Synchrony between midbrain gene transcription and dopamine terminal regulation is modulated by chronic alcohol drinking.

Alcohol use disorder is marked by disrupted behavioral and emotional states which persist into abstinence. The enduring synaptic alterations that remain despite the absence of alcohol are of interest for interventions to prevent relapse. Here, 28 male rhesus macaques underwent over 20 months of alcohol drinking interspersed with three 30-day forced abstinence periods. After the last abstinence period, we paired direct sub-second dopamine monitoring via ex vivo voltammetry in nucleus accumbens slices with RNA-sequencing of the ventral tegmental area. We found persistent augmentation of dopamine transporter function, kappa opioid receptor sensitivity, and dynorphin release - all inhibitory regulators which act to decrease extracellular dopamine. Surprisingly, though transcript expression was not altered, the relationship between gene expression and functional readouts of these encoded proteins was highly dynamic and altered by drinking history. These results outline the long-lasting synaptic impact of alcohol use and suggest that assessment of transcript-function relationships is critical for the rational design of precision therapeutics.

Medial prefrontal dopamine dynamics reflect allocation of selective attention.

The mesocortical dopamine system is comprised of midbrain dopamine neurons that predominantly innervate the medial prefrontal cortex (mPFC) and exert a powerful neuromodulatory influence over this region 1,2 . mPFC dopamine activity is thought to be critical for fundamental neurobiological processes including valence coding and decision-making 3,4 . Despite enduring interest in this pathway, the stimuli and conditions that engage mPFC dopamine release have remained enigmatic due to inherent limitations in conventional methods for dopamine monitoring which have prevented real-time in vivo observation 5 . Here, using a fluorescent dopamine sensor enabling time-resolved recordings of cortical dopamine activity in freely behaving mice, we reveal the coding properties of this system and demonstrate that mPFC dopamine dynamics conform to a selective attention signal. Contrary to the long-standing theory that mPFC dopamine release preferentially encodes aversive and stressful events 6-8 , we observed robust dopamine responses to both appetitive and aversive stimuli which dissipated with increasing familiarity irrespective of stimulus intensity. We found that mPFC dopamine does not evolve as a function of learning but displays striking temporal precedence with second-to-second changes in behavioral engagement, suggesting a role in allocation of attentional resources. Systematic manipulation of attentional demand revealed that quieting of mPFC dopamine signals the allocation of attentional resources towards an expected event which, upon detection triggers a sharp dopamine transient marking the transition from decision-making to action. The proposed role of mPFC dopamine as a selective attention signal is the first model based on direct observation of time-resolved dopamine dynamics and reconciles decades of competing theories.

Evaluation of putative signatures of consciousness using specific definitions of responsiveness, connectedness, and consciousness.

BACKGROUND: Understanding the neural correlates of consciousness has important ramifications for the theoretical understanding of consciousness and for clinical anaesthesia. A major limitation of prior studies is the use of responsiveness as an index of consciousness. We identified a collection of measures derived from unresponsive subjects and more specifically their association with consciousness (any subjective experience) or connectedness (specific experience of environmental stimuli). METHODS: Using published data generated through the UNderstanding Consciousness Connectedness and Intra-Operative Unresponsiveness Study (NCT03284307), we evaluated 10 previously published resting-state EEG-based measures that were derived using unresponsiveness as a proxy for unconsciousness. Measures were tested across dexmedetomidine and propofol sedation and natural sleep. These markers represent the complexity, connectivity, cross-frequency coupling, graph theory, and power spectrum measures. RESULTS: Although many of the proposed markers were associated with consciousness per se (reported subjective experience), none were specific to consciousness alone; rather, each was also associated with connectedness (i.e. awareness of the environment). In addition, multiple markers showed no association with consciousness and were associated only with connectedness. Of the markers tested, loss of normalised-symbolic transfer entropy (front to back) was associated with connectedness across all three experimental conditions, whereas the transition from disconnected consciousness to unconsciousness was associated with significant decreases in permutation entropy and spectral exponent (P<0.05 for all conditions). CONCLUSIONS: None of the proposed EEG-based neural correlates of unresponsiveness corresponded solely to consciousness, highlighting the need for a more conservative use of the term (un)consciousness when assessing unresponsive participants. CLINICAL TRIAL REGISTRATION: NCT03284307.

Structured tracking of alcohol reinforcement (STAR) for basic and translational alcohol research.

