Martin Sarter's research while affiliated with University of Michigan and other places

Turning on cue or stopping at a red light requires the detection of such cues to select action sequences, or suppress action, in accordance with cue-associated action rules. Cortico-striatal projections are an essential part of the brain attention-motor interface. Glutamate-sensing microelectrode arrays were used to measure glutamate transients in the dorsomedial striatum (DMS) of male and female rats walking a treadmill and executing cued turns and stops. Prelimbic-DMS projections were chemogenetically inhibited to determine their behavioral necessity and the cortico-striatal origin of cue-evoked glutamate transients. Furthermore, we investigated rats exhibiting preferably goal-directed (goal trackers, GTs) versus cue-driven attention (sign trackers, STs), to determine the impact of such cognitive-motivational biases on cortico-striatal control. GTs executed more cued turns and initiated such turns more slowly than STs. During turns, but not missed turns or cued stops, cue-evoked glutamate concentrations were higher in GTs than in STs. In conjunction with turn cue-evoked glutamate spike levels, the presence of a single spike rendered GTs to be almost twice as likely to turn than STs. In contrast, multiple glutamate spikes predicted GTs to be less likely to turn than STs. In GTs, but not STs, inhibition of prelimbic-DMS projections attenuated turn rates, turn cue-evoked glutamate peaks, and increased the number of spikes. These findings suggest that turn cue-evoked glutamate release in GTs is tightly controlled by cortico-striatal neuronal activity. In contrast, in STs, glutamate release from DMS glutamatergic terminals may be regulated by other striatal circuitry, preferably mediating cued suppression of action and reward tracking.

Sign-tracking (ST) describes the propensity to approach and contact a Pavlovian reward cue. By contrast, goal-trackers (GTs) respond to such a cue by retrieving the reward. These behaviors index the presence of opponent cognitive-motivational traits, with STs exhibiting attentional control deficits, behavior dominated by incentive motivational processes, and vulnerability for addictive drug taking. Attentional control deficits in STs were previously attributed to attenuated cholinergic signaling, resulting from deficient translocation of intracellular choline transporters (CHTs) into synaptosomal plasma membrane. Here, we investigated a posttranslational modification of CHTs, poly-ubiquitination, and tested the hypothesis that elevated cytokine signaling in STs contributes to CHT modification. We demonstrated that intracellular CHTs, but not plasma membrane CHTs, are highly ubiquitinated in male and female sign-tracking rats when compared with GTs. Moreover, levels of cytokines measured in cortex and striatum, but not spleen, were higher in STs than in GTs. Activation of the innate immune system by systemic administration of the bacterial endotoxin lipopolysaccharide (LPS) elevated ubiquitinated CHT levels in cortex and striatum of GTs only, suggesting ceiling effects in STs. In spleen, LPS increased levels of most cytokines in both phenotypes. In cortex, LPS particularly robustly increased levels of the chemokines CCL2 and CXCL10. Phenotype-specific increases were restricted to GTs, again suggesting ceiling effects in STs. These results indicate that interactions between elevated brain immune modulator signaling and CHT regulation are essential components of the neuronal underpinnings of the addiction vulnerability trait indexed by sign-tracking.

