2.3.2. Dopamine synthesis: the tyrosine hydroxylase gene

The rate-limiting step of dopamine biosynthesis is the conversion of tyrosine into dihydroxyphenylalanine (DOPA) by tyrosine hydroxylase (TH), which is then turned into dopamine by aromatic amino acid decarboxylase (Figure 1). A 4 bp repeat sequence (TCAT tetranucleotide repeat) in the first intron has been thoroughly investigated by Meloni s laboratory. Their latest in vitro study showed a quantitative silencing effect of the TCAT-repeat on TH gene expression, with the 3-, 5-, 8-, and 10-repeat alleles exhibiting a step-wise inhibition on transcriptional activity (Albanese et al., 2001). Association studies refer to these intronic microsatellite alleles as K1–K5, numbering them in a decreasing order, i.e., from K1 = 10-repeat to K5 = 6-repeat (other alleles are rare). Dopamine is converted to norepinephrine by dopamine beta hydroxylase (DBH) in the synaptic vesicles of noradrenergic neurons; therefore, polymorphisms of the DBH gene belong to the noradrenergic system. Interestingly, the DBH enzyme can be measured in the plasma, which allows for in vivo measurements of its genetic variants (reviewed by Gizer et al., 2009).

2.3.3. Dopamine clearance: the dopamine and norepinephrine transporter genes

Dopamine neurotransmission is efficiently terminated via dopamine transporter (DAT) in subcortical regions by taking back dopamine from the synapse to the presynaptic neuron (Figure 1A). DAT is the site of action of stimulant drugs such as cocaine, amphetamine, and methylphenidate, which is used in ADHD treatment. Coding region variants of the DAT gene (DAT1, official symbol SLC6A3) are rare; therefore, VNTRs and SNPs of the non-coding regions have been studied in association studies. There is a common 40 bp VNTR in the 3′ untranslated region (UTR) with repeat numbers between 3 and 13. The most frequent allele has 10 repeats, followed by the 9-repeat allele; the others are rare variants. The results of comparative studies of the most frequent variants are controversial using either reporter gene assays (Fuke et al., 2001; Miller and Madras, 2002; Mill et al., 2005a) or post-mortem brain expression data (Mill et al., 2002a; Wonodi et al., 2009). SPECT analyses appear to have more congruent results: Two independent studies have reported a similar difference in striatal transporter density when studying large (N = 96 and N = 79), healthy populations. Participants with at least one copy of the 9-repeat allele (9/9 and 9/10 genotype) had a significantly higher transporter density compared to the 10/10 genotype group (van Dyck et al., 2005; van de Giessen et al., 2009). Studying a 30 bp VNTR in intron 8 (the most frequent alleles were originally named as 2-repeat and 3-repeat, but recent association studies refer to them as 5-repeat and 6-repeat), Guindalini et al. (2006) showed that the 3-repeat allele had a reduced basal expression compared to the 2-repeat allele, however, this allele had a 3-fold higher induction compared to the 2-repeat allele in response to a KCl and forskolin challenge. The higher level of transcriptional activity of the 3-repeat allele was also demonstrated in post-mortem midbrain tissues (Brookes et al., 2007). Constructing haplotypes from the two DAT1 VNTRs are preferred in recent ADHD studies (Asherson et al., 2007; Rommelse et al., 2008; Stevens et al., 2009; Franke et al., 2010).

As mentioned in the previous section, cortical DAT availability is very low compared to that of the striatum, however, dopamine can be taken up by the norepinephrine transporter (NET) in the cortex, resulting in a different dopamine elimination in this brain region (Figure 1B). Similarly to the DAT1 gene, the NET gene (official symbol SLC6A2) does not contain many common non-synonymous polymorphisms, therefore, synonymous SNPs from exons (e.g., rs5569, also called as G1287A from exon 9) and intronic SNPs (e.g., rs2242447 from intron 13) have been selected to cover the whole gene in comprehensive association studies. Unfortunately, the different workgroups did not select the same sets of SNPs, making the comparison of single marker and haplotype analyses hard (Gizer et al., 2009). The only SNP indicated to have functional relevance is located 3081 bp upstream from the transcription start site (−3081 A/T, rs28386840). The −3081 T-allele showed decreased promoter activity compared to the A-allele (Kim et al., 2006).