Prenatal and Adolescent Alcohol Exposure Programs Immunity Across the Lifespan: CNS-mediated Regulation

Overview of innate immunity

The ability to defend against a seemingly endless onslaught of rapidly mutating, foreign pathogens has led to the evolution of an enormously complex immune system in mammalian species. For this reason, the immune system has evolved both a general, “innate” immune system that provides a universal, first line of defense against most major categories of infectious disease, as well as a more specific, “adaptive” immune response that allows for targeted defense and durable immunity against pathogen re-infection for the long term. These two distinct branches (or phases) of immunological function work together to ensure robust host defense. Importantly, the CNS orchestrates host immunity through a wide variety of neural and hormonal pathways (Maier & Watkins, 1998; Glaser & Kiecolt-Glaser, 2005), at least in part because of the relative immunological privilege of the CNS conferred by the Blood-Brain Barrier (BBB; see Vore & Deak, 2022 for a recent review). For instance, key immunological organs in the body such as the liver, spleen, and lymph nodes are all densely innervated, providing a direct pathway for neural-immune communication (Dantzer, 2018), while at the same time, many cranial nerves are surrounded by Nerve-Associated Lymphoid Tissue (NALT), which express receptors for cytokines and transduce immunological signals into neural impulses (Wulfing et al., 2018). Alternatively, circulating immune cells and the cytokines they produce can bind to endothelial cells of the cerebrovasculature, and propagate immunological signaling cascades across the vascular wall into the brain parenchyma, alerting the CNS of a peripheral infection (Serrats et al., 2010). To a lesser extent, immune-to-brain communication also occurs through active transport of cytokines across the BBB, or through passive diffusion of cytokines in areas where the BBB is weak or absent (Banks, 2005). Finally, activation of immune-signaling pathways within the CNS activates the hypothalamic-pituitary-adrenal axis leading to rapid release of corticosteroids (Turnbull and Rivier, 1999), which have potent immunomodulatory properties (Cain & Cidlowski, 2017). Thus, when a localized region of the body encounters a pathogen, the infection is detected rapidly by the CNS through a variety of mechanisms, and the CNS coordinates a set of physiological, neuroendocrine and behavioral alterations referred to as the “sickness syndrome” that is, ideally, proportionate to the magnitude of the threat. Behavioral components of the sickness syndrome include anorexia, fatigue, hyperalgesia, and fever, which together optimize immune function for effective host defense (Dantzer, 2004; Saper et al., 2012; see Figure 2A for a visual representation of the relationships between these mechanisms).

In many cases, the innate immune system is activated through Pattern Recognition Receptors (PRRs). Toll-Like Receptors (TLRs) are a family of PRRs that can be embedded in the cell membrane (e.g. TLR2, TLR4) or on intracellular endosomes (e.g. TLR 3,7,8). TLRs recognize general molecular motifs that are common to classes of pathogens, termed Pathogen-Associated Molecular Patterns (PAMPs; see Evans et al., 2015 for review). For instance, viral-derived double-stranded RNA (dsRNA) is detected by TLR3, whereas ssRNA is detected by TLR7 and TLR8 (Iwasaki et al., 2014), triggering the expression of classic proinflammatory cytokines (TNFα, IL-1β, IL-6) as well as Type 1 Interferons (predominantly IFNα and IFNβ). Other members of the TLR family are sensitive to distinct pathogen motifs. For instance, TLR4 recognizes lipopolysaccharide (LPS), a component of the cell wall of gram-negative bacteria (Molteni et al., 2016), whereas TLR5 recognizes bacterial flagellin (Yoon et al., 2012). When bound to either TLR3 or TLR4, signaling cascades through NFkB as well as TRIF/Myd88 pathways are activated (Erridge, 2010; Kawasaki and Kawai, 2014), eliciting downstream induction of immune-related genes essential for coordination of host defense processes. Transcription factors downstream of the Myd88 and TRIF-dependent pathways include cytokines and chemokines (e.g. TNFα, IL6, IL-1β). In addition, cytokines forward propagate the induction of other inflammatory mediators in order to defend the host through inflammatory processes (Saper et al., 2012). In this way, the initial response to pathogen exposure is detected by TLRs, which elicit rapid induction and release of pro-inflammatory cytokines and a system-wide reaction referred to as the Acute Phase Response.