Sign tracking versus goal tracking in rats indicate vulnerability and resistance, respectively, to Pavlovian cue-evoked addictive drug taking and relapse. Here, we tested hypotheses predicting that the opponent cognitive-behavioral styles indexed by sign tracking versus goal tracking include variations in attentional performance which differentially depend on basal forebrain projection systems. Pavlovian Conditioned Approach (PCA) testing was used to identify male and female sign-trackers (STs) and goal-trackers (GTs), as well as rats with an intermediate phenotype (INTs). Upon reaching asymptotic performance in an operant task requiring the detection of visual signals (hits) as well as the reporting of signal absence for 40 min per session, GTs scored more hits than STs, and hit rates across all phenotypes correlated with PCA scores. STs missed relatively more signals than GTs specifically during the last 15 min of a session. Chemogenetic inhibition of the basal forebrain decreased hit rates in GTs but was without effect in STs. Moreover, the decrease in hits in GTs manifested solely during the last 15 min of a session. Transfection efficacy in the horizontal limb of the diagonal band (HDB), but not substantia innominate (SI) or nucleus basalis of Meynert (nbM), predicted the behavioral efficacy of chemogenetic inhibition in GTs. Furthermore, the total subregional transfection space, not transfection of just cholinergic neurons, correlated with performance effects. These results indicate that the cognitive-behavioral phenotype indexed by goal tracking, but not sign tracking, depends on activation of the basal forebrain-frontal cortical projection system and associated biases toward top-down or model-based performance.

Sign-tracking (ST) describes the propensity to approach and contact a Pavlovian reward cue. By contrast, goal-trackers (GTs) respond to such a cue by retrieving the reward. These behaviors index the presence of opponent cognitive-motivational traits, with STs exhibiting attentional control deficits, behavior dominated by incentive motivational processes, and vulnerability for addictive drug taking. Attentional control deficits in STs were previously attributed to attenuated cholinergic signaling, resulting from deficient translocation of intracellular choline transporters (CHTs) into synaptosomal plasma membrane. Here we investigated a post-translational modification of CHTs – poly-ubiquitination - and tested the hypothesis that elevated cytokine signaling in STs contributes to CHT modification. We demonstrated that intracellular CHTs, but not plasma membrane CHTs, are highly ubiquitinated in male and female sign-tracking rats when compared with GTs. Moreover, levels of cytokines measured in cortex and striatum, but not spleen, were higher in STs than in GTs. Activation of the innate immune system by systemic administration of the bacterial endotoxin lipopolysaccharide (LPS) elevated ubiquitinated CHT levels in cortex and striatum of GTs only, suggesting ceiling effects in STs. In spleen, LPS increased levels of most cytokines in both phenotypes. In cortex, LPS particularly robustly increased levels of the chemokines CCL2 and CXCL10. Phenotype-specific increases were restricted to GTs, again suggesting ceiling effects in STs. These results indicate that interactions between elevated brain immune modulator signaling and CHT regulation are essential components of the neuronal underpinnings of the addiction vulnerability trait indexed by sign-tracking. Significance Statement Sign-tracking rats (STs) have emerged as a fruitful animal model for determining the neuro-behavioral foundations, including the cholinergic-attentional control deficits, which heighten the risk for the manifestation of addictive behaviors. The results from the present experiments suggest that, in STs, elevated levels of neuro-immune signaling interact with a post-translational modification of choline transporters that yields a relatively low transporter capacity. Furthermore, the effects of activation of the innate immune system, by administrating the endotoxin lipopolysaccharide, mechanistically supported such an interaction. The results extend research on the neuronal vulnerabilities for addictive behaviors to the role of neuro-immune signaling and suggests a new role of neuro-immune modulators in influencing complex cognitive-motivational traits.

Transport of choline via the neuronal high-affinity choline transporter (CHT; SLC5A7) is essential for cholinergic terminals to synthesize and release acetylcholine (ACh). In humans, we previously demonstrated an association between a common CHT coding substitution (rs1013940; Ile89Val) and reduced attentional control as well as attenuated frontal cortex activation. Here, we used a CRISPR/ Cas9 approach to generate mice expressing the I89V substitution and assessed, in vivo, CHT-mediated choline transport, and ACh release. Relative to wild-type (WT) mice, CHT-mediated clearance of choline in male and female mice expressing one or two Val89 alleles was reduced by over 80% in cortex and over 50% in striatum. Choline clearance in CHT Val89 mice was further reduced by neuronal inactivation. Deficits in ACh release, 5 and 10min after repeated depolarization at a low, behaviorally relevant frequency, support an attenuated reloading capacity of cholinergic neurons in mutant mice. The density of CHTs in total synaptosomal lysates and neuronal plasma-membrane-enriched fractions was not impacted by the Val89 variant, indicating a selective impact on CHT function. When challenged with a visual disruptor to reveal attentional control mechanisms, Val89 mice failed to adopt a more conservative response bias. Structural modeling revealed that Val89 may attenuate choline transport by altering conformational changes of CHT that support normal transport rates. Our findings support the view that diminished sustained cholinergic signaling capacity underlies perturbed attentional performance in individuals expressing CHT Val89. The CHT Val89 mouse serves as a valuable model to study heritable risk for cognitive disorders arising from cholinergic dysfunction.

