SPECIAL ISSUE: ORIGINAL ARTICLE THE COGNITIVE GENETICS OF ATTENTION DEFICIT HYPERACTIVITY DISORDER (ADHD): SUSTAINED ATTENTION AS A CANDIDATE PHENOTYPE Mark A. Bellgrove1,2,3, Ziarih Hawi2, Michael Gill2 and Ian H. Robertson1 (1Department of Psychology and Trinity College Institute of Neuroscience, Trinity College Dublin, Dublin, Ireland; 2Departments of Psychiatry and Genetics and Trinity College Institute of Neuroscience, Trinity College Dublin, Dublin, Ireland; 3Cognitive Neuroscience Laboratory, University of Melbourne, Victoria, Australia) ABSTRACT Here we describe the application of cognitive genetics to the study of attention deficit hyperactivity disorder (ADHD). Cognitive genetics owes much to the pioneering work of cognitive neuropsychologists such as John Marshall, whose careful observations of cognitive dissociations between brain-lesioned patients greatly advanced the theoretical understanding of normal cognitive function. These theories have in turn helped to constrain linkages between candidate genes and cognitive processes and thus help to drive the relatively new field of cognitive genetics in a hypothesis-driven fashion. We examined the relationship between sustained attention deficits in ADHD and genetic variation in a catecholamine-related gene, dopamine beta hydroxylase (DβH). DBH encodes the enzyme that converts dopamine to noradrenaline and is crucial to catecholamine regulation. A polymorphism with the DBH gene has been associated with ADHD. In fifty-two children with ADHD, we examined whether variation in the Taq I DBH gene polymorphism was related to sustained attention performance. Participants performed the Sustained Attention to Response Test (SART). Performance on the SART discriminates ADHD from control children, and in imaging work, is associated with right frontoparietal activation. A significant effect of DBH genotype was found on SART performance measures. Children possessing two copies of the ADHD-associated risk allele (A2) had significantly poorer sustained attention than those ADHD children who did not possess this allele or a non-genotyped control group. The DBH gene may contribute to the susceptibility for ADHD, in part because of its varying effects on the development of brain mechanisms mediating sustained attention. Key words: genetics, attention, sustained attention, dopamine, ADHD INTRODUCTION In recent times there has been increased interest within cognitive neuroscience in the role played by genetics in cognitive processes. This paper does not seek to explicitly review the application of molecular genetics to cognitive neuroscience- for this the reader may refer to a number of recent and excellent summaries (Goldberg and Weinberger, 2004; Parasuraman and Greenwood, 2004). Our purpose is, however, to provide an example of this approach from our own recent work on attention deficit hyperactivity disorder (ADHD). In our view the application of molecular genetics to studies of cognition – cognitive genetics – follows strongly in the tradition of cognitive neuropsychology. In the latter, the effects of localised cerebral lesions on specific cognitive functions have thrown considerable light on the cognitive architecture of the normally functioning human brain (e.g., Broca, 1824-1880; Wernicke, 1848-1905). For example, the syndrome of unilateral spatial neglect, in which patients display deficits in detecting and acting upon contralesional stimuli is most frequent and severe following lesions centred on the posterior parietal lobe of the right cerebral hemisphere (Robertson and Marshall, 1993). The central role played by this region in human spatial attention has been confirmed by Cortex, (2006) 42, 838-845 numerous neuroimaging studies with healthy participants that have shown activity in the right temporo-parietal junction, particularly when an observer must shift their attention from an invalidly cued location to detect a target at a contralateral location (Corbetta et al., 2000; Corbetta and Shulman, 2002). In the case of cognitive genetics, specific allelic variants of candidate genes (see Table I) take the place of specific cerebral lesions, and so a linkage is made between high level cognitive functions on the one hand, and molecular genetics on the other. Such linkages are essential to test contemporary models of attention (Corbetta and Shulman, 2002; Posner and Peterson, 1990) which include variables across very different levels of analysis – cognitive, anatomical, neurochemical and now genetic. For example, Parasuraman et