The Acute Phase Response entails a series of systemic events designed to protect the organism from further tissue damage or infection. Pathogenic intruders typically enter the organism through the skin or mucosal membranes. TLR binding on or within intrinsic immune cells (e.g., macrophages, mast cells, dendritic cells, etc.) induces rapid expression and secretion of chemokines and other cytokines to attract distal immune cells to the site of injury. To facilitate leukocyte recruitment, local histamine release disrupts tight junctions between endothelial cells, promoting extravasation of immune cells from the bloodstream into the infected tissue, and ultimately contributing to edema (swelling; Presta et al., 2018). Increased expression of adhesion molecules such as ICAM1 on the blood side of the vasculature facilitates capture of circulating immune cells for extravasation into the infected or damaged tissue (Muller, 2002; Harjunpaa et al., 2019). As cytokine concentrations rise and spread into systemic circulation, circulating cytokines reach the liver, where they initiate a rapid shift in liver metabolic activity. Under normal conditions, the liver is responsible for the vast majority of plasma protein production, especially albumin and other carrier proteins (Schreiber, 1978). During the acute phase response, however, Kupffer cells (i.e., resident macrophages of the liver) produce IL-6 and other cytokines that bind to cognate receptors on liver hepatocytes through paracrine signaling, causing hepatocytes to curtail production of carrier proteins in favor of producing a family of proteins collectively referred to as acute phase proteins (Norris et al., 2014). Thus, in this scenario, the rapid drop in normative carrier proteins such as albumin and corticosteroid binding globulin classifies these protein alterations as acute phase negative proteins, whereas other proteins that rapidly increase during the acute phase response are referred to as acute phase positive proteins, including C-reactive protein (CRP), Serum Amyloid-A, α1-acid glycoprotein, Mannose binding lectin, haptoglobin, and various complement proteins, etc. (Jain et al., 2011). The general role of these acute phase positive proteins is to opsonize foreign material, targeting it for fast recognition, phagocytosis, and clearance by immune cells. The liver is also densely innervated by the vagus nerve and the celiac ganglion, two nerves that transduce cytokine signals into neural impulses (Goehler et al., 2000). Indeed, the vagus nerve appears to be particularly valuable in immune-to-brain communication (a) early in the propagation of an inflammatory response (Pavlov and Tracey, 2012); (b) during insults that directly involve the liver and other visceral organs innervated by the vagus (Romanovsky et al., 2000); and (c) under low-to-moderate pathogenic conditions where circulating cytokines remain modest (Hansen et al., 2001; Goehler et al., 1999). In these ways, the liver plays an instrumental role in host defense processes, and physiological conditions which disrupt liver function -- such as chronic use and misuse of alcohol -- sets the stage for impaired host defense.

