A plethysmographic method for measuring function in locally irradiated mouse lung.

A plethysmograph has been developed to measure pulmonary function in mice after single doses of X rays to both lungs. The apparatus consists of a whole-body airtight chamber fitted with a Lavalier microphone. The microphone acts as a sensitive electrical capacitance manometer converting pressure changes in the chamber into an electrical signal which is electronically processed and recorded on a pen recorder. Two parameters of lung function were simultaneously monitored, breathing rate and amplitude. Lung function has been tested in male CBA mice aged two to six months and in animals which have received graded X-ray doses to both lungs. No diurnal rhythm or agerelated increase has been observed up to six months in control mice. The two lung-function parameters exhibited a dose-dependent response in irradiated lungs tested 16 weeks after irradiation; the response was reproducible in successive experiments. Respiration rate was increased above a threshold dose of 11 Gy (1100 rad), while amplitude decreased, also with a threshold at 11 Gy. These changes were observed before histological evidence of fibrosis became apparent and before pulmonary insufficiency led to deaths in the higher dose groups. The measurement of lung function by plethysmography is an alternative to lethality for assessing radiation damage in the lungs of small animals. The technique is non-destructive, responding to lower doses than LDso, and allows quantitative assessment of sequential changes in the lungs in each mouse over long post-irradiation times. New quantitative endpoints for the assessment of normal tissue injury after irradiation are being developed by an increasing number of investigators (Chauser et al., 1976; Glatstein et al., 1975; Hayashi and Suit, 1971; Hirst et al., 1977; Hopewell, 1975; Stewart, 1977; Van der Kogel et al., 1977; Vatistas and Hornsey, 1966; White and Hornsey, 1977; Withers, 1967; Withers and Elkind, 1969). Because radiation damage to normal tissues is the doselimiting factor in radiotherapy it is important that endpoints should be established which enable injury, particularly late injury, to be quantitated. Many of the techniques presently available are invasive or involve sacrifice of the animal. Functional assays which permit sequential assessment of injury in the same groups of animals over a period of time are available for only a few organs in small laboratory animals. The present study reports a new non-invasive technique of measuring changes in pulmonary function after irradiation of both lungs of mice. MATERIALS AND METHODS Experimental animals and radiation procedures Lung function was tested in mice in which both lungs were irradiated and in sham-irradiated controls. Male CBA/Ht mice, 10-12 weeks old, were given single doses of X rays ranging from 8 to 20 Gy. Radiation was delivered with a 250 kV X-ray unit, operated at 240 kV with an HVT of 1.3 mm Cu at a target skin distance of 20 cm. The dose rate was 3.65 Gy per min. Four mice were irradiated simultaneously on a specially constructed jig. The total dose was delivered within four to ten minutes in four equal increments with the jig rotated 90 deg after each increment to allow for a constant 10% dose-difference between the four irradiation positions. Both lungs were irradiated through two anterior 20x15 mm apertures in a 3.0 mm thick lead shield which protected the remainder of the body. Structures in the thoracic mid-line were also protected by a 1.0 mm wide strip in the lead shield separating the two lung fields. Perspex pillars fixed to the base of the jig kept the mouse in a supine nonrotated position and held the rib cage in position with respect to the radiation fields. This positioning was checked in some animals by radiographs which showed that less than 5% of the total volume of lung was protected by the mid-line shield. Dose distribution in the lung was checked using thermoruminescent dosimeters in dead mice. The mice were anaesthetized before irradiation with sodium pentobarbital (60 mg/kg) given intraperitoneally. Megimide (15 mg) was given to each mouse after irradiation to speed recovery from the anaesthetic.