Neuroplasticity in brain reward circuitry following a history of ethanol dependence

Abstract

Mitogen-activated and extracellular regulated kinase (MEK) and extracellular signal-regulated protein kinase (ERK) pathways may underlie ethanol-induced neuroplasticity. Here, we used the MEK inhibitor 1,4-diamino-2,3-dicyano-1,4-bis(2-aminophenylthio)butadiene (UO126) to probe the role of MEK/ERK signaling for the cellular response to an acute ethanol challenge in rats with or without a history of ethanol dependence. Ethanol (1.5 g/kg, i.p.) induced expression of the marker genes c-fos and egr-1 in brain regions associated with both rewarding and stressful ethanol actions. Under non-dependent conditions, ethanol-induced c-fos expression was generally not affected by MEK inhibition, with the exception of the medial amygdala (MeA). In contrast, following a history of dependence, a markedly suppressed c-fos response to acute ethanol was found in the medial pre-frontal/orbitofrontal cortex (OFC), nucleus accumbens shell (AcbSh) and paraventricular nucleus (PVN). The suppressed ethanol response in the OFC and AcbSh, key regions involved in ethanol preference and seeking, was restored by pre-treatment with UO126, demonstrating a recruitment of an ERK/MEK-mediated inhibitory regulation in the post-dependent state. Conversely, in brain areas involved in stress responses (MeA and PVN), an MEK/ERK-mediated cellular activation by acute ethanol was lost following a history of dependence. These data reveal region-specific neuroadaptations encompassing the MEK/ERK pathway in ethanol dependence. Recruitment of MEK/ERK-mediated suppression of the ethanol response in the OFC and AcbSh may reflect devaluation of ethanol as a reinforcer, whereas loss of an MEK/ERK-mediated response in the MeA and PVN may reflect tolerance to its aversive actions. These two neuroadaptations could act in concert to facilitate progression into ethanol dependence.

Figure 1

Schematic representation of the sampled areas for the densitometric evaluation of mRNAs in a coronal section through the rat forebrain at Bregma levels +2 to −3 mm. Cingulate cortex (cg); frontal motor cortex (M1); primary sensory cortex (S1); infralimbic cortex (IL); orbitofrontal cortex (OFC); nucleus accumbens core (AcbC); nucleus accumbens shell (AcbSh); central amygdaloid nucleus (CeA); medio amygdaloid nucleus (MeA); basolateral amygdaloid nucleus (BLA); dorsal hippocampal subregions (CA: Cornus Ammon areas, CA1 to CA4; dentate gyrus, (DG); supraoptic nucleus (SO); hypothalamic paraventricular nucleus (PVN).

Figure 2

Right: Bar graph illustrating c-fos expression in the medial amygdala after UO 126 (1, 2.5 and 5 nmol, icv) and EtOH (1.5 g/kg, i.p.) treatment in naïve Wistar rats. Corrected p-values: ***p < 0.001 vs veh-sal group, #p < 0.05 vs veh-EtOH control group. Left: Bright-field microphotographs from autoradiograms of in situ hybridization of c-fos mRNA in the amygdala region after UO126 (5 nmol, icv) and EtOH (1.5 g/kg, i.p.) treatment in naïve Wistar rats. Arrows indicate c-fos mRNA in MeA and CeA. c-fos mRNA levels are increased in both MeA and CeA after ethanol challenge and decreased in MeA after UO126 treatment. UO 126 treatment show no effect on ethanol-c-fos in CeA, scale bar = 1 mm, for abbreviations see figure 1, for details on treatment, see Material and Methods

Figure 3

Ethanol induced c-fos (upper panel) and egr-1 (lower panel) expression in different forebrain regions of Wistar rats. Bar graphs illustrating c-fos and egr-1 expression 45 minutes after ethanol (EtOH, 1.5 g/kg, i.p., black bar) or saline (sal, i.p., white bar) injection in rats pretreated with vehicle (veh, 4 % DMSO, icv). Data are expressed as percent of control group (% veh-sal group, mean ± S.E.M.). Statistical analysis were performed by one-way ANOVA followed by Holms corrected Bonferroni's post-hoc test, n = 4−6/group, corrected p-values: *p<0.05, **p<0.01, ***p<001; for abbreviations see figure 1, for details on treatment, see Material and Methods.

Figure 4

A: Bar graphs are illustrating the effects of MEK inhibitor UO126 (2.5 nmol, icv) or vehicle (4 % DMSO, icv) on ethanol (1.5 g/kg, i.p)-induced c-fos mRNA in different forebrain regions of rats with a history of ethanol dependence and age-matched controls. Corrected p-values: *p < 0.05, **p < 0.01 vehicle treated ethanol exposed vs vehicle treated control group, #p < 0.05, ###p < 0.001 UO treated group vs corresponding ethanol exposed or control group. B: Bright-field microphotographs from autoradiograms of in situ hybridization are showing the effects of MEK inhibitor UO126 (2.5 nmol, icv) on ethanol (1.5 g/kg, i.p.)-induced c-fos mRNA levels in the OFC, SO and PVN region of rats with a history of ethanol dependence and vehicle treated age-matched control rats, scale bar = 1 mm; for abbreviations see figure 1, for details on treatment, see Material and Methods.

Figure 5

Schematic representation summarizing the results on ethanol-induced c-fos expression and its interaction with the ERK1/2 kinase signaling pathway in brain regions related to the extended amygdala (adapted from(Heimer, 2003)). Only those regions are shown that react in non-dependent rats (left) with increased c-fos to an acute ethanol challenge. Under non-dependent conditions, ethanol-induced c-fos expression was generally not affected by MEK inhibition, with the exception of the MeA (red circles), and to a lesser extend the PVN. Post-dependent rats (right) show reduced c-fos induction in the prefrontal cortex (Cg, IL, OFC) and the PVN upon ethanol challenge (indicated by short bold arrows) compared to non-dependent controls which probably is a correlate of tolerance to the drug. UO126 markedly increased c-fos expression in OFC and AcbSh, key components of circuitry mediating positive drug reinforcement, demonstrating a recruitment of an ERK mediated inhibitory regulation in the post-dependent state. Thus, positive MEK/ERK-ethanol interactions are related to the central division and negative interactions to the medial division of the extended amygdala. Beside anatomical evidences of a division of the extended amygdala into a central- and medial part (Alheid, 2003; Heimer, 2003) our results support the idea of a corresponding functional divisions, i.e. devaluation of ethanol as a reinforcer and tolerance to its aversive actions, respectively, which may both take part in the development of ethanol dependence. Brain regions anatomically or functionally related to the central or the medial divisions of the extended amygdala are colored in yellow or in blue, respectively, and their efferents/afferents are indicated as arrows with respective color. Black arrows indicate other connections between the regions. For abbreviations see figure 1, and for details on treatment, see Material and Methods. ¹(van Dongen et al, 2005), ²(Hoover & Vertes, 2007), ³(McDonald et al, 1996), ⁴(Reynolds & Zahm, 2005), ⁵(Schoenbaum & Setlow, 2003), ⁶(Silverman et al, 1981), ⁷(Vertes, 2004), ⁸(Vertes, 2006)