Similarly, the gastrointestinal (GI) tract and the human microbiome at large have gained attention over recent years for their collective role in building and maintaining innate immunity. The human GI tract houses an estimated 10¹⁴ microorganisms of more than 2,000 different species (Lazar et al., 2018) and begins developing upon passage through the birth canal, with subsequent honing through the acquisition of other microbes, immune cells, metabolites, etc. via breastfeeding (Belkaid and Hand, 2014). The symbiotic relationship between these commensal species and the host has evolved to prime and maintain immune responsiveness at baseline and during infection and does so through a variety of mechanisms. The GI tract, along with the skin and the respiratory system, is constantly exposed to external pathogens and microorganisms, and therefore actively carries out host defense processes in order to maintain homeostasis and control inflammation (Yuan and Walker, 2004). Thus, maintenance of the gut epithelium under normal conditions is crucial to the containment of the commensal microbiome and the sequestration of invading pathogens. This reinforcement begins early in life, as the interaction of endothelial PRRs with microbial products like LPS effectively ‘trains’ the gut barrier to distinguish between resident bacteria and foreign pathogens (Belkaid and Hand, 2014; Lazar et al., 2018). Intestinal epithelial cells also produce microbial peptides, such as α-defensins and RegIIIγ, that contribute to barrier integrity via the direct execution of antimicrobial processes or simply accumulating in the mucus to create a physical barrier (Belkaid and Hand, 2014). Regulation of commensal species is similarly achieved through the actions of Secretory Immunoglobulin A (sIgA), which is produced in lymphoid tissue and contributes to both the compartmentalization of these bacteria and the host’s inflammatory response to their presence (Macpherson and Slack, 2007). While the aforementioned mechanisms contribute to the maintenance of steady-state protection against commensal bacteria, these species themselves also play an important role in protection of the host organism against infection. Commensals can aid during infection by (1) competing with pathogens for nutrients (referred to as colonization resistance), (2) creating a hostile environment for pathogen replication, and (3) secreting antimicrobial peptides that inhibit pathogen survival (Belkaid and Hand, 2014). Microbiota in the GI tract have also been shown to interact directly with immune cells; for example, the interaction of monocytes with the gut microbiome has been shown to stimulate the production of PGE₂ and in turn activate neutrophils during acute infection (Grainger et al., 2013). Thus, an appropriately fortified intestinal barrier and a healthy microbiome are imperative to both baseline homeostatic function and the execution of the innate immune response to infection.

Immune dysfunction in individuals diagnosed with AUD

A significant challenge for studies examining health outcomes associated with alcohol consumption across the lifespan is the wide variability of alcohol consumption patterns across individuals. The Diagnostic and Statistical Manual of Mental Disorders (DSM-V) characterizes AUD as the uncontrolled consumption of alcohol that clinically impairs an individual from healthy functioning (American Psychiatric Association, 2013). A diagnosis requires the manifestation of at least two of eleven criteria defining occupational dysfunction, alcohol cravings, tolerance, and withdrawal (see Witkiewitz, Litten, and Leggio, 2019 for further review). The number of criteria met determines a subclassification of mild, moderate, or severe AUD. These diagnostic categories begin to acknowledge the wide variation in alcohol use patterns across individuals, the quantity and pattern of which may determine the subsequent effects on immune function (Barr et al., 2015; Sureshchandra et al., 2019). Thus, a challenge for the field has been a lack of consensus for definitions of varying intensities of drinking among human alcohol studies. For instance, moderate alcohol consumption may include up to one drink per day for females and one drink per day for males according to the Dietary Guidelines for Americans (USDA and USDHHS, 2020). The NIAAA defines binge drinking as consumption of four drinks for females and five drinks for males in a two hour period elevating blood alcohol concentrations (BAC) to at least 0.08%. The Substance Abuse and Mental Health Services Administration (SAMHSA, 2021) defines heavy drinking as binge drinking on five or more days in the past 30 days. Most commonly, clinical alcohol studies utilize the Alcohol Use Disorder Identification Test (AUDIT; Babor et al., 2001) as a tool for screening alcohol use patterns and AUD. Further complicating the matter is the general inaccuracy of self-reported measures of lifetime alcohol use, variety in definitions of different modes of drinking, and failure to report age of drinking onset in clinical alcohol use studies (Calvo et al., 2020).

