Assignment #4 Journal Article Review

Frodl, T., Koutsouleris, N., Bottlender, R., Born, C., Jager, M, Morgenthaler, M., et al. (2008). "Reduced gray matter brain volumes are associated wtih variants of the serotonin transporter gene in major depression." Molecular Psychiatry, 1, 1093-1101.

Introduction

Major depression is suspected to be caused by an imbalance in neuronal serotonin level, and many studies have demonstrated this relationship. The serotonergic system and brian derived neurotrophic factor (BDNF) have been seen to reciprocally regulate one another, and together are responsible for early CNS development, as well as hippocampal neurogenesis in adults. Evidence of low serotonin levels in patients suffering from major depression have led Frodl et al. to suspect that dysfunctional neuronal plasticity is a major contributor to the psychopathology of mood disorders like depression. Recent studies have shown patients suffering from major depression to have generally reduced hypocampus volumes, but the manual measuring techniques used in those studies give inconsistent results for brain regions without an obvious morphologically defined border.

In addition, there have been three allelic polymorphisms (triallelic polymorphism) of the 5-HTTLPR gene responsible for encoding components of the serotonergic system; S-allele (normal), and L-allele (subtypes La and Lg) showing increased 5-HHT expression and increased 5-HT reuptake. Lg subtypes have been seen to have the same 5-HT reuptake efficiency as S-allelic individuals, therefore they have been grouped in this experiment as ‘equal’ to the S-allele. Therefore, this triallelic polymorphism is expressed as homozygous = SS/LgLg/LgS and heterozygous = LaS or LaLg.
In this experiment, Frodl et al. use Voxel-based morphology (VBM) to clearly measure the gray matter volumes from magnetic resonance images in mulitple brain regions of patients with different genotypes in the 5-HTTLPR gene, to see if patients suffering from major depression and who are homozygous for the L-allele have reduced brain gray matter in the regions explored.

Methods

Population:

N=77

middle aged inpatients suffering from major depression, diagnosed using criteria outlined in the DSM-IV. During this experiment, patients taking antidepressants (Selective serotonin reuptake inhibitors [SSRIs], tricyclic antidepressants, and others) continued taking their antidepressants as suggested by their physician.

77 healthy individuals were also analyzed, and served as a control comparison group.

Gray matter volume data was obtained using magnetic resonance imaging.

Brain regions analyzed:

1. Hippocampus
2. Amygdale
3. Anterior gyrus cinguli (ACC)
4. Dorsolateral prefrontal cortex (DLPFC)
5. Dorsomedial prefrontal cortex (DMPFC)

Patient genotypes were determined by executing DNA sequencing on patient blood samples, using the ‘QIAamp Blood Isolation Kit’ (QIAGEN) to isolate DNA, followed by PCR and gel electrophoresis of PCR products.

Comparison of whole-brain gray matter volumes between control and experimental groups were tested for significant difference at a tolerance for Type 1 error of 5%. Voxel-based morphology data comparisons between the control and experimental group for each brain region of interest was tested for significant difference, at tolerance for Type 1 error of 10%.

Results & Conclusions:

In the healthy control group, there was a strong association between 5-HTTLPR polymorphism and GMVs in all investigated brain regions
5-HTTLPR contributes to brain structure development

Smallest gray matter volumes found in healthy group were of those individuals homozygous for S allele. This difference in gray matter volume was observed in all brain regions analyzed.

Reduction in gray matter volumes of S homozygous individuals of the patient group were only seen in the subregions of the hippocampus.
La homozygous patients, relative to La homozygous healthy controls show the largest volume reduction in cortical brain regions, as well as the amygdale and interior hippocampus.

Figure 1: Magnetic Resonance Images of gray matter in patients in relation to control groups, measured on a scale of density. See colour legend at bottom left of figure for density reduction scores.

Overall, the greatest gray matter volume alteration (difference between control and depressed patient) was seen in individuals homozygous for La.
Homozygous La individuals have higher expression of serotonin transporter (5-HTT) and therefore lower synapse serotonin levels. Since the serotonergic system upregulates the expression of BDNF (through upregulation of cAMP), patients with lower serotonin levels may be more vulnerable to abnormal hippocampal neurogenesis.
It is therefore likely that decrease in serotonin levels, induced by those factors contributing to depression, may cause serotonin levels to drop so severely that structural changes result in the brain. These individuals could therefore be more at risk for development of major depression.

Problems with this experiment:

The sample sizes were relatively small; larger sample sizes would have allowed Frodl et al to draw more specific conclusions.

There is too much variation with the experimental group (patients suffering from depression). Patients are on different medications, and have grown up in different environments.

