1. Procedural Description
Subcutaneously inject 1 μl / g body weight of 0.5 mg / ml atropine sulfate monohydrate (Cat# 11330, Fluka, USA) into each mouse to prevent the salivary
	secretions from disrupting the movement of the infusions.
Anesthetize each mouse with 1 μl / g body weight of a 100 mg / ml ketamine and 20 mg / ml xylazine (Phoenix Scientific, AR, USA) solution 
	injected intra-peritoneal (I.P.)
Place the mouse on a custom-made plastic platform in the ventral position: the maxillary incisors are locked on a metal wire, and the mandibular incisors are hooked on an elastic string in order to hold the mouth open.
Use an Insulin syringe with a 29-gauge needle (Cat# 600145, Tyco Healthcare, MA, USA) placed inside a 0.58 mm diameter polyethylene tube (Cat# 427410, Becton 
	Dickinson, MD, USA; Fig 2). The polyethylene tube is inserted 5mm over the tip of a 29-gauge needle. The needle is used as a rigid guide to move the 
	polyethylene tube into the Wharton's duct. The polyethylene tube is inserted 3-5 mm inside the duct. Note that only the polyethylene tube enters the salivary gland 
	duct while the needle remains outside the duct. Inject 50ul of filtered 0.4% trypan blue stain solution (Cat# 15250, Gibco, USA) into the submandibular gland via the 
	Wharton's duct, which is located right beneath the tongue. The infusion is delivered steadily with positive pressure on the syringe piston; the finger remains on the 
	piston even after all of the infusion has been delivered to the gland, in order to prevent a backflow of the infusion intra-orally. The trypan blue solution is used in 
	the video as a dye to demonstrate the technique. fter the injection of the trypan blue solution, the mouse was euthanized prior to dissection. The salivary gland is surgically exposed in order to determine 
	how much of the infusions have actually remained in the gland.
2. Representative Results:
After the infusions were injected, the mouse was dissected to surgically expose the post-cannulated salivary gland. As shown on Image-2, the dark coloration of the Wharton's duct provides evidence that the trypan blue solution was infused successfully. 
imgsrc://cloudfront.jove.com/files/ftp_upload/3074/3074fig1.jpg
Figure 1. Cannulation of mouse submandibular gland.
imgsrc://cloudfront.jove.com/files/ftp_upload/3074/3074fig2.jpg
Figure 2. Polyethylene tube is encased around the tip of a 29-gauge needle. The syringe needle is used as a rigid guide to move the polyethylene tube into the Wharton's duct. The polyethylene tube is inserted (cannulated) 3-5 mm inside the duct. Note that only the polyethylene tube enters the salivary gland duct while the needle remains outside the duct.Subscription Required. Please recommend JoVE to your librarian.