The mTOR signaling pathway in the prefrontal cortex is compromised in major depressive disorder

Recent studies demonstrate that rapid antidepressant response to ketamine is mediated by activation of the mammalian target of rapamycin (mTOR) signaling pathway, leading to increased synaptic proteins in the prefrontal cortex (PFC) of rats. Our postmortem studies indicate robust deficits in prominent postsynaptic proteins including N-methyl-d-aspartate (NMDA) receptor subunits (NR2A, NR2B), metabotropic glutamate receptor subtype 5 (mGluR5) and postsynaptic density protein 95 kDa (PSD-95) in the PFC in major depressive disorder (MDD). We hypothesize that deficits in the mTOR-dependent translation initiation pathway contribute to the molecular pathology seen in the PFC of MDD subjects, and that a rapid reversal of these abnormalities may underlie antidepressant activity. The majority of known translational regulation occurs at the level of initiation. mTOR regulates translation initiation via its downstream components: p70-kDa ribosomal protein S6 kinase (p70S6K), and eukaryotic initiation factors 4E and 4B (eIF4E and eIF4B). In this study, we examined the expression of mTOR and its core downstream signaling targets: p70S6K, eIF4E, and eIF4B in the PFC of 12 depressed subjects and 12 psychiatrically healthy controls using Western blot. Levels of eIF4E phosphorylated at serine 209 (p-eIF4E-Ser209) and eIF4B phosphorylated at serine 504 (p-eIF4B-Ser504) were also examined. Adjacent cortical tissue samples from both cohorts of subjects were used in our previous postmortem analyses. There was a significant reduction in mTOR, p70S6K, eIF4B and p-eIF4B protein expression in MDD subjects relative to controls. No group differences were observed in eIF4E, p-eIF4E or actin levels. Our findings show deficits in mTOR-dependent translation initiation in MDD particularly via the p70S6K/eIF4B pathway, and indicate a potential association between marked deficits in synaptic proteins and dysregulation of mTOR signaling in MDD.     

Effect of chronic fluoxetine treatment on male and female rat erythrocyte and prefrontal cortex fatty acid composition

Omega-3 (n-3) polyunsaturated fatty acids (PUFA) and fluoxetine (FLX) have additive effects in the treatment of major depressive disorder, and FLX up-regulates genes that regulate fatty acid biosynthesis in vitro. Although these data suggest that FLX may augment n-3 fatty acid biosynthesis, the in vivo effects of FLX treatment on PUFA biosynthesis and peripheral and central membrane compositions are not known. In the present study, male and female rats were treated with FLX (10 mg/kg/day) through their drinking water for 30 days (P60–P90). Plasma FLX and norfluoxetine (NFLX) concentrations were determined by liquid chromatography tandem mass spectrometry, and erythrocyte and prefrontal cortex (PFC) fatty acid composition determined by gas chromatography. To confirm central effects of FLX, serotonin turnover in the PFC was determined by high performance liquid chromatography. Chronic FLX treatment resulted in clinically-relevant plasma FLX concentrations in male and female rats, and significantly decreased serotonin turnover in the PFC. After correcting for multiple comparisons, chronic FLX treatment did not significantly alter erythrocyte fatty acid composition in male or female rats. Chronic FLX treatment significantly and selectively increased docosapentaenoic acid (22:5n-6) in the PFC of female rats, but not in male rats. These preclinical findings do not support the hypothesis that chronic FLX treatment increases n-3 fatty acid biosynthesis or membrane composition.     

High-frequency rTMS treatment increases left prefrontal myo-inositol in young patients with treatment-resistant depression

Background

Neuroimaging studies suggest that the prefrontal cortex (PFC) is involved in the pathophysiology of major depression. Repetitive transcranial magnetic stimulation (rTMS) as an antidepressant intervention has increasingly been investigated in the last two decades. In this study metabolic changes within PFC of severely depressed patients before and after rTMS were evaluated by proton magnetic resonance spectroscopy (1H-MRS).

