Hostname: page-component-586b7cd67f-t7fkt Total loading time: 0 Render date: 2024-11-24T12:49:17.180Z Has data issue: false hasContentIssue false
  • English
  • Français

Cyclic AMP reversal of hypoxanthine-arrested preimplantation mouse embryos is EDTA-dependent

Published online by Cambridge University Press:  26 September 2008

Mary K. Dienhart  and
Stephen M. Downs
Mary K. Dienhart
Affiliation:
Biology Department, Marquette University, Milwaukee, Wisconsin, USA.
Stephen M. Downs*
Affiliation:
Biology Department, Marquette University, Milwaukee, Wisconsin, USA.
*
Stephen M. Downs, Biology Department Marquette University, P.O. Box 1881, Milwaukee, WI 53201-1881, USA. Telephone: +1(414) 288-1698. Fax: +1 (414) 288-7357.
Get access Rights & Permissions [Opens in a new window]

Summary

Hypoxanthine can block preimplantation mouse embryo development in vitro at the 2- to 4-cell stages, and this has recently been shown to be reversed by cAMP-elevating agents. However, the extent of this hypoxanthine-induced arrest is determined by the culture conditions and strain of mouse. Whitten's and KSOM/AA are two embryo culture media that support preimplantation development to the blastocyst stage. This study was undertaken to examine the influence of several components in these media on hypoxanthine-arrested preimplantation mouse embryos and to test the hypothesis that reversal of the hypoxanthine block by cAMP-elevating agents requires cooperative interaction with the chelator, EDTA. Initial experiments demonstrated that embryo development was blocked in the presence of hypoxanthine in Whitten's medium but not in KSOM/AA; furthermore, removal of EDTA from KSOM/AA rendered this medium incapable of supporting high levels of development to blastocyst (9%), whereas high numbers of blastocysts (80%) formed in Whitten's medium, which does not contain the chelator. Consequently, Whitten's medium was used to test our hypothesis. It has previously been demonstrated that the phosphodiesterase inhibitor, IBMX, can reverse the developmental arrest imposed by hypoxanthine in EDTA-supplemented Earle's basic salt solution, but in the present study the addition of IBMX to Whitten's medium resulted in a block to development and failed to reverse the hypoxanthine arrest. These disparate effects can be explained by the presence or absence of EDTA. Supplementing Whitten's medium with EDTA reverses the IBMX effect, but not the hypoxanthine-induced block. While IBMX alone is unable to reverse the hypoxanthine block in Whitten's medium, development is greatly enhanced by the simultaneous addition of EDTA and IBMX. Similar results were obtained with the cAMP analogue, 8-AHA-cAMP. The data therefore support our hypothesis that the reversal of the hypoxanthine-induced arrest by cAMP-elevating agents is critically dependent on the presence of EDTA. We contrast this with the situation in mouse oocytes, where the hypoxanthine-induced meiotic arrest is not reversed by the addition of EDTA and/or cAMP-elevating agents.

Keywords

Cyclic AMPEDTAEmbryoHypoxanthine
Type
Article
Information
Zygote , Volume 4 , Issue 2 , May 1996 , pp. 129 - 137
Copyright
Copyright © Cambridge University Press 1996

