Glutamate transporters: A key piece in the glutamate puzzle of major depressive disorder
Introduction
More than 60 years after the introduction of pharmacological therapies, Major Depression Disorder (MDD) still has a significant impact as a major public health issue. The development of antidepressant medications has traditionally focused on monoamine related compounds, but still a large group of patients are considered non-responsive (Connolly and Thase, 2012). Therefore, a considerable interest in non-monoaminergic approaches to the treatment of MDD has gained momentum, and glutamate-modifying therapies are emerging as a valuable new path that could quicken the recovery of patients through fast acting compounds (Bunney and Bunney, 2012). Since glutamatergic neurotransmission is a very complex system, exploring the role of its different components and how they contribute to the pathophysiology of mood disorders is of great importance to understand the effects of novel therapeutic compounds.
The amino acid glutamate is a ubiquitous molecule that acts as the major excitatory neurotransmitter in the nervous system (Dingledine and McBain, 1999). Numerous projection fibers from cortical and subcortical structures utilize glutamate as their main neurotransmitter (Cotman and Monaghan, 1986), and its stimulatory effect is essential to maintain the activity of other neurotransmitter systems (Michaelis, 1998). However, it is also recognized that the excitatory properties of glutamate should be kept under a delicate balance or it would otherwise exert damage that can range from cellular malfunction to extensive tissue injury, a condition known as excitotoxicity (Thomas, 1995; Dong et al., 2009).
In order to act as a neurotransmitter, presynaptic glutamate is stored in vesicles to be released after calcium influx triggered by action potentials. The process of storing glutamate is performed by the vesicular glutamate transporters (VGLUTs). The three vesicular transporters, VGLUT1, 2 and 3 are differentially distributed in the brain; in the hippocampus VGLUT1 is the most abundantly expressed (Fremeau et al., 2004; van der Hel et al., 2009). Following synaptic release glutamate interacts with both ionotropic (NMDA, AMPA, kainate) and metabotropic (mGlu) receptors (Greenamyre and Porter, 1994). Neurotransmitter activity ends when the amino acid is removed from the synaptic cleft by specific high affinity glutamate transporters (Danbolt et al., 1994; Amara and Fontana, 2002). In humans the excitatory amino acid transporters (EAAT) family of proteins is composed of five members, namely EAAT1 through 5, encoded by the gene family of solute carriers SLC1A. Table 1 summarizes the names and aliases of the genes and proteins that constitute this group (Torp et al., 1994; Danbolt et al., 1998; Dehnes et al., 1998; Danbolt, 2001).
Acute or chronic excitotoxicity caused by excessive glutamate release has been extensively reviewed and is known to be the main cause of tissue damage in acute events such as ischemia, epilepsy, traumatic brain injury, and is also considered an essential cause of damage in neurodegenerative diseases such as Alzheimer's disease and amyotrophic lateral sclerosis (ALS) (Campiani et al., 2003; Dong et al., 2009). The possibility of impaired glutamate uptake in acute excitotoxic conditions has been repeatedly postulated (Mathern et al., 1999; Demarque et al., 2004; Camacho and Massieu, 2006; Bjornsen et al., 2007), and the likelihood of altered transport in chronic degenerative diseases has also been explored. When a specific family of high affinity carriers was characterized a specific role for the glutamate transporters as the source of glutamatergic dysfunction was first considered (Danbolt, 2001; Gegelashvili et al., 2001; Maragakis and Rothstein, 2001; Maragakis and Rothstein, 2004).
The involvement of glutamate in the physiopathology of mental disease was first suggested by considering glutamatergic hypofunction as a possible mechanism in schizophrenia (Carlsson and Carlsson, 1990; Bunney, 1995; Carlsson et al., 1999) as the use of NMDA blockers such as ketamine induce schizophrenia-like symptoms (Kegeles et al., 2000; Sinner and Graf, 2008). Following this premise, the possibility of alterations of the diverse components of the glutamatergic system in psychiatric disorders has been explored (Harrison et al., 2003). These studies initially focused on the expression and function of glutamate receptors, with special interest in the NMDA receptor. Nowak et al. (1995) showed decrease of high affinity binding of NMDA receptors in the frontal cortex of suicide victims. Feyissa et al. (2009) described a reduction of NMDA receptor subunits NR2A and NR2B in the prefrontal cortex of depressed patients. An in situ hybridization study demonstrated that in the perirhinal cortex (but not hippocampus) several glutamate receptor subunits were significantly lower in MDD patients compared to controls, including the NR2A and NR2B subunits of NMDA receptor as well as the GluR1, GluR3, and GluR5 subunits of AMPA receptors (Beneyto et al., 2007). Noga et al. (1997) reported increased AMPA receptor binding in the caudate nucleus of suicide subjects. Specific changes in the hippocampus were observed by Law and Deakin (2001) who reported reduced NR1 mRNA levels in MDD group as compared to controls.
