Effects of intra-PVN infusion of RU486 on behavior in rats that were chronically treated with CORT
We investigated the effects of RU486 on depressive- and anxiety-like behaviors and the levels of GRs, MRs, and CRF in the PVN in rats that were treated with CORT + vehicle. On day 16 during CORT consumption at 08:00, the rats received an intra-PVN infusion of RU486 for 7 days. As shown in Fig. 3, rats that were treated with CORT + vehicle also exhibited depressive-like behaviors in the FST (F 3, 23 = 42.47, p < 0.01, Fig. 3A) and SPT (F 3, 23 = 7.599, p < 0.01, Fig. 3B) and anxiety-like behaviors in the SIT (F 3, 23 = 24.03, p < 0.01, Fig. 3C) and NSFT (F 3, 23 = 6.225, p< 0.01, Fig. 3D). The intra-PVN infusion of RU486 did not influence CORT-induced depressive- or anxiety-like behaviors (Fig. 3A-D). Western blot revealed that the intra-PVN infusion of RU486 restored GR expression in the PVN (F 3, 16 = 15.88, p < 0.01, Fig. 3F) without marked improvement in MR levels in the PVN (Fig. 3E) and significantly decreased the MR/GR ratio (F 3, 16 = 8.628, p < 0.01, Fig. 3G). Unexpectedly, intra-PVN RU486 administration augmented the increase in CRF levels compared with rats that were treated with CORT + vehicle (F 3, 16 = 94.75, p < 0.01, Fig. 3H). These data suggest that although GR levels in the PVN in rats that were treated with CORT + vehicle rats were restored to normal by the intra-PVN infusion of RU486 alone, the MR/GR ratio was still imbalanced, and CRF levels remained high. Thus, depressive- and anxiety-like behaviors in rats that were chronically treated with CORT were not inhibited.
Effects of intra-PVN infusion of spironolactone on behavior in rats that were chronically treated with CORT
To elucidate the involvement of MRs in the PVN in the behavioral alterations that were induced by chronic CORT exposure, the MR antagonist spironolactone was microinjected into the PVN on day 16 of CORT consumption at 08:00 for 7 days. Depressive-like behavior, including immobility time in the FST (Fig. 4A) and sucrose preference in the SPT (Fig. 4B), were not reversed by intra-PVN spironolactone administration. In contrast, the intra-PVN infusion of spironolactone in CORT-exposed rats exerted anxiolytic-like effects, reflected by an increase in social interaction in the SIT (F 3, 22= 18.76, p < 0.01, Fig. 4C) and a shorter feeding latency in the NSFT (F 3, 22 = 19.07, p< 0.01, Fig. 4D). The intra-PVN injection of spironolactone reversed the decrease in MR levels in the PVN that was induced by chronic CORT exposure (F 3, 16 = 26.15, p < 0.01, Fig. 4E). Chronic CORT exposure significantly altered GR and CRF expression, and these changes were unaffected by intra-PVN spironolactone administration (Fig. 4F, H). Because MR levels returned to normal but GRs levels failed to recover, the MR/GR ratio was two-times higher than normal and notably out of balance (F 3, 16 = 7.344, p < 0.01, Fig. 4G). Based on these results, the CORT-induced decrease in MR levels in the PVN may be associated with anxiety-like behavior, and intra-PVN spironolactone administration restored MR levels to normal, which may be related to its anxiolytic effect. Meanwhile, the failure of GR and CRF levels and the MR/GR ratio to return to normal in the PVN in CORT-treated rats may explain why the intra-PVN infusion of spironolactone did not produce an antidepressant effect.
Effects of co-administration of RU486 and spironolactone in the PVN on behavior and GR, MR and CRF levels in rats that were chronically treated with CORT
Accumulating evidence indicates that MRs cooperate with GRs in the stress response (Spencer et al., 1998; Mattsson et al., 2009; Jadavji et al., 2011). In this experiment, we co-administered the GR antagonist RU486 and MR antagonist spironolactone in the PVN. RU486 was microinjected in the PVN once daily beginning on day 16 of CORT consumption at 08:00 for 7 days. Spironolactone was administered on the same days, 40 min after RU486 administration. Combined GR and MR blockade in the PVN in rats that were chronically treated with CORT restored behavior in the FST (F 3,22 = 15.78,p < 0.01, Fig. 5A), SPT (F 3,22 = 9.56, p < 0.01, Fig. 5B), SIT (F 3,22 = 16.68, p < 0.01, Fig. 5C) and NSFT (F 3,22 = 18.99, p < 0.01; Fig. 5D) and the levels of MRs (F 3,16 = 14.64, p < 0.01, Fig. 5E), GRs (F 3,16 = 33.93, p < 0.01, Fig. 5F) and CRF (F 3,16 = 22.60, p < 0.01, Fig. 5H) in the PVN. Altogether, these data suggested co-administration of the MR and GR antagonists in the PVN restored the levels of MRs, GRs, and CRF and the MR/GR ratio in the PVN to normal in CORT-exposed rats, thus inhibiting depressive- and anxiety-like behaviors.