Relationship between development, metabolism, and mitochondrial organization in 2-cell hamster embryos in the presence of low levels of phosphate.
The effect of low concentrations of inorganic phosphate (P(i)) on development, metabolic activity, and mitochondrial organization in the same cohorts of cultured hamster embryos was evaluated. Two-cell embryos were collected from eCG-stimulated golden hamsters and cultured in HECM-10 with 0.0 (control), 1.25, 2.5, or 5.0 microM KH(2)PO(4). Glucose utilization through the Embden-Meyerhof pathway (EMP) and tricarboxylic acid (TCA)-cycle activity were determined following 5 h of culture. Mitochondrial organization in living embryos was evaluated using multiphoton microscopy at 6 h of culture. Development was assessed at 27 h (on-time 8-cell stage) and 51 h (on-time blastocyst stage) of culture. Total cell numbers, as well as cell allocation to the trophectoderm and inner cell mass were determined for morula- and blastocyst-stage embryos. Culture with P(i) did not alter TCA-cycle activity. However, culture with > or 2.5 microM P(i) significantly increased (P < 0.01) EMP activity compared to control. Mitochondrial organization was significantly (P < 0.01) disrupted by P(i) in a dose-dependent manner. Development to the 8-cell, morula/blastocyst, and blastocyst stages was significantly reduced (P < 0.05) in the presence of > or =2.5 microM P(i) compared to both control and 1.25 microM P(i). This study clearly demonstrates that, for hamster embryos, inclusion of even exceptionally low concentrations of P(i) in culture medium dramatically alters embryo physiology. Additionally, although 2-cell embryos can tolerate some structural disruption without concomitant, detrimental effects on development or metabolic activity, metabolic disturbance is associated with decreased developmental competence.
Ludwig
T E
TE
Department of Animal Health and Biomedical Sciences, Laboratory of Molecular Biology, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA. ludwig@ahabs.wisc.edu
Squirrell
J M
JM
Palmenberg
A C
AC
Bavister
B D
BD
D000818:Animals; D001755:Blastocyst / Q000187:drug effects / Q000502:physiology; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D002952:Citric Acid Cycle; D006224:Cricetinae; D003470:Culture Media; D046508:Culture Techniques; D004622:Embryo, Mammalian / Q000187:drug effects* / Q000378:metabolism* / Q000648:ultrastructure; D005314:Embryonic and Fetal Development / Q000187:drug effects*; D005947:Glucose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008928:Mitochondria / Q000187:drug effects* / Q000648:ultrastructure; D009028:Morula / Q000187:drug effects / Q000502:physiology; D009994:Osmolar Concentration; D010710:Phosphates / Q000008:administration & dosage* / Q000494:pharmacology
D006063:Chorionic Gonadotropin; D003470:Culture Media; D010710:Phosphates; D005947:Glucose