With these definitions in mind, Figure 1 reports the prevalence of drinking across the lifespan as reported in the most recent data from SAMHSA (SAMHSA, 2020), which raises several issues we feel are critically relevant for determining the influence of alcohol on host immunity. First and foremost, the literature has struggled to develop universal operational definitions for casual drinking, leading to difficulties comparing studies across recent decades. However, using guidelines from the American Psychological Association (APA) and the preponderance of studies published to date, it seems reasonable to conclude that most studies evaluating the impact of alcohol consumption on host immunity (and other physiological processes, for that matter) have focused on high dose alcohol effects, defined as heavy drinking, AUD diagnosis, or regular binge drinking, which seem to produce a wide range of inflammatory disorders. Figure 1 highlights these risk factors in the “red zone” to recognize the clinically-defined, highest risk groups for disrupted host immunity. Yet, a substantial number of individuals consume alcohol intermittently and may experience more subtle or more transient effects on host immunity, falling into a risk group in which vulnerability to alcohol-mediated disruption of immunity may be less evident, termed the “gray zone”. Thus, the overall influence of alcohol on host immunity may be under-estimated substantially by focusing on the most severe cases of alcohol misuse. As a related matter, we would note that studies examining alcohol effects on host immunity do not readily discriminate between the cumulative impact of ethanol consumption across the lifespan, and the acute biological effects of alcohol that might influence host immunity at the time of infection. In reviewing studies, we have attempted to be clear on these issues in our description of study outcomes.

With this in mind, chronic alcohol use has been shown to affect most major aspects of immunological function, though many of the mechanisms underlying these changes remain unclear. Alcohol induces expression of TLR3 and 4 through long-term exposure, creating lasting changes in PRR expression. Increased expression of TLR-3, 2, 4, and HMGB1 (a PAMP that binds to and activates TLR4) have been found in postmortem tissue from the orbitofrontal cortex and hippocampus (Crews et al., 2013) of alcoholics and is associated with greater alcohol dependence (Ponomarev et al., 2012; Crews et al., 2013). Furthermore, activation of TLR4 through an alcohol-dependent increase in HMGB1 following a lifetime history of alcohol exposure seems to be responsible for increased neuronal hyperexcitability and cell death seen in those with alcohol-related brain damage (Crews and Vetreno, 2016). In this sense, it is notable that HMGB1 is released from activated neurons and promotes inflammation through TLR4-mediated mechanisms (Yang et al., 2021). Increased TLR4 potentiated activation of the NFkB pathway in human alcoholics (Crews et al., 2013; Zou and Crews, 2006). In addition to increased risk of alcohol-related brain damage, oxidation of ethanol by CYP2E1 increases synthesis of reactive oxygen species (ROS) (Koop, 2006). Increased levels of ROS may ultimately lead to adverse inflammatory consequences such as cancer, atherosclerosis, inflammation, cirrhosis (see Barr et al., 2015 for review). The increased activity of these innate immune molecules leads to the gradual accumulation of ROS and may contribute to the lasting neuroinflammation and neurodegeneration in alcoholics (Liu et al., 2017).

There seems to be a bi-directional relationship between the cellular components of the innate immune system and increased alcohol intake, where increased consumption of alcohol activates the innate immune system and specific components of the innate immune system increase alcohol consumption (see Robinson et al., 2014 for review). Due to increased expression of TLR4 following chronic alcohol consumption (Liu et al., 2011), it was initially proposed that TLR4 activation would lead to increased consumption of alcohol (Wu et al., 2012; Bajo et al., 2016), but subsequent studies do not concur with these initial findings (Harris et al., 2017; Blednov et al., 2017), raising questions about the precise role of TLR4 as a moderator of alcohol consumption. Other studies have also examined a potential role for TLR3 as a moderator of alcohol consumption, showing that activation or inhibition of TLR3 produced bi-directional alterations in ethanol self-administration (McCarthy et al., 2018; Randall et al., 2019; Warden, 2019a; Warden, 2019b). Chronic ethanol consumption increased TIR-domain-containing adaptor-inducing interferon-β (TRIF) pathway components in mice (McCarthy et al., 2018). Additionally, activation of the TRIF pathway increased self-administration of ethanol in Myd88 −/− mice (Warden 2019b). Together, these findings support a feed-forward framework for alcohol misuse, where chronic alcohol consumption increases TLR3 and TLR4 expression, and exposure to antigens/pathogens that activate these PRRs would propel further drinking, and so on. This underscores the importance of understanding how alcohol exposure alters immune function, and in particular, whether there are specific developmental periods (prenatal, adolescent) during which alcohol exposure might exert long-lasting effects on immune function.