al. (2005) have recently shown that visuospatial attention is substantially influenced by allelic variation in a polymorphism of the gene, CHRNA4, encoding a nicotinic acetylcholine receptor subunit (alpha-4 subunit). Focus on this candidate gene was warranted since pharmacological studies of primates show that visuospatial attention can be modulated by acetylcholine (Witte et al., 1997) and impairments in visuospatial attention are prominent in neurodegenerative diseases affecting basal forebrain cholinergic pathways (e.g., Alzheimer’s disease). Sustained attention, genetics and ADHD 839 TABLE I Glossary of terms Term Allele Candidate gene Intron Single nucleotide polymorphism (SNP) Linkage disequilibrium (LD) Definition An alternative form of a gene that can occur at a given locus on a gene. Alleles differ in their nucleotide sequence. A single allele for each locus of a gene is inherited from each parent. A gene located on a chromosomal region that is thought to be involved in a trait (e.g., cognitive process) or a disorder (e.g., ADHD). Candidate genes are normally chosen based upon prior physiological, genetic or biochemical characterisation that leads one to suspect that this gene is involved in the trait. A nucleotide sequence in a gene that does not code for the gene product. Introns can be compared to exons which are the protein coding portion of a gene. Different versions of a gene, due to differences in nucleotides on DNA, are called polymorphisms. The most common form of genetic variation, the SNP, reflects a change in a single base in the DNA that differs from the usual base at that position. LD occurs when alleles at different loci occur together within an individual at a greater than chance frequency. Specifically, Parasuraman et al. (2005) found that allelic variation in CHRNA4 affected both the pattern of benefits and costs elicited by an endogenous cuing manipulation. The study by Parasuraman et al. (2005) provides important first insights that individual differences in visuospatial attention may be at least partially explained by underlying genetic influences. Before turning our attention to the cognitive genetics of ADHD, it is necessary to review the empirical work that bears directly upon the neuroanatomical models that guide our selection of both candidate genes and candidate cognitive processes in ADHD. A Neuroanatomical Model of Sustained Attention Our own neuropsychological and rehabilitation research on the attentional sequelae of traumatic brain injury (TBI), stroke, and more recently ADHD, has been heavily influenced by neuroanatomical models of attention that propose distinct attentional systems for the control of spatial attention and non-spatial alertness (Posner and Peterson, 1990; Pardo et al., 1991; Wilkins et al., 1987). We have defined non-spatial alertness or sustained attention, as the internal control of alertness in the absence of exogenous support. Spatial selectivity, on the other hand, serves to enhance perception at certain locations in space. A number of sources of information converge to suggest a predominant role of the right prefrontal cortex in human sustained attention. First, it is well established that right frontal lesions lead to an increase in reaction time when an imperative stimulus is not preceded by a warning signal (Posner et al., 1987). Frontal patients do nevertheless benefit from a warning signal indicating that it is the intrinsic and not phasic aspects of alertness that are impaired with right frontal lesions (Posner et al., 1987; Sturm et al., 1999). Wilkins et al. (1987) demonstrated that patients with right frontal lobe lesions were specifically impaired when required to perform a simple tone counting task of a slow event rate. These authors concluded that a monotonous task demand impaired the right frontal patient’s ability to sustain attention. A right frontal focus for sustained attention is also consistent with the cortical distribution of noradrenergic neurons arising predominantly from the locus coeruleus (Marrocco et al., 1994). These neurons contain DβH which is the enzyme responsible for the conversion of dopamine to noradrenaline. Accordingly, drugs that suppress NA function also impair sustained attention in humans (Smith and Nutt, 1996) but these deficits can be reversed by the provision of phasic alerts that extrinsically release NA from the locus coeruleus. Functional imaging studies have generally confirmed this model by showing that sustained attention is achieved through the reciprocal