Rodent Brain Perfusion

Last Review Date: November 29, 2017

 

Perfusion is considered the means of euthanasia when an animal is anesthetized for immediate perfusion that results in death.  When this is the case, the following step must be taken: 

  • The lab must record that the animal was adequately anesthetized prior to opening the body cavity for perfusion or prior to initiating percutaneous perfusion.

It is known that transcardial perfusion to replace blood in rat brain with saline solution can be used to maximize the health of brain slices by minimizing excitotoxicity and thereby improve experimental outcomes. In particular, this procedure is important for maintaining healthy rat brain slices in experiments involving regions in the basal ganglia.

Perfusion Methodology

  1. The rat will be anesthetized in a chemical fume hood using isoflurane until unresponsive to tail/toe pinches.
  2. The rat is secured in the supine position (lying on the back with face upward) by gently taping the forepaws and hindpaws to a pinnable styrofoam inside a chemical fume hood. Kimwipes are placed into a 15mL falcon tube, and 3 drops of isoflurane are added onto the kimwipes. The Falcon tube is then placed on the nose of the rat nose to maintain the rat anesthetized.
  3. An incision is made through the skin with surgical scissors along the thoracic midline from just beneath the xiphoid process to the clavicle. Two additional skin incisions are made from the xiphoid process along the base of the ventral ribcage laterally.
  4. The two flaps of skin are gently folded rostrally and laterally, making sure to expose the thoracic field completely.
  5. The cartilage of the xiphoid process is grasped with blunt forceps and raised slightly to insert pointed scissors. The thoracic musculature and ribcage is cut between the breastbone and medial rib insertion points and the incision is extended rostrally to the level of the clavicles.
  6. The diaphragm is separated from the chest wall on both sides with scissor cuts.
  7. The reflected ribcage is pinned laterally to expose the heart (and other thoracic organs).
  8. The pericardial sac is gently torn with blunt forceps.
  9. The beating heart is secured with blunt forceps and the left ventricle is poked with a feeding needle.
  10. The right atrium is immediately cut with scissors and infusion of saline solution begins at the first sign of blood flow.
  11. Perfusion continues until the fluid exiting the right atrium is entirely clear.
  12. The rat brain is dissected and submerged in ice-cold saline solution.