| Summary |
This study further examined the association between ADHD and the DRD4, DAT1, and 5HTT genes by testing their association with multivariate phenotypes derived from continuous measures of ADHD symptom severity. DNA was collected in 202 families consisting of at least one ADHD proband and at least one parent or sibling. VNTR polymorphisms of the DRD4 and DAT1 genes were significantly associated with the continuous ADHD phenotype. The association with DRD4 was driven by both inattentive and hyperactive symptoms, while the association with DAT1 was driven primarily by inattentive symptoms. These results use novel methods to build upon important connections between dopamine genes and their final behavioral manifestation as symptoms of ADHD. |
| Total Sample |
The current sample includes 202 families consisting of at least one ADHD proband and at least one parent, dizygotic twin, or sibling. |
| Sample Collection |
Participants completed the procedures described below as part of the Colorado Learning Disabilities Research Center (CLDRC) twin study, an ongoing study of the etiology of learning disabilities, ADHD, and other related disorders (e.g., DeFries et al. 1997; Willcutt et al. 2003). Recruitment and testing procedures have been described previously (i.e. Willcutt et al. 2003; Bidwell et al. 2007). |
| Diagnosis Description |
The Disruptive Behavior Rating Scale (DBRS; Barkley and Murphy 1998) was used to obtain parent and teacher ratings of the 18 symptoms of DSM-IV ADHD. Each symptom on the DBRS was rated on a four point scale (never or rarely, sometimes, often, and very often). Items rated as often or very often were scored as positive symptoms and items rated as never or rarely or sometimes were scored as negative symptoms, consistent with the procedure used in previous studies of similar rating scales (e.g., Pelham et al. 1992). The parent and teacher ratings of each of the 18 ADHD symptoms were coded (never or rarely = 0, sometimes = 1, often = 2, very often = 3) and combined using a modified 'or rule' algorithm, similar to that used in the DSM-IV field trials for disruptive behavior disorders (Lahey et al. 1994), by taking the higher of the two ratings. Because the focus of the overall project is on the etiology and correlates of familial ADHD, potential participants with a documented brain injury, significant hearing or visual impairment, or other rare genetic or environmental etiology (e.g., Down syndrome, Fragile X syndrome, or other sex chromosome anomalies) were excluded from the sample. In addition, participants were excluded if their Full Scale IQ score was below 75 on the Wechsler Intelligence Scale for Children, Revised (Wechsler 1974). |
| Technique |
DNA was obtained from blood samples, buccal swabs, or saliva (Oragene; DNAGenotek, Kanata, Ontario, Canada), and extraction was done using the appropriate Gentra PureGene kit (Qiagen, Valencia, CA) for the blood and buccal samples and with the Oragene procedure for saliva samples. Genotyping for the DRD4 and 5HTTLPR polymorphisms was done using a touchdown polymerase chain reaction (PCR) procedure. Genotyping for the DAT1 30 bp polymorphism was done by standard PCR. For more details about the genetic data acquisition and genotyping, please refer to the original publication. |
| Analysis Method |
Family Based Association Tests (FBAT) is a non-parametric extension of the transmission equilibrium test (TDT), which is robust to population stratification. FBAT used family data to test for an association between a genetic polymorphism and a disease or a quantitative trait. FBAT-GEE is an FBAT test that allows for continuous multivariate phenotypic data and tests multiple phenotypes simultaneously, and thus allows for testing of continuous phenotypes and avoids multiple testing. FBAT-GEE is robust to missing parental genotypes and assumptions about the distribution of the phenotype (Laird and Lange 2006). In order to take advantage of the available multivariate symptom data, the FBAT-GEE test (implemented in PBAT Version 3.5) was used to evaluate association with each candidate gene and the continuous phenotypes based on the DSM-IV ADHD symptoms. All genotypes that included a minimum of 20 families were tested. For more details about the phenotypes tested in the current study, please refer to the original publication. |
| Result Description |
FBAT-GEE analyses for association with the common ADHD phenotype: When testing the Total ADHD phenotype, there was a statistically significant association with the DRD4 gene, with the 4-repeat allele significantly associated (p=0.007) with higher ADHD symptom scores. A significant association was also found with the DAT1 gene with the 10-repeat allele significantly associated (p=0.02) with higher symptoms. When testing for association with 5HTT, no significant associations were found. FBAT-GEE analyses for association with symptom dimensions: Significant associations were found with the 4-repeat allele of the DRD4 gene and both the inattentive and hyperactive-impulsive symptom dimensions (p=0.007 and 0.003, respectively). With regard to DAT1, the 10-repeat (p=0.005) allele showed significant association for the inattentive symptoms, but not for the hyperactive-impulsive symptoms. |
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