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Serotonin Transporter Linked Polymorphic Region: From Behavior to Neural Mechanisms
Commentary
Biological Psychiatry
Serotonin Transporter Linked Polymorphic Region: From Behavior to Neural Mechanisms Lukas Pezawas The clinical efficacy of selective serotonin reuptake inhibitors that exert their therapeutic action via the serotonin transporter protein (SERT), which is coded by a single gene (SLC6A4), has fueled research over the past 3 decades. The first report on a functional degenerate repeat polymorphic region within SLC6A4 (serotonin-transporter-linked polymorphic region [5HTTLPR]), associated with anxiety-related personality traits (1), made this genetic variant subject to numerous studies in animals, healthy humans, and psychiatric patients. A mechanistic account of how 5-HTTLPR might moderate psychiatric disease was first demonstrated by a study (2) that showed that the less transcriptionally efficient short (S) allele of 5-HTTLPR amplifies depressive symptoms in the presence of environmental adversity. However, despite the overwhelming number of positive reports, subsequent meta-analytical studies have battled over the reality of such a gene-environment interaction in patients. Thus, a collaborative meta-analysis to draw final conclusions has been launched. Similarly, the originally reported association between 5HTTLPR and anxiety-related personality traits has been challenged several times and in its current state is inconclusive at best. This lack of robust reproducibility of the original findings has been frustrating to many researchers in the field because it questioned the idea of a simple relationship between behavior and a promising candidate gene for psychiatric illness. However, one might also find it intriguing that an association between a molecular biological feature, such as 5-HTTLPR, and a phenomenologically operationalized psychiatric nosology or a psychological construct based on subjective ratings has been found in the first place, given the absence of any biological criterion within their definitions. Genes do not code for psychopathology or personality, but for proteins that may have a direct impact on cell function. Therefore, an intermediate phenotype approach (3) was developed using genetic data as independent variables and imaging measures as dependent variables. This research strategy aimed to establish a more biologically meaningful relationship between a psychiatric risk gene and psychopathology by measuring neural circuit function instead of psychopathology itself. An important reason for this procedure was that behavioral genetics has taught us that changes in brain function related to genetic variation may or may not result in behavioral readouts. Initial studies were conducted primarily in healthy control subjects because of the suspected biological heterogeneity of patient samples and disease-related epiphenomena that might affect neural circuit function (e.g., medication, highly variable symptoms, social deprivation), among other reasons. This research strategy was intended to enable the development of novel and more complex disease models
that consider the individual variation of neural wiring and function as opposed to oversimplified biological traitbehavior models. The aforementioned approach has been tremendously successful and highlighted several effects of 5-HTTLPR on a brain systems level, such as frequently replicated increased amygdala reactivity in S-allele carriers using fearful faces as stimuli (4), which is in line with work in animals demonstrating the importance of the amygdala in fear conditioning. Given the developmental role of serotonin in shaping mood circuitry, it is not surprising that the S-allele of 5-HTTLPR has also been shown to decrease subgenual anterior cingulate cortex and amygdala volume, possibly leading to increased amygdala reactivity via a corticolimbic circuitry encompassing both structures (5). The importance of this mood circuitry has been highlighted in numerous human and animal studies investigating emotion processing and treatment over the years. In addition, 5-HTTLPR effects have been shown in various brain regions or systems that were previously related to fear or emotion processing using different functional paradigms and resting state in healthy subjects and patients. However, the increasing number of 5-HTTLPR imaging studies has demonstrated that the overall reported effect size of 5-HTTLPR across diverse samples and study protocols has been small. Although meta-analytical evidence was able to verify initial reports in healthy subjects, results derived from patient studies are still inconclusive. Consequently, many groups have launched studies with far more complicated genetic models to address the apparent complexity of genetic effects on neural systems, including epistasis, epigenetics (6), and