Erratum: Murine Left Pulmonary Hilar Clamp Model of Lung Ischemia Reperfusion Injury

Summary

An erratum was issued for: Murine Left Pulmonary Hilar Clamp Model of Lung Ischemia Reperfusion Injury. The Protocol and Representative Results sections were updated.

Abstract

This corrects the article 10.3791/66232

Protocol

An erratum was issued for: Murine Left Pulmonary Hilar Clamp Model of Lung Ischemia Reperfusion Injury. The Protocol and Representative Results sections were updated.

Section 1 of the Protocol has been updated from:

1. Anesthesia and intubation

1. Select a mouse that weighs at least 25 g. This will facilitate easier intubation due to the larger orifice between the vocal cords.
2. Inject the mouse intraperitoneally with a mixture of ketamine (dose 100 mg/kg of body weight) and xylazine (dose 10 mg/kg of body weight). This mixture should be loaded into a ½ cc syringe with a ½ inch 29G needle. Wait about 5 min for ketamine to take effect – the mouse should not exhibit spontaneous movement and does not respond to a toe pinch, which confirms adequate anesthesia.
3. Inject buprenorphine (dose 0.05 mg/kg of body weight) subcutaneously prior to surgery for additional pain control.
4. Apply non-medicated ophthalmic ointment over both eyes to avoid corneal desiccation during surgery.
5. Shave the mouse with a clipper over the left chest and back (see Figure 1A), extending onto the abdomen if laparotomy is planned (see step 5.3).
6. Once the mouse is anesthetized, perform the intubation and remainder of the procedure under a microscope, over a warmed mat at ~37 °C (to maintain normothermia in the mouse).
7. Suspend the mouse vertically by applying an atraumatic clamp on the tip of its tongue, with its back facing toward the surgeon. The rump of the mouse can rest on the mat (see Figure 1B and Supplementary Figure 1A).
8. Pull on the whiskers to expose the oropharynx. Use the prongs of a curved mosquito clamp to depress the tongue and spread the oropharynx in the anterior-posterior axis. When the oropharynx is sufficiently opened, there should be a clear view of the vocal cords (through the microscope objective). The vocal cords of an anesthetized mouse should still open and close spontaneously.
9. Insert a homemade introducer (see Supplementary Figure 1B) with a curve on the end into a 1 inch 20G angiocatheter, which is used as the endotracheal tube (ETT). Use the non-dominant hand to keep the oropharynx open with a curved mosquito (see Figure 1C). With a clear view of the vocal cords, use the dominant hand to guide the tip of the introducer and then the ETT between and past the vocal cords.
   NOTE: It is important to visualize the ETT going through the vocal cords to avoid false intubation into the esophagus. Care should be taken to avoid injury to the vocal cords during intubation (i.e., the number of attempts of intubation should be limited to 5 and the ETT should not be advanced if resistance is met).
10. Once the ETT is inserted, remove the introducer, and connect the ETT to a small animal ventilator. Observe for symmetric chest rise to confirm correct endotracheal intubation. The ventilator settings should be 100-105 breaths per minute and fraction of inspired oxygen of 100%. Tidal volume is 0.35 mL and positive end-expiratory pressure is 1 cm H₂O.
    NOTE: Ensure that the chest rather than abdomen is rising as accidental esophageal intubation will lead to death if not promptly corrected.
11. Once confirmed to be in position, secure the ETT with a 5 cm strip of 1 inch silk tape around the mouse nose, ensuring there is adequate contact with the ETT and nose so that the ETT will not slide out of the mouth (see Figure 1E).
12. To maintain adequate anesthesia throughout the surgery, administer 1%-1.5% isoflurane in-line with the oxygen flow.

to:

1. Anesthesia and intubation

1. Select a mouse that weighs at least 25 g. This will facilitate easier intubation due to the larger orifice between the vocal cords.
2. Inject the mouse intraperitoneally with a mixture of ketamine (dose 100 mg/kg of body weight) and xylazine (dose 10 mg/kg of body weight). This mixture should be loaded into a ½ cc syringe with a ½ inch 29G needle. Wait about 5 min for ketamine to take effect – the mouse should not exhibit spontaneous movement and does not respond to a toe pinch, which confirms adequate anesthesia.
3. Inject buprenorphine (dose 0.05 mg/kg of body weight) subcutaneously prior to surgery for additional pain control.
4. Apply non-medicated ophthalmic ointment over both eyes to avoid corneal desiccation during surgery.
5. Shave the mouse with a clipper over the left chest and back (see Figure 1A), extending onto the abdomen if laparotomy is planned (see step 5.3).
6. Once the mouse is anesthetized, perform the intubation using a preferred intubation set-up and the remainder of the procedure under a microscope over a warmed mat at ~37 °C (to maintain normothermia in the mouse).
7. After achieving an adequate view of the vocal cords, insert a homemade introducer (see Supplementary Figure 1) with a curve on the end into a 1-inch 20G angiocatheter, which is used as the endotracheal tube (ETT). Guide the tip of the introducer and then the ETT between and past the vocal cords.
   NOTE: It is important to visualize the ETT going through the vocal cords to avoid false intubation into the esophagus. Care should be taken to avoid injury to the vocal cords during intubation (i.e., the number of attempts of intubation should be limited to 5 and the ETT should not be advanced if resistance is met).
8. Once the ETT is inserted, remove the introducer, and connect the ETT to a small animal ventilator. Observe for symmetric chest rise to confirm correct endotracheal intubation. The ventilator settings should be 100-105 breaths per minute and fraction of inspired oxygen of 100%. Tidal volume is 0.35 mL and positive end-expiratory pressure is 1 cm H₂O.
   NOTE: Ensure that the chest rather than abdomen is rising as accidental esophageal intubation will lead to death if not promptly corrected.
9. Once confirmed to be in position, secure the ETT with a 5 cm strip of 1 inch silk tape around the mouse nose, ensuring adequate contact with the ETT and nose so that the ETT will not slide out of the mouth (see Figure 1B-C).
10. To maintain adequate anesthesia throughout the surgery, administer 1%-1.5% isoflurane in-line with the oxygen flow.

Figure 1 in the Representative Results has been updated to:

Figure 1: Mouse intubation. (A) Mouse with left chest and abdomen shaved. (B) Endotracheal tube connected to ventilator tubing. (C) Once intubation is confirmed, tape the tube circumferentially to the nose of the mouse. (D) Intubated mouse with inflow and outflow tubing from and to the small animal ventilator. Please click here to view a larger version of this figure.

Supplementary Figure 1 in the Representative Results has been updated from:

Supplementary Figure 1: (A) Clamp set-up for intubation. (B) Introducer for endotracheal tube. Please click here to download this File.

to:

Supplementary Figure 1: Introducer for endotracheal tube. Please click here to download this File.