interaction between cortical and sub-cortical areas. Specifically, the right dorsolateral prefrontal cortex, anterior cingulate and inferior parietal lobe act to exert top-down (endogenous) control, via thalamic nuclei, over the locus coeruleus to promote NA release (Manly et al., 2003; Pardo et al., 1991; Sturm et al., 1999; Sturm and Willmes, 2001; Sturm et al., 2004; Coull et al., 1999). It follows that the sustained attention deficits of right frontal patients may stem from a failure to endogenously regulate alertness in a top-down fashion, and that activating intact phasic alertness systems may therefore ameliorate these deficits. In support of this relationship we have argued that the sustained attention deficits in TBI arise from a failure to endogenously maintain alertness (Robertson et al., 1997a; Manly et al., 2003) and have shown that phasic alerting cues can improve the ability of TBI patients to effectively maintain task goals (Manly et al., 2002). More recently, we have also demonstrated that sustained attention deficits in children with ADHD (O’Connell et al., 2004) can be ameliorated by the provision of random phasic alerts designed to heighten activity within RH networks (O’Connell et al., in press). Posner and Peterson (1990) argued that the alertness, or sustained attention system, could coactivate the spatial attentional system of the parietal lobe, either directly or via brainstem nuclei. Clinical support for this notion comes from the existence of 840 Mark A. Bellgrove and Others deficits in non-spatial alertness in patients with right parietal lesions and the neglect syndrome. Working with 44 RH patients, Robertson et al. (1997b) demonstrated that impairments on a simple tone counting task were a significant predictor of spatial bias, as assessed using a range of measures of unilateral spatial neglect. While deficits in sustained and spatial attention could both be a consequence of RH damage and co-exist independently, additional evidence from both clinical and non-clinical groups suggests a more dynamic interplay. First, Robertson et al. (1995, 1998) demonstrated that the extent of chronic left unilateral neglect can be substantially reduced by sustained attention training or by the provision of random phasic alerts. Second, taking an individual differences approach, we have demonstrated that variation in the capacity for sustained attention is accompanied by variation in the attentional bias seen on clinical tests of neglect, in both healthy children (Dobler et al., 2005) and adults (Bellgrove et al., 2004). For example, Bellgrove et al. (2004) asked healthy undergraduates to perform both the Sustained Attention to Response Task (SART), in which participants must withhold their response to infrequent no-go events, and the Greyscales Task, a perceptual measure of attentional bias. Performance on the SART correlates highly with everyday lapses of attention and distinguishes TBI patients (Robertson et al., 1997a; Manly et al., 2003) and ADHD children from controls (O’Connell et al., 2004). Much research with the Greyscales Task has shown a reliable leftward bias that is consistent with the phenomenon of pseudoneglect (Nicholls et al., 1999; Mattingley et al., 1994). In Bellgrove et al. we found that participants who performed poorly on the SART also had a significantly attenuated leftward bias (i.e., as in neglect, the orienting bias of the RH was weakened), relative to those who performed well on the SART. We suggested that the attenuated leftward bias on the part of the poor sustained attenders, relative to the good sustained attenders, arose from individual differences in the efficiency with which fronto-parietal networks of the RH control visual attention. Finally, Dobler et al. (2003) have also documented a case of developmental unilateral neglect in a boy referred to clinical services for attentional problems. Objective testing confirmed the existence of both a profound sustained attention deficit and a left-sided inattention. Strikingly, the extent of left-sided inattention was markedly reduced by the same phasic alerting procedures used in adult patients with RH lesions and neglect (Robertson et al., 1997a). The Cognitive Genetics of ADHD ADHD is a childhood disorder that is defined by age-inappropriate levels of inattention, hyperactivity and/or impulsivity (American Psychiatric Association, 1995). Family, twin and adoption studies all suggest that the disorder is strongly heritable (see Kirley et al., 2002 for review). Stimulant medications, such as