Antidepressants inhibit serotonin reuptake, therefore increasing overall serotonin in the neuronal synapses. Since increasing serotonin levels in the brain helps stimulate cell growth and propogation in brain regions like the hypothalamus, patients taking antidepressants may not represent the morphological phenotype (ie: gray matter volume) that is true to the genotype. . Frodl et al, however do recognize these additional variables and suggest that further studies should be designed to explore the interactions of these variables.

Comments/Critique:

This paper was commendable because it focused on the interaction between biological factors and psychological factors in order to explain their results. Frodl’s hypothesis was of interest, because if his hypothesis holds true, it could help define a biological marker for the diagnosis of depression, or the risk in an individual to develop depression. Many researchers have attempted to explain depression through biological causal agents and therefore have designed their statistical models to include only biologically based explanatory variables. Since depression is an illness affecting the human mind, designing experiments while ignoring the complex nature of depression due to the interaction of biological agents with psychological factors renders these experiments weak.

In terms of the paper’s presentation of the results, the figures gave a good, comprehensive presentation of the MRI images taken. They underwent very thorough analysis of the data, and presented the key aspects of their statistical analysis in ANOVA tables, which allows for the public to confirm the validity of their conclusions. The positioning of the tables in the article is slightly awkward, because a few of them are located in the discussion section.

Hopefully Frodl et al. will conduct further experiments of similar nature with the addition that patients be selected to have similar life histories (if possible) or at least be taking the same type of antidepressant (or none at all, if possible).

To access this journal article through NCBI (PubMed), please visit the following link:

http://www.ncbi.nlm.nih.gov/pubmed/19008895?ordinalpos=1&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum

Full text pdf or HTML files are also available through the Journal of Molecular Psychiatry:

http://www.nature.com/mp/journal/v13/n12/abs/mp200862a.html

Thanks for visiting :)

Assignment #3 - Serotonin Function and Pathology

Serotonin is a neurotransmitter that has been described as being the molecule with the most diverse and numerous functions in the human body. Since serotonin helps control appetite, sleep, memory/learning, temperature regulation, mood, behaviour, cardiovascular function, muscle contraction, endocrine regulation and depression, any type of pathology related to serotonin will emerge in illnesses affecting any combination of these functions.

Serotonin is produced in the Pineal gland, located in the centre of the human brain. Serotonin produced in the Pineal gland is stored and released by dense bodies of neurons, called Raphe Nuclei within the medulla, pons and midbrain. These serotonergic neurons have axons that project to many parts of the brain, and it is believed serotonin’s diverse behavioural effects are a consequence of this physical feature.

Figure 1: Broad dispersion of Serotonin from raphe nuclei in the brain
during a panic attack.

Depression

DSM-IV-TR definition!

There are many illnesses that are believed to be related to low levels of serotonin. Amongst serotonin related illnesses, depression is by far the most prevalent of serotonin related illness in North America. Depression is characterized by a mixture of symptoms including of sadness, worthlessness, loss of interest or motivation, insomnia that occurs chronically within an individual. It is estimated to affect 12-17% of the population at some point during the lifetime of an individual. (Malberg, 2000)

Depression is believed to be caused by serotonin deficiency in the body. Though this pathology has been studied extensively, there has yet to be a biological mechanism described for depression and the complexity of the human mind may not be a function of biological factors alone. Researchers have measured decreased concentrations of serotonin metabolites in the cerebrospinal fluid and brain tissues of depressed people, and this serves as the most concrete evidence relating depression to serotonin.

To combat depression, antidepressants function to raise the level of serotonin within in synapse.
There are many types of antidepressants available on the market, and they can be grouped based on their mode of action. (Healy, 1997)

The three most common classes of antidepressants are

• Tricyclic Antidepressants


• Monoamine Oxidase Inhibitors (MAOIs), and


• 5-HT Reuptake Inhibitors (Also known as Selective Serotonin Reuptake Inhibitors, SSRIs) {Most commonly prescribed (eg: Prozac)}. (Healy, 1997)
  
  

1 Tricyclic Antidepressants


These compounds have a three-ring (tricyclic) central structure, and serve more so as a sedative rather than a stimulant. First type of antidepressants “discovered” (imipramine, Tofranil), interest in the compound as an antihistamine. (Healy, 1997) This class of compounds are usually secondarily prescribed in cases where SSRI have not served as a successful treatment of depression, because motality due to tricyclic antidepression overdose is a well recognized problem (Barbey 1998).

Figure 2: Chemical structure of the Tricyclic Antidepressant, Imipramine

2 Monoamine Oxidase Inhibitors (MAOIs):


This class of Antidepressants work to increase the levels of serotonin in the synaptic clefts by inhibiting the enzyme monoamine oxidase. Monoamine oxidase, along with the enzyme catchecol-O-methyl transferase are located in the synaptic cleft, and function to degrade serotonin after its release from the presynaptic cell.




