Ovariectomy and estrogen supplementation

Bilateral ovariectomy was performed under isoflurane anesthesia, as previously described (56) and performed in our laboratory (57). Estradiol (E2) was administered via subcutaneous implantation of Silastic tubing (10 mm/100 g body weight), filled with a solution of E2 in sesame oil (182.4 μg E2/1 mL) to reach circulating E2 levels comparable to proestrus (58). Ovariectomized (Ovx) rats were implanted 1 week following ovariectomy. Diestrous rats were implanted on day one of diestrus. Control rats were implanted with Silastic tubing containing vehicle (sesame oil). All rats were killed 48 hours after implantation.