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Abramczuk, J., Solter, D. & Koprowski, H. (1977). The beneficial effects of EDTA on development of mouse one–cell embryos in chemically defined medium. Dev. Biol. 61, 378–83.CrossRefGoogle ScholarPubMed
Alexiou, M. & Leese, H.J. (1992). Purine utilisation, de novo synthesis and degradation in mouse preimplantation embryos. Development 114, 185–92.CrossRefGoogle ScholarPubMed
Alexiou, M. & Leese, H.J. (1994). Enzymes of purine salvage and catabolism in the mouse preimplantation embryo measured by high performance liquid chromatography. J. Reprod. Fertil. 101. 151–8.CrossRefGoogle ScholarPubMed
Anbari, K. & Schultz, R.M. (1993). Effect of sodium and betaine in culture media on development and relative rates of protein synthesis in preimplantation mouse embryos in vitro. Mol. Reprod. Dev. 35, 24–8.CrossRefGoogle ScholarPubMed
Bastias, M.C., McGee, Belser S.T., Bryan, S.H. & Vasquez, J.M. (1993). in vitro deleterious effect of hypoxanthine in Ham's Nutrient Mixture F-10 culture medium on human oocyte fertilization and early embryonic development. Fertil. Steril. 60, 876–80.CrossRefGoogle ScholarPubMed
Bavister, B.D. (1995). Culture of preimplantation embryos: facts arid artifacts. Hum. Reprod. Update. 1, 91–148.CrossRefGoogle Scholar
Biggers, J.D., Lawitts, J.A. & Lechene, C.P. (1993). The protective action of betaine on the deleterious effects of NaCl on preimplantation mouse embryos in vitro. Mol. Reprod. Dev. 34, 380–90.CrossRefGoogle ScholarPubMed
Brinster, R.L. (1965). Studies on the development of mouse embryos in vitro. IV. Interaction of energy sources. J. Reprod. Fertil. 10, 227–40.CrossRefGoogle Scholar
Brown, J.J.G. & Whittingham, D.G. (1991). The roles of pyruvate, lactate and glucose during preimplantation development of embryos from F1 hybrid mice in vitro. Development 112, 99–105.CrossRefGoogle ScholarPubMed
Chatot, C.L., Ziomek, C.A., Bavister, B.D., Lewis, J.L. & Torres, I. (1989). An improved culture medium supports development of random-bred 1-cell mouse embryos in vitro. J. Reprod. Fertil. 86, 679–88.CrossRefGoogle Scholar
Chatot, C.L., Lewis, J.L., Torres, I. & Ziomek, C.A. (1990). Development of 1-cell embryos from different strains of mice in CZB medium. Biol. Reprod. 42, 432–40.CrossRefGoogle Scholar
Chatot, C.L., Lewis, Williams J.L., Torres, I. & Ziomek, C.A. (1994). One-minute exposure of 4-cell mouse embryos to glucose overcomes the morula block in CZB medium. Mol. Reprod. Dev. 37, 407–12.CrossRefGoogle ScholarPubMed
Downs, S.M. (1995). The influence of glucose, cumulus cells, and metabolic coupling on ATP levels and meiotic control in the isolated mouse oocyte. Dev. Biol. 167, 502–12.CrossRefGoogle ScholarPubMed
Downs, S.M. & Dow, M.P.D. (1991). Hypoxanthine-maintamed two-cell block in mouse embryos: dependence on glucose and effect of hypoxanthine phosphoribosyltransferase inhibitors. Biol. Reprod. 44, 1025–39.CrossRefGoogle ScholarPubMed
Downs, S.M., Coleman, D.L., Ward, Bailey P.F. & Eppig, J.J. (1985). Hypoxanthine is the principal inhibitor of murine oocyte maturation in a low molecular weight fraction of porcine follicular fluid. Proc. Natl. Acad. Sci. USA 82, 454–8.CrossRefGoogle Scholar
Downs, S.M., Daniel, S.A.J., Bornslaeger, E.A., Hoppe, P.C. & Eppig, J.J. (1989). Maintenance of meiotic arrest in mouse oocytes by purines: modulation of cAMP levels and cAMP phosphodiesterase activity. Gamete Res. 23, 323–34.CrossRefGoogle ScholarPubMed
Du, Z.F. & Wales, R.G. (1993). Some effects of genotype and composition of the culture medium on the development of mouse zygotes in vitro. Reprod. Fertil Dev. 5, 405–15.CrossRefGoogle Scholar
Eppig, J.J., Ward-Bailey, P.F. & Coleman, D.L. (1985). Hypoxanthine and adenosine in murine ovarian follicular fluid: concentrations and activity in maintaining oocyte meiotic arrest. Biol. Reprod. 33, 1041–9.CrossRefGoogle ScholarPubMed
Erbach, G.T., Lawitts, J.A., Papaioannou, V.E. & Biggers, J.D. (1994). Differential growth of the mouse preimplantation embryo in chemically defined media. Biol. Reprod. 50, 1027–333.CrossRefGoogle ScholarPubMed