There is inherent tension between methodologies developed to address basic research questions in model species and those intended for preclinical to clinical translation: basic investigations require flexibility of experimental design as hypotheses are rapidly tested and revised, whereas preclinical models emphasize standardized protocols and specific outcome measures. This dichotomy is particularly relevant in alcohol research, which spans a diverse range of basic sciences in addition to intensive efforts towards understanding the pathophysiology of alcohol use disorder (AUD). To advance these goals there is a great need for approaches that facilitate synergy across basic and translational areas of nonhuman alcohol research. In male and female mice, we establish a modular alcohol reinforcement paradigm: Structured Tracking of Alcohol Reinforcement (STAR). STAR provides a robust platform for quantitative assessment of AUD-relevant behavioral domains within a flexible framework that allows direct crosstalk between translational and mechanistically oriented studies. To achieve cross-study integration, despite disparate task parameters, a straightforward multivariate phenotyping analysis is used to classify subjects based on propensity for heightened alcohol consumption and insensitivity to punishment. Combining STAR with extant preclinical alcohol models, we delineate longitudinal phenotype dynamics and reveal putative neuro-biomarkers of heightened alcohol use vulnerability via neurochemical profiling of cortical and brainstem tissues. Together, STAR allows quantification of time-resolved biobehavioral processes essential for basic research questions simultaneous with longitudinal phenotyping of clinically relevant outcomes, thereby providing a framework to facilitate cohesion and translation in alcohol research.

Systemic kappa opioid receptor antagonism accelerates reinforcement learning via augmentation of novelty processing in male mice.

Selective inhibition of kappa opioid receptors (KORs) is highly anticipated as a pharmacotherapeutic intervention for substance use disorders and depression. The accepted explanation for KOR antagonist-induced amelioration of aberrant behaviors posits that KORs globally function as a negative valence system; antagonism thereby blunts the behavioral influence of negative internal states such as anhedonia and negative affect. While effects of systemic KOR manipulations have been widely reproduced, explicit evaluation of negative valence as an explanatory construct is lacking. Here, we tested a series of falsifiable hypotheses generated a priori based on the negative valence model by pairing reinforcement learning tasks with systemic pharmacological KOR blockade in male C57BL/6J mice. The negative valence model failed to predict multiple experimental outcomes: KOR blockade accelerated contingency learning during both positive and negative reinforcement without altering innate responses to appetitive or aversive stimuli. We next proposed novelty processing, which influences learning independent of valence, as an alternative explanatory construct. Hypotheses based on novelty processing predicted subsequent observations: KOR blockade increased exploration of a novel, but not habituated, environment and augmented the reinforcing efficacy of novel visual stimuli in a sensory reinforcement task. Together, these results revise and extend long-standing theories of KOR system function.

Pavlovian-conditioned opioid tolerance.

Opioid tolerance develops as a learned response to drug-associated cues and is thus a dynamic effect modulated by the interaction between drug and environment.

Recurrent activity within microcircuits of macaque dorsolateral prefrontal cortex tracks cognitive flexibility.

Human and non-human primate data clearly implicate the dorsolateral prefrontal cortex (dlPFC) as critical for advanced cognitive functions 1,2 . It is thought that intracortical synaptic architectures within dlPFC are the integral neurobiological substrate that gives rise to these processes, including working memory, inferential reasoning, and decision-making 3-7 . In the prevailing model, each cortical column makes up one fundamental processing unit composed of dense intrinsic connectivity, conceptualized as the 'canonical' cortical microcircuit 3,8 . Each cortical microcircuit receives sensory and cognitive information from a variety of sources which are represented by sustained activity within the microcircuit, referred to as persistent or recurrent activity 4,9 . Via recurrent connections within the microcircuit, activity can propagate for a variable length of time, thereby allowing temporary storage and computations to occur locally before ultimately passing a transformed representation to a downstream output 4,5,10 . Competing theories regarding how microcircuit activity is coordinated have proven difficult to reconcile in vivo where intercortical and intracortical computations cannot be fully dissociated 5,9,11,12 . Here, we interrogated the intrinsic features of isolated microcircuit networks using high-density calcium imaging of macaque dlPFC ex vivo . We found that spontaneous activity is intrinsically maintained by microcircuit architecture, persisting at a high rate in the absence of extrinsic connections. Further, using perisulcal stimulation to evoke persistent activity in deep layers, we found that activity propagates through stochastically assembled intracortical networks, creating predictable population-level events from largely non-overlapping ensembles. Microcircuit excitability covaried with individual cognitive performance, thus anchoring heuristic models of abstract cortical functions within quantifiable constraints imposed by the underlying synaptic architecture.

