Diethylstilbestrol Versus Estradiol: MATERIALS AND METHODS(2)
Ovariectomy and placement of E2 pellets were performed at 21 days of age under pentobarbital-induced anesthesia. The pellets were inserted s.c. between the shoulder blades and consisted of a plugged Silastic (Dow Corning Corp., Midland, MI) tube (open lumen length, 1.0 cm; inner diameter, 1.57 mm; outer diameter, 2.41 mm) filled with crystalline E2. According to previous determinations, this procedure maintains serum E2 levels at approximately 200 pg/ml for at least 5 mo. At all the ages studied, at least three animals from each treatment group were anesthetized with CO2, weighed, killed, and immediately processed as described below.
Tissue Harvesting, Processing, and Histological Analysis
The bodies of prepubertal animals were eviscerated, and the lower torsos were immersed in ice-cold fixative for at least 24 h before reproductive tracts were excised, trimmed of adhering fat and mesentery under a dissecting microscope, and placed back in fixative. The fixative consisted of 4% paraformaldehyde in Dulbecco PBS (pH 7.4). For the adult animals, reproductive tracts were immediately excised and then also placed in fixative for at least 24 h before trimming and placement back in fixative.