Excellent reviews of alcohol effects on immune function are available already (Szabo & Saha, 2012), so our focus here will be to briefly frame the key mechanisms, as a means to provide a foundation for understanding age-specific vulnerabilities to alcohol. Under normal conditions (no alcohol present), circulating monocytes recognize and bind LPS through TLR4 and are activated to form mature macrophages once bound. MyD88 pathways downstream of TLR4 ultimately signal transcription for the nuclear factor kappa B (NFκB) whose activation leads to enhanced transcription of pro-inflammatory cytokines and chemokines (Blackwell and Christman, 1997; Hoffman et al., 2006). Importantly, these natural functions of monocytes/macrophages are severely impacted by either acute or chronic alcohol consumption, effects that appear to occur in both a dose- and time-dependent manner (Barr et al., 2015; Sureshchandra et al., 2019). In response to alcohol, both in vitro and in vivo studies of isolated peripheral blood mononuclear cells (PBMC) from healthy humans or those affected by AUD begin to help identify mechanisms by which different intensities of alcohol exposure will affect lifetime immunity. Isolated PBMCs from healthy males and females (no history of alcohol abuse; < 6 drinks/week) that were incubated in 25 or 50 mM of ethanol expressed endotoxin tolerance where pro-inflammatory cytokine release was inhibited and release of heat shock proteins was increased (Muralidharan et al., 2014). Even when challenged with LPS, PBMCs from healthy humans inhibited the release of pro-inflammatory cytokines (IL-1β; Mandrekar et al., 2009) after acute exposure to 25mM of ethanol. These findings are indicative of the general pattern of effects often observed in studies of alcohol effects on host immune function.

Consistent with the effects of alcohol on circulating monocytes/macrophages, humans with a history of chronic heavy drinking were more likely to develop a wide range of organ-related pathologies, at least in part due to actions of ethanol on monocytes/macrophages resident to individual organs such as the liver (Edmondson, 1980). Other vulnerabilities to the liver arise from alcohol metabolites such as acetate and acetaldehyde, both of which can increase risk for hepatocellular carcinoma (HCC; Grewal and Viswanathen, 2016) and alcoholic liver disease (ALD; Szabo and Bala, 2010). Inflammation derived from LPS in microflora of the gut evidently induced steatosis, inflammation, and ultimately cirrhosis in those with ALD (see Arab, Arrese, and Shah, 2020 for review). Alcohol increased gut permeability in even healthy humans following an acute heavy drinking episode (Bode, Kugler, Bode, 1987; Bala et al., 2014; de Jong et al., 2015). Chronic alcohol consumption increased the quantity of gut-derived microflora and their subsequent LPS production (Bode et al., 1984; Hauge, Persson, and Danielsson, 1997). Indeed, chronic alcohol exposure has been shown to increase endotoxin in blood (Walter et al., 2017), though other studies have reported no effect of acute alcohol challenge on plasma endotoxin (Doremus-Fitzwater et al., 2015), suggesting that these effects may be dependent on the dose and/or route of ethanol administration. Alcohol abstinence programs of 12, 6, and 3 weeks have been found to decrease baseline plasma LPS levels among excessive drinkers; however, there is conflicting evidence as to whether such programs also lead to a decrease in baseline pro-inflammatory cytokines (Leclercq et al., 2012; Liangpunsakul et al., 2017; Donnadieu et al., 2018). Importantly, the disrupted gut barrier following chronic heavy drinking promoted activation of Kupffer cells which led to subsequent increases in TNFα and superoxide (see Wheeler, 2003 for review). Ultimately, chronic activation of Kupffer cells through a gut-derived, LPS-induced proinflammatory state seems to be responsible for the pathophysiology of ALD (Takei et al., 2004). Alcoholic hepatitis seems to develop from similar microbial translocation-mediated mechanisms (see Mathurin and Lucey, 2012 for review).