Dynamic causal modelling of auditory surprise during disconnected consciousness: The role of feedback connectivity.

The neural mechanisms through which individuals lose sensory awareness of their environment during anesthesia remains poorly understood despite being of vital importance to the field. Prior research has not distinguished between sensory awareness of the environment (connectedness) and consciousness itself. In the current study, we investigated the neural correlates of sensory awareness by contrasting neural responses to an auditory roving oddball paradigm during consciousness with sensory awareness (connected consciousness) and consciousness without sensory awareness (disconnected consciousness). These states were captured using a serial awakening paradigm with the sedative alpha2 adrenergic agonist dexmedetomidine, chosen based on our published hypothesis that suppression of noradrenaline signaling is key to induce a state of sensory disconnection. High-density electroencephalography was recorded from 18 human subjects before and after administration of dexmedetomidine. By investigating event-related potentials and taking advantage of advances in Dynamic Causal Modeling (DCM), we assessed alterations in effective connectivity between nodes of a previously established auditory processing network. We found that during disconnected consciousness, the scalp-level response to standard tones produced a P3 response that was absent during connected consciousness. This P3 response resembled the response to oddball tones seen in connected consciousness. DCM showed that disconnection produced increases in standard tone feedback signaling throughout the auditory network. Simulation analyses showed that these changes in connectivity, most notably the increase in feedback from right superior temporal gyrus to right A1, can explain the new P3 response. Together these findings show that during disconnected consciousness there is a disruption of normal predictive coding processes, so that all incoming auditory stimuli become similarly surprising.

The protective effects of escitalopram on synaptic plasticity in the CA1 region of chronically stressed and non-stressed male rats.

INTRODUCTION: Stress impairs cognitive processes and escitalopram affects them in various ways. The present study has compared the protective effects of two escitalopram doses on neural excitability and synaptic plasticity in the CA1 region of chronically stressed and non-stressed male rats. METHODS: Forty-nine rats were randomly allocated into seven groups: Control (Co), stress (St), sham (Sh), escitalopram 10 and 20 mg/kg (Esc10 and Esc20), and stress-escitalopram 10 and 20 mg/kg (St-Esc10 and St-Esc20). Induction of restraint stress (6 h/day) and escitalopram injections were performed for 14 days. The fEPSP slope and amplitude were measured according to input-output functions and after the LTP induction in the hippocampal CA1 region. Also, serum corticosterone levels were evaluated in all experimental groups. RESULTS: The fEPSP slope and amplitude decreased significantly in the St group and increased significantly in the Esc10 group compared with the Co group. In non-stressed states, significant increases in slope and amplitude occurred in the Esc10 group compared with the Esc20 group. Notably, these values were also significantly enhanced by both escitalopram doses under stressed conditions. Moreover, serum corticosterone levels significantly elevated in the St group although its levels decreased in both St-Esc groups compared with the St. CONCLUSION: Stress significantly attenuated neural excitability and long-term plasticity in the CA1 area. Only escitalopram 10 mg/kg improved synaptic excitability, as well as LTP induction and maintenance in non-stressed subjects even more than normal levels. However, under stress conditions, both escitalopram doses enhanced neural excitability and memory probably due to reduced serum corticosterone levels.

Distinct EEG signatures differentiate unconsciousness and disconnection during anaesthesia and sleep.

