Lecture 4 Flashcards
How do high temperatures impact maturing oocytes and embryos differently?
Maturing Oocytes: Highly sensitive to high temperatures due to their limited ability to produce protective proteins
Embryos: More tolerant of high temperatures as they can produce heat shock proteins for protection
What are the potential consequences of exposing an oocyte to temperatures below 37°C?
Disruption of microtubules, potentially irreversible
Impaired enzyme function
Difficulty maintaining homeostasis
Reduced embryo viability
What are the main challenges associated with maintaining optimal osmolality in embryo culture media?
Evaporation: Microdrops are highly susceptible to evaporation, especially during preparation and on heated microscope stages. Oil covers slow down but don’t completely prevent evaporation
Time-lapse incubators: The use of shallow dishes, small drop sizes, thin oil layers, and lack of humidification in many time-lapse incubators further increase the risk of evaporation and osmolality changes
Why is a low oxygen environment (5% O2) considered superior to a high oxygen environment (21% O2) for embryo culture?
Reduces oxidative damage by minimizing the production of harmful reactive oxygen species (ROS)
Supports optimal mitochondrial function, ensuring efficient energy production
Promotes the use of the more efficient TCA cycle for energy production in early cleavage-stage embryos
Reduces the incidence of aneuploidy and mosaicism in mouse embryos
Explain the role of bicarbonate and HEPES/MOPS buffers in embryo culture media.
Bicarbonate (NaHCO3): The primary buffer system used in media for incubator culture. It interacts with CO2 to regulate pH, maintaining a physiological range of 7.2-7.4
HEPES/MOPS: Synthetic buffers used in media designed for manipulations outside the incubator. They maintain a stable pH in atmospheric CO2 conditions, preventing harmful pH fluctuations during procedures
How does the composition of culture media change to support the different metabolic needs of embryos at various stages?
Fertilization Media: High glucose to support energy-intensive fertilization and early cell division
Cleavage Media: Low glucose but high lactate and pyruvate to cater to the embryo’s preference for the TCA cycle during this stage
Blastocyst Media: High glucose and low lactate/pyruvate to align with the embryo’s increased reliance on glycolysis
What are the two main approaches to providing stage-specific culture media?
Sequential Media: Involves changing the media at different stages (fertilization, cleavage, blastocyst) to provide the optimal nutrient composition for each stage
Single-Step Media: Uses one media formulation for the entire culture period, offering a “buffet” of nutrients for the embryo to choose from
Name the three primary components of embryo culture media and their key functions.
Salts: Provide essential ions for osmotic balance, pH regulation, and cellular function
Metabolites: Supply energy sources (glucose, lactate, pyruvate) and building blocks for growth and development14
Buffers: Maintain a stable pH, crucial for embryo survival and development. Bicarbonate is used for incubator culture, while HEPES/MOPS are used for manipulations outside the incubator
What is the significance of phenol red in embryo culture media?
Phenol red acts as a pH indicator, allowing for visual assessment of pH changes in the media12 . Some manufacturers avoid using it due to concerns about potential estrogenic effects