Fissore, R.A., Jackson, K.V. & Kiessling, A.A. (1989). Mouse zygote development in culture medium without protein in the presence of ethylenediaminetetraacetic acid. Biol. Reprod. 41, 835–41.CrossRefGoogle ScholarPubMed
Fissore, R.A., O'Keefe, S. & Kiessling, A.A. (1992). Purineinduced block to mouse embryo cleavage is reversed by compounds that elevate cyclic adenosine monophosphate. Biol. Reprod. 47. 1105–12.CrossRefGoogle ScholarPubMed
Ho, Y., Doherty, A.S. & Schultz, R.M. (1994). Mouse preimplantation embryo development in vitro: effect of sodium concentration in culture media on RNA synthesis and accumulation and gene expression. Mol. Reprod. Dev. 38, 131–41.CrossRefGoogle ScholarPubMed
Ho, Y., Wigglesworth, K., Eppig, J.J. & Schultz, R.M. (1995). Preimplantation development of mouse embryos in KSOM: augmentation by amino acids and analysis of gene expression. Mol. Reprod. Dev. 41, 232–8.CrossRefGoogle ScholarPubMed
Hoppe, P.C. (1985). Technique of fertilization in vitro. In Reproductive Toxicology, ed. Dixon, R.L.. pp. 191–9. New York: Raven Press.Google Scholar
Hoshi, M. & Toyoda, Y. (1985). Effect of EDTA on the preimplantation development of mouse embryos in vitro. Jpn. J. Zootech Soc. 56, 931–7.Google Scholar
Lawitts, J.A. & Biggers, J.D. (1991 a). Optimization of mouse embryo culture media using simplex methods. J. Reprod. Fertil. 91, 543–56.CrossRefGoogle Scholar
Lawitts, J.A. & Biggers, J.D. (1991 b). Overcoming the 2-cell block by modifying standard components in a mouse embryo culture medium. Biol. Reprod. 45, 245–51.CrossRefGoogle Scholar
Lawitts, J.A. & Biggers, J.D. (1992). Joint effects of sodium chloride, glutamine, and glucose in mouse preimplantation embryo culture media. Mol. Reprod. Dev. 31, 189–94.CrossRefGoogle Scholar
Loutradis, D., John, D. & Kiessling, A.A. (1987). Hypoxanthine causes a 2-cell block in random-bred mouse embryos. Biol. Reprod. 37, 311–16.CrossRefGoogle ScholarPubMed
Loutradis, D.C., Kiessling, A.A., Kallianidis, K., Siskos, K., Creatsas, G., Michalas, S. & Aravantinos, D. (1993). A preliminary trial of human zygote culture in Ham's F-10 without hypoxanthine. J. Assist Reprod. Genet. 10, 271–5.CrossRefGoogle ScholarPubMed
Martin, K.L. & Leese, H.J. (1995). Role of glucose in mouse preimplantation embryo development. Mol. Reprod. Dev. 40, 436–43.CrossRefGoogle ScholarPubMed
Nasr-Esfahani, M.M. & Johnson, M.H. (1991). The origin of reactive oxygen species in mouse embryos cultured in vitro. Development 113, 551–60.CrossRefGoogle Scholar
Nasr-Esfahani, M., Johnson, M.H. & Aitken, R.J. (1990). The effect of iron and iron chelators on the in-vitro block to development of the mouse preimplantation embryo: BAT6 a new medium for improved culture of mouse embryos in vitro. Hum. Reprod. 5, 997–1003.CrossRefGoogle ScholarPubMed
Nasr, Esfahani M.H., Winston, N.J. & Johnson, M.H. (1992). Effects of glucose, glutamine, ethylenediaminetetraacetic acid and oxygen tension on the concentration of reactive oxygen species and on development of the mouse preimplantation embryo in vitro. J. Reprod. Fertil. 96, 219–31.CrossRefGoogle Scholar
Nureddin, A., Epsaro, E. & Kiessling, A.A. (1990). Purines inhibit the development of mouse embryos in vitro. J. Reprod. Fertil. 90, 455–64.CrossRefGoogle ScholarPubMed
Poueymirou, W.T. & Schultz, R.M. (1987). Differential effects of activators of cAMP-dependent protein kinase and protein kinase C on cleavage of one-cell mouse embryos and protein synthesis and phosphorylation in one- and two-cell embryos. Dev. Biol. 121, 489–98.CrossRefGoogle ScholarPubMed
Rieger, D. (1992). Relationships between energy metabolism and development of early mammalian embryos. Theriogenology 37, 75–93.CrossRefGoogle Scholar
Toyoda, Y., Azuma, S. & Takeda, S. (1989). Effects of chelating agents on preimplantation development of mouse embryos in vitro. In Development of Preimplantation Embryos and Their Environment, ed. Yoshinaga, K & Mori, I. pp. 171–9. New York: Alan R Liss.Google Scholar
Whitten, W.K. (1957). Culture of tubal ova. Nature 179, 1081–2.CrossRefGoogle ScholarPubMed