Figure 3: Serotonin is released from presynaptic cell vessicles into the synaptic cleft, where they are eventually removed by reuptake and degredation by enzymes such as monoamine oxidase.


The discovery of monoamine oxidase inhibitors is a complex and important story. The first reported MOAI was isoniazid and iproniazid synthesized in 1912, but isoniazids resynthesis in 1951 is of much interest. During WWII, V2 rockets were initially propelled using liquid oxygen and ethanol. Nearing the end of the war, as stocks of these compounds decreased, the rockets were propelled by a compound called hydrazine. When the war ended, production companies found themselves holding a surplus of hydrazine, and these were taken over by chemical companies, many of which who were in the process of developing new pharmaceutical compounds. A series of hydrazine derivatives were formed, one including iproniazid in 1951, and it was found to be an effective anti-tuberculous drug. Physicians, however noticed mental side effect in their tuberculosis patients, including a generally favourable affect on patient demeanor. Newspapers reported patients to be dancing in the wards, regardless of the holes (lesions) in their lungs, and from then on, interest in MOAIs skyrocketed. (Healy,1997)





3 Selective Serotonin Reuptake Inhibitors (SSRIs)


Of all the antidepressants described here, SSRIs are by far the most prevalently prescribed to individuals with depression. Not only do they appear to be effective in broad range of individuals, but this class of antidepressants are thought to be safer in overdose (Barbey, 1998). SSRIs have a mode of action of inhibiting the reuptake of serotonin from the synaptic cleft into the presynaptic cell.

Though antidepressants are commonly prescribed for individuals suffering from depression, anxiety and other affective disorders, the cellular and molecular mechanism by which antidepressants give their therapeutic effect is not well understood.




Serotonin and cell differentiation in the hippocampus

In addition to direct inhibition of serotonin reuptake, chronic administration of antidepressants are suspected to have a mode of action affecting cell differentiation within the hippocampus (Malberg, 2000)

The hippocampus is one of the only brain regions to continue cellular reproduction during the lifetime. Clinical research prior to this study showed that stress and depression had effects on the hippocampus, seen by morphological changes such as atrophy and loss of pyramidal neurons (Malberg, 2000). Antidepressants have been shown to ‘undo’ the morphological changes seen in the hippocampus by upregulating the expression of brain-derived neurotrophic factor (BDNF) in the hippocampus. BDNF has been shown to promote cellular differentiation as well as cell survival during development in the adult brain (also shown in vitro). Chronic administration of antidepressants has also been shown to block the stress-induced downregulation of BNDF gene expression (Malberg, 2000).

Seasonal Affective Disorder

A mood disorder related to reduced sunlight in the winter months.

DSM-IV-TR definition

In fruitflies, it has been demonstrated that serotonin decreases the sensitivity of the body to light, and chronic exposure to darkness results in decreased serotonin levels in fruit fly brains (Sehgal et al, 2005). Normally, a protein in flies called cryptochrome controls the response of the body’s circadian rhythm to light. Another protein called GSK3bβ has the ability to affect the sequential events leading to the ‘resetting’ of the circadian clock. Serotonin receptor (5-HT1B) binding leads to activation of GSK3bβ, allowing the body circadian rhythm to be reset (Sehgal et al, 2005).

Figure 5: Binding of serotonin to its receptor affects the cascade involved with expression of compounds resetting the body's circadian rhythm.

Further study of serotonin’s relationship with Seasonal Affective Disorder will help increase the etiology and potential treatments for SAD. Currently, those experiencing SAD may choose to take antidepressants, in attempt to raise their serotonin brain levels and alleviate symptoms (Sehgal et al, 2005). Individuals suffering from depression often exhibit symptoms of insomnia and irregular sleep patterns, which are most likely related to serotonin’s contribution in the regulation of circadian rhythm.

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References:


[1] Barbey, J. T., Roose, S. P. (1998). SSRI Safety in Overdose. Journal of Clinical Psychiatry. (59)15:42-28

[2] DSM-IV-TR

[3] Healy, David. (1997). The Antidepressant Era. Cambridge Massachusettes, USA:Harvard University Press

[4] Malberg, J. E et al. (2000). Chronic Antidepressant Treatment Increases Neurogenesis in Adult Rat Hippocampus. Journal of Neuroscience, 20(24):9104-9110

[5] Rosling, Claire. Serotonin: A Molecule of Happiness.
Retrieved November 3, 2008, from
http://www.chm.bris.ac.uk/motm/serotonin/home1.htm

[6] University Of Pennsylvania School Of Medicine. “Penn Researchers Determine Role
Of Serotonin In Modulating Circadian Rhythm." ScienceDaily 11 July
2005. 5 November 2008