Changes in gene expression in regions of the extended amygdala of alcohol-preferring rats after binge-like alcohol drinking

Abstract

The objective of this study was to determine time-course changes in gene expression within two regions of the extended amygdala after binge-like alcohol drinking by alcohol-preferring (P) rats. Adult male P rats were given 1-h access to 15 and 30% ethanol three times daily for 8 weeks. Rats (n = 10/time point for ethanol and n = 6/time point for water) were killed by decapitation 1, 6, and 24 h after the last drinking episode. RNA was prepared from individual micropunch samples of the nucleus accumbens shell (ACB-shell) and central nucleus of the amygdala (CeA); analyses were conducted with Affymetrix Rat Genome 230.2 GeneChips. Ethanol intakes were 1.5-2 g/kg for each of the three sessions. There were no genes that were statistically different between the ethanol and water control groups at any individual time point. Therefore, an overall effect, comparing the water control and ethanol groups, was determined. In the ACB-shell and CeA, there were 276 and 402 probe sets for named genes, respectively, that differed between the two groups. There were 1.5-3.6-fold more genes with increased expression than with decreased expression in the ethanol-drinking group, with most differences between 1.1- and 1.2-fold. Among the differences between the ethanol and water control groups were several significant biological processes categories that were in common between the two regions (e.g., synaptic transmission, neurite development); however, within these categories, there were few genes in common between the two regions. Overall, the results indicate that binge-like alcohol drinking by P rats produces region-dependent changes in the expression of genes that could alter transcription, synaptic function, and neuronal plasticity in the ACB-shell and CeA; within each region, different mechanisms may underlie these alterations because there were few common ethanol-responsive genes between the ACB-shell and CeA.

Fig. 1

Ethanol intakes (g/kg) by P rats for each of the three one-hr access periods to 15 and 30% ethanol during the dark cycle. Ethanol was given consecutive 5 days each week over the 8 weeks of access. Data are presented in 5-day blocks for each 1-hr period of access. Data are the means ± SEM (n = 30).

Fig. 2

Abridged Ingenuity® Pathways Analysis of effects of ethanol drinking in the nucleus accumbens shell of P rats showing increased expression of genes in calcium/calmodulin and glutamate signaling pathways. Red indicates increased expression, green indicates reduced expression (only Wipf1 had reduced expression), and clear symbol indicates genes that were not identified as differentially expressed, but were linked to multiple genes that had changed significantly. Abbreviations: Akap9 – A kinase anchor protein; Basp1 - brain abundant signal protein; Cacna1h – calcium channel T type; Calm1,2,3 – calmodulin 1,2,3; Camk – calcium/calmodulin-dependent protein kinase; Clic5 – chloride intracellular channel; Fbxo2 – F-box protein; Gria1 – AMPA1 receptor; Grin1 – NMDA1 receptor; Kcnj16 – K+ inwardly-rectifying channel; Myo5a – myosin Va; Myrip – myosin & Rab interacting protein; Nrxn1 – neurexin 1; Pcnt –pericantrin; Pka – protein kinase; Scf - (Skp1-cullen-1, F-box) ubiquitin ligase; Skp1 – S-phase kinase protein; Snta – syntrophin acidic; Wipf1 – Wiskott-Aldrich syndrome-like interacting protein. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 3

Abridged Ingenuity® Pathways Analysis of effects of ethanol drinking in the central nucleus of the amygdala of P rats showing increased expression of genes in glutamate signaling pathways and alterations in gene expression in related pathways. Red indicates increased expression, green indicates decreased expression, and clear symbol indicates genes that were not identified as differentially expressed, but were linked to multiple genes that had changed significantly. Abbreviations: 14-3-3 – tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein; Ampar – AMPA receptors; C1qbp – complement component 1, q subcomponent binding protein; Cybb – cytochrome b-245, beta polypeptide; Eif2s2 – eukaryotic translation initiation factor 2, subunit 2 (beta); Gabar-a – GABA receptor associated protein; Gabra1 – gamma-aminobutyric acid (GABA-A) receptor, subunit alpha 1; Gabrg1 – gamma-aminobutyric acid (GABA-A) receptor, subunit gamma 1; Gabrg2 – gamma-aminobutyric acid (GABA-A) receptor, subunit gamma 2; Gnai2 – guanine nucleotide binding protein (G protein), alpha inhibiting 1; Gri – glutamate receptor, ionotropic; Gria1, 2, 3 – glutamate receptor ionotropic AMPA 1 (alpha 1, 2, 3); Grik2 – glutamate receptor, ionotropic, kainate 2 (beta 2); Grik5 – glutamate receptor, ionotropic, kainate 5 (gamma 2); Mek1/2 – mitogen-activated protein kinase kinase ½; Mgmt – O-6-methylguanine-DNA methyltransferase; Nmnat1 – nicotinamide nucleotide adenyltransferase 1; Nsf –N-ethylmaleimide sensitive fusion protein; P2ry1 – purinergic receptor P2Y, G-protein coupled 1; Pkca – protein kinase C alpha; Rgs2 – regulator of G protein signaling 2; Rgs 7 – regulator of G protein signaling 7; Stx3 – syntaxin 3. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)