Six-week-old female rats were anesthetized with isoflurane (5% isoflurane, 95% O2) and bilateral ovariectomy was performed. The test group of female rats underwent ovariectomy (ovaries removed by surgical method) and the other group underwent sham surgery (the skin was cut and sewed back in place). The surgery consisted of a dorsolateral incision of the skin between the last rib and pelvis and muscle dissection to expose periovarian fat. Forceps were used to find the ovaries surrounded by variable amount of fat. The ovary was pulled out and the junction between the fallopian tube and uterine horn was cut. Bleeding was usually light and stopped soon. The horn and periovarian fat were returned into the abdominal cavity. The muscle wound was first sutured shut and the skin incision was closed. In sham surgery, the rats underwent the same incision but no ovaries were removed. To reduce pain, ketoprofen (2.5 mg/kg IM) was administered after surgery and the rats recovered within 3 days.

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