geneenvironment interactions (7), which revealed stronger effect sizes than initial studies associating a single genetic variant to neural networks. In this issue of Biological Psychiatry, Klumpers et al. (8) report novel data on the effects of 5-HTTLPR on brain function using two functional magnetic resonance imaging paradigms, classic fear conditioning and instructed fear, in two independent samples of healthy subjects. They report strongly overlapping activation increases in both paradigms encompassing the dorsal anterior cingulate cortex and adjacent dorsomedial prefrontal cortex, anterior insula, ventral striatum, and thalamus among other areas, but not the amygdala. The latter finding is supported by meta-analytical evidence of human imaging studies, the results of which contradict animal work. An investigation of 5-HTTLPR-moderated changes of corticolimbic circuitries has not been pursued by Klumpers et al. The authors argue that 5-HTTLPR moderates activation of the salience network, which is anchored in the dorsal anterior cingulate cortex and anterior insula and further encompasses
SEE CORRESPONDING ARTICLE ON PAGE 582
522 & 2015 Society of Biological Psychiatry Biological Psychiatry October 15, 2015; 78:522–524 www.sobp.org/journal
http://dx.doi.org/10.1016/j.biopsych.2015.08.010 ISSN: 0006-3223
Biological Psychiatry
Commentary
subcortical structures such as the amygdala, ventral striatum, and substantia nigra/ventral tegmental area. This neural system has shown to be robustly engaged in most imaging studies in healthy subjects and psychiatric patients using cognitive or emotion-related paradigms or resting state. Apart from its function in salience detection, it is thought to facilitate switching between large-scale brain networks such as the central executive and default mode network. The authors further demonstrate the convergence of their findings on a psychophysiologic level. They conclude that 5-HTTLPR affects the interindividual variability in anticipatory responses to threat via dorsomedial prefrontal cortex and speculate that this mechanism might be relevant for stress-related psychopathology. Although Klumpers et al. put forth a compelling argument, their argument raises several important questions for the field of fear and threat research, which have broad implications for imaging genetics studies investigating 5-HTTLPR. The first question concerns reproducibility. Using their paradigms, the authors did not detect any 5-HTTLPR-moderated amygdala activation, which is surprising given the large body of available evidence of amygdala hyperreactivity and corticolimbic dysfunction in animals and human imaging genetics studies. Engaging robust amygdala activation, specifically in the context of genetic studies, is still a challenging task for many research groups. The reasons for this may simply be related to magnetic resonance imaging hardware and software issues (e.g., resolution of image acquisition, choice of imaging sequence), physiologic artifacts (e.g., vessels), or the design of the applied paradigm itself. At the present time, no data repository is available where researchers can easily retrieve information on reliability of imaging measures regarding a specific paradigm, which would facilitate the use of the most robust paradigms available today. Similarly, reproducibility has been recognized as a general problem in the neuroimaging field and is likely to become more important in the near future (e.g., the Stanford Center for Reproducible Neuroscience). Another question concerns comparability. Preprocessing and statistical analyses of imaging data have been rapidly evolving into a large number of highly sophisticated procedures that are used as decided on by the scientist conducting research. This situation has an impact on results and undermines the overall comparability of imaging genetics studies, of which 5-HTTLPR meta-analyses are one example. Given the abundance of practices in reporting and performing preprocessing as well as statistics, standardization and consensus are urgently needed. Consequently, the Organization of Human Brain Mapping has installed the Committee on Best Practice in Data Analysis to address these issues in the future. A third question concerns genetics. Work in animals strongly supports the importance of serotonin signaling and gene function of SERT for behavioral pathogenesis. Likely the most convincing evidence in humans stems primarily from rare variants occurring in patients with autism that produce SERT proteins that are no longer sensitive to intracellular signaling pathways controlling SERT trafficking and activity (9). There are several largely unexplored possibilities of how genetic variants affecting SERT expression could affect neural function, such as whether genetic variation within the serotoninergic transcription regulatory network alters neural function (9).