methylphenidate have efficacy in treating the cognitive and behavioural features of the disorder. Stimulants act by inhibiting the re-uptake and promoting the release of catecholamines (including dopamine and noradrenaline) (Krause et al., 2003). In support of catecholamine hypotheses, candidate gene studies have found associations with variants at a number of genetic loci including the dopamine transporter gene (Gill et al., 1997; Daly et al., 1999; Cook et al., 1995; Waldman et al., 1998), dopamine receptor genes (e.g., DRD4 and DRD5) (Daly et al., 1999; Faraone et al., 2001) and the gene (DBH) encoding DßH (Daly et al., 1999). Each of these variants may confer a small part of the genetic susceptibility to ADHD. A limited number of studies have examined whether genetic variants that are thought to confer susceptibility to ADHD are also associated with cognitive impairment (Langley et al., 2004; Swanson et al., 2000; Manor et al., 2002; Bellgrove et al., 2005a, 2005b, 2005c, 2005d). The approach of linking a genetic risk factor to an alternative, or intermediate, cognitive phenotype has much intuitive appeal. Rather than seeking associations between a gene and a subjective diagnostic category, here association is sought between a gene and a well-operationalised, objectively measured cognitive process. In the pursuit of such linkages it is worth bearing in mind that “a poorly-defined phenotype is unlikely to facilitate identification of the underlying genotype” (Baddeley, 1996, p.186). When, however, the cognitive process itself is well understood in cognitive-neuroanatomical terms, then progress may be made in determining how the gene contributes to variation in the development of brain mechanisms modulating that cognitive process. Associations of this type will prove powerful in elucidating the brain-behaviour basis of conditions, such as ADHD. In our own genotype/phenotype studies of ADHD, we have employed a sustained attention phenotype. As reviewed above, sustained attention is thought to be achieved throughout a frontoparietal network and to be modulated by catecholamines. A large body of evidence supports the existence of sustained attention deficits in children, adolescents and adults with ADHD (Seidman et al., 1998; Epstein et al., 2001; Shallice et al., 2002; O’Connell et al., 2004). Further, such deficits are ameliorated by methylphenidate (Solanto, 1998). Groot et al. (2004) recently examined familial influences on sustained attention in preschoolers using the Amsterdam Neuropsychological Tasks. Correlations of performance measures were generally higher in monozygotic than dizygotic twin pairs suggesting genetic influence. Heritability 841 Sustained attention, genetics and ADHD estimates for sustained attention measures were, however, generally moderate (.46-.72) and modelfitting suggested the influence of both genetic and unique environmental effects. Using an affectedsibling pair design within an ADHD cohort, SlaatsWillemse et al. (2005) recently showed significant sib-pair correlations for aspects of sustained attention. These studies provide preliminary evidence for genetic influences on sustained attention in ADHD and non-ADHD samples. Although the heritability of sustained attention may be less than for the ADHD diagnosis itself, sustained attention measures may still be informative for molecular genetic studies if the magnitude of the association between the gene and cognitive phenotype is larger than between the gene and the clinical phenotype (Doyle et al., 2005). In our work we have used the SART to examine sustained attention in ADHD. The SART is a short (5 minute) go/no-go test in which participants are presented, for example, with a fixed sequence of 1-9 digits and are required to respond to the presentation of digits (go-trials), except when the digit “3” (no-go trial) is presented. Withholding to a rare target, as opposed to responding to a rare target in the case of CPT-like tasks, shifts the automatic response set to the nontargets and thus successful withholding places greater demands on the sustained attention system in order to overcome the pre-potent non-target response (Robertson et al., 1997a). This is particularly true in a fixed sequence version of the SART, where the no-go target (‘3’) occurs predictably in the 1, 2, 3… 9 cycle. This version is sensitive to frontal lobe dysfunction (Manly et al., 2003) and minimizes the inhibitory components of the task that may be apparent in, for example, a random sequence (Fassbender et al., 2004) (see