Claires lecture 2

Cryodiluent: Buffer for PH and osmolarity (320OSM), an energy source (glucose/fructose), protective cooling agent(Egg yolk), cryoprotective agent (glycerol), antibiotics(pen and strep)

Protocol: one or two-step, no advantage of 2-step for rams. 30-degree water bath, cool to 5 degrees over 2 hours, freeze, into pellets or straws 

Pellets: better PT motility, easier and quicker freezing technique, slightly higher pregnancy and fertility rates using laparo AI

Straws: more precise storage and labelling system, better ID system, ease of transport

Dilution rate: depends on site of AI, number motile sperm, number of AI doses needed and conc. of sperm

Seminal plasma contains both toxic and protective factors

Balance is required where toxic factors are diluted to minimise harm, and protective factors are in a conc. allowing them to remain protective

Rams 10-25% seminal plasma is optimal

Cryopreservation can lead to capacitation and acrosome reaction, reducing sperm longevity

Adding SP to sperm PT reverses capacitation and improves longevity

During respiration, oxygen is reduced to give water

Incomplete reduction of oxygen forms ROS : chemical entities with powerful oxidising properties

Products of basal cellular metabolism: exercise, mitochondria, phagocytes, reactions with iron and other metals.1-4% 

Oxygen is a free radical with 2 unpaired electrons

As oxygen scavenges electrons it creates ROS

ROS cause oxidative damage to biological molecules such as fatty acids, DNA, proteins, enzymes

Oxidative stress occurs when imbalance between ROS and antioxidants 

Vitamins and minerals which together with enzymes, neutralise free radicals. 

Minerals associated with antioxidant enzymes- deficiency affects enzyme function, active within cells 

vitamins circulate in bloodstream reducing free radical presence 

SP contains AO- freezing dilutes this out and reduced AO enzyme activity 

Low ROS concentrations stimulate capacitation, hyperactivation and the acrosome reaction

Excessive ROS causes impaired sperm function and lipid peroxidation- takes electrons from lipids in cell membranes, ram sperm have high lipid concentration