The use of gene expression data in imaging studies is currently in development (10) and will definitely promote a better understanding of the effects of genetic variants that impact serotonin signaling. Apart from genes affecting SERT expression, it might also be worthwhile to investigate the neural consequences of genes that regulate intracellular packing and trafficking of SERT. Finally, the use of genetic risk scores in imaging studies investigating the complex genetic interplay between 5-HTTLPR and other variants provides the opportunity to account for additive genetic effects on neural function (7). In conclusion, Klumpers et al. present an important study that expands the current understanding of the specific role of 5-HTTLPR in fear conditioning and anticipation of threat by relating the salience network and specifically dorsomedial prefrontal cortex function to these behavioral outcomes. Despite the large body of convincing imaging literature on 5HTTLPR, there is ample room for future progress, which will likely come from methodologic improvements, such as increased reproducibility and comparability of imaging studies, as well as imaging genetics studies addressing the full complexity of SERT expression and protein-level transport to its final functional presynaptic transmembrane location.
Acknowledgments and Disclosures The author reports no biomedical financial interests or potential conflicts of interest.
Article Information From the Department of Psychiatry and Psychotherapy, Medical University of Vienna, Vienna, Austria. Address correspondence to Lukas Pezawas, M.D., Department of Psychiatry and Psychotherapy, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria; E-mail: lukas.pezawas@meduniwien. ac.at. Received Aug 11, 2015; accepted Aug 12, 2015.
References 1.
2.
3.
4.
5.
6.
7.
8.
Lesch KP, Bengel D, Heils A, Sabol SZ, Greenberg BD, Petri S, et al. (1996): Association of anxiety-related traits with a polymorphism in the serotonin transporter gene regulatory region. Science 274:1527–1531. Caspi A, Sugden K, Moffitt TE, Taylor A, Craig IW, Harrington H, et al. (2003): Influence of life stress on depression: moderation by a polymorphism in the 5-HTT gene. Science 301:386–389. Meyer-Lindenberg A, Weinberger DR (2006): Intermediate phenotypes and genetic mechanisms of psychiatric disorders. Nat Rev Neurosci 7: 818–827. Hariri AR, Mattay VS, Tessitore A, Kolachana B, Fera F, Goldman D, et al. (2002): Serotonin transporter genetic variation and the response of the human amygdala. Science 297:400–403. Pezawas L, Meyer-Lindenberg A, Drabant EM, Verchinski BA, Munoz KE, Kolachana BS, et al. (2005): 5-HTTLPR polymorphism impacts human cingulate-amygdala interactions: A genetic susceptibility mechanism for depression. Nat Neurosci 8:828–834. Nikolova YS, Koenen KC, Galea S, Wang CM, Seney ML, Sibille E, et al. (2014): Beyond genotype: Serotonin transporter epigenetic modification predicts human brain function. Nat Neurosci 17: 1153–1155. Rabl U, Meyer BM, Diers K, Bartova L, Berger A, Mandorfer D, et al. (2014): Additive gene-environment effects on hippocampal structure in healthy humans. J Neurosci 34:9917–9926. Klumpers F, Kroes MC, Heitland I, Everaerd D, Akkermans SE, Oosting RS, et al. (2015): Dorsomedial prefrontal cortex mediates
Biological Psychiatry October 15, 2015; 78:522–524 www.sobp.org/journal
523
Biological Psychiatry
Commentary
9.
524
the impact of serotonin transporter linked polymorphic region genotype on anticipatory threat reactions. Biol Psychiatry 78:582–589. Deneris ES, Wyler SC (2012): Serotonergic transcriptional networks and potential importance to mental health. Nat Neurosci 15:519–527.
10.
Richiardi J, Altmann A, Milazzo AC, Chang C, Chakravarty MM, Banaschewski T, et al. (2015): Brain Networks. Correlated gene expression supports synchronous activity in brain networks. Science 348:1241–1244.