Dysfunction and degeneration of CNS cholinergic systems is a significant component of multi-system pathology in Parkinson's disease (PD). We review the basic architecture of human CNS cholinergic systems and the tools available for studying changes in human cholinergic systems. Earlier post-mortem studies implicated abnormalities of basal forebrain corticopetal cholinergic (BFCC) and pedunculopontine-laterodorsal tegmental (PPN-LDT) cholinergic projections in cognitive deficits and gait-balance deficits, respectively. Recent application of imaging methods, particularly molecular imaging, allowed more sophisticated correlation of clinical features with regional cholinergic deficits. BFCC projection deficits correlate with general and domain specific cognitive deficits, particularly for attentional and executive functions. Detailed analyses suggest that cholinergic deficits within the salience and cingulo-opercular task control networks, including both neocortical, thalamic, and striatal nodes, are a significant component of cognitive deficits in non-demented PD subjects. Both BFCC and PPN-LDT cholinergic projection systems, and striatal cholinergic interneuron (SChI), abnormalities are implicated in PD gait-balance disorders. In the context of experimental studies, these results indicate that disrupted attentional functions of BFCC and PPN-LDT cholinergic systems underlie impaired gait-balance functions. SChI dysfunction likely impairs intra-striatal integration of attentional and motor information. Thalamic and entorhinal cortex cholinergic deficits may impair multi-sensory integration. Overt degeneration of CNS systems may be preceded by increased activity of cholinergic neurons compensating for nigrostriatal dopaminergic deficits. Subsequent dysfunction and degeneration of cholinergic systems unmasks and exacerbates functional deficits secondary to dopaminergic denervation. Research on CNS cholinergic systems dysfunctions in PD requires a systems-level approach to understanding PD pathophysiology.

Theta and gamma rhythms and their cross-frequency coupling play critical roles in perception, attention, learning, and memory. Available data suggest that forebrain acetylcholine (ACh) signaling promotes theta-gamma coupling, although the mechanism has not been identified. Recent evidence suggests that cholinergic signaling is both temporally and spatially constrained, in contrast to the traditional notion of slow, spatially homogeneous, and diffuse neuromodulation. Here, we find that spatially constrained cholinergic stimulation can generate theta-modulated gamma rhythms. Using biophysically-based excitatory-inhibitory (E-I) neural network models, we simulate the effects of ACh on neural excitability by varying the conductance of a muscarinic receptor-regulated K+ current. In E-I networks with local excitatory connectivity and global inhibitory connectivity, we demonstrate that theta-gamma-coupled firing patterns emerge in ACh modulated network regions. Stable gamma-modulated firing arises within regions with high ACh signaling, while theta or mixed theta-gamma activity occurs at the peripheries of these regions. High gamma activity also alternates between different high-ACh regions, at theta frequency. Our results are the first to indicate a causal role for spatially heterogenous ACh signaling in the emergence of localized theta-gamma rhythmicity. Our findings also provide novel insights into mechanisms by which ACh signaling supports the brain region-specific attentional processing of sensory information.