also Bellgrove et al., 2005d). The specificity of the SART for indexing a sustained attention deficit in children and adolescents with ADHD, relative to matched controls has been demonstrated (Shallice et al., 2002; O’Connell et al., 2004). Here we examined the performance of children and adolescents with ADHD on the SART, in relation to DBH genotype. DßH is the enzyme catalysing the conversion of dopamine to noradrenaline and is critical to catecholamine regulation in the brain. An association between ADHD and the A2 allele of a Taq I polymorphism (maps to intron 5) of the DBH gene has been reported (Daly et al., 1999; Roman et al., 2002). Since activity within fronto-parietal networks subserving sustained attention is modulated by input from DβH-containing noradrenergic neurons, we predicted that DBH genotype would relate to sustained attention deficits in ADHD. No studies have examined the relationship between DBH genotype and a quantitative sustained attention phenotype either in normality or in pathological conditions, such as ADHD. We predicted that sustained attention would be affected by DBH genotype, with ADHD children possessing two copies of the A2 allele of the Taq I polymorphism displaying greater impairment than those not possessing this allele. MATERIALS AND METHODS Subjects Fifty-two children and adolescents with ADHD (45 male, 48 right-handed) were recruited as part of ongoing genotype/phenotype studies within our laboratory. The mean age of this sample was 12.2 years (SD = 2.3) and the mean IQ, as assessed by the WISC-III, was 97 (SD = 11.2)1. Exclusion criteria included known neurological conditions including pervasive developmental disorders and epilepsy. All children met DSM-IV criteria for ADHD. Diagnostic details can be found in Kirley et al. (2002). 75% of these participants met diagnostic criteria for ADHD-Combined Type (ADHD-CT), 15% for ADHD-Predominantly Inattentive Type (ADHD-In) and 10% for ADHDPredominantly Hyperactive-Impulsive Type (ADHD-H/I). 73% of the ADHD probands met diagnostic criteria for other disorders such as Oppositional Defiant Disorder (ODD) and Conduct Disorder (CD). Stimulant medication was withdrawn at least 24 hours prior to the neuropsychological testing. These 52 participants were divided according to the presence of the A2 allele of the Taq I DBH gene polymorphism, yielding three DBH genotype groups, henceforth referred to as “No High-Risk” (n = 15), “One High-Risk” (n = 17) and “Two High-Risk” (n = 20), respectively. These groups did not differ in terms of Age [F (2, 49) = .72, p = .49], IQ [F (2, 48) = 1.29, p = .28], frequency of the three ADHD Sub-types (ADHD-CT: χ22 = 3.9, p = .14; ADHD-In: χ22 = .55, p = .76; ADHD-H/I: χ22 = 4.35, p = .11) or in terms of parental ratings of symptomatology as assessed by the Conners rating scale (Conners, 1998). A comparable group of 21 healthy control children (18 = male, 20 = right-handed) was also recruited from schools in and around Dublin. Control children had a mean age of 12 years (SD = 1.29) and a mean IQ of 103 (SD = 11.5). There were no significant differences between the control children and the ADHD DBH-genotype groups in terms of age [F (3, 69) = .63, p = .60] or IQ [F (3, 68) = 2.13, p = .10]. DBH Genotyping DNA was extracted from blood samples or buccal cells from the ADHD proband in each family. Primer 1Note that IQs were not available for 1 of the ADHD participants. 842 Mark A. Bellgrove and Others sequence and amplification conditions can be found elsewhere (Daly et al., 1999). The Sustained Attention to Response Task (SART) Briefly, the SART is a short (5 minute) test in which participants are presented, for example, with a fixed sequence of 1-9 digits and are required to respond to the presentation of digits (go-trials), except when the digit “3” (no-go trial) is presented. In order not to disadvantage ADHD children with an impulsive response style, participants were required to respond upon the presentation of a predictably timed response cue. Participants performed 225 trials, incorporating 25 no-go trials. Dependent measures included: a) Commission errors (i.e., failure to withhold to the no-go ‘3’); b) Omission errors (i.e., failures to respond to the go digits); c) Total errors (a composite of commission and omission errors); d) Mean RT (msec) to the go digits (GoRT) and e) Response variability (defined as the SDGoRT/MeanGoRT). The latter measure adjusts for differences in RT making it suitable for comparing intra-individual variability between patient and