Biological Psychiatry October 15, 2015; 78:522–524 www.sobp.org/journal
Biological Psychiatry
Serotonin Transporter Linked Polymorphic Region: From Behavior to Neural Mechanisms Lukas Pezawas The clinical efficacy of selective serotonin reuptake inhibitors that exert their therapeutic action via the serotonin transporter protein (SERT), which is coded by a single gene (SLC6A4), has fueled research over the past 3 decades. The first report on a functional degenerate repeat polymorphic region within SLC6A4 (serotonin-transporter-linked polymorphic region [5HTTLPR]), associated with anxiety-related personality traits (1), made this genetic variant subject to numerous studies in animals, healthy humans, and psychiatric patients. A mechanistic account of how 5-HTTLPR might moderate psychiatric disease was first demonstrated by a study (2) that showed that the less transcriptionally efficient short (S) allele of 5-HTTLPR amplifies depressive symptoms in the presence of environmental adversity. However, despite the overwhelming number of positive reports, subsequent meta-analytical studies have battled over the reality of such a gene-environment interaction in patients. Thus, a collaborative meta-analysis to draw final conclusions has been launched. Similarly, the originally reported association between 5HTTLPR and anxiety-related personality traits has been challenged several times and in its current state is inconclusive at best. This lack of robust reproducibility of the original findings has been frustrating to many researchers in the field because it questioned the idea of a simple relationship between behavior and a promising candidate gene for psychiatric illness. However, one might also find it intriguing that an association between a molecular biological feature, such as 5-HTTLPR, and a phenomenologically operationalized psychiatric nosology or a psychological construct based on subjective ratings has been found in the first place, given the absence of any biological criterion within their definitions. Genes do not code for psychopathology or personality, but for proteins that may have a direct impact on cell function. Therefore, an intermediate phenotype approach (3) was developed using genetic data as independent variables and imaging measures as dependent variables. This research strategy aimed to establish a more biologically meaningful relationship between a psychiatric risk gene and psychopathology by measuring neural circuit function instead of psychopathology itself. An important reason for this procedure was that behavioral genetics has taught us that changes in brain function related to genetic variation may or may not result in behavioral readouts. Initial studies were conducted primarily in healthy control subjects because of the suspected biological heterogeneity of patient samples and disease-related epiphenomena that might affect neural circuit function (e.g., medication, highly variable symptoms, social deprivation), among other reasons. This research strategy was intended to enable the development of novel and more complex disease models
that consider the individual variation of neural wiring and function as opposed to oversimplified biological traitbehavior models. The aforementioned approach has been tremendously successful and highlighted several effects of 5-HTTLPR on a brain systems level, such as frequently replicated increased amygdala reactivity in S-allele carriers using fearful faces as stimuli (4), which is in line with work in animals demonstrating the importance of the amygdala in fear conditioning. Given the developmental role of serotonin in shaping mood circuitry, it is not surprising that the S-allele of 5-HTTLPR has also been shown to decrease subgenual anterior cingulate cortex and amygdala volume, possibly leading to increased amygdala reactivity via a corticolimbic circuitry encompassing both structures (5). The importance of this mood circuitry has been highlighted in numerous human and animal studies investigating emotion processing and treatment over the years. In addition, 5-HTTLPR effects have been shown in various brain regions or systems that were previously related to fear or emotion processing using different functional paradigms and resting state in healthy subjects and patients. However, the increasing number of 5-HTTLPR imaging studies has demonstrated that the overall reported effect size of 5-HTTLPR across diverse samples and study protocols has been small. Although meta-analytical evidence was able to verify initial reports in healthy subjects, results derived from patient studies are still inconclusive. Consequently, many groups have launched studies with far more complicated genetic models to address the apparent complexity of genetic effects on neural systems, including epistasis, epigenetics (6), and geneenvironment interactions (7), which