Fig. 1. Structure of Icariin were first trained to consume a 1% sucrose solution after the 3-d adaptation period. The test of sucrose solution intake baseline was performed 9 times (once every three days) for each animal. The test involved an 18 h period of food and water deprivation followed by offering the 1% sucrose solution for 1 h. Sucrose-intake was measured by weighting bottle weights containing sucrose solution before and after the test respectively. The sucrose intake was expressed in relation to animals' body weights (g/kg).CMS Procedure After the sucrose intake baseline test, the animals were divided into two groups with similar average intake. Twenty-one rats in one group were housed under normal condition (as non-stressed animals) and twenty-eight rats in the other group were subjected to CMS procedure (as CMS-treated animals) throughout. Subsequently, sucrose consumption was monitored weekly for the next experimental period (12 weeks). The CMS procedure was carried out for 6 weeks using the method described by Willner et al. with small modification. 19,20) Each experiment cycle extended 7 d and included the following steps:Low intensity stroboscopic illumination (150 flashes/min) for 6 h; tilted cage (45°) for 14 h; intermittent illumination (2 h light/2 h dark cycle) for 12 h; random partnership for 12 h; wet cage (200 ml water in 100 g sawdust bedding) for 12 h; continuous light for 12 h; low intensity stroboscopic illumination (150 flashes/min) for 10 h; water and food deprivation for 14 h; noise (6300 Hz tone, approx. 90 dB) for 6 h; tilted cage (45°) for 18 h; low intensity stroboscopic illumination (150 flashes/min) for 12 h; continuous light for 12 h; noise (6300 Hz tone, approx. 90 dB) for 9 h; water and food deprivation for 18 h; sucrose test for 1 h.All of the stressors were applied individually and continuously, day and night. Non-stressed rats were housed in separate rooms away from CMS-stressed animals. Food and water were freely available in the home cage except that they were deprived of both before each sucrose test.Drug Administration Normal control rats and stressed rats were further divided into subgroups on the basis of sucrose intake scores after the CMS procedure. The nonstressed animal groups were divided into three subgroups and they received normal saline (1 ml/kg, normal control, nϭ7), icariin (35 mg/kg, nϭ7), icariin (70 mg/kg, nϭ5) and fluoxetine (7 mg/kg, nϭ7), respectively. The CMS-treated animal groups were further divided into four subgroups (7 rats/subgroup) and they received normal saline (1 ml/kg, stressed control), icariin (35 mg/kg), icariin (70 mg/kg), and fluoxetine (7 mg/kg), respectively. All drugs were suspended in 0.9% normal saline, and were orally administered once daily for 6 weeks.Blood Collection After 6-week administration and the last sucrose intake test, all animals were left without any treatment until the following morning. The next day, they were etherized and decapitated for bleeding between 09:00 a.m. and 09:30 a.m. Blood was collected on ic...