Toward a feline-optimized culture medium: impact of ions, carbohydrates, essential amino acids, vitamins, and serum on development and metabolism of in vitro fertilization-derived feline embryos relative to embryos grown in vivo.
The objective of this study was to define the physiologic needs of domestic cat embryos to facilitate development of a feline-specific culture medium. In a series of factorial experiments, in vivo-matured oocytes (n = 2040) from gonadotropin-treated domestic cats were inseminated in vitro to generate embryos (n = 1464) for culture. In the initial study, concentrations of NaCl (100.0 vs. 120.0 mM), KCl (4.0 vs. 8.0 mM), KH(2)PO(4) (0.25 vs. 1.0 mM), and the ratio of CaCl(2) to MgSO(4)-7H(2)O (1.0:2.0 mM vs. 2.0:1.0 mM) in the medium were evaluated during Days 1-6 (Day 0: oocyte recovery and in vitro fertilization [IVF]) of culture. Subsequent experiments assessed the effects of varying concentrations of carbohydrate (glucose, 1.5, 3.0, or 6.0 mM; l-lactate, 3.0, 6.0, or 12.0 mM; and pyruvate, 0.1 or 1.0 mM) and essential amino acids (EAAs; 0, 0.5, or 1.0x) in the medium during Days 1-3 and Days 3-6 of culture. Inclusion of vitamins (0 vs. 1.0x) and fetal calf serum (FCS; 0 vs. 5% [v/v]) in the medium also was evaluated during Days 3-6. Development and metabolism of IVF embryos on Day 3 or Day 6 were compared to age-matched in vivo embryos recovered from naturally mated queens. A feline-optimized culture medium (FOCM) was formulated based on these results (100.0 mM NaCl, 8.0 mM KCl, 1.0 mM KH(2)PO(4), 2.0 mM CaCl(2), 1.0 mM MgSO(4), 1.5 mM glucose, 6.0 mM L-lactate, 0.1 mM pyruvate, and 0x EAAs with 25.0 mM NaHCO(3), 1.0 mM alanyl-glutamine, 0.1 mM taurine, and 1.0x nonessential amino acids) with 0.4% (w/v) BSA from Days 0-3 and 5% FCS from Days 3-6. Using this medium, ~70% of cleaved embryos developed into blastocysts with profiles of carbohydrate metabolism similar to in vivo embryos. Our results suggest that feline embryos have stage-specific responses to carbohydrates and are sensitive to EAAs but are still reliant on one or more unidentified components of FCS for optimal blastocyst development.
Herrick
Jason R
JR
Center for Conservation and Research of Endangered Wildlife, Cincinnati Zoo and Botanical Garden, Cincinnati, Ohio 45220, USA. jason.herrick@cincinnatizoo.org
Bond
Jennifer B
JB
Magarey
Genevieve M
GM
Bateman
Helen L
HL
Krisher
Rebecca L
RL
Dunford
Susan A
SA
Swanson
William F
WF
D000601:Amino Acids, Essential / Q000494:pharmacology*; D000818:Animals; D001755:Blastocyst / Q000187:drug effects; D002241:Carbohydrates / Q000494:pharmacology*; D002415:Cats / Q000502:physiology*; D002452:Cell Count; D003470:Culture Media / Q000737:chemistry*; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D047108:Embryonic Development / Q000187:drug effects; D005260:Female; D005307:Fertilization in Vitro; D007477:Ions / Q000737:chemistry; D008297:Male; D011247:Pregnancy; D012661:Semen / Q000502:physiology; D044967:Serum; D014815:Vitamins / Q000494:pharmacology*
D000601:Amino Acids, Essential; D002241:Carbohydrates; D003470:Culture Media; D007477:Ions; D014815:Vitamins