Associate editor: D.M. Lovinger
Intracellular signaling pathways that regulate behavioral responses to ethanol

https://doi.org/10.1016/j.pharmthera.2005.07.004Get rights and content

Abstract

Recent evidence indicates that ethanol modulates the function of specific intracellular signaling cascades, including those that contain cyclic adenosine 3′, 5′-monophosphate (cAMP)-dependent protein kinase A (PKA), protein kinase C (PKC), the tyrosine kinase Fyn, and phospholipase D (PLD). In some cases, the specific components of these cascades appear to mediate the effects of ethanol, whereas other components indirectly modify responses to ethanol. Studies utilizing selective inhibitors and genetically modified mice have identified specific isoforms of proteins involved in responses to ethanol. The effects of ethanol on neuronal signaling appear restricted to certain brain regions, partly due to the restricted distribution of these proteins. This likely contributes specificity to ethanol's actions on behavior. This review summarizes recent work on ethanol and intracellular signal transduction, emphasizing studies that have identified specific molecular events that underlie behavioral responses to ethanol.

Introduction

Alcohol abuse and alcoholism are significant public health issues throughout the world. In the United States alone, they affect roughly 14 million people, costing ∼ US$184 billion a year (Shalala, 2000). Despite the scale of the problem, there are few pharmacological treatments for alcohol use disorders. An important approach to rational medications development is to understand the mechanisms by which ethanol alters nervous system function. Most research has focused on ethanol modulation of cell surface proteins and much knowledge has been gained regarding the actions of ethanol at specific receptor- and voltage-gated ion channels and transporters. Less well studied are ethanol's effects on intracellular signaling cascades. Early experiments in this field were predominantly focused upon in vitro treatment of cultured cells with relatively high concentrations of ethanol (up to 200 mM), based upon blood alcohol concentrations found in human alcoholics (Diamond and Messing, 1994, Messing and Diamond, 1997). These cultured cells were then treated with pharmacological kinase inhibitors, many of which have subsequently been shown to exhibit a broader target range than originally thought (Alessi, 1997, Leemhuis et al., 2002, Voutilainen-Myllyla et al., 2003, Thomas et al., 2004, Dokladda et al., 2005). Recent research has taken advantage of more sophisticated molecular techniques to focus attention on individual proteins and their response to alcohol in specific brain regions of transgenic and normal animals. In this review, we will examine research aimed at understanding how ethanol's effects on intracellular signaling cascades are transformed into specific outputs, emphasizing research that encompasses both in vitro model systems and in vivo behavioral studies.

Section snippets

Adenylyl cyclases and protein kinase A

Several neurotransmitters and neuromodulators act at G-protein coupled receptors (GPCRs), which either increase (Gs-coupled) or decrease (Gi-coupled) the activity of adenylyl cyclase (AC), depending on the type of G protein that they stimulate (reviewed in Mailliard & Diamond, 2004). AC generates cyclic adenosine 3′, 5′-monophosphate (cAMP), which is required to activate cAMP-dependent protein kinase A (PKA). Increased PKA activity results in the phosphorylation of downstream targets, including

Fyn kinase

Another mechanism that modulates ethanol inhibition of NMDA receptors involves the Src family tyrosine kinase Fyn (Fig. 2). In hippocampal slices, acute tolerance to ethanol inhibition of NMDA receptors begins ∼ 15 min after an initial exposure to ethanol. This is associated with increased tyrosine phosphorylation of the NR2B subunit of NMDA receptors, which enhances NMDA receptor function and counters the inhibitory effects of ethanol (Miyakawa et al., 1997). Both ethanol-stimulated NR2B

Protein kinase C

Protein kinase C (PKC) is a family of phospholipid-dependent kinases that transduce signals involving lipid second messengers (Nishizuka, 2001, Olive and Messing, 2004). The family is encoded by 9 genes and is divided into 3 subclasses based on differences in structure and response to second messengers. Conventional cPKCs (α, β, and γ) are activated by calcium and diacylglycerol, novel nPKCs (δ, ε, η, and θ) are activated by diacylglycerol but not by calcium, and atypical aPKCs (ζ and τ/λ) are

Phospholipase D

Although phospholipase D (PLD) is one of very few enzymes that can utilize ethanol as a substrate, there are few studies examining the interaction between PLD and ethanol in neuronal systems, and none that have examined the potential consequences of this on behavior. Under normal conditions, stimulation of PLD generates phosphatidic acid (PA) through the hydrolysis of membrane phospholipids, mainly phosphatidylcholine. PA is a second messenger for several proteins, including Raf-1, aPKCζ,

Discussion

It is clear from in vitro studies in cultured cells that ethanol modulates signal transduction through cascades that involve PKA and PKC. In vivo studies with genetically engineered mice have confirmed that components of these signaling cascades modulate behavioral responses to ethanol. In the brain, ethanol's effects are observed in specific brain regions depending on the presence of specific isozymes and other signaling molecules. This is most clearly demonstrated for down-regulation of NMDA

Acknowledgments

This work was supported by US Public Health Service Grants AA08117 and AA13588 from the National Institute on Alcohol Abuse and Alcoholism to R. O. Messing and by funds provided by the State of California for medical research on alcohol and substance abuse through the University of California at San Francisco.

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      Conversely, a reduction in PKC-dependent GABAAR phosphorylation disinhibits receptor binding to AP-2 to enable GABAAR endocytosis (Terunuma et al., 2008). Interestingly, EtOH alters PKA expression and translocation (Diamond and Gordon, 1994; Newton and Messing, 2006) and chronic PKA activation in cerebellar granule cells increases α1 subunit membrane expression (Ives et al., 2002), suggesting that PKA might also regulate GABAAR membrane localization after chronic EtOH exposure. Recently, the Morrow laboratory reported that either PKA activation or PKC inhibition prevented ethanol-induced increases in α4 subunit expression and decreases in the decay of GABA mIPSCs, whereas PKA inhibition had no effect (Bohnsack et al., 2016; Carlson et al., 2016a).

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