Elsevier

NeuroImage

Volume 53, Issue 3, 15 November 2010, Pages 970-977
NeuroImage

Neural response to working memory load varies by dopamine transporter genotype in children

https://doi.org/10.1016/j.neuroimage.2009.12.104Get rights and content

Abstract

Inheriting two (10/10) relative to one (9/10) copy of the 10-repeat allele of the dopamine transporter genotype (DAT1) is associated with Attention Deficit Hyperactivity Disorder, a childhood disorder marked by poor executive function. We examined whether functional anatomy underlying working memory, a component process of executive function, differed by DAT1 in 7–12 year-old typically developing children. 10/10 and 9/10 carriers performed a verbal n-back task in two functional magnetic resonance imaging (fMRI) runs varying in working memory load, high (2-back vs. 1-back) and low (1-back vs. 0-back). Performance accuracy was superior in 9/10 than 10/10 carriers in the high but not low load runs. Examination of each run separately revealed that frontal–striatal–parietal regions were more activated in 9/10 than 10/10 carriers in the high load run; the groups did not differ in the low load run. Examination of load effects revealed a DAT1 × Load interaction in the right hemisphere in the caudate, our a priori region of interest. Exploratory analysis at a more liberal threshold revealed this interaction in other basal ganglia regions (putamen, and substantial nigra/subthalamic nuclei – SN/STN) and in medial parietal cortex (left precuneus). The striatal and parietal regions were more activated in 9/10 carriers under high than low load, and DAT1 differences (9/10 > 10/10) were evident only under high load. In contrast, SN/STN tended to be more activated in 10/10 carriers under low than high load and DAT1 differences (10/10 > 9/10) were evident only under low load. Thus, 10-repeat homozygosity of DAT1 was associated with reduced performance and a lack of increased basal ganglia involvement under higher working memory demands.

Introduction

Individual differences in executive function, the ability to control thoughts and actions in a goal-directed manner, are attributable at least in part, to genetic variation (reviewed in Goldberg and Weinberger, 2004). One approach to identifying sources of genetic variation has been to examine association with heritable disorders that include executive dysfunction. This approach has been productive in a common disorder of childhood, Attention Deficit Hyperactivity Disorder (ADHD), that is defined by symptoms of inattention, impulsivity, and hyperactivity that significantly impede executive function [e.g., Tower of London (Nigg et al., 2002)] and its component processes such as working memory (Nigg et al., 2002) and response inhibition (DeVito et al., 2009). Molecular genetic studies of ADHD found a small [< 4% (Waldman et al., 1998)] but significant association with a variable number of tandem repeat sequences (VNTR) in the 3′-untranslated region (UTR) of the gene (SLC6A3) coding for the dopamine transporter (DAT1) (first reported by Cook et al., 1995, and meta-analysis by Yang et al., 2007). ADHD was more prevalent in homozygous carriers of the 10-repeat allele (10/10) relative to heterozygous carriers (9/10) and number of hyperactive-impulsive symptoms increased with the number of 10-repeat alleles (Waldman et al., 1998). Furthermore, 10/10 typically developing children exhibited more hyperactivity symptoms (Mill et al., 2005). Thus, a functional polymorphism of DAT1 contributes to variability in phenotypic expression that is relevant to executive function.

The effect of the DAT1 VNTR on executive task performance is not conclusively known at present. As ADHD is associated with poor executive function and 10-repeat homozygosity is associated with ADHD, 10/10 carriers ought to perform worse on tasks of executive function or its component processes than 9/10 carriers. Indeed, typically developing 10/10 children had worse inhibitory performance [TEA-Ch Opposite Worlds task (Cornish et al., 2005), more errors of commission on the Continuous Performance Test (Loo et al., 2003)] relative to 9/10 children. Furthermore, 10/10 adults made more errors of commission on the Continuous Performance Test relative to 9 carriers [combined 9/9 and 9/10 carriers (Caldu et al., 2007)]. However, in a study of ADHD children, 10/10 carriers performed better on various tasks tapping working memory (e.g., Self-ordered pointing, Arithmetic and Digit Span subtests of the WISC-III) relative to 9/10 carriers (Karama et al., 2008). In contrast to findings showing DAT1 differences, 10/10 and 9-carrier adults did not differ in working memory performance (e.g., n-back task) (Bertolino et al., 2006, Bertolino et al., 2009, Caldu et al., 2007). Results from neuropsychological tasks of executive function also reveal a mixed picture. While 10/10 ADHD children performed better on the Tower of London than 9/10 carriers (Karama et al., 2008), performance on the Wisconsin Card Sorting Task did not differ by DAT1 (Barkley et al., 2006). These mixed findings cannot be explained by developmental stage (adults vs. children), diagnosis (ADHD vs. controls), or type of task. Thus, although 10-repeat homozygosity ought to be disadvantageous for executive function, task performance does not reveal a consistent pattern of results.