revealed stronger effect sizes than initial studies associating a single genetic variant to neural networks. In this issue of Biological Psychiatry, Klumpers et al. (8) report novel data on the effects of 5-HTTLPR on brain function using two functional magnetic resonance imaging paradigms, classic fear conditioning and instructed fear, in two independent samples of healthy subjects. They report strongly overlapping activation increases in both paradigms encompassing the dorsal anterior cingulate cortex and adjacent dorsomedial prefrontal cortex, anterior insula, ventral striatum, and thalamus among other areas, but not the amygdala. The latter finding is supported by meta-analytical evidence of human imaging studies, the results of which contradict animal work. An investigation of 5-HTTLPR-moderated changes of corticolimbic circuitries has not been pursued by Klumpers et al. The authors argue that 5-HTTLPR moderates activation of the salience network, which is anchored in the dorsal anterior cingulate cortex and anterior insula and further encompasses
SEE CORRESPONDING ARTICLE ON PAGE 582
522 & 2015 Society of Biological Psychiatry Biological Psychiatry October 15, 2015; 78:522–524 www.sobp.org/journal
http://dx.doi.org/10.1016/j.biopsych.2015.08.010 ISSN: 0006-3223
Biological Psychiatry
Commentary
subcortical structures such as the amygdala, ventral striatum, and substantia nigra/ventral tegmental area. This neural system has shown to be robustly engaged in most imaging studies in healthy subjects and psychiatric patients using cognitive or emotion-related paradigms or resting state. Apart from its function in salience detection, it is thought to facilitate switching between large-scale brain networks such as the central executive and default mode network. The authors further demonstrate the convergence of their findings on a psychophysiologic level. They conclude that 5-HTTLPR affects the interindividual variability in anticipatory responses to threat via dorsomedial prefrontal cortex and speculate that this mechanism might be relevant for stress-related psychopathology. Although Klumpers et al. put forth a compelling argument, their argument raises several important questions for the field of fear and threat research, which have broad implications for imaging genetics studies investigating 5-HTTLPR. The first question concerns reproducibility. Using their paradigms, the authors did not detect any 5-HTTLPR-moderated amygdala activation, which is surprising given the large body of available evidence of amygdala hyperreactivity and corticolimbic dysfunction in animals and human imaging genetics studies. Engaging robust amygdala activation, specifically in the context of genetic studies, is still a challenging task for many research groups. The reasons for this may simply be related to magnetic resonance imaging hardware and software issues (e.g., resolution of image acquisition, choice of imaging sequence), physiologic artifacts (e.g., vessels), or the design of the applied paradigm itself. At the present time, no data repository is available where researchers can easily retrieve information on reliability of imaging measures regarding a specific paradigm, which would facilitate the use of the most robust paradigms available today. Similarly, reproducibility has been recognized as a general problem in the neuroimaging field and is likely to become more important in the near future (e.g., the Stanford Center for Reproducible Neuroscience). Another question concerns comparability. Preprocessing and statistical analyses of imaging data have been rapidly evolving into a large number of highly sophisticated procedures that are used as decided on by the scientist conducting research. This situation has an impact on results and undermines the overall comparability of imaging genetics studies, of which 5-HTTLPR meta-analyses are one example. Given the abundance of practices in reporting and performing preprocessing as well as statistics, standardization and consensus are urgently needed. Consequently, the Organization of Human Brain Mapping has installed the Committee on Best Practice in Data Analysis to address these issues in the future. A third question concerns genetics. Work in animals strongly supports the importance of serotonin signaling and gene function of SERT for behavioral pathogenesis. Likely the most convincing evidence in humans stems primarily from rare variants occurring in patients with autism that produce SERT proteins that are no longer sensitive to intracellular signaling pathways controlling SERT trafficking and activity (9). There are several largely unexplored possibilities of how genetic variants affecting SERT expression could affect neural function, such as whether genetic variation within the serotoninergic transcription regulatory network alters neural function (9).
