As an alternative, we hypothesized that nicotine administration altered the DA and GABA responses to alcohol through a neuroendocrine signal (Armario, 2010). Stress-related hormones, such as glucocorticoids, cause long-term homeostatic changes in neural function and influence DA and GABA transmission (Barrot et al., 2000, Butts et al., 2011 and Joëls and Baram, 2009). Nicotine activates the HPA axis to increase plasma levels of corticosterone (Lutfy et al., 2012), the principle glucocorticoid

ABT-737 manufacturer in rodents, which we confirmed (Figure S3). To determine whether glucocorticoid receptor activation during nicotine pretreatment contributes to subsequent alterations in ethanol-induced DA release, we systemically blocked glucocorticoid receptors with RU486 (Cadepond et al., 1997) prior to nicotine pretreatment. Pretreatment with RU486 (Figure 5A, blue circles) prevented the inhibitory effect of nicotine on ethanol-induced DA release (group × time: F(10,240) = 4.75, p < 0.01). Lonafarnib order This increased DA response to ethanol after RU486 and nicotine pretreatment

was not distinguishable from the control rats pretreated with saline alone or RU486 alone ( Figure 5A, dashed trace). These results suggested that stress hormone receptor activation within the VTA, after nicotine pretreatment, attenuated the subsequent DA response to ethanol. To test this hypothesis, we blocked glucocorticoid receptors locally in the VTA with RU486 prior to nicotine pretreatment. The control group that received a local intra-VTA microinfusion of vehicle followed by nicotine pretreatment showed a decreased DA response to ethanol 15 hr later (Figure 5B, red circles), consistent with our previous data (see Figure 1). This inhibitory effect of nicotine pretreatment was prevented by intra-VTA microinfusion of RU486 prior to nicotine pretreatment (Figure 5B, blue circles) (group × time: F(10,140) =

2.43, p < 0.05). We should note that the intra-VTA RU486 did not completely reverse secondly the effect of nicotine pretreatment. A post hoc comparison indicated a significant difference between the saline control ( Figure 5B, dashed line) and the group pretreated with intra-VTA RU486 + Nic (F(10,220) = 2.01, p < 0.05). The microinfusion sites were dispersed mainly in the more ventral VTA, including the anterior and posterior regions ( Figure 5C). There was no consistent relationship between the microinfusion site and the individual DA responses to ethanol in either group. As a negative control, microinfusion of RU486 outside and adjacent to the VTA did not reverse the inhibitory effect of nicotine pretreatment (n = 3).