A similar monocyte/macrophage-mediated cascade occurs due to actions of ethanol on alveolar macrophages in the lung, contributing to acute respiratory distress syndrome (ARDS; Thakur et al., 2009). These alcohol-induced changes in viability of immune cells of the lung seem to be controlled by the increased expression of cytokines and ROS responsible for oxidative stress damage to the lungs (Liang et al 2012; Mehta and Guidot, 2012), as well as the increased recruitment of neutrophils by alveolar macrophages providing additional damage (Dorio and Forman, 1988). Additionally, heavy alcohol consumption may play a critical role in lung cancer diagnosis, in a dose-dependent manner (Korte, 2002), even after controlling for cigarette use (see Bandera, Freudenheim and Vera, 2001 for review). Damage to the lungs following chronic heavy drinking not only decreases integrity of lung function, but leaves the host more vulnerable to risk of other respiratory infections (see Simet and Sisson, 2015 for an extensive review on consequences of chronic alcohol consumption on lung health). Considering the critical role of the liver in initiation of the acute phase response of host defense, as well as the lungs as a point of entry for respiratory pathogens, alcohol-mediated modification of intrinsic immune cells of these organs are particularly problematic, and must be taken into account when considering host defense processes.

Damage to internal organs resulting from chronic heavy drinking, in turn, promotes susceptibility to both bacterial and viral infections (eg. HIV, viral respiratory infections, tuberculosis, and pneumonia; Rehm et al., 2009; Baum et al., 2010; Szabo and Saha, 2012; Justice et al., 2016; Osna et al., 2022). For example, cases of Mycobacterium tuberculosis, Klebsiella and Streptococcus pneumoniae are more prevalent among chronic alcoholics (Rehm et al., 2009; Bhatty et al., 2011; Jong et al., 1995). Hepatitis C (HCV) is more prevalent among alcoholics, and seems to further propagate HCV replication in hepatocytes (Hou et al., 2013). The progression of liver disease induced by hepatitis B (HBV) diagnosis has been found to be further accelerated by chronic alcohol use (Gao and Bataller, 2011). Reviews for chronic alcohol use and subsequent impact on HIV diagnosis and trajectory are readily available (Hahn and Samet, 2010; Monnig, 2016; Molina et al., 2018). Endotoxin and other bacterial components in circulation provide an avenue for repeated or continuous stimulation of the innate immune system via TLR4. Studies have shown that overactivation of TLR4 leads to microglial dysfunction, significant neuroinflammation and neuronal death (Blanco et al., 2005; Alfonso-Loeches et al., 2010; Fernandez-Lizarbe et al., 2009 & 2013; see Henriques et al., 2018 for further review of the specific role of microglia in alcohol-related neurotoxicity and Yang et al., 2014 for a review of the microglia-alcohol relationship across the lifespan).

