Intranasal Drug Administration in Mice via Pipette and Reverse-Cannulation for Nose-to-Brain Drug Delivery

Abstract

Source: Kanazawa, T., et al. Novel Methods for Intranasal Administration Under Inhalation Anesthesia to Evaluate Nose-to-Brain Drug Delivery. J. Vis. Exp. (2018)

This video demonstrates the intranasal administration of radiolabeled drugs in an anesthetized mouse to achieve nose-to-brain drug delivery. It explains how drugs cross the nasal epithelium and travel via neuronal pathways to reach the brain, as well as how ciliary motion influences drug retention and clearance. Additionally, the video describes an alternative reverse cannulation method, which involves inserting a cannula through the esophagus to deliver drugs against the ciliary motion, enhancing retention and improving transport efficiency.

Protocol

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

1. Animals Used for Intranasal Administration Under Inhalation Anesthesia

1. House the experimental mice in stainless-steel cages under a 12-h light/dark cycle (light on 8:00 AM–8:00 PM), with a controlled temperature maintained at 23 ± 1°C, humidity of 50% ± 10%, and ad libitum access to food and water.
2. Prior to experimentation, anesthetize the mice via inhalation of 2% isoflurane, following initiation at a concentration of 4%. Confirm the requisite level of anesthetization by checking for the disappearance of surface righting.

2. Preparation of the Administration Solution

1. Prepare an administration solution of [¹⁴C]-inulin (50 μM, 0.5 μCi/mL per mouse) by diluting in phosphate-buffered saline, and store at 4 °C until use.

3. Intranasal Administration for Mice

1. Micropipette method using a temporarily openable inhalation mask (Figure 1)
   1. Fix mice in the supine position on a corkboard by taping their limbs under inhalation anesthesia with 2% isoflurane (Figure 1A).
   2. Administer a total volume of 25-μL administration solution to each mouse at 30-s intervals, via 1- to 2-μL doses alternatively administered into the left and right nostrils while the mice are fixed under inhalation anesthesia (Figure 1B and C).
   3. Note: Waste anesthetic gas is scavenged by active means (fume hood, hard-ducted biosafety cabinet, vacuum, etc.) when the mask is opened and by passive means(waste anesthetic gas cannister) when closed.
2. Reverse cannulation method from the airway side through the esophagus (Figure 2)
   1. Fix mice in the supine position on a corkboard by taping their limbs under inhalation anesthesia with 2% isoflurane.
   2. Hair in the neck is shaved and prepped via betadine or chlorhexidine application, followed by an alcohol rinse.
   3. Expose the trachea and esophagus by expanding the skin below the throat with forceps after making a small incision (1.5 cm) with scissors.
   4. Make an incision (1 mm) in the trachea using scissors.
   5. Insert a cannula (internal diameter: 0.58 mm, exterior diameter: 0.965 mm) to a length of 1.2 cm and attach the opposite end of the cannula to the inside of the inhalation mask.
   6. Make an incision (1 mm) in the esophagus using scissors, insert a cannula (internal diameter: 0.28 mm, exterior diameter: 0.61 mm) to a length of 1.4 cm toward the posterior part of the nasal cavity, and ligate it (Figure 2A and B).Note: Procedures 3.2.2 to 3.2.4 were carried out under a stereoscopic microscope at ×10 magnification.
   7. Attach a needle (27G × 1/2) to a 1-mL syringe filled with an administration solution and connect to a programmable micro-syringe pump.
   8. Connect the above needle to the cannula that had been inserted into the esophagus at 3.2.5 (Figure 2C).
   9. Administer a total volume of 25 μL [¹⁴C]-inulin solution at a constant rate (5 μL/min) (Figure 2C and 2D).

Results

Figure 1: Intranasal administration using a micropipette in conjunction with a temporarily openable inhalation mask...

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
ddY mouse	Japan SLC, Inc.		Male, 4-6 weeks, 20-30 g
Isoflurane	Pfizer	v002139
Isoflurane setup	SHINANO manufacturing CO. LTD.	SN-487-OTAir, SN-489-4
Isoflurane mask	SHINANO manufacturing CO. LTD.		For small rodents
Isoflurane mask (Opneable type)	SHINANO manufacturing CO. LTD.		Special orders
Anesthesia Box	SHINANO manufacturing CO. LTD.	SN-487-85-02
Animal experiments scissors-1	NATSUME SEISAKUSHO CO., LTD.	B-27H
Animal experiments scissors-2	NATSUME SEISAKUSHO CO., LTD.	B-13H
Tweezers-1	FINE SCIENCE TOOLS Inc.	11272-30	Dumont #7 Dumoxel
Tweezers-2	NATSUME SEISAKUSHO CO., LTD.	A-12-1
Cannula tube (PE-50)	Becton, Dickinson and Company.	5069773	I.D.: 0.58 mm, O.D.: 0.965 mm
Cannula tube (SP-10)	NATSUME SEISAKUSHO CO., LTD.	KN-392	I.D.: 0.28 mm, O.D.: 0.61 mm
Shaver	MARUKAN, LTD.	DC-381
Stereoscopic microscope	Olympus Corporation	SZ61
Needle 27G 1/2 in 13 mm	TERUMO CORPORATION	NN-2738R
1 mL syringe	TERUMO CORPORATION	SS-01T
Syringe pump	Neuro science	NE-1000
Micro spatula	Shimizu Akira Inc.	91-0088
Micropipette (0.5-10 uL)	Eppendorf AG	Z368083
Pipette chip	Eppendorf AG	0030 000.811
Tape	TimeMed Labeling System, Inc.	T-534-R	For fixing mouse
[¹⁴C]-Inulin	American Radiolabeled Chemicals Inc.	ARC0124A	0.1 mCi/mL
EtOH	Wako Pure Chemical Industries, Ltd.	054-00461