Dopamine D1-Receptor Organization Contributes to Functional Brain Architecture.

Recent work has recognized a gradient-like organization in cortical function, spanning from primary sensory to transmodal cortices. It has been suggested that this axis is aligned with regional differences in neurotransmitter expression. Given the abundance of dopamine D1-receptors (D1DR), and its importance for modulation and neural gain, we tested the hypothesis that D1DR organization is aligned with functional architecture, and that inter-regional relationships in D1DR co-expression modulate functional cross talk. Using the world's largest dopamine D1DR-PET and MRI database (N = 180%, 50% female), we demonstrate that D1DR organization follows a unimodal-transmodal hierarchy, expressing a high spatial correspondence to the principal gradient of functional connectivity. We also demonstrate that individual differences in D1DR density between unimodal and transmodal regions are associated with functional differentiation of the apices in the cortical hierarchy. Finally, we show that spatial co-expression of D1DR primarily modulates couplings within, but not between, functional networks. Together, our results show that D1DR co-expression provides a biomolecular layer to the functional organization of the brain.

Widespread fMRI BOLD Signal Overactivations during Cognitive Control in Older Adults Are Not Matched by Corresponding Increases in fPET Glucose Metabolism.

A common observation in fMRI studies using the BOLD signal is that older adults, compared with young adults, show overactivations, particularly during less demanding tasks. The neuronal underpinnings of such overactivations are not known, but a dominant view is that they are compensatory in nature and involve recruitment of additional neural resources. We scanned 23 young (20-37 years) and 34 older (65-86 years) healthy human adults of both sexes with hybrid positron emission tomography/MRI. The radioligand [18F]fluoro-deoxyglucose was used to assess dynamic changes in glucose metabolism as a marker of task-dependent synaptic activity, along with simultaneous fMRI BOLD imaging. Participants performed two verbal working memory (WM) tasks: one involving maintenance (easy) and one requiring manipulation (difficult) of information in WM. Converging activations to the WM tasks versus rest were observed for both imaging modalities and age groups in attentional, control, and sensorimotor networks. Upregulation of activity to WM-demand, comparing the more difficult to the easier task, also converged between both modalities and age groups. For regions in which older adults showed task-dependent BOLD overactivations compared with the young adults, no corresponding increases in glucose metabolism were found. To conclude, findings from the current study show that task-induced changes in the BOLD signal and synaptic activity as measured by glucose metabolism generally converge, but overactivations observed with fMRI in older adults are not coupled with increased synaptic activity, which suggests that these overactivations are not neuronal in origin.SIGNIFICANCE STATEMENT Findings of increased fMRI activations in older compared with younger adults have been suggested to reflect increased use of neuronal resources to cope with reduced brain function. The physiological underpinnings of such compensatory processes are poorly understood, however, and rest on the assumption that vascular signals accurately reflect neuronal activity. Comparing fMRI and simultaneously acquired functional positron emission tomography as an alternative index of synaptic activity, we show that age-related overactivations do not appear to be neuronal in origin. This result is important because mechanisms underlying compensatory processes in aging are potential targets for interventions aiming to prevent age-related cognitive decline.

Impact of simulated reduced injected dose on the assessment of amyloid PET scans.

PURPOSE: To investigate the impact of reduced injected doses on the quantitative and qualitative assessment of the amyloid PET tracers [18F]flutemetamol and [18F]florbetaben. METHODS: Cognitively impaired and unimpaired individuals (N = 250, 36% Aß-positive) were included and injected with [18F]flutemetamol (N = 175) or [18F]florbetaben (N = 75). PET scans were acquired in list-mode (90-110 min post-injection) and reduced-dose images were simulated to generate images of 75, 50, 25, 12.5 and 5% of the original injected dose. Images were reconstructed using vendor-provided reconstruction tools and visually assessed for Aß-pathology. SUVRs were calculated for a global cortical and three smaller regions using a cerebellar cortex reference tissue, and Centiloid was computed. Absolute and percentage differences in SUVR and CL were calculated between dose levels, and the ability to discriminate between Aß- and Aß + scans was evaluated using ROC analyses. Finally, intra-reader agreement between the reduced dose and 100% images was evaluated. RESULTS: At 5% injected dose, change in SUVR was 3.72% and 3.12%, with absolute change in Centiloid 3.35CL and 4.62CL, for [18F]flutemetamol and [18F]florbetaben, respectively. At 12.5% injected dose, percentage change in SUVR and absolute change in Centiloid were < 1.5%. AUCs for discriminating Aß- from Aß + scans were high (AUC ≥ 0.94) across dose levels, and visual assessment showed intra-reader agreement of > 80% for both tracers. CONCLUSION: This proof-of-concept study showed that for both [18F]flutemetamol and [18F]florbetaben, adequate quantitative and qualitative assessments can be obtained at 12.5% of the original injected dose. However, decisions to reduce the injected dose should be made considering the specific clinical or research circumstances.

