos recovered their sizes mostly not fully relative to their initial sizes in the intact state in solutions of the appropriate osmolarity. Similar osmotic behavior was also observed in individual cells of deblastomerised embryos.
The experimental data presented in the article make it possible to predict the behavior of embryos during micromanipulations, to determine the sequence, technique and conditions of operations with them, as well as the types, shape and sizes of microtools necessary for this.


  1. Kauffold, P., Tamm, I., Shihov, I. Ya., Erig, F., Falge, R., & Rommel, P. (1990). Ocenka kachestva embrionov krupnogo rogatogo skota: Rukovodstvo po peresadke embrionov [Assessing the Quality of Bovine Embryos: A Guide to Embryo Transfer]. Moskva : Agropromizdat, 56 [in Russian]
  2. Yang, X., Chen, Y. Q., Chen, J. B., & Foote, R. H. (1990). Potential of hypertonic medium treatment for embryo micromanipulation: I. Survival of rabbit embryos in vitro and in vivo following sucrose treatment. Mol. Reprod. Dev. 27. 110-117.
  3. Yang, X., Chen, J. B., Chen, Y. Q., & Foote, R. H. (1990). Improved developmental potential of rabbit oocytes fertilized by sperm microinjection into perivitelline space enlarged by hypertonic media. J.Exp.Zool. 255(1). 114-119.
  4. Yang, X., Zhang, L., Kovaks, A., Tobback, C., & Foote, R. H. (1990). Potential of hypertonic medium treatment for embryo micromanipulation: II. Assessment of nuclear transplantation methodology, isolation, subzona insertion, and electrofusion of blastomeres to intact or functionally enucleated oocytes in rabbits. Mol. Reprod. Dev., 27. 118-129.
  5. Liu, J. L., Sung, L. Y., Barber, M., & Yang, X. (2002). Hypertonic Medium Treatment for Localization of Nuclear Material in Bovine Metaphase II Oocytes. Biol. of Reprod. 66(5). 1342-1349.
  6. van Os, H. C., & Zeilmaker, G. H. (1986). Volumetric behavior and survival of mouse zygotes and embryos in hyperosmotic media. Hum. Reprod. 1(2). 95-98.
  7. A.s.1727822 1А SSSR, MKI A61D 19/04, C12Q 1/24. Sposob provedenia mikrohirurgicheskih operaciy s yaycekletkami i embrionami [Method for performing microsurgical operations with oocytes and embryos] Ostashko F. I., Bezugly N. D., Gordienko N. A. (SSSR). №4832540/15; Zayavl. 12.04.1992; Opubl. 23.04.1992; Bul.№ 15. 3 s. [in Russian]
  8. Gordienko, N. A., Bezugly, N. D., & Bondarenko ,A. I. (1992). Ispolzovanie neizotonicheskih rastvorov dla provedenia mikrohirurgicheskih operaciy s yaycekletkami i embrionami mlekopitayuschih [The use of non-isotonic solutions for microsurgical operations with mammallian oocytes and embryos]. Tezisy dokladov Rossiysko-Germanskogo simpoziuma “Uspehi Biotehnologii v razmnozhenii selskohoziaystvennyh zhivotnyh” [in Russian]
  9. Pat.17876A Ukraina, MKV6 A61D 19/04 Sposib otrymannia hymernyh embrioniv [The method of obtaining chimeric embryos] Bezugly M.D., Gordienko N.O., Lisin V.I. (Ukraina). № 96083085; Zayavl. 01.08.1996; Opubl. 03.06.1997. Bul. №5. 4 р. [in Ukrainian]
  10. Lisin, V. I. (2006). Tehnika mikrohirurgicheskoy denudacii yaycekletok i embrionov mlekopitayuschih [The technique of microsurgical denudation of mammalian oocytes and embryos]. Naukovo-tekhnichnyi biuleten Instytutu tvarynnytstva NAAN – Scientific and Technical Bulletin of the Institute of Animal Science of the National Academy of Agrarian Science of Ukraine. Kharkiv, № 94. P. 207-213. [in Russian]
  11. Baranovskiy, D. I., Lubeckiy, M. D., Prudnikov, V. G., & Tischenko, S. I. (1988). Metodicheskiye rekomendacii dla samostoyatelnogo izuchenia osnov biometrii [Guidelines for self-study of the basics of biometrics]. Harkov, HZVI, 87 s. [in Russian]
  12. Lakin, G. F. (1990). Biometria: Ucheb. posobiye dla biol. spec. VUZov [Biometrics: Textbook for Biological Specialties of Universities]. Moskva : Vysshaya shkola, 352 [in Russian].