Loss of Basal Forebrain Cholinergic Neurons Following Adolescent Binge Ethanol Exposure: Recovery With the Cholinesterase Inhibitor Galantamine

Abstract

Binge drinking and alcohol abuse are common during adolescence and cause both cognitive deficits and lasting cholinergic pathology in the adult basal forebrain. Acetylcholine is anti-inflammatory and studies using the preclinical adolescent intermittent ethanol (AIE; 5.0 g/kg, i.g., 2 day on/2 day off from postnatal day [P]25 to P54) model of human adolescent binge drinking report decreased basal forebrain cholinergic neurons (BFCNs) and induction of proinflammatory genes that persist long into adulthood. Recent studies link AIE-induced neuroimmune activation to cholinergic pathology, but the underlying mechanisms contributing to the persistent loss of BFCNs are unknown. We report that treatment with the cholinesterase inhibitor galantamine (4.0 mg/kg, i.p.) administered during AIE (i.e., P25–P54) or following the conclusion of AIE (i.e., P57–P72) recovered the persistent loss of cholinergic neuron phenotype markers (i.e., ChAT, TrkA, and p75NTR) and somal shrinkage of residual ChAT + neurons known to persist in AIE-exposed adults. Galantamine treatment also recovered the AIE-increased expression of the proinflammatory receptors TLR4 and RAGE, the endogenous TLR4/RAGE agonist HMGB1, and the transcription activation marker pNF-κB p65. Interestingly, we find BFCNs express TLR4 and RAGE, and that AIE treatment increased pNF-κB p65 expression in adult ChAT + IR neurons, consistent with intracellular HMGB1-TLR4/RAGE signaling within BFCNs. AIE increased epigenetic transcription silencing markers (i.e., H3K9me2 and H3K9me3) in the adult basal forebrain and H3K9me2 occupancy at cholinergic phenotype gene promoters (i.e., ChAT and TrkA). The finding of no AIE-induced changes in total basal forebrain NeuN + neurons with galantamine reversal of AIE-induced ChAT + neuron loss, TLR4/RAGE-pNF-κB p65 signals, and epigenetic transcription silencing markers suggests that AIE does not cause cell death, but rather the loss of the cholinergic phenotype. Together, these data suggest that AIE induces HMGB1-TLR4/RAGE-pNF-κB p65 signals, causing the loss of cholinergic phenotype (i.e., ChAT, TrkA, and p75NTR) through epigenetic histone transcription silencing that result in the loss of the BFCN phenotype that can be prevented and restored by galantamine.

Galantamine prevents and reverses neuroimmune induction and loss of adult hippocampal neurogenesis following adolescent alcohol exposure

“…Loss of these cholinergic projections after AIE could play a role in hippocampal induction of inflammatory responses due to loss of acetylcholine activation of nicotinic and muscarinic receptors on both neurons and glia. Recent evidence suggests that AIE-induced loss of expression of cholinergic markers in the basal forebrain can be reversed by chronic treatment with galantamine [19]. These findings support the hypothesis that AIEinduced loss of cholinergic anti-inflammatory signaling contributes to the induction of hippocampal proinflammatory genes and the consequential loss of hippocampal neurogenesis.…”

“…Rats were weighed on the first of every 2 consecutive days of gavage treatment to ensure accurate dosing as adolescence is a period of rapid body growth and weight gain. All rats gained weight across adolescence, but neither AIE nor galantamine impacted bodyweight in either the prevention study (study 1, p = 0.3 and 0.2, respectively) or restoration study (study 2, p = 0.07, 0.8, respectively) [for more details see [19]].…”

Adolescent Intermittent Ethanol (AIE) Enhances the Dopaminergic Response to Ethanol within the Mesolimbic Pathway during Adulthood: Alterations in Cholinergic/Dopaminergic Genes Expression in the Nucleus Accumbens Shell

“…Daily administrations of an α7 NAchR agonist (AR-R1779) during early adolescence (only 6 days of exposure) resulted in adult EtOH consumption similar to rats given AIE exposure (enhanced acquisition and relapse [31]). Our results were conceptually replicated with the recent report that pretreatment with the cholinesterase inhibitor galantamine (countering the activity of the α7 receptor) prevented ABAE-induced increase in the expression of TLR4, RAGE, HMGB1, and pNF-κB p65 during adulthood [62]. Donepezil can reverse AIE-induced decreases in dendritic spine density and expression of the Fmr1 gene in the hippocampus [14].…”

Cholinergic and Neuroimmune Signaling Interact to Impact Adult Hippocampal Neurogenesis and Alcohol Pathology Across Development

“…For example, NFĸB p65 increases gene transcription of a variety of proinflammatory genes, including TLR4, IL-1β, TNFα, CCL2, and COX-2. However, in the presence of HMGB1, NFĸB p50 forms a repressome complex with G9a (Abhimanyu et al, 2021), driving H3K9 methylation at the ChAT and TrkA promotors, reducing cholinergic gene transcription and suppressing the cholinergic neuronal phenotype (Vetreno et al, 2020;Crews et al, 2021). and it is unknown whether these findings extend to females.…”