Dissociations between glucose metabolism and blood oxygenation in the human default mode network revealed by simultaneous PET-fMRI.

The finding of reduced functional MRI (fMRI) activity in the default mode network (DMN) during externally focused cognitive control has been highly influential to our understanding of human brain function. However, these negative fMRI responses, measured as relative decreases in the blood-oxygenation-level-dependent (BOLD) response between rest and task, have also prompted major questions of interpretation. Using hybrid functional positron emission tomography (PET)-MRI, this study shows that task-positive and -negative BOLD responses do not reflect antagonistic patterns of synaptic metabolism. Task-positive BOLD responses in attention and control networks were accompanied by concomitant increases in glucose metabolism during cognitive control, but metabolism in widespread DMN remained high during rest and task despite negative BOLD responses. Dissociations between glucose metabolism and the BOLD response specific to the DMN reveal functional heterogeneity in this network and demonstrate that negative BOLD responses during cognitive control should not be interpreted to reflect relative increases in metabolic activity during rest. Rather, neurovascular coupling underlying BOLD response patterns during rest and task in DMN appears fundamentally different from BOLD responses in other association networks during cognitive control.

When functional blurring becomes deleterious: Reduced system segregation is associated with less white matter integrity and cognitive decline in aging.

Healthy aging is accompanied by progressive decline in cognitive performance and concomitant changes in brain structure and functional architecture. Age-accompanied alterations in brain function have been characterized on a network level as weaker functional connections within brain networks along with stronger interactions between networks. This phenomenon has been described as age-related differences in functional network segregation. It has been suggested that functional networks related to associative processes are particularly sensitive to age-related deterioration in segregation, possibly related to cognitive decline in aging. However, there have been only a few longitudinal studies with inconclusive results. Here, we used a large longitudinal sample of 284 participants between 25 to 80 years of age at baseline, with cognitive and neuroimaging data collected at up to three time points over a 10-year period. We investigated age-related changes in functional segregation among two large-scale systems comprising associative and sensorimotor-related resting-state networks. We found that functional segregation of associative systems declines in aging with exacerbated deterioration from the late fifties. Changes in associative segregation were positively associated with changes in global cognitive ability, suggesting that decreased segregation has negative consequences for domain-general cognitive functions. Age-related changes in system segregation were partly accounted for by changes in white matter integrity, but white matter integrity only weakly influenced the association between segregation and cognition. Together, these novel findings suggest a cascade where reduced white-matter integrity leads to less distinctive functional systems which in turn contributes to cognitive decline in aging.

Balance between Transmitter Availability and Dopamine D2 Receptors in Prefrontal Cortex Influences Memory Functioning.

Insufficient or excessive dopaminergic tone impairs cognitive performance. We examine whether the balance between transmitter availability and dopamine (DA) D2 receptors (D2DRs) is important for successful memory performance in a large sample of adults (n = 175, 64-68 years). The Catechol-O-Methyltransferase polymorphism served as genetic proxy for endogenous prefrontal DA availability, and D2DRs in dorsolateral prefrontal cortex (dlPFC) were measured with [11C]raclopride-PET. Individuals for whom D2DR status matched DA availability showed higher levels of episodic and working-memory performance than individuals with insufficient or excessive DA availability relative to the number of receptors. A similar pattern restricted to episodic memory was observed for D2DRs in caudate. Functional magnetic resonance imaging data acquired during working-memory performance confirmed the importance of a balanced DA system for load-dependent brain activity in dlPFC. Our data suggest that the inverted-U-shaped function relating DA signaling to cognition is modulated by a dynamic association between DA availability and receptor status.

Dedifferentiation of caudate functional connectivity and striatal dopamine transporter density predict memory change in normal aging.

