Apical Resection Neonatal Mouse Model: A Surgical Procedure to Amputate the Ventricular Apex to Study Regenerative Potential in a Neonate Mouse Heart

Protocolo

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Apical resection operation in neonatal 1-day-old Cx3cr1 ^GFP/+ mouse (C57BL/6 background)

1. Anesthesia
   1. Take all the mouse pups out from the cages and put them in a clean dry box.
   2. Embed each mouse in the ice for about 2-3 minutes. Ensure that there is a thin barrier such as latex or gauze between the ice and the mouse pup to prevent frostbite. 
   3. Identify sufficient anesthesia by observing for the following signs: pale skin, lack of limb movement, and lack of pedal reflex.
2. Thoracotomy
   1. Precool the bronze operating platform overnight at -20 °C to help maintain anesthesia.
   2. Take the anesthetized mouse out of the ice box and put it on a bronze operating platform. Anchor the mouse on the operating platform in a supine position using medical adhesive tape.
   3. Put the operating platform with the mouse on it under a stereoscope.
   4. Disinfect the mouse chest using a prep pad soaked with betadine and 70% alcohol or as consistent with institutional policy.
   5. Incise the skin with a 1 cm cut at the fourth intercostal area of the chest cavity and then separate the intercostal muscles using microsurgical scissors until the heart is accessed.
   6. Alternately press the chest and abdomen with the help of two forceps until the heart is exteriorized out of chest without any mechanical damage.
   7. Set the ribs below and above the small incision as a natural fixation to immobilize the heart.
3. Ventricular apex resection
   1. Locate the apex of the left ventricle. Cut a 1 mm diameter of ventricular apex tissue using iridectomy scissors under a stereoscope.
   2. Confirm that the left ventricular chamber is exposed and begin oozing.
   3. Gently press the heart back to chest cavity using a cotton swab.
   4. Suture the muscles, ribs, and skin using 8-0 Prolene sutures.
   5. Clean the mouse thoroughly after the operation.
4. Post-operation care
   1. Transfer the mouse from the operating platform to a 37 °C heating blanket to warm up the body immediately after the operation.
   2. Confirm anabiosis of the mouse by observing for the following signs: spontaneous respiration restoration, skin color change from pale to pink, and movement of limbs.
   3. Take the operated mouse back to its mother once it has recovered.
   4. Mingle the operated mouse with its mother's nesting materials if necessary.
      NOTE: It is important to remove all the 1-day-old pups from the mother at once and then return them all at once after all the pups are recovered.

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Materiais

Name	Company	Catalog Number	Comments
Neonatal Heart Dissociation Kit	Miltenyi Bio Tech, Teterow, Germany	130-098-373