Fig 1. Experimental procedure.

In Experiment 1, mice received a single s.c. injection of either oil vehicle (control, 0.1 ml sesame oil) or various doses (0.5 μg/0.1 ml, 5 μg/0.1 ml, or 50 μg/0.1 ml) of EB 7 days post-ovariectomy. Fear conditioning was conducted 2 days after EB treatment, and the mice were tested for their conditioned fear responses the following day. In Experiment 2, mice were ovariectomized and implanted s.c. with a Silastic capsule (I.D. 1.98 × 20.0 mm) containing either vehicle or various doses (0.05 μg, 0.5 μg, 5 μg, 50 μg/0.1 ml) of EB. Two weeks post-implantation, fear conditioning and testing were conducted. In the conditioning phase, 3 min after being placed in the chamber, mice were administered three consecutive foot shocks (duration: 2 s, 0.8 mA) with 30 s intershock intervals. The day after the last behavioral test, the animals were sacrificed using a pentobarbital overdose. The uteri were collected and the wet weights were recorded. In Experiment 3, mice were ovariectomized and implanted (s.c.) with a Silastic capsule (I.D. 1.98 × 20.0 mm) containing either vehicle or various doses (0.05 μg, 0.5 μg, 5 μg, 50 μg/0.1 ml) of EB. Two weeks post-implantation, the animals were decapitated and their trunk blood was collected for the hormonal assay.