Age-related changes in striatal function are potentially important for predicting declining memory performance over the adult life span. Here, we used fMRI to measure functional connectivity of caudate subfields with large-scale association networks and positron emission tomography to measure striatal dopamine transporter (DAT) density in 51 older adults (age 65-86 years) who received annual cognitive testing for up to 7 years (mean = 5.59, range 2-7 years). Analyses showed that cortical-caudate functional connectivity was less differentiated in older compared with younger adults (n = 63, age 18-32 years). Unlike in younger adults, the central lateral caudate was less strongly coupled with the frontal parietal control network in older adults. Older adults also showed less "decoupling" of the caudate from other networks, including areas of the default network (DN) and the hippocampal complex. Contrary to expectations, less decoupling between caudate and the DN was not associated with an age-related reduction of striatal DAT, suggesting that neurobiological changes in the cortex may drive dedifferentiation of cortical-caudate connectivity. Reduction of specificity in functional coupling between caudate and regions of the DN predicted memory decline over subsequent years at older ages. The age-related reduction in striatal DAT density also predicted memory decline, suggesting that a relation between striatal functions and memory decline in aging is multifaceted. Collectively, the study provides evidence highlighting the association of age-related differences in striatal function to memory decline in normal aging.

Dopamine D2/3 Binding Potential Modulates Neural Signatures of Working Memory in a Load-Dependent Fashion.

Dopamine (DA) modulates corticostriatal connections. Studies in which imaging of the DA system is integrated with functional imaging during cognitive performance have yielded mixed findings. Some work has shown a link between striatal DA (measured by PET) and fMRI activations, whereas others have failed to observe such a relationship. One possible reason for these discrepant findings is differences in task demands, such that a more demanding task with greater prefrontal activations may yield a stronger association with DA. Moreover, a potential DA-BOLD association may be modulated by task performance. We studied 155 (104 normal-performing and 51 low-performing) healthy older adults (43% females) who underwent fMRI scanning while performing a working memory (WM) n-back task along with DA D2/3 PET assessment using [11C]raclopride. Using multivariate partial-least-squares analysis, we observed a significant pattern revealing positive associations of striatal as well as extrastriatal DA D2/3 receptors to BOLD response in the thalamo-striatal-cortical circuit, which supports WM functioning. Critically, the DA-BOLD association in normal-performing, but not low-performing, individuals was expressed in a load-dependent fashion, with stronger associations during 3-back than 1-/2-back conditions. Moreover, normal-performing adults expressing upregulated BOLD in response to increasing task demands showed a stronger DA-BOLD association during 3-back, whereas low-performing individuals expressed a stronger association during 2-back conditions. This pattern suggests a nonlinear DA-BOLD performance association, with the strongest link at the maximum capacity level. Together, our results suggest that DA may have a stronger impact on functional brain responses during more demanding cognitive tasks.SIGNIFICANCE STATEMENT Dopamine (DA) is a major neuromodulator in the CNS and plays a key role in several cognitive processes via modulating the blood oxygenation level-dependent (BOLD) signal. Some studies have shown a link between DA and BOLD, whereas others have failed to observe such a relationship. A possible reason for the discrepancy is differences in task demands, such that a more demanding task with greater prefrontal activations may yield a stronger association with DA. We examined the relationship of DA to BOLD response during working memory under three load conditions and found that the DA-BOLD association is expressed in a load-dependent fashion. These findings may help explain the disproportionate impairment evident in more effortful cognitive tasks in normal aging and in those suffering dopamine-dependent neurodegenerative diseases (e.g., Parkinson's disease).

Longitudinal Evidence for Increased Functional Response in Frontal Cortex for Older Adults with Hippocampal Atrophy and Memory Decline.

The functional organization of the frontal cortex is dynamic. Age-related increases in frontal functional responses have been shown during various cognitive tasks, but the cross-sectional nature of most past studies makes it unclear whether these increases reflect reorganization or stable individual differences. Here, we followed 130 older individuals' cognitive trajectories over 20-25 years with repeated neuropsychological assessments every 5th year, and identified individuals with stable or declining episodic memory. Both groups displayed significant gray matter atrophy over 2 successive magnetic resonance imaging sessions 4 years apart, but the decline group also had a smaller volume of the right hippocampus. Only individuals with declining memory demonstrated increased prefrontal functional responses during memory encoding and retrieval over the 4-year interval. Regions with increased functional recruitment were located outside, or on the borders of core task-related networks, indicating an expansion of these over time. These longitudinal findings offer novel insight into the mechanisms behind age-associated memory loss, and are consistent with a theoretical model in which hippocampus atrophy, past a critical threshold, induces episodic-memory decline and altered prefrontal functional organization.