Method

Thirty-four young depressed patients with treatment-resistant unipolar depression were enrolled in a double-blind, randomized study〔active ((n = 19) vs. sham(n = 15)), and the PFC was investigated before and after high-frequency (15 Hz) rTMS using 3-tesla proton magnetic resonance spectroscopy. Response was defined as a 50% reduction of the Hamilton depression rating scale. The results were compared with 28 age- and gender-matched healthy controls.

Results

In depressive patients a significant reduction in myo-inositol (m-Ino) was observed pre-rTMS (p < 0.001). After successful treatment, m-Ino increased significantly in left PFC and the levels no longer differed from those of age-matched controls. In addition to a positive correlation between clinical improvement and an increment in m-Ino ratio, a correlation between clinical improvement and early age onset was observed.

Conclusions

Our results support the notion that major depressive disorder is accompanied by state-dependent metabolic alterations, especially in myo-inositol metabolism, which can be partly reversed by successful rTMS.     

Omega-3 fatty acid deficiency during perinatal development increases serotonin turnover in the prefrontal cortex and decreases midbrain tryptophan hydroxylase-2 expression in adult female rats: dissociation from estrogenic effects

A dysregulation in central serotonin neurotransmission and omega-3 fatty acid deficiency have been implicated in the pathophysiology of major depression. To determine the effects of omega-3 fatty acid deficiency on indices of serotonin neurotransmission in the adult rat brain, female rats were fed diets with or without the omega-3 fatty acid precursor α-linolenic acid (ALA) during perinatal (E0-P90), post-weaning (P21-P90), and post-pubescent (P60-130) development. Ovariectomized (OVX) rats and OVX rats with cyclic estrogen treatment were also examined. Serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) content, and fatty acid composition were determined in the prefrontal cortex (PFC), and tryptophan hydroxylase-2 (TPH-2), serotonin transporter, and 5-HT_1A autoreceptor mRNA expression were determined in the midbrain. ALA deficiency during perinatal (−62%, p = 0.0001), post-weaning (−34%, p = 0.0001), and post-pubertal (−10%, p = 0.0001) development resulted in a graded reduction in adult PFC docosahexaenoic acid (DHA, 22:6n−3) composition. Relative to controls, perinatal DHA-deficient rats exhibited significantly lower PFC 5-HT content (−65%, p = 0.001), significant greater 5-HIAA content (+15%, p = 0.046), and a significant greater 5-HIAA/5-HT ratio (+73%, p = 0.001). Conversely, post-weaning DHA-deficient rats exhibited significantly greater PFC 5-HT content (+12%, p = 0.03), no change in 5-HIAA content, and a significantly smaller 5-HIAA/5-HT ratio (−9%, p = 0.01). Post-pubertal DHA-deficient and OXV rats did not exhibit significant alterations in PFC 5-HT or 5-HIAA content. Only perinatal DHA-deficient rats exhibited a significant reduction in midbrain TPH-2 mRNA expression (−29%, p = 0.03). These preclinical data support a causal link between perinatal omega-3 fatty acid deficiency and reduced central serotonin synthesis in adult female rats that is independent of ovarian hormones including estrogen.     

Tryptophan hydroxylase 2 gene is associated with major depression and antidepressant treatment response

Tryptophan hydroxylase-2 (TPH2) is the rate-limiting biosynthetic isoenzyme for serotonin that is preferentially expressed in the brain and has been implicated in the pathogenesis of major depressive disorder (MDD) and in the mechanism of antidepressant action. This study aimed to investigate whether common genetic variation in the TPH2 gene is associated with MDD and therapeutic response to antidepressants in a Chinese population. A total of 508 MDD patients and 463 unrelated controls were recruited. Among the MDD patients, 187 accepted selective serotonin reuptake inhibitor (fluoxetine or citalopram) antidepressant treatment for 8 weeks with therapeutic evaluation before and after treatment. Five TPH2 polymorphisms were genotyped and their association with MDD or treatment response was assessed by haplotype and single-marker analysis. In single-marker-based analysis, the rs17110747-G homozygote polymorphism was found to be more frequent in the MDD patients than in the controls (P = 0.002). Genotype analysis in responders (defined as those with a 50% reduction in baseline Hamilton score) and non-responders after 8 weeks of antidepressant treatment showed that the proportion of rs2171363 heterozygote carriers was higher in the responders than the non-responders (P = 0.009). No significant association with MDD or antidepressant therapeutic response was discovered in haplotype analyses. Our findings show that TPH2 genetic variants may play a role in MDD susceptibility and in acute therapeutic response to selective serotonin reuptake inhibitors.     