The use of gene expression data in imaging studies is currently in development (10) and will definitely promote a better understanding of the effects of genetic variants that impact serotonin signaling. Apart from genes affecting SERT expression, it might also be worthwhile to investigate the neural consequences of genes that regulate intracellular packing and trafficking of SERT. Finally, the use of genetic risk scores in imaging studies investigating the complex genetic interplay between 5-HTTLPR and other variants provides the opportunity to account for additive genetic effects on neural function (7). In conclusion, Klumpers et al. present an important study that expands the current understanding of the specific role of 5-HTTLPR in fear conditioning and anticipation of threat by relating the salience network and specifically dorsomedial prefrontal cortex function to these behavioral outcomes. Despite the large body of convincing imaging literature on 5HTTLPR, there is ample room for future progress, which will likely come from methodologic improvements, such as increased reproducibility and comparability of imaging studies, as well as imaging genetics studies addressing the full complexity of SERT expression and protein-level transport to its final functional presynaptic transmembrane location.
Acknowledgments and Disclosures The author reports no biomedical financial interests or potential conflicts of interest.
Article Information From the Department of Psychiatry and Psychotherapy, Medical University of Vienna, Vienna, Austria. Address correspondence to Lukas Pezawas, M.D., Department of Psychiatry and Psychotherapy, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria; E-mail: lukas.pezawas@meduniwien. ac.at. Received Aug 11, 2015; accepted Aug 12, 2015.
References 1.
2.
3.
4.
5.
6.
7.
8.
Lesch KP, Bengel D, Heils A, Sabol SZ, Greenberg BD, Petri S, et al. (1996): Association of anxiety-related traits with a polymorphism in the serotonin transporter gene regulatory region. Science 274:1527–1531. Caspi A, Sugden K, Moffitt TE, Taylor A, Craig IW, Harrington H, et al. (2003): Influence of life stress on depression: moderation by a polymorphism in the 5-HTT gene. Science 301:386–389. Meyer-Lindenberg A, Weinberger DR (2006): Intermediate phenotypes and genetic mechanisms of psychiatric disorders. Nat Rev Neurosci 7: 818–827. Hariri AR, Mattay VS, Tessitore A, Kolachana B, Fera F, Goldman D, et al. (2002): Serotonin transporter genetic variation and the response of the human amygdala. Science 297:400–403. Pezawas L, Meyer-Lindenberg A, Drabant EM, Verchinski BA, Munoz KE, Kolachana BS, et al. (2005): 5-HTTLPR polymorphism impacts human cingulate-amygdala interactions: A genetic susceptibility mechanism for depression. Nat Neurosci 8:828–834. Nikolova YS, Koenen KC, Galea S, Wang CM, Seney ML, Sibille E, et al. (2014): Beyond genotype: Serotonin transporter epigenetic modification predicts human brain function. Nat Neurosci 17: 1153–1155. Rabl U, Meyer BM, Diers K, Bartova L, Berger A, Mandorfer D, et al. (2014): Additive gene-environment effects on hippocampal structure in healthy humans. J Neurosci 34:9917–9926. Klumpers F, Kroes MC, Heitland I, Everaerd D, Akkermans SE, Oosting RS, et al. (2015): Dorsomedial prefrontal cortex mediates
Biological Psychiatry October 15, 2015; 78:522–524 www.sobp.org/journal
523
Biological Psychiatry
Commentary
9.
524
the impact of serotonin transporter linked polymorphic region genotype on anticipatory threat reactions. Biol Psychiatry 78:582–589. Deneris ES, Wyler SC (2012): Serotonergic transcriptional networks and potential importance to mental health. Nat Neurosci 15:519–527.
10.
Richiardi J, Altmann A, Milazzo AC, Chang C, Chakravarty MM, Banaschewski T, et al. (2015): Brain Networks. Correlated gene expression supports synchronous activity in brain networks. Science 348:1241–1244.
Biological Psychiatry October 15, 2015; 78:522–524 www.sobp.org/journal