2.4. Echocardiography

To evaluate diastolic and systolic function and left ventricular geometry, transthoracic cardiac echography was performed. Rats were anesthetized in an induction cage with 3% isoflurane and then maintained to steady-state sedation level with 2% isoflurane by the nose connected to the anesthesia system. The desired depth of anesthesia was confirmed in animals by the absence of reflex after toe pinch. Rats were placed in a decubitus position on a warming pad, the chest was shaved, and hair removal cream was used to remove the remaining body hairs. A gel (Ocry-gel, TVM) was applied to both eyes to prevent drying of the sclera and a lubricated rectal probe was inserted to continuously monitor body temperature in order to maintain it in a range of 37°C ± 0.5. ECG was recorded from suitable electrodes connected to Affiniti 50 (Philips). A layer of preheated gel was applied over the chest. Two-dimensional, M-mode and pulse wave Doppler were obtained from a 15-MHz ultrasound probe, while tissue Doppler at mitral annulus level was recorded from a 12-MHz ultrasound probe. Video were recorded and then analyzed off-line in Q-Station (Philips). All measures were taken from five cardiac cycles and then averaged.