Three lines of evidence suggest that the influence of DAT1 VNTR on executive function is mediated by the striatum and its cortical projections. First, the dopamine transporter (DAT) reuptakes dopamine from extracellular space following release, and therefore, plays a role in regulating synaptic dopamine levels. It is expressed most abundantly in the striatum and other basal ganglia regions where it is the primary means of dopamine clearance from the synapse (Madras et al., 2005). Relative to the basal ganglia, DAT is found in lower concentrations in other regions such as parietal cortex (Lewis et al., 2001), hippocampus (Lewis et al., 2001), and cerebellum (Dahlin et al., 2007). It is minimally expressed in the prefrontal cortex where the main mechanism for dopamine clearance is the catechol-O-methyltrasferase (COMT) enzyme rather than DAT (Karoum et al., 1994). Thus, functional effects of individual variability in DAT expression are associated with the striatum most directly, relative to any other brain region.

Second, striatal DAT expression differs as a function of DAT1 alleles. In vitro studies found that higher DAT expression was associated with the 10-repeat allele (Fuke et al., 2001, Mill et al., 2002, VanNess et al., 2005). Indeed, DAT expression in the caudate measured by ligand-based in vivo imaging was higher in children with ADHD who were 10/10 carriers relative to 9/10 carriers (Cheon et al., 2005). However, findings from in vivo studies of typically developing subjects are mixed, showing greater (Heinz et al., 2000), reduced (Jacobsen et al., 2000, van de Giessen et al., 2009, van Dyck et al., 2005), or no different (Krause et al., 2006, Martinez et al., 2001) DAT availability in the caudate in 10/10 relative to 9/10 carriers. Higher striatal DAT expression may be functionally deleterious because it is likely to result in reduced dopamine signaling due to enhanced clearance. Indeed, higher DAT expression in the caudate has been observed in ADHD relative to control subjects [reviewed in (Spencer et al., 2005)] and is related to reduced attentional function mediated by parietal cortex (Tomasi et al., 2009). Together, these findings suggest that effects of DAT1 alleles on DAT expression are observed in the caudate and furthermore, differences in striatal DAT expression are functionally relevant.

Third, functional magnetic resonance imaging (fMRI) studies found that prefrontal–striatal activation during tasks tapping component processes of executive function was reduced in 10/10 relative to 9 carriers. During response inhibition, 10/10 ADHD children and their unaffected siblings had reduced activation in the caudate relative to 9/10 carriers (Durston et al., 2008). This difference was not observed in control children. In normal adults, prefrontal regions showed reduced activation in 10/10 relative to 9 carriers during n-back working memory performance (Bertolino et al., 2006, Bertolino et al., 2009). As DAT expression is minimal in the prefrontal cortex, these results may reflect differences associated with striatal projections to the prefrontal cortex. Parallel measurement of functional activation by fMRI and dopamine signaling by ligand-based positron emission tomography indicated that prefrontal–striatal activation during working memory relates to dopaminergic activity (Landau et al., 2009). Thus, prefrontal–striatal functional activation revealed by fMRI is sensitive to DAT1 allelic differences.