Neurocognitive Profiles of Older Adults with Working-Memory Dysfunction.

Individuals differ in how they perceive, remember, and think. There is evidence for the existence of distinct subgroups that differ in cognitive performance within the older population. However, it is less clear how individual differences in cognition in old age are linked to differences in brain-based measures. We used latent-profile analysis on n-back working-memory (WM) performance to identify subgroups in a large sample of older adults (n = 181; age = 64-68 years). Our analysis identified one larger normal subgroup with higher performance (n = 113; 63%), and a second smaller subgroup (n = 55; 31%) with lower performance. The low-performing subgroup showed weaker load-dependent BOLD modulation and lower connectivity within the fronto-parietal network (FPN) as well as between FPN and striatum during n-back, along with lower FPN connectivity at rest. This group also exhibited lower FPN structural integrity, lower frontal dopamine D2 binding potential, inferior performance on offline WM tests, and a trend-level genetic predisposition for lower dopamine-system efficiency. By contrast, this group exhibited relatively intact episodic memory and associated brain measures (i.e., hippocampal volume, structural, and functional connectivity within the default-mode network). Collectively, these data provide converging evidence for the existence of a group of older adults with impaired WM functioning characterized by reduced cortico-striatal coupling and aberrant cortico-cortical integrity within FPN.

Dopamine D2 receptor availability is linked to hippocampal-caudate functional connectivity and episodic memory.

D1 and D2 dopamine receptors (D1DRs and D2DRs) may contribute differently to various aspects of memory and cognition. The D1DR system has been linked to functions supported by the prefrontal cortex. By contrast, the role of the D2DR system is less clear, although it has been hypothesized that D2DRs make a specific contribution to hippocampus-based cognitive functions. Here we present results from 181 healthy adults between 64 and 68 y of age who underwent comprehensive assessment of episodic memory, working memory, and processing speed, along with MRI and D2DR assessment with [(11)C]raclopride and PET. Caudate D2DR availability was positively associated with episodic memory but not with working memory or speed. Whole-brain analyses further revealed a relation between hippocampal D2DR availability and episodic memory. Hippocampal and caudate D2DR availability were interrelated, and functional MRI-based resting-state functional connectivity between the ventral caudate and medial temporal cortex increased as a function of caudate D2DR availability. Collectively, these findings indicate that D2DRs make a specific contribution to hippocampus-based cognition by influencing striatal and hippocampal regions, and their interactions.

BOLD Variability is Related to Dopaminergic Neurotransmission and Cognitive Aging.

Dopamine (DA) losses are associated with various aging-related cognitive deficits. Typically, higher moment-to-moment brain signal variability in large-scale patterns of voxels in neocortical regions is linked to better cognitive performance and younger adult age, yet the physiological mechanisms regulating brain signal variability are unknown. We explored the relationship among adult age, DA availability, and blood oxygen level-dependent (BOLD) signal variability, while younger and older participants performed a spatial working memory (SWM) task. We quantified striatal and extrastriatal DA D1 receptor density with [(11)C]SCH23390 and positron emission tomography in all participants. We found that BOLD variability in a neocortical region was negatively related to age and positively related to SWM performance. In contrast, BOLD variability in subcortical regions and bilateral hippocampus was positively related to age and slower responses, and negatively related to D1 density in caudate and dorsolateral prefrontal cortex. Furthermore, BOLD variability in neocortical regions was positively associated with task-related disengagement of the default-mode network, a network whose activation needs to be suppressed for efficient SWM processing. Our results show that age-related DA losses contribute to changes in brain signal variability in subcortical regions and suggest a potential mechanism, by which neocortical BOLD variability supports cognitive performance.

Multiple Brain Markers are Linked to Age-Related Variation in Cognition.

Age-related alterations in brain structure and function have been challenging to link to cognition due to potential overlapping influences of multiple neurobiological cascades. We examined multiple brain markers associated with age-related variation in cognition. Clinically normal older humans aged 65-90 from the Harvard Aging Brain Study (N = 186) were characterized on a priori magnetic resonance imaging markers of gray matter thickness and volume, white matter hyperintensities, fractional anisotropy (FA), resting-state functional connectivity, positron emission tomography markers of glucose metabolism and amyloid burden, and cognitive factors of processing speed, executive function, and episodic memory. Partial correlation and mediation analyses estimated age-related variance in cognition shared with individual brain markers and unique to each marker. The largest relationships linked FA and striatum volume to processing speed and executive function, and hippocampal volume to episodic memory. Of the age-related variance in cognition, 70-80% was accounted for by combining all brain markers (but only ∼20% of total variance). Age had significant indirect effects on cognition via brain markers, with significant markers varying across cognitive domains. These results suggest that most age-related variation in cognition is shared among multiple brain markers, but potential specificity between some brain markers and cognitive domains motivates additional study of age-related markers of neural health.