BACKGROUND: How conscious experience becomes disconnected from the environment, or disappears, across arousal states is unknown. We sought to identify the neural correlates of sensory disconnection and unconsciousness using a novel serial awakening paradigm. METHODS: Volunteers were recruited for sedation with dexmedetomidine i.v., propofol i.v., or natural sleep with high-density EEG monitoring and serial awakenings to establish whether subjects were in states of disconnected consciousness or unconsciousness in the preceding 20 s. The primary outcome was classification of conscious states by occipital delta power (0.5-4 Hz). Secondary analyses included derivation (dexmedetomidine) and validation (sleep/propofol) studies of EEG signatures of conscious states. RESULTS: Occipital delta power differentiated disconnected and unconscious states for dexmedetomidine (area under the curve [AUC] for receiver operating characteristic 0.605 [95% confidence interval {CI}: 0.516; 0.694]) but not for sleep/propofol (AUC 0.512 [95% CI: 0.380; 0.645]). Distinct source localised signatures of sensory disconnection (AUC 0.999 [95% CI: 0.9954; 1.0000]) and unconsciousness (AUC 0.972 [95% CI: 0.9507; 0.9879]) were identified using support vector machine classification of dexmedetomidine data. These findings generalised to sleep/propofol (validation data set: sensory disconnection [AUC 0.743 {95% CI: 0.6784; 0.8050}]) and unconsciousness (AUC 0.622 [95% CI: 0.5176; 0.7238]). We identified that sensory disconnection was associated with broad spatial and spectral changes. In contrast, unconsciousness was associated with focal decreases in activity in anterior and posterior cingulate cortices. CONCLUSIONS: These findings may enable novel monitors of the anaesthetic state that can distinguish sensory disconnection and unconsciousness, and these may provide novel insights into the biology of arousal. CLINICAL TRIAL REGISTRATION: NCT03284307.

Association between plasma tau and postoperative delirium incidence and severity: a prospective observational study.

BACKGROUND: Postoperative delirium is associated with increases in the neuronal injury biomarker, neurofilament light (NfL). Here we tested whether two other biomarkers, glial fibrillary acidic protein (GFAP) and tau, are associated with postoperative delirium. METHODS: A total of 114 surgical patients were recruited into two prospective biomarker cohort studies with assessment of delirium severity and incidence. Plasma samples were sent for biomarker analysis including tau, NfL, and GFAP, and a panel of 10 cytokines. We determined a priori to adjust for interleukin-8 (IL-8), a marker of inflammation, when assessing associations between biomarkers and delirium incidence and severity. RESULTS: GFAP concentrations showed no relationship to delirium. The change in tau from preoperative concentrations to postoperative Day 1 was greater in patients with postoperative delirium (P<0.001) and correlated with delirium severity (ρ=0.39, P<0.001). The change in tau correlated with increases in IL-8 (P<0.001) and IL-10 (P=0.0029). Linear regression showed that the relevant clinical predictors of tau changes were age (P=0.037), prior stroke/transient ischaemic attack (P=0.001), and surgical blood loss (P<0.001). After adjusting for age, sex, preoperative cognition, and change in IL-8, tau remained significantly associated with delirium severity (P=0.026). Using linear mixed effect models, only tau (not NfL or IL-8) predicted recovery from delirium (P<0.001). CONCLUSIONS: The change in plasma tau was associated with delirium incidence and severity, and resolved over time in parallel with delirium features. The impact of this putative perioperative neuronal injury biomarker on long-term cognition merits further investigation. CLINICAL TRIAL REGISTRATION: NCT02926417 and NCT03124303.

Cohort study of electroencephalography markers of amyloid-tau-neurodegeneration pathology.

Electroencephalography signatures of amyloid-ß, tau and neurodegenerative pathologies would aid in screening for, tracking progression of, and critically, understanding the pathogenesis of dementia. We hypothesized that slowing of the alpha peak frequency, as a signature of hyperpolarization-activated cyclic nucleotide gated 'pacemaker' channel activity, would correlate with amyloid and tau pathology burden measured by amyloid (Pittsburgh Compound B) and tau (MK-6240) positron emission tomography or CSF biomarkers. We also hypothesized that EEG power would be associated with neurodegeneration (CSF neurofilament light and hippocampal volume). Wakeful high-density EEG data were collected from 53 subjects. Both amyloid-ß and tau pathology were associated with slowing in the alpha peak frequency [Pittsburgh Compound B (+) vs. Pittsburgh Compound B (-) subjects, P = 0.039 and MK-6240 (+) vs. MK-6240 (-) subjects, P = 0.019]. Furthermore, slowing in the peak alpha frequency correlated with CSF Aß42/40 ratio (r 2 = 0.270; P = 0.003), phosphoTau (pTau181, r 2 = 0.290; P = 0.001) and pTau181/Aß42 (r 2 = 0.343; P < 0.001). Alpha peak frequency was not associated with neurodegeneration. Higher CSF neurofilament light was associated with lower total EEG power (r 2 = 0.136; P = 0.018), theta power (r 2 = 0.148; P = 0.014) and beta power (r 2 = 0.216; P = 0.002); the latter was also associated with normalized hippocampal volume (r 2 = 0.196; P = 0.002). Amyloid-tau and neurodegenerative pathologies are associated with distinct electrophysiological signatures that may be useful as mechanistic tools and diagnostic/treatment effect biomarkers in clinical trials.