In the present study, we used fMRI to examine whether activation during the n-back working memory task differs by the DAT1 VNTR in 7–12 year-old typically developing children. This study fills two gaps in current knowledge about the role of DAT1 in executive function: First, it is not known whether DAT1 differences influence brain activation during typical development. Caudate activation differences observed in ADHD children and their unaffected siblings in Durston et al.'s (2008) study appear to reflect disorder-related and inherited alterations in DAT expression because they were not observed in control children. However, the small sample size of each genotype group in controls (n = 4) could have limited the ability to reveal activation differences in typical development. Furthermore, DAT1 differences observed in typically developing adults may not extend to childhood as they could depend upon maturation of prefrontal–striatal circuits through adolescence. Indeed, developmental functional imaging studies of working memory indicate that involvement of prefrontal and associated regions increases from childhood to adulthood (Crone et al., 2006, Klingberg et al., 2002). Second, it is not known whether neural response to added working memory load differs by DAT1 VNTR. fMRI studies show that prefrontal and striatal involvement increases with working memory demands (Chang et al., 2007, Wager and Smith, 2003). Studies in animals and humans suggest that those functional changes are mediated by load-dependent changes in dopaminergic signaling (Aalto et al., 2005, Floresco and Phillips, 2001). Thus, it is possible that individual differences in load-dependent functional activation relate to DAT1 allelic status.

Children in the present study performed a verbal n-back task under high and low working memory loads. The n-back task requires a response to the current trial if the stimulus letter matches that on a trial occurring n trials ago (e.g., 0, 1, 2). Thus, performance accuracy depends upon maintaining and updating of task-relevant stimuli and inhibiting interference from irrelevant stimuli, as trials proceed. Furthermore, varying n provides a manipulation for working memory load as maintenance, updating, and inhibitory demands increase with the size of n (e.g., low for 0, medium for 1, and high for 2). The low load fMRI run included n = 1 (termed 1-back) relative to n = 0 (termed 0-back) conditions and the high load fMRI run included n = 2 (termed 2-back) relative to 1-back conditions. We selected the n-back task for three reasons: (1) its requirements evoke working memory processes relevant to executive function; (2) it has been used successfully with fMRI in children as young as 8 years (Libertus et al., 2009); and (3) it is sensitive to DAT1 differences in past fMRI studies of adults reviewed above. We hypothesized that caudate activation would be lower in 10/10 than 9/10 children based upon Durston et al.'s (2008) findings, and furthermore, this difference would be enhanced under high working memory load (DAT1 × Load interaction) as more demanding cognition ought to be more sensitive to reduced dopaminergic signaling in those with higher DAT expression. While our hypothesis focused on the caudate because it is a striatal region with high DAT expression that is reliably engaged during working memory, we also explored DAT1 × Load interaction in the rest of the brain. Based upon the association of DAT1 VNTR and ADHD, we expected n-back accuracy to be worse in 10/10 than 9/10 children.

Section snippets

Participants

Twenty typically developing children between 7 and 12 years of age were recruited through advertisements in the Washington DC area and were paid for participation. Ten children were carriers of two copies of the 10-repeat allele (termed “10/10”) and 10 were carriers of one copy of the 10-repeat and 9-repeat allele (termed “9/10”). Children were genotyped following recruitment and consecutively enrolled into the study until equal samples of the two genotypes were obtained. The 9/10 group (7

Behavioral results

For each run, mean percent accuracy [percent hits (number of correct targets/total number of targets) minus percent false alarms (number of presses for non-targets/total number non-targets)] and mean reaction time (in ms) for hits were computed for each subject. Data from each run were analyzed separately to match the fMRI contrasts of interest, with a mixed ANOVA including DAT1 (9/10, 10/10) as a between-subject factor and Condition (Low WM Run: 1-back vs. 0-back; High WM Run: 2-back vs.

Discussion

The present results showed that the neural response to working memory load varied by DAT1 VNTR in typically developing 7–12 year-old children. n-back accuracy was superior in 9/10 than 10/10 carriers under high but not low working memory demands. Examination of each run separately revealed that a prefrontal–striatal–parietal network of regions was more activated in 9/10 than 10/10 carriers in the high load run; no DAT1 differences were observed in the low load run. Examination of load effects

Acknowledgments

This work was supported by grants from the National Institute of Mental Health MH065395-01 to CJV and MS MH70564 to MAS, and a Canadian Institutes for Health Research (CIHR) Doctoral Research Award to MS. We would like to thank Nick Parrott and Devon Shook for assistance with fMRI analyses.

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