An Intracerebroventricular Injection to Analyze Murine Cytomegalovirus Distribution in a Neonatal Mouse Brain

Abstract

Source: Kawasaki, H., et al. Intracerebroventricular and Intravascular Injection of Viral Particles and Fluorescent Microbeads into the Neonatal Brain. J. Vis. Exp. (2016).

This video demonstrates the intracerebroventricular injection process for studying murine cytomegalovirus (MCMV) distribution in the neonatal mouse brain. MCMV, engineered to express an enhanced green fluorescent protein (EGFP) for visibility, is injected into the lateral ventricle of a mouse pup's brain. The injected virus circulates through the cerebrospinal fluid, infecting cells in the periventricular region, including the marginal zone, choroid plexus, and subventricular zone.

Protocol

1. ICV Injection of MCMV and Fluorescent Microbeads into Neonatal Mice

1. Maintain normal pregnant ICR mice in a temperature-controlled facility under a 12 hr light/dark cycle. The neonates are designated as P 0.5 on the day of birth.
2. Sterilize a 10-µl syringe and a 27 G needle with 70% alcohol.
3. Load the injection solution (5 µl) containing MCMV or fluorescent microbeads into the needle by carefully pulling the plunger of the syringe.
4. Restrain neonatal mouse (P 0.5) by putting the mouse on ice for 3 - 4 min. Once the animal is under anesthesia, use the toe-pinch response method to determine the depth of....

Results

Figure 1: Injection Sites of the Neonatal Mouse. (A, B) ICV injection of 5 µl MCMV (approximately 5 × 10⁵ PFU) or of 5 µl microbeads (approximately 5.75 × 10⁹ particles). (A) The injection site i.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
SPHERO TM Fluorescent Polystyrene Nile Red 0.04-0.06	Spherotech, Inc.	FP-00556-2
SPHERO TM Fluorescent Polystyrene Nile Red 0.1-0.3	Spherotech, Inc.	FP-0256-2
SPHERO TM Fluorescent Polystyrene Nile Red 1.7-2.2	Spherotech, inc.	FP-2056-2
10% mouse serum	DAKO	X0910
C57BL/6 mouse	SLC, Inc.
ICR mouse	SLC, Inc.
Modified Microliter Syringes (7000 Series)	Hamilton company
35-gauge needle	Saito Medical
pEGFP-N1 vector	Clontech	#6085-1