Effects of venlafaxine and fluoxetine on lymphocyte subsets in patients with major depressive disorder: A flow cytometric analysis

Background

Studies have yielded conflicting results concerning flow cytometric lymphocyte analyses in patients with depression. Data about the effect of antidepressants on lymphocyte subsets are also contradictory. The aim of this study was to determine effects of venlafaxine versus fluoxetine on lymphocyte subsets in depressive patients.

Methods

Sixty-nine patients diagnosed with major depressive disorder (MDD) according to DSM-IV and 36 healthy controls are included in the study. Sixty-nine patients were randomized to take fluoxetine (FLX) (n = 33) or venlafaxine (VEN) (n = 36). Serum lymphocyte subsets included CD3, CD4, CD8, CD16/56, CD19, CD45, Anti-HLA-DR which were measured by flow cytometric analyses at baseline and 6 weeks after the start of treatment. The severity of depression was evaluated with Hamilton rating scale for depression.

Results

At baseline, patients with MDD had significantly lower CD16/56 ratio and higher CD45 ratio compared to the controls. Although numerically higher in the VEN treated patients, treatment response rates between the FLX (53%) and the VEN (75%) groups were not different statistically. CD45 values decreased significantly in the VEN group at the end of the 6 week treatment period whereas no difference was observed in the FLX group. By the 6th week, treatment responders showed a significantly higher CD16/56 ratio than non-responders. Baseline severity of depression and anxiety was positively correlated with baseline CD45 ratio and negatively correlated with baseline CD16/56 ratio. We did not observe consistent changes in the absolute number of circulating B or T cells, nor in the helper/inducer (CD4) or suppressor/cytotoxic (CD8) subsets.

Conclusions

CD16/56 was lower in patients with MDD and increased in treatment responders at 6th week. CD45 ratio was higher in patients with MDD than healthy subjects; it decreased with antidepressant treatment and was positively correlated with the severity of depression. Antidepressant treatment contributes to immune regulation in patients with major depressive disorder.     

Metabotropic glutamate receptor protein expression in the prefrontal cortex and striatum in schizophrenia

We investigated the expression of metabotropic glutamate receptors (mGluR) in the prefrontal cortex (PFC) and striatum in schizophrenia. mGluRs modulate the release and reuptake of synaptic glutamate and mediate some molecular correlates of neuroplasticity, including long-term potentiation. The mGluRs are expressed widely in the PFC and striatum, regions often implicated in the pathophysiology of schizophrenia. Thus, we hypothesized that abnormal expression of mGluRs might contribute to glutamatergic dysfunction observed in the PFC and striatum in schizophrenia. Accordingly, we measured the expression of metabotropic glutamate receptors (mGluRs) in Brodmann areas 9, 11, 32, and 46 in the prefrontal cortex (PFC) and the caudate, putamen, and nucleus accumbens in schizophrenia (16 cases, 9 controls) by Western blot analysis. We found an increase in the expression of mGluR1a and mGluR2/3 immunoreactivity in the PFC in schizophrenia, while no changes in the expression of mGluR4a or mGluR5 were detected in this region. In the striatum we found no changes in the expression of any of the mGluRs studied. These results suggest that alterations of mGluR1a and mGluR2/3 expression in the PFC may contribute to the pathophysiology of schizophrenia, and support targeting these receptors for the generation of novel treatment modalities for this disabling illness.     

