Novel 5-HTTLPR Allele Associates with Higher Serotonin Transporter Binding in Putamen: A [ 11 C] DASB Positron Emission Tomography Study Nicole Praschak-Rieder, James Kennedy, Alan A. Wilson, Douglas Hussey, Anahita Boovariwala, Matthaeus Willeit, Nathalie Ginovart, Subi Tharmalingam, Mario Masellis, Sylvain Houle, and Jeffrey H. Meyer Background: The serotonin transporter (5-HTT)-linked polymorphic region (5-HTTLPR) has two frequent alleles, designated long (L), and short (S). The S allele is associated with lower levels of 5-HTT mRNA and lower 5-HTT expression in human cell lines. A functional single nucleotide variant was detected within L, designated L A and L G . Only L A is associated with high levels of in vitro 5-HTT expression, whereas L G is low expressing and more similar to S. We examined the possible influence of the long (A/G) variant on 5-HTT density in the living human brain using 3-(11)C-amino-4-(2-dimethylaminomethylphenyl-sulfanyl) benzonitrile ([ 11 C]DASB) positron emission tomography. Methods: The 5-HTT binding potential (5-HTT BP), an index of 5-HTT density, was found in 43 healthy subjects genotyped for 5-HTTLPR long (A/G), and in an ethnically homogenous subsample of 30 Caucasian-Canadians. Results: The L A /L A was associated with higher 5-HTT BP in putamen (p = .026, not corrected). This association became stronger in the Caucasian subsample (p = .004) and was significant even after correcting for multiple comparisons. Conclusions: The 5-HTTLPR long (A/G) polymorphism influences 5-HTT density leading to higher putamen 5-HTT BP in healthy L A /L A carriers of Caucasian ancestry. This finding extends the role of this polymorphism from in vitro reports of higher 5-HTT expression with the L A /L A genotype into in vivo brains of healthy human subjects. Key Words: [ 11 C] DASB, depression, healthy subjects, positron emission tomography, serotonin transporter, triallelic 5-HTTLPR T he serotonin transporter (5-HTT) modulates extracellular serotonin levels, and abnormal regulation of 5-HTT has been associated with a number of psychiatric diagnoses and psychopathological symptoms, such as depression, anxiety, impulsivity, and aggression (Mann et al 2000; Meyer et al 2004a; Neumeister et al 2004; Owens and Nemeroff 1994; Schloss and Henn 2004; Willeit et al 2000). The serotonin transporter-linked polymorphic region (5-HTTLPR) is a common polymorphism on chromosome 17q12 that impacts 5-HTT mRNA transcrip- tion in human cell lines (Heils et al 1995, 1996; Lesch et al 1996). The 5-HTTLPR is commonly subdivided into lesser expressing short (S) alleles, and greater expressing long (L) alleles. Even though some associations between 5-HTTLPR and psychiatric illness (Anguelova et al 2003; Hoefgen et al 2005; Lotrich and Pollock 2004) or psychiatric symptoms (Bondy et al 2006; Haberstick et al 2006; Hasler et al 2006; Heinz et al 2005; Willeit et al 2003) have been reported, 5-HTT density in the living human brain and postmortem was not consistently related to the differential transcriptional activity of 5-HTTLPR L and S alleles (Lim et al 2006; Willeit et al 2001; Shioe et al 2003; Mann et al 2000; Heinz et al 2000; Little et al 1998). Nakamura et al (2000) had described several novel variants of S and L. A single nucleotide variant (A to G) was detected at the sixth nucleotide within the first of two extra 20-23bp repeats of the 5-HTTLPR L allele, designated long A and long G (also designated  and  by Nakamura et al [2000], and rs25531 by Kraft et al [2005], respectively). Interestingly, only long A (L A ) is associated with high levels of 5-HTT mRNA transcription in vitro, whereas long G (L G ) is more similar to S with low levels of 5-HTT mRNA (Hu et al 2006). Thus, long (A/G), together with S, comprise a triallelic locus. The triallelic polymorphism is also preferentially associated with selective serotonin reuptake inhib- itor response (Kraft et al 2005; Wendland et al 2006). Given the triallelic nature of 5-HTTLPR, it can be speculated that 5-HTTLPR genotypes incorporating the L G variant will also better predict regional 5-HTT binding potential (5-HTT BP), an index for 5-HTT density, in the living human brain. However, a recent study found no association of the triallelic 5-HTTLPR polymorphism on 5-HTT BP using the 5-HTT radio- tracer [ 11 C] -(+)- McN 5652 and positron emission tomography (PET; Parsey et al 2006). Some of the limitations of [ 11 C]-(+)-McN 5652 PET were raised in this report and it was acknowledged that [ 11 C] DASB PET would have methodological advantages. [ 11 C]- (+)-McN 5652 has very high nonspecific uptake relative to specific binding and 5-HTT BP values are only detectable in the thalamus, and possibly the basal ganglia and midbrain. Thus, the sensitivity of this imaging technique may be low for detecting effects. Moreover, to our best knowledge, detailed reliability data for regional 5-HTT BP found with [ 11 C]-(+)-McN 5652 PET have not been published. The main advantages of [ 11 C] DASB are that it binds reversibly to 5-HTT with high affinity and high selectivity (Wilson et al 2000, 2002), has a much higher ratio of specific binding to free and nonspecific binding in vivo, and shows reliable measurements in multiple brain regions under test-retest conditions, including prefrontal cortex (Ginovart et al 2001; Houle et al 2000; Meyer et al 2004a, 2004b; Praschak-Rieder et al 2005). As a result [ 11 C] DASB PET is currently the optimal method for quantification of the 5-HTT BP in humans. Here we examine From the Vivian M. Rakoff Positron Emission Tomography Imaging Centre (NP-R, AAW, DH, AB, MW, NG, SH, JHM), and the Neurogenetics Section (JK, ST, MM), Centre for Addiction and Mental Health, University of Toronto, Toronto, Canada; and the Department of General Psychiatry (NP-R, MW), University of Vienna, Austria. Address reprint requests to Jeffrey Meyer, M.D., Ph.D., Vivian M. Rakoff PET Imaging Centre, Centre for Addiction and Mental Health, and Depart- ment of Psychiatry, University of Toronto, 250 College Street, Toronto, ON, M5T 1R8, Canada; E-mail: jeff.meyer@camhpet.ca. Received June 16, 2006; revised September 21, 2006; accepted September 25, 2006. BIOL PSYCHIATRY 2007;xx:xxx 0006-3223/07/$32.00 doi:10.1016/j.biopsych.2006.09.022 © 2007 Society of Biological Psychiatry ARTICLE IN PRESS