In addition to increased risk of organ-related pathologies, humans with a history of chronic heavy drinking also experience impaired wound healing and prolonged recovery following an operative procedure (see Tonnesen and Kehlet, 1999 for review). Preoperative alcohol abuse predicted longer recovery time following operative procedures due to increased wound infections and cardiopulmonary insufficiency (Tonessen and Kehlet, 1999). Increased circulating levels of adrenocorticotropic hormone (ACTH) following abstinence from alcohol may be a potential mechanism by which humans with AUD have delayed postoperative recovery (Tonessen et al., 1992). Although, a study employing one month of abstinence prior to surgery in chronic alcohol misusers (< 60g ethanol/day) reduced postoperative complications when compared to alcoholic controls who did not receive the intervention (31% v. 74%; Tonessen et al., 1999). Chronic alcoholics also show impaired bone recovery following fractures and are more likely to develop osteopenia (Chakkalakal, 2005; Bikle, Genant, and Cann, 1985). Dermal wound healing was also found to be profoundly inhibited by alcohol consumption (see Jung et al., 2011 for an extensive review), where acute binge episodes leading to BACs of at least 200 mg/dL increased wound related infections by 2.6 fold (Gentilello et al., 1993). Together, these findings suggest that heavy alcohol consumption may not only increase risk of injury, but also increase risk of extended complications following injury.

Developmental sensitivity to alcohol exposure

Given the extensive health consequences of chronic heavy drinking in human adults, it is imperative to evaluate and understand the effects of alcohol across the lifespan. The developmental period during which the onset of alcohol consumption occurs may be critical in assessing the risk for and severity of later-life drinking and immune dysfunction.

Adolescent onset of alcohol abuse as a unique period of vulnerability for adverse immune-related impact in later life.

Binge drinking emerges in adolescent humans as early as 12 years of age, and becomes especially prevalent around the age of 16 (Spear, 2000), yet the long-lasting effects of adolescent binge-drinking on host defense processes has only recently been tested. A few clinical studies have specifically assessed long-term immune dysfunction following adolescent alcohol exposure, though it is admittedly difficult to dissociate alcohol exposure effects that result from specific sensitivity or vulnerability of the adolescent period from life-long effects of alcohol consumption that was initiated during adolescence. In a study where both 20 year-old males and females who had engaged in binge drinking for at least 2 years were compared to age-matched controls, all individuals who engaged in binge drinking had elevated levels of gut-derived LPS and upregulated transcription of inflammatory markers such as IL-1β, IL-6, and MCP-1 in PMBCs. Females with a history of binge drinking during adolescence had higher markers of inflammation in PBMCs than their male counterparts (Orio et al., 2018), indicating sex-specificity in exposure effects. Opposite effects on adolescent cortical morphology between male and female binge drinkers aged 16-19 have also been reported (Squeglia et al., 2012). Still, further clinical studies are needed to more thoroughly assess the effects of alcohol exposure during adolescence rather than later in life.

Preclinical studies that have employed adolescent alcohol exposure models have found significant alterations in adult neuroimmune function, including residual activation of microglia (Sanchez-Alavez et al., 2019a and b), increased expression of TLRs and activity of NfκB signaling pathways (Crews et al., 2017), and enhanced neuroimmune gene expression when challenged with ethanol during adulthood (Vore et al., 2021). Other studies have demonstrated that Adolescent Intermittent Ethanol (AIE) exposure also exerts effects on peripheral immune function. Male rats that underwent an AIE exposure procedure exhibited impaired cytokine induction within circulating leukocytes evoked by injection of LPS in adulthood, an effect that was only observed in males (Vore et al., 2017). Interestingly, a similar impairment in extracellular cytokine release was observed in the hippocampus following the same AIE exposure (Gano et al., 2019). These findings complement studies that have examined neuroimmune function in AIE-exposed, adult male rats by measuring the fever response, rather than cytokine gene expression, after LPS exposure. Exposure to 2 weeks of a liquid ethanol diet impaired the fever response to LPS (Taylor et al., 2002) in adult males. In a rat model, Telles et al (2017) showed impairment in LPS-induced fever after 8 intraperitoneal injections of ethanol during mid-adolescence. Fever in this model was also diminished when rats were challenged with intracerebroventricular (i.c.v.) administration of PGE₂ (considered the final common mediator of febrile responses) instead of LPS, suggesting a more global impairment in thermogenic responses that could be mediated by ethanol-induced changes in Brown Adipose Tissue (BAT; Cruz et al., 2020). Although this impaired responsiveness to i.c.v. PGE₂ predicts that fever responses to any pathogen should be compromised in rats with a history of adolescent ethanol exposure, since BAT metabolism is critical to the exothermic reactions that generate heat during fever, this has not been tested. Furthermore, it is somewhat paradoxical that expression of multiple TLRs are increased in the CNS after chronic ethanol in both humans and rodent models, which would seem to suggest that fever responses should be sensitized (rather than inhibited) in adolescent alcohol exposed subjects that are later challenged with LPS. However, other competing or compensatory mechanisms could be at play, such as increased circulating endotoxin due to impaired intestinal barrier function (Tamai et al., 2000; Keshavarzian et al., 2009), increased circulating LPS binding protein (Meng et al., 2021), decoupling of TLRs from their cognate signaling pathways, or a shift in central versus peripheral immunoregulatory mechanisms. Finally, virtually no studies have utilized adult animals exposed to a comparable regimen of alcohol exposure to test for age-specific vulnerability effects. Clearly, these issues require directed efforts to fully understand the extent to which adolescence may reflect a critical and/or sensitive period for the untoward effects of alcohol on host immune defense mechanisms. Indeed, an underwhelming amount of literature examining long-term effects of adolescent alcohol exposure exists relative to PAE, despite adolescence being a similar time of sensitivity to external influences that shape the trajectory of brain development through adulthood (Spear 2013; Lees et al., 2020), and the high prevalence rates of adolescent binge drinking.