Transport of choline via the neuronal high-affinity choline transporter (CHT; SLC5A7 ) is essential for cholinergic terminals to synthesize and release acetylcholine (ACh). In humans, we previously demonstrated an association between a common CHT coding substitution (rs1013940; Ile89Val) and reduced attentional capacity as well as attenuated frontal cortex activation. Here, we used a CRISPR/Cas9 approach to generate mice expressing the I89V substitution and assessed, using in vivo cortical choline biosensing, CHT-mediated choline transport, and ACh release. CHT-mediated clearance of choline in mice expressing one or two Val89 alleles was reduced by over 7-fold relative to wild type (WT) mice, suggesting dominant-negative effects. Choline clearance in CHT Val89 mice was further reduced by neuronal inactivation. Deficits in ACh release, 5 and 10 min after repeated depolarization at a low, behaviorally relevant frequency, support an attenuated reloading capacity of cholinergic neurons in mutant mice. The density of CHTs in total synaptosomal lysates and neuronal plasma-membrane-enriched fractions was not impacted by the Val89 variant, indicating a selective impact on CHT function. Consistent with this hypothesis, structural modeling revealed that Val89 may attenuate choline transport by changing the ability of choline to induce conformational changes of CHT that support normal transport rates. Our findings suggest that diminished, sustained cholinergic signaling capacity in the frontal cortex underlies perturbed attentional performance in individuals expressing CHT Val89. Our work supports the utility of the CHT Val89 mouse model as a valuable model to study heritable risk for cognitive disorders arising from cholinergic dysfunction. Significance Statement Acetylcholine (ACh) signaling depends on the capacity of the neuronal choline transporter (CHT) to import choline into presynaptic terminals for ACh synthesis. Previous research demonstrated that humans expressing the common CHT coding variant Val89 exhibit attentional vulnerabilities and attenuated fronto-cortical activation during attention. Here, we find that mice engineered to express the Val89 variant exhibit reduced CHT-mediated choline clearance in frontal cortex and a diminished capacity to sustain neuronal ACh release. These findings provide a mechanistic framework for interpreting the attentional phenotype associated with the human Val89 variant and establish a mouse model that permits a more invasive interrogation of CNS effects as well as the development of therapeutic strategies tailored to those, including Val89 carriers, with presynaptic cholinergic perturbations.

Objective Attentional deficits following degeneration of brain cholinergic systems contribute to gait‐balance deficits in Parkinson disease (PD). As a step towards assessing if α4β2* nicotinic acetylcholine receptor (nAChR) stimulation improves gait‐balance function, we assessed target engagement of the α4β2* nAChR partial agonist varenicline. Methods Non‐demented PD participants with cholinergic deficits were identified with [18F]fluoroethoxybenzamicol positron emission tomography (PET). α4β2* nAChR occupancy after subacute oral varenicline treatment was measured with [18F]flubatine PET. With a dose selected from the nAChR occupancy experiment, varenicline effects on gait, balance, and cognition were assessed in a double‐masked placebo‐controlled crossover study. Primary endpoints were normal pace gait speed and a measure of postural stability. Results Varenicline doses (0.25 mg per day, 0.25 mg b.i.d., 0.5 mg b.i.d., and 1.0 mg b.i.d.) produced 60% ‐ 70% receptor occupancy. We selected 0.5 mg po b.i.d for the crossover study. Thirty‐three participants completed the crossover study with excellent tolerability. Varenicline had no significant impact on the postural stability measure and caused slower normal pace gait speed. Varenicline narrowed the difference in normal pace gait speed between dual task and no dual task gait conditions, reduced dual task cost, and improved sustained attention test performance. We obtained identical conclusions in 28 participants with treatment compliance confirmed by plasma varenicline measurements. Interpretation Varenicline occupied α4β2* nicotinic acetylcholine receptors, was tolerated well, enhanced attention, and altered gait performance. These results are consistent with target engagement. α4β2* agonists may be worth further evaluation for mitigation of gait and balance disorders in PD. This article is protected by copyright. All rights reserved.