br Materials and methods br Results br Discussion Consistent
Materials and methods
Discussion Consistent with previous reports (Barr et al., 2010, Vuong et al., 2010), the current study demonstrates that rats treated with amphetamine exhibit heightened anxiety states during withdrawal. Furthermore, icv. infusion of a CRF2 receptor antagonist decreased anxiety-like behaviors of rats in the EPM during amphetamine withdrawal, without affecting locomotion. Intra-dRN infusion of a CRF2 receptor antagonist attenuates anxiety-like behaviors of rats during amphetamine withdrawal and also following early life stress, with CRF1 receptor antagonism having little effect (Vuong et al., 2010, Bledsoe et al., 2011). The current study adds to this by demonstrating that global central blockade of CRF2 receptors also reduces anxiety-like behaviors in an animal model of heightened anxiety, highlighting the therapeutic potential of CRF2 receptor antagonism. Given that CRF2 receptors are found in the periphery, specifically in the heart, gastrointestinal (GI) tract, lung, skeletal muscle, and vasculature (as reviewed by Bale and Vale, 2004), future research should assess the effects of systemically administered CRF2 receptor antagonists to further determine the feasibility of targeting CRF2 receptors in the treatment of anxiety. In ITE to amphetamine pre-treated rats, icv. infusion of ASV had an anxiogenic effect on saline pre-treated rats. This observation is consistent with findings from CRF2 knockout mice, which exhibit increased anxiety behavior (Bale et al., 2000), but is in contrast to pharmacological findings. Takahashi et al. (2001) show that icv. infusion of ASV (in a concentration range used by the current study) decreases anxiety-like behaviors in rats not subjected to any pre-treatment. Similarly, we demonstrate here that rats not exposed to pre-treatment injection procedures exhibit decreased anxiety-like behaviors following icv. infusion of ASV. The most likely explanation for our results is that the mild stress induced by prior handling and injection was sufficient to alter behavioral responses of saline-treated rats to CRF2 receptor antagonism in mildy anxiogenic environments. Combined, previous and current findings suggest that while CRF2 receptors mediate production of anxiety-like behaviors in rodents, central pharmacological blockade will have differential effects on behavior depending on prior handling experience. These findings emphasize the idea that there are occasions in which a non-handled control group might need to serve as a ‘control’ for the manipulated vehicle-treated animals to assist in interpretation of pharmacological and behavioral findings. Furthermore, these results suggest that the ability of mild pre-stress to induce anxiety-like behavior when combined with CRF2 receptor antagonism may increase drug seeking behavior and relapse, a possibility that requires further testing. One of the potential mechanisms by which ASV has differential effects on behavior as a function of handling experience could be related to changes in regionally specific CRF receptor expression. In contrast to icv. infusion, direct infusion of CRF1 or CRF2 receptor antagonists into the dRN has no effect on anxiety-like behaviors of saline pre-treated or control rats (Vuong et al., 2010, Bledsoe et al., 2011). This suggests that the dRN was not the locus of the ASV-induced anxiogenic effect in saline pre-treated rats. Therefore, we examined CRF receptor expression in three other brain regions (LS, CeA and BNST) in which CRF2 receptors are associated with fear or anxiety-like reponses (Radulovic et al., 1999, Liu et al., 2004, Henry et al., 2006, Skórzewska et al., 2011, Takahashi et al., 2011, Lebow et al., 2012, Ventura-Silva et al., 2012, Elharrar et al., 2013). One of the major differences in CRF receptor expression in saline pre-treated rats as compared to both un-treated and amphetamine pre-treated rats was observed as a decrease in CRF1 receptors within the LS. However, a role for LS CRF1 receptors in mediating anxiety is not clear as CRF2 receptors in the LS have been more often implicated in increased anxiety-like behaviors (e.g. Radulovic et al., 1999, Henry et al., 2006, Bakshi et al., 2002). Interestingly, activation of CRF1 and CRF2 receptors have opposing effects on glutamatergic transmission in the LS (Liu et al., 2004). Specifically, glutamatergic transmission in the LS is facilitated by CRF1 receptor activity but dampened by CRF2 receptor activation (Liu et al., 2004). Thus, when CRF2 receptors are blocked by ASV as in the current study, CRF actions on CRF1 receptors in the LS would be attenuated in saline pre-treated rats as compared to un-treated and amphetamine pre-treated rats, potentially reducing glutamatergic transmission.