control groups (Manly et al., 2000; Stuss et al., 2003). RESULTS Performance of the ADHD participants was standardized relative to that of the control group. Z-scores for each of the DBH-genotype groups for each of the dependent variables are presented in Table II. Z-scores were compared across the four groups (Control vs. No High-Risk vs. One High-Risk vs. Two High-Risk DBH groups) using univariate analysis of variance (ANOVA). Post-hoc comparisons were performed using the GamesHowell Test. There was a significant effect of Group on errors of commission [F (3, 69) = 3.16, p = .03, η2 = .12], that was driven by the significant difference TABLE II Fixed SART performance as a function of ADHD DBH genotype groups. Performance of each of the ADHD DBH genotype groups on each of the Fixed SART measures are displayed as mean (SD) of z-scores. Performance of the control sample is not shown since this group, by definition, had a mean of zero (SD = 1) DBH genotype group (Number of copies of allele 2) 0 (N = 15) 1 (N = 17) 2 (N = 19) Fixed SART Mean (SD) Mean (SD) Mean (SD) Commission Errors Omission Errors Total Errors Mean Go-RT Mean Response Variability .03 (1.06) .54 (1.53) 1.05 (1.27) .67 (1.99) 3.59 (4.17) 3.01 (3.78) .36 (1.53) 2.11 (2.71) 2.19 (2.40) .71 (.88) .44 (1.54) .62 (1.14) .58 (1.03) 1.37 (1.95) 1.21 (1.70) between the control group and the Two High-Risk DBH group (p = .028). The Two High-Risk DBH group made more errors of commission than the No High-Risk DBH group (p = .06). There was a significant effect of Group on omission errors [F (3, 69) = 6.27, p = .001, η2 = .21]. Post-hoc comparisons revealed that the controls differed significantly from both the One and Two High-Risk DBH groups (p = .014, p = .012, respectively). After controlling for Group, there was a significant correlation between errors of commission and omission [r2 = .28, p = .001]. There was a significant effect of Group on the composite measure of total errors (commission plus omission) [F (3, 69) = 6.14, p = .001, η2 = .21]. Post-hoc comparisons indicated that controls differed significantly from both the One (p = .03) and Two High-Risk DBH (p = .004) groups. Additionally, the Two High-Risk DBH group made significantly more total errors than the No HighRisk DBH group (p = .045). There was no effect of Group for Go RT [F (3,69) = 1.41, p = .25, η2 = .06] but there was for response variability [F (3, 69) = 3.50, p = .02, η2 = .13]. Post-hoc comparisons indicated that controls differed significantly from the Two HighRisk DBH group. Using the scores of the ADHD participants alone, regression was performed to examine the parametric effect of the A2 allele of the Taq I DBH polymorphism. Significant effects were observed for errors of commission [F (1, 50) = 5.45, p = .02, r2 = .08] and total errors [F (1, 50) = 4.97, p = .03, r2 = .07]. DISCUSSION In this paper we have outlined an approach whereby discrete cognitive processes may be linked to specific genes. We have termed this approach cognitive genetics and have provided one example of its application in ADHD research. In the example given, we investigated the relationship between sustained attention and DBH genotype in ADHD. We presented preliminary data that the A2 allele of the Taq 1 polymorphism of the DBH gene is associated with sustained attention deficits in children and adolescents with ADHD. ADHD participants who possessed one or two copies of this allele performed more poorly than nongenotyped control children. Comparisons within the ADHD sample indicated that possession of two copies of the A2 allele was associated with significantly poorer sustained attention than nonpossession of this allele. The observation that sustained attention in ADHD is affected by a DBH gene variant is significant for several reasons. First, the results suggest a neurocognitive phenotype that is related 843 Sustained attention, genetics and ADHD to variation in the DBH gene in ADHD. A number of studies have now demonstrated an association between ADHD and a number of susceptibility loci (Hawi et al., 2003a) including the Taq I DBH gene polymorphism (Daly et al., 1999; Roman et al., 2002). An important innovation within such studies is to use continuously quantifiable phenotypes that might relate more closely to dysfunction within discrete neural systems, than do symptom-based categories. The use of such refined phenotypes may increase the power of genetic studies to detect associations with genes of small effect (Castellanos and Tannock, 2002). We have provided