Chemogenetic inhibition of corticotropin-releasing factor neurons in the central amygdala alters binge-like ethanol consumption in male mice

Abstract

Repetitive bouts of binge drinking can lead to neuroplastic events that alter ethanol's pharmacologic effects and perpetuate excessive consumption. The corticotropin-releasing factor (CRF) system is an example of ethanol-induced neuroadaptations that drive excessive ethanol consumption. Our laboratory has previously shown that CRF antagonist, when infused into the central amygdala (CeA), reduces binge-like ethanol consumption. The present study extends this research by assessing the effects of silencing CRF-producing neurons in CeA on binge-like ethanol drinking stemming from "Drinking in the Dark" (DID) procedures. CRF-ires-Cre mice underwent surgery to infuse G_i/o-coupled Designer Receptors Exclusively Activated by Designer Drugs (DREADD) virus or a control virus into either the CeA or basolateral amygdala (BLA). G_i/o-DREADD-induced CRF-neuronal inhibition in the CeA resulted in a 33% decrease in binge-like ethanol consumption. However, no effect on ethanol consumption was seen after DREADD manipulation in the BLA. Moreover, CeA CRF-neuronal inhibition had no effect on sucrose consumption. The effects of silencing CRF neurons in the CeA on ethanol consumption are not secondary to changes in motor function or anxiety-like behaviors as assessed in the open-field test (OFT). Finally, the DREADD construct's functional ability to inhibit CRF-neuronal activity was demonstrated by reduced ethanol-induced c-Fos following DREADD activation. Together, these data suggest that the CRF neurons in the CeA play an important role in binge ethanol consumption and that inhibition of the CRF-signaling pathway remains a viable target for manipulating binge-like ethanol consumption. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Figure 1.

Mice used in behavioral studies (A) underwent surgery in the BLA or CeA receiving either G_i/o-DREADD or control virus after a baseline was established in the DID with ethanol. After a recovery period, the impact of G_i/o-DREADD activation on ethanol consumption was determined in 2 cycles of DID with CNO. To ensure the CNO did not have an independent effect, a third DID cycle was conducted using vehicle on the test day. As a follow-up, mice that were used in the CeA studies also underwent 1 cycle of sucrose DID consumption with CNO. Finally, the same mice were tested in an OFT with CNO to compare the G_i/o-DREADD vs control viral impact on locomotor activity and anxiety after CNO injection. In a separate cohort for the c-Fos studies (B), viral constructs were injected into the CeA and activated with CNO. Ethanol was administered ip before animals were euthanized.

Figure 2.

Silencing of CRF neurons in the CeA reduced binge-like ethanol consumption (A) and BECs (B) in the G_i/o-DREADD mice (n=11) compared with mice with control virus (n=10). In the absence of CNO, however, viral condition did not alter ethanol consumption (C) and therefore had no effect on BECs (D). Finally, no effect of silencing CRF neurons was observed on sucrose consumption (E) compared with the control virus condition group. (*p<0.05 compared to control virus group). All data are presented as mean ± SEM.

Figure 3.

G_i/o-DREADD viral administration (n=8) within the BLA had no effect on either ethanol consumption (A) or BECs (B) compared with the control group (n=7). All data are presented as mean ± SEM.

Figure 4.

Photomicrographs of mice with either control (A; n=9) or G_i/o (B; n=12) DREADD show a depression of ethanol-induced c-Fos IR in mice with activated G_i/o DREADD. Direct comparisons show that CRF neuronal inhibition within CeA reduced c-Fos expression in G_i/o animals compared with controls (C). Panel D is a multi-panel image capturing the amygdala indicating that DREADD expression was confined to the CeA. Panel E is a colabel showing the overlap of CRF+ (green) and DREADD+ cells (red) within the CeA.