Dopamine D1 Binding Potential Predicts Fusiform BOLD Activity during Face-Recognition Performance.

The importance of face memory in humans and primates is well established, but little is known about the neurotransmitter systems involved in face recognition. We tested the hypothesis that face recognition is linked to dopamine (DA) activity in fusiform gyrus (FFG). DA availability was assessed by measuring D1 binding potential (BP) during rest using PET. We further assessed blood-oxygen-level-dependent (BOLD) signal change while subjects performed a face-recognition task during fMRI scanning. There was a strong association between D1 BP and BOLD activity in FFG, whereas D1 BP in striatal and other extrastriatal regions were unrelated to neural activity in FFG. These results suggest that D1 BP locally modulates FFG function during face recognition. Observed relationships among D1 BP, BOLD activity, and face-recognition performance further suggest that D1 receptors place constraints on the responsiveness of FFG neurons. SIGNIFICANCE STATEMENT: The importance of face memory in humans and primates is well established, but little is known about the neurotransmitter systems involved in face recognition. Our work shows a role for a specific neurotransmitter system in face memory.

Dopamine transporter availability in clinically normal aging is associated with individual differences in white matter integrity.

Aging-related differences in white matter integrity, the presence of amyloid plaques, and density of biomarkers indicative of dopamine functions can be detected and quantified with in vivo human imaging. The primary aim of the present study was to investigate whether these imaging-based measures constitute independent imaging biomarkers in older adults, which would speak to the hypothesis that the aging brain is characterized by multiple independent neurobiological cascades. We assessed MRI-based markers of white matter integrity and PET-based marker of dopamine transporter density and amyloid deposition in the same set of 53 clinically normal individuals (age 65-87). A multiple regression analysis demonstrated that dopamine transporter availability is predicted by white matter integrity, which was detectable even after controlling for chronological age. Further post-hoc exploration revealed that dopamine transporter availability was further associated with systolic blood pressure, mirroring the established association between cardiovascular health and white matter integrity. Dopamine transporter availability was not associated with the presence of amyloid burden. Neurobiological correlates of dopamine transporter measures in aging are therefore likely unrelated to Alzheimer's disease but are aligned with white matter integrity and cardiovascular risk. More generally, these results suggest that two common imaging markers of the aging brain that are typically investigated separately do not reflect independent neurobiological processes. Hum Brain Mapp 37:621-631, 2016. © 2015 Wiley Periodicals, Inc.

Influences of a DRD2 polymorphism on updating of long-term memory representations and caudate BOLD activity: magnification in aging.

A number of genetic polymorphisms are related to individual differences in cognitive performance. Striatal dopamine (DA) functions, associated with cognitive performance, are linked to the TaqIA polymorphism of the DRD2/ANKK1 gene. In humans, presence of an A1 allele of the DRD2/ANKK1-TaqIA polymorphism is related to reduced density of striatal DA D2 receptors. The resource-modulation hypothesis assumes that aging-related losses of neurochemical and structural brain resources modulate the extent to which genetic variations affect cognitive functioning. Here, we tested this hypothesis using functional MRI during long-term memory (LTM) updating in younger and older carriers and noncarriers of the A1-allele of the TaqIa polymorphism. We demonstrate that older A1-carriers have worse memory performance, specifically during LTM updating, compared to noncarriers. Moreover, A1-carriers exhibited less blood oxygen level-dependent (BOLD) activation in left caudate nucleus, a region critical to updating. This effect was only seen in older adults, suggesting magnification of genetic effects on functional brain activity in aging. Further, a positive relationship between caudate BOLD activation and updating performance among non-A1 carriers indicated that caudate activation was behaviorally relevant. These results demonstrate a link between the DRD2/ANKK1-TaqIA polymorphism and neurocognitive deficits related to LTM updating, and provide novel evidence that this effect is magnified in aging.

A multivariate analysis of age-related differences in functional networks supporting conflict resolution.