preliminary evidence that the association of ADHD with the A2 allele of the Taq I DBH gene polymorphism may be related, in part, to its effects on sustained attention. Second, while noradrenaline has been proposed as the dominant neuromodulator of the sustained attention network, it seems likely that other catecholamines may also play an important role. In the context of ADHD research, genetic variants within the dopamine system have also been found to be associated with sustained attention deficits (Bellgrove et al., 2005c, 2005d; Loo et al., 2003; Manor et al., 2002). Given the central role of DBH in the regulation of catecholamines in the brain, it seems unlikely that changes within the noradrenergic system would not be associated with concomitant changes within the dopaminergic system. Since the prefrontal cortex is exquisitely sensitive to fluctuations in both noradrenaline or dopamine (Arnsten, 1998), dysfunction within either system may impair the reciprocal relationship between top-down and bottom-up influences that seems necessary for sustained attention. There are several limitations to the preliminary study described herein. A critique of these limitations may prove instructive in highlighting methodological issues in cognitive genetics. While we have demonstrated in this study that the Taq I DBH polymorphism is relevant to sustained attention in ADHD, we have not demonstrated that it is relevant to sustained attention in healthy populations. We have, however, recently examined the effect of this polymorphism on sustained attention in healthy undergraduate students and have observed an association between the A2 allele of the Taq I polymorphism and relatively poorer sustained attention (Bellgrove et al., in preparation). Given the small sample size of the current study and the moderate heritability of sustained attention, our results should nevertheless be viewed as preliminary until replicated in an independent sample. While the Taq I DBH polymorphism has been associated with ADHD within our Irish studies, it is unlikely to be a functional variant given its intronic location. A number of other markers exist within the DBH gene that are thought to be functional; that is allelic variation within a polymorphism located within the DBH gene leads to variation in the expression of the gene’s protein product in the brain. In the case of DBH, genetic variation may lead to an alteration in the expression of DβH, the enzyme converting dopamine to noradrenaline. DβH occurs in the plasma as a heritable trait and is strongly related to variation in the DBH gene (Cubells and Zabetian, 2004). Linking plasma DβH levels to a DBH gene variant therefore helps to establish that variant’s functional candidacy. Parasuraman et al. (2005) recently reported that a single nucleotide polymorphism (SNP), a G to A substitution at 444 within exon 2 of the DBH gene, associated with spatial working memory in controls. The A allele of this variant is associated with lower plasma and cerebrospinal fluid (CSF) levels of DβH, relative to the G allele. The G444A polymorphism is in linkage disequilibrium with the ADHD-associated Taq I polymorphism within the Irish population (D’ = .85; Hawi et al., 2003b). Linkage disequilibrium refers to two markers in close physical proximity such that a specific allele at one locus predicts the existence of specific alleles at other loci (Cubells and Zabetian, 2004). Given that sustained attention may be a separable factor in maintaining the integrity of behavioural goals in working memory (Robertson, 2004), the results of Parasuraman et al. (2005) therefore overlap somewhat with those reported herein. A plausible biological hypothesis for ADHD might therefore be that polymorphisms within the DBH gene lower the expression of DβH by diminishing DBH gene transcription. This scenario would lead to a relative elevation in the dopamine to noradrenaline ratio and thus contribute to sustained attention deficits. Future work in ADHD should therefore examine multiple functional markers within the DBH gene in order to elucidate its role in the sustained attention deficits in ADHD. In this endeavour a polymorphism within the promoter region of the gene (– 1021C-T) has recently been identified as the main variant controlling plasma DβH levels (Cubells and Zabetian, 2004). In summary, we have provided an example of a cognitive genetic approach to ADHD by showing that sustained attention deficits can be predicted by DBH genotype. 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