Quantitative microinjection of trehalose into mouse oocytes and zygotes, and its effect on development

Ali Eroglu, Joel A. Lawitts, Mehmet Toner, Thomas L. Toth

Research output: Contribution to journalArticle

53 Citations (Scopus)

Abstract

Sugars such as trehalose are effectively used by various organisms as protective agents to undergo anhydrobiosis and cryobiosis. The objective of this study was first to establish a method for quantitative delivery of trehalose as a model sugar into oocytes, and then to evaluate its effect on development of mouse zygotes. To this end, a quantitative microinjection technique was developed using volumetric response of microdroplets suspended in dimethylpolysilaxene. To verify accuracy of this technique, both microdroplets and oocytes were microinjected with fluorophore-labeled dextran. Thereafter, injection volumes were calculated from fluorescence intensity, and volumetric responses of both microdroplets and oocytes. Comparison of calculated injection volumes revealed that this technique reflects microinjection into oocytes with pL-accuracy. The next series of experiments focused on toxicity of injection buffers (i.e., 10mM Tris and 15mM Hepes) and trehalose. Microinjection of Hepes and Tris buffer in the presence of 0.1M trehalose resulted in blastocyst rates of 86 and 72%, respectively, without a significant difference when compared to controls (86%). In subsequent experiments, Hepes was used as the injection buffer, and embryonic development of zygotes was studied as a function of intracellular trehalose concentrations. Microinjection of trehalose up to 0.15M resulted in development to blastocyst stage similar to controls (85 and 87%, respectively) while the blastocyst rate was significantly decreased (43%) in the presence of 0.20M intracellular trehalose. When transferred to foster mothers, trehalose-injected zygotes (0.1M) implanted and developed to day 16 fetuses similar to controls, healthy pups were born. The findings of this study suggest that trehalose at effective intracellular concentrations does not impair development of mouse zygotes.

Original languageEnglish (US)
Pages (from-to)121-134
Number of pages14
JournalCryobiology
Volume46
Issue number2
DOIs
StatePublished - Jan 1 2003
Externally publishedYes

Fingerprint

Trehalose
Zygote
Microinjections
zygote
trehalose
Oocytes
oocytes
mice
Blastocyst
blastocyst
injection
Injections
buffers
Sugars
quantitative analysis
Buffers
sugars
Protective Agents
Tromethamine
Fluorophores

Keywords

  • Development
  • Egg
  • Hypertonic medium
  • Mouse oocyte
  • Quantitative microinjection
  • Toxicity
  • Trehalose

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Quantitative microinjection of trehalose into mouse oocytes and zygotes, and its effect on development. / Eroglu, Ali; Lawitts, Joel A.; Toner, Mehmet; Toth, Thomas L.

In: Cryobiology, Vol. 46, No. 2, 01.01.2003, p. 121-134.

Research output: Contribution to journalArticle

Eroglu, Ali ; Lawitts, Joel A. ; Toner, Mehmet ; Toth, Thomas L. / Quantitative microinjection of trehalose into mouse oocytes and zygotes, and its effect on development. In: Cryobiology. 2003 ; Vol. 46, No. 2. pp. 121-134.
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