The results of these studies point to region-dependent alterations of glutamate related molecules in different brain areas in mood disorders. Also a large body of evidence emphasizes the role of the hippocampus in emotion and cognition and its contribution to psychiatric symptoms (for a review see Drevets et al., 2008), and this being a predominantly glutamatergic circuit (Storm-Mathisen, 1981) calls for a thorough assessment of the molecules related to that system. Still, the available number of human postmortem publications in this topic is limited and further studies involving the different components of the glutamatergic system in the hippocampus are needed.
Although the possibility of altered glutamate uptake as part of the pathophysiology of mental illness has been considered for over a decade very little progress has been made in the field. Only in recent years was microarray technology used to estimate glutamate transporters mRNA expression. Using these methods Choudary et al. (2005) described a decrease of SLC1A2 and SLC1A3 (the genes for EAAT2 and EAAT1) in the anterior cingulate and dorsolateral prefrontal cortex of MDD patients; a similar result was reported by Bernard et al. (2011) in the locus coeruleus in MDD but not BPD subjects.
In this study we used microarray technology and in situ hybridization to explore the possibility of alterations in the expression of the glial glutamate transporters SLC1A2 (EAAT2) and SLC1A3 (EAAT1) and the vesicular transporter SLC17A7 (VGLUT) in the hippocampus of depressed patients.
Section snippets
Brain samples
Human brain samples from major depression disorder (MDD), bipolar disorder (BPD) and control subjects were obtained through the Pritzker Neuropsychiatric Consortium from the Brain Donor Program at the University of California, Irvine with consent of the next of kin, whom were also interviewed to obtain information on the subjects' physical health, medication and recreational substance use, to be added to the medical records and coroner's investigation information. MDD and BPD diagnosis were
LCM and microarray assay results
We carried out gene expression profiling studies using laser capture microdissection combined with microarray to evaluate global changes in basal gene expression in samples of whole hippocampus.
Disease-specific gene expression differences from controls were evaluated using Student's t-tests. Genes were considered to be significantly altered if the p-values were p = 0.05 or lower. Fold change (FC) values were also obtained to assess the magnitude of the change in the disease subjects (Table 3).
Discussion
Currently the monoamine hypothesis is the most commonly accepted insight in to the pathophysiology of mood disorders. This premise was “reverse engineered” from the investigation of the mechanisms of action of early therapeutic agents such as monoamine oxidase inhibitors and tricyclic antidepressants (Mathews et al., 2012). However, to this day MDD is still a leading cause of disability (Murray and Lopez, 1996) and the rates of treatment success are less than satisfactory. Therefore it is
Contributors
Adriana Medina: study design, data analysis, writing of the first draft of the manuscript and final revision of manuscript.
Sharon Burke: laboratory procedures and final revision of the manuscript.
Robert Thompson: laboratory procedures and final revision of the manuscript.
William Bunney Jr.: supervised sample collection and final revision of the manuscript.
Richard Myers: final revision of the manuscript.
Alan Shatzberg: final revision of the manuscript.
Huda Akil and Stanley J Watson: supervised
Role of the funding source
None.
Conflict of interest
The authors do not have any conflict of interest.
Acknowledgments
The authors are members of the Pritzker Neuropsychiatric Disorders Research Consortium, which is supported by the Pritzker Neuropsychiatric Disorders Research Fund L.L.C. A shared intellectual property agreement exists between this philanthropic fund and the University of Michigan, Stanford University, the Weill Medical College of Cornell University, HudsonAlpha Institute of Biotechnology and the University of California at Irvine, to encourage the development of appropriate findings for
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