Mouse breathalyzer

Abstract

Background: The development of a relatively simple, noninvasive method for estimating blood ethanol concentrations in mice will be useful in behavioral studies related to alcoholism. This study validated such a method.

Methods: The apparatus consists of a body chamber fitted with a head stock through which the mouse head protrudes. This was fitted against a water-jacketed head-space chamber surrounding the mouse's head. Rebreathed air maintained at 37 degrees C in the head-space chamber was removed using a peristaltic pump and loaded into a 1-ml injection loop. Ethanol in the sample was quantified using gas chromatography. To validate this method, ethanol levels in breath samples were compared against those in tail blood samples collected immediately after the breath samples. Breath samples were collected at 5, 10, 20, 40, 80, 120, and 160 minutes after ethanol (0.4, 0.8, 1.2, 1.6, 2.4, and 3.2 g/kg) was administered to male C57BL/6J mice.

Results: Breath and blood ethanol levels were well correlated (r(2) = 0.96) across time points on the descending ethanol-time curve at doses below 2.4 g/kg. Correlation for these doses on the ascending portion of the curve had greater variance, but was still well correlated (r(2) = 0.92).

Conclusions: The mouse breathalyzer is an accurate, convenient, noninvasive and well-tolerated method for estimating blood ethanol concentrations in mice across a range of behaviorally relevant concentrations below 2.4 g/kg, especially on the descending limb of the ethanol-time curve. Although this procedure requires a gas chromatograph in the animal facility, the ability to estimate ethanol concentrations quickly and easily will be especially useful in behavioral studies where repeated blood sampling is not feasible.

Fig. 1

Illustration of the mouse breathalyzer apparatus. Mice are placed into the lower part of the head stock and the upper portion is placed so that the subject can breathe normally, but cannot withdraw its head. The thumb screw secures the stock in place. The body chamber is secured by a thumb screw to keep the subject’s body in line with the head. The mouse is then placed so that his head is completely inside the head chamber. The water jacket maintains the head chamber at 37°C. A tunnel through the water jacket allows tubing from the injection loop to enter the head chamber for sampling.

Fig. 2

Time course of ethanol levels in breath (A) and blood (B) following ethanol administration at zero minutes in mice (n = 4 per dose). Points represent mean levels expressed as integrated peak area in breath or g/dl in blood. Error bars represent the SEM.

Fig. 3

Time course of ethanol levels in breath (A,C) and blood (B,D) following 2.4 g/kg (A,B) or 3.2 g/kg (C,D) ethanol administration at zero minutes in mice (n = 4 per dose). Each symbol represents an individual mouse at a given dose.

Fig. 4

Breath ethanol levels expressed as a function of blood ethanol concentration and the resulting correlation. (A) Data for all doses of ethanol. Gray circles are reproduced from panel A (doses <2.4 g/kg), downward triangles are points following 2.4 g/kg, and upright triangles are points following 3.2 g/kg. The gray line is the regression through all the data points. (B) Data for doses less up to and including 1.6 g/kg. Each point represents a pair of breath and blood ethanol samples at 1 time point following 1 dose of ethanol in 1 subject. Open circles represent points on the ascending limb of the ethanol–time curve, while black circles represent points on the descending limb of the ethanol–time curve. The solid gray line is the regression through all the points, the dashed gray line is the regression through points on the ascending limb, and the solid black line is the regression for points on the descending limb.