Effects of Actinomycin D and Cycloheximide on RNA and Protein Synthesis in Cleavage Stage Mouse Embryos

Tasca, Richard J.; Hillman, Nina

doi:10.1038/2251022a0

Cited by 90 publications

(20 citation statements)

References 16 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Exogenous amino acid consumption and incorporation by preimplantation murine and bovine embryos increases throughout development, particularly after the 8-cell and compaction stages (Tasca & Hillman 1970, Brinster 1971, Epstein & Smith 1973, Kaye et al 1982. Although beneficial effects on preimplantation development may be obtained through the addition of amino acids in vitro, their spontaneous breakdown above that resulting from metabolic deamination can compromise development through increases in ammonium concentration (Gardner & Lane 1993, Katchadourian et al 1996.…”

Section: Introductionmentioning

confidence: 99%

Ammonium exposure and pyruvate affect the amino acid metabolism of bovine blastocysts in vitro

Orsi¹,

Leese²

2004

Reproduction

View full text Add to dashboard Cite

The accumulation of ammonium is a major artefact of in vitro embryo culture. This study has examined ammonium production and potential mechanisms of disposal in preimplantation bovine blastocysts. Embryos were produced by in vitro maturation and fertilisation of oocytes, and cultured in synthetic oviduct fluid containing amino acids and BSA (SOFaaBSA). Ammonium/urea concentrations were determined enzymatically. Amino acid appearance/disappearance 'profiles' of single blastocysts were determined at 0, 1.25 and 2.5 mM NH 4 Cl (with or without 0.33 mM pyruvate), and with or without 10 mM dipicolinic acid (DPCA; a glutamate dehydrogenase (GLDH) inhibitor) or 2 mM amino-oxyacetate (AOA; a transaminase inhibitor). Free ammonium was produced at a rate of 4.281 (6 0.362) pmol/embryo/h, while urea production was undetectable. The presence/absence of pyruvate affected amino acid profiles, especially alanine appearance (P < 0.001), glutamate disappearance (P < 0.05) and overall turnover (the sum of appearance and disappearance) (P < 0.001). GLDH inhibition with DPCA had no effect on amino acid overall disappearance, but glutamate disappearance increased, while that of arginine decreased (P < 0.05). The transaminase inhibitor, AOA, depressed turnover (P < 0.05), aspartate and glutamate disappearance, and alanine appearance. Thus, bovine blastocysts release ammonium as free ions or fix them, not as urea, but as alanine, possibly glutamine and, less likely, arginine. An active role for GLDH and transaminases in regulating blastocyst amino acid metabolism was demonstrated.

show abstract

Section: Introductionmentioning

confidence: 99%

Ammonium exposure and pyruvate affect the amino acid metabolism of bovine blastocysts in vitro

Orsi¹,

Leese²

2004

Reproduction

View full text Add to dashboard Cite

show abstract

“…A number of experiments with the inhibitor of RNA transcription, actinomycin D, have demonstrated that early cleavage shows partial resistance to the drug, and that protein synthetic levels are suppressed by only 50% (Monesi & Molinaro, 1970;Tasca & Hillman, 1970;Golbus, Calarco & Epstein, 1973). These observations suggest that in the mammal, unlike amphibians and sea-urchins, some but not all of the protein synthesis of early embryos is dependent on new mRNA.…”

Section: The Maternal Environmentmentioning

confidence: 86%

Intrinsic and extrinsic factors in preimplantation development

Johnson¹

1979

Reproduction

View full text Add to dashboard Cite

“…The primary effect of actinomycin D in mouse embryos is inhibition of RNA synthesis (Monesi, Molinaro, Spalleta & Davoli, 1970;Tasca & Hillman, 1970;Golbus, Calarco & Epstein, 1973), especially ribosomal RNA synthesis (Pikó, 1970), which in turn impairs protein synthesis. For example, a dose of 10" * µg/ml almost completely inhibits RNA synthesis and reduces protein synthesis by 20% (Monesi et al, 1970 (Pedersen & Spindle, 1976 …”

mentioning

confidence: 99%

Differential inhibition of trophoblast outgrowth and inner cell mass growth by actinomycin D in cultured mouse embryos

Glass¹,

Spindle²,

Pedersen³

1976

Reproduction

View full text Add to dashboard Cite

Summary. Treatment of preimplantation mouse embryos in vitro with 10\ p=n-\ 3 to 10\ p=n-\ 1\ g=m\ g actinomycin D/ml for 2 hr showed that (i) (BSS) and were cultured in modified Eagle's basal medium (BME) (Goldstein et al, 1975).A stock solution of actinomycin D (Calbiochem, La Jolla, California) was made with distilled water at a concentration of 10 µg/ml, kept at 4°C in the dark, and diluted to appropriate concentrations in modified BME before use. Embryos were incubated for 2 hr in medium containing actinomycin D, washed 5 times in modified Hanks' BSS, and transferred to 8-chamber culture slides (Lab Tek, Naperville, Illinois) containing modified BME. They were allowed to grow for 6 days (morulae), 5 days (early blastocysts) or 4 days (late blastocysts). The response of the embryos to actinomycin D treatment was evaluated at the end of the culture period by counting the number of embryos that developed to the following postimplantation endpoints: (1) trophoblast outgrowth, (2) growth of a well defined inner cell mass on the trophoblast outgrowth, and (3) development of this inner cell mass into 2 germ layers (endoderm and ectoderm) recognizable under the dissecting microscope.Each treatment was repeated 4-12 times and the combined results were analysed with the 2 test for significant differences (1 d.f., <0-05 or <0-01).

show abstract

Effects of Actinomycin D and Cycloheximide on RNA and Protein Synthesis in Cleavage Stage Mouse Embryos

Cited by 90 publications

References 16 publications

Ammonium exposure and pyruvate affect the amino acid metabolism of bovine blastocysts in vitro

Ammonium exposure and pyruvate affect the amino acid metabolism of bovine blastocysts in vitro

Intrinsic and extrinsic factors in preimplantation development

Differential inhibition of trophoblast outgrowth and inner cell mass growth by actinomycin D in cultured mouse embryos

Contact Info

Product

Resources

About