Conclusions

Overall, effective host immunity begins with a robust and well-organized innate immune response involving both the CNS and multiple peripheral end organs including organs in which infections commonly originate (skin, lung, gut, and mucosal membranes) as well as those essential to processing and propagation of the immune response (liver, brain, spleen). Binge drinking, chronic alcohol use, and diagnosis with an AUD are associated with acute and, in some cases, long-lasting changes in host immunity. Although a great deal of granular, mechanistic findings exist, most published studies focus on one specific feature of alcohol effects on host immunity, tending to focus exclusively on one organ system (CNS, liver, spleen), failing to recognize the importance of bi-directional, immune-to-brain communication pathways that are critical to effective coordination of immunological processes and ultimately host defense. Alcohol has a powerful, seemingly direct effect on neuroimmune function, which can in turn influence peripheral immunity through a top-down approach in which neural, endocrine, or humoral mediators modulate circulating immune cell function. Additionally, direct actions of ethanol (or its metabolites) on peripheral organs (liver, gut, spleen) may also influence neuroimmune processes through bottom-up signaling mechanisms (see Figure 2B). The bi-directionality of these immune-brain networks underscores the importance of acknowledging whole-organism physiology when considering the influence of alcohol on host defense processes. A major goal of this review was to highlight this bi-directionality, and to call for more integrative studies in both human and pre-clinical models that take into account the full range of systems-level interactions that subserve effective host defense.

To complicate matters further, one must also consider that AUDs have deep roots in development, with both prenatal and adolescent periods serving as the initial ages during which alcohol exposure or use (respectively) occurs, and that age of first alcohol use is highly correlated with later AUD diagnosis (DeWit et al., 2000; Miller and Spear, 2006). Though our developmental biology appears to be capable of substantial resilience, these early developmental epochs may be critical and/or sensitive periods to developmental programming events that can produce life-long changes in host immunity and tissue repair processes, including to the effects of alcohol. In this way, the rapidly changing neural, hormonal, and immune systems during early development may be particularly vulnerable to long-lasting effects of alcohol exposure that occurs during these periods. Studies in the literature specifically on long-lasting effects of adolescent binge drinking (humans) or exposure (preclinical models) are sorely lacking. Delineating the mechanisms of prenatal and adolescent alcohol effects on neural-immune interactions that utilizes an integrative, whole-organism perspective may ultimately yield important insight into not just AUD per se, but also into the wide range of health-related maladies that result from early life exposure to alcohol.