Functional neuroimaging studies demonstrate age-related differences in recruitment of a large-scale attentional network during interference resolution, especially within dorsolateral prefrontal cortex (DLPFC) and anterior cingulate cortex (ACC). These alterations in functional responses have been frequently observed despite equivalent task performance, suggesting age-related reallocation of neural resources, although direct evidence for a facilitating effect in aging is sparse. We used the multi-source interference task and multivariate partial-least-squares to investigate age-related differences in the neuronal signature of conflict resolution, and their behavioral implications in younger and older adults. There were interference-related increases in activity, involving fronto-parietal and basal ganglia networks that generalized across age. In addition an age-by-task interaction was observed within a distributed network, including DLPFC and ACC, with greater activity during interference in the old. Next, we combined brain-behavior and functional connectivity analyses to investigate whether compensatory brain changes were present in older adults, using DLPFC and ACC as regions of interest (i.e. seed regions). This analysis revealed two networks differentially related to performance across age groups. A structural analysis revealed age-related gray-matter losses in regions facilitating performance in the young, suggesting that functional reorganization may partly reflect structural alterations in aging. Collectively, these findings suggest that age-related structural changes contribute to reductions in the efficient recruitment of a youth-like interference network, which cascades into instantiation of a different network facilitating conflict resolution in elderly people.

Dopamine release in human associative striatum during reversal learning.

The dopaminergic system is firmly implicated in reversal learning but human measurements of dopamine release as a correlate of reversal learning success are lacking. Dopamine release and hemodynamic brain activity in response to unexpected changes in action-outcome probabilities are here explored using simultaneous dynamic [11C]Raclopride PET-fMRI and computational modelling of behavior. When participants encounter reversed reward probabilities during a card guessing game, dopamine release is observed in associative striatum. Individual differences in absolute reward prediction error and sensitivity to errors are associated with peak dopamine receptor occupancy. The fMRI response to perseverance errors at the onset of a reversal spatially overlap with the site of dopamine release. Trial-by-trial fMRI correlates of absolute prediction errors show a response in striatum and association cortices, closely overlapping with the location of dopamine release, and separable from a valence signal in ventral striatum. The results converge to implicate striatal dopamine release in associative striatum as a central component of reversal learning, possibly signifying the need for increased cognitive control when new stimuli-responses should be learned.

Longitudinal monitoring of the mouse brain reveals heterogenous network trajectories during aging.

The human aging brain is characterized by changes in network efficiency that are currently best captured through longitudinal resting-state functional MRI (rs-fMRI). These studies however are challenging due to the long human lifespan. Here we show that the mouse animal model with a much shorter lifespan allows us to follow the functional network organization over most of the animal's adult lifetime. We used a longitudinal study of the functional connectivity of different brain regions with rs-fMRI under anesthesia. Our analysis uncovers network modules similar to those reported in younger mice and in humans (i.e., prefrontal/default mode network (DMN), somatomotor and somatosensory networks). Statistical analysis reveals different patterns of network reorganization during aging. Female mice showed a pattern akin to human aging, with de-differentiation of the connectome, mainly due to increases in connectivity of the prefrontal/DMN cortical networks to other modules. Our male cohorts revealed heterogenous aging patterns with only one group confirming the de- differentiation, while the majority showed an increase in connectivity of the somatomotor cortex to the Nucleus accumbens. In summary, in line with human work, our analysis in mice supports the concept of de-differentiation in the aging mammalian brain and reveals additional trajectories in aging mice networks.

Assessment of circulating apoE4 levels from dried blood spot samples in a large survey setting.

INTRODUCTION: The apolipoprotein E (APOE) ε4 allele is associated with high risk for Alzheimer's disease. It is unclear whether individual levels of the circulating apoE4 protein in ε4 carriers confer additional risk. Measuring apoE4 protein levels from dried blood spots (DBS) has the potential to provide information on genetic status as well as circulating levels and to include these measures in large survey settings. METHODS: We developed a multiplex immunoassay to detect apoE4 protein levels in DBS from 15,974 participants, aged 50+ from Wave 6 of the Survey of Health, Ageing and Retirement in Europe (SHARE). RESULTS: The apoE4 protein signal was presented in two separable distributions. One distribution corresponded to carriers of at least one copy of the ε4 allele. Fieldwork cofounders affected protein levels but did not explain individual differences. DISCUSSION: Future research should investigate how genotype and apoE4 level interact with lifestyle and other variables to impact cognitive aging.