Cohort study into the neural correlates of postoperative delirium: the role of connectivity and slow-wave activity.

BACKGROUND: Delirium frequently affects older patients, increasing morbidity and mortality; however, the pathogenesis is poorly understood. Herein, we tested the cognitive disintegration model, which proposes that a breakdown in frontoparietal connectivity, provoked by increased slow-wave activity (SWA), causes delirium. METHODS: We recruited 70 surgical patients to have preoperative and postoperative cognitive testing, EEG, blood biomarkers, and preoperative MRI. To provide evidence for causality, any putative mechanism had to differentiate on the diagnosis of delirium; change proportionally to delirium severity; and correlate with a known precipitant for delirium, inflammation. Analyses were adjusted for multiple corrections (MCs) where appropriate. RESULTS: In the preoperative period, subjects who subsequently incurred postoperative delirium had higher alpha power, increased alpha band connectivity (MC P<0.05), but impaired structural connectivity (increased radial diffusivity; MC P<0.05) on diffusion tensor imaging. These connectivity effects were correlated (r2=0.491; P=0.0012). Postoperatively, local SWA over frontal cortex was insufficient to cause delirium. Rather, delirium was associated with increased SWA involving occipitoparietal and frontal cortex, with an accompanying breakdown in functional connectivity. Changes in connectivity correlated with SWA (r2=0.257; P<0.0001), delirium severity rating (r2=0.195; P<0.001), interleukin 10 (r2=0.152; P=0.008), and monocyte chemoattractant protein 1 (r2=0.253; P<0.001). CONCLUSIONS: Whilst frontal SWA occurs in all postoperative patients, delirium results when SWA progresses to involve posterior brain regions, with an associated reduction in connectivity in most subjects. Modifying SWA and connectivity may offer a novel therapeutic approach for delirium. CLINICAL TRIAL REGISTRATION: NCT03124303, NCT02926417.

Postoperative delirium is associated with increased plasma neurofilament light.

While delirium is associated with cognitive decline and dementia, there is limited evidence to support causality for this relationship. Clarification of how delirium may cause cognitive decline, perhaps through evidence of contemporaneous neuronal injury, would enhance plausibility for a causal relationship. Dose-dependence of neuronal injury with delirium severity would further enhance the biological plausibility for this relationship. We tested whether delirium is associated with neuronal injury in 114 surgical patients recruited to a prospective biomarker cohort study. Patients underwent perioperative testing for changes in neurofilament light, a neuronal injury biomarker, as well as a panel of 10 cytokines, with contemporaneous assessment of delirium severity and incidence. A subset of patients underwent preoperative MRI. Initially we confirmed prior reports that neurofilament light levels correlated with markers of neurodegeneration [hippocampal volume (ΔR2 = 0.129, P = 0.015)] and white matter changes including fractional anisotropy of white matter (ΔR2 = 0.417, P < 0.001) with similar effects on mean, axial and radial diffusivity) in our cohort and that surgery was associated with increasing neurofilament light from preoperative levels [mean difference (95% confidence interval, CI) = 0.240 (0.178, 0.301) log10 (pg/ml), P < 0.001], suggesting putative neuronal injury. Next, we tested the relationship with delirium. Neurofilament light rose more sharply in participants with delirium compared to non-sufferers [mean difference (95% CI) = 0.251 (0.136, 0.367) log10 (pg/ml), P < 0.001]. This relationship showed dose-dependence, such that neurofilament light rose proportionately to delirium severity (ΔR2 = 0.199, P < 0.001). Given that inflammation is considered an important driver of postoperative delirium, next we tested whether neurofilament light, as a potential marker of neurotoxicity, may contribute to the pathogenesis of delirium independent of inflammation. From a panel of 10 cytokines, the pro-inflammatory cytokine IL-8 exhibited a strong correlation with delirium severity (ΔR2 = 0.208, P < 0.001). Therefore, we tested whether the change in neurofilament light contributed to delirium severity independent of IL-8. Neurofilament light was independently associated with delirium severity after adjusting for the change in inflammation (ΔR2 = 0.040, P = 0.038). These data suggest delirium is associated with exaggerated increases in neurofilament light and that this putative neurotoxicity may contribute to the pathogenesis of delirium itself, independent of changes in inflammation.