Serum IL-7 and G-CSF in major depressive disorder

Major depressive disorder (MDD) has been associated with dysregulated immune systems and impaired T cell function, but data on depression-related alterations in the levels of immunomodulatory growth factors are scarce. In order to further clarify the mechanisms underlying immune system dysregulation in depressed subjects, we examined the associations between MDD and serum levels of two immunomodulatory growth factors, interleukin (IL)-7 and granulocyte-colony stimulating factor (G-CSF), in 122 subjects (MDD with long-term symptomatology, n = 61; controls, n = 61). The MDD subjects had lowered levels of IL-7. In a model adjusted for age, gender and body mass index, subjects in the lowest tertile of IL-7 had a 3.4-fold increased likelihood for MDD (p = 0.010). Further adjustments for sleep disturbances, alcohol use, smoking, and metabolic syndrome did not alter these findings. Moreover, the exclusion of subjects with rheumatoid arthritis, coronary heart disease, or the use of non-steroidal anti-inflammatory medications or oral corticosteroids only slightly attenuated the findings. The G-CSF levels did not differ between the two groups. The lowering of the serum levels of IL-7, a regulator of T cell homeostasis, in MDD subjects may underlie the depression-related impaired T cell function.     

A synthetic cannabinoid agonist promotes oligodendrogliogenesis during viral encephalitis in rats

Chronic CNS infection by several families of viruses can produce deficits in prefrontal cortex (PFC) and striatal function. Cannabinoid drugs have been long known for their anti-inflammatory properties and their ability to modulate adult neuro and gliogenesis. Therefore, we explored the effects of systemic administration of the cannabinoid agonist WIN55,212-2(WIN) on prefrontal cortex (PFC) and striatal cytogenesis in a viral model of CNS injury and inflammation based on Borna Disease (BD) virus encephalitis. Active BrdU⁺ progenitor populations were significantly decreased 1 week after BrdU labeling in BD rats [p < 0.001 compared to uninfected (NL) controls] while less than 5% of BrdU⁺ cells colabeled for BDV protein. Systemic WIN (1 mg/kg i.p. twice daily × 7 days) increased the survival of BrdU⁺ cells in striatum (p < 0.001) and PFC of BD rats, with differential regulation of labeled oligodendroglia precursors vs microglia/macrophages. WIN increased the percentage of BrdU⁺ oligodendrocyte precursor cells and decreased BrdU⁺ ED-1-labeled phagocytic cells, without producing pro- or antiviral effects. BDV infection decreased the levels of the endocannabinoid anandamide (AEA) in striatum (p < 0.05 compared to NL rats), whereas 2-AG levels were unchanged. Our findings indicate that: 1) viral infection is accompanied by alterations of AEA transmission in the striatum, but new cell protection by WIN appears independent of its effect on endocannabinoid levels; and 2) chronic WIN treatment alters the gliogenic cascades associated with CNS injury, promoting oligodendrocyte survival. Limiting reactive gliogenesis and macrophage activity in favor of oliogodendroglia development has significance for demyelinating diseases. Moreover, the ability of cannabinoids to promote the development of biologically supportive or symbiotic oligodendroglia may generalize to other microglia-driven neurodegenerative syndromes including NeuroAIDS and diseases of aging.     

The SIGMAR1 gene is associated with a risk of schizophrenia and activation of the prefrontal cortex

Several studies have identified the possible involvement of sigma non-opioid intracellular receptor 1 (SIGMAR1) in the pathogenesis of schizophrenia. The Gln2Pro polymorphism in the SIGMAR1 gene has been extensively examined for an association with schizophrenia. However, findings across multiple studies have been inconsistent. We performed a meta-analysis of the association between the functional Gln2Pro polymorphism and schizophrenia using combined samples (1254 patients with schizophrenia and 1574 healthy controls) from previously published studies and our own additional samples (478 patients and 631 controls). We then used near-infrared spectroscopy to analyze the effects of the Gln2Pro genotype, a schizophrenia diagnosis and the interaction between genotype and diagnosis on activation of the prefrontal cortex (PFC) during a verbal fluency task (127 patients and 216 controls). The meta-analysis provided evidence of an association between Gln2Pro and schizophrenia without heterogeneity across studies (odds ratio = 1.12, p = 0.047). Consistent with previous studies, patients with schizophrenia showed lower bilateral activation of the PFC when compared to controls (p < 0.05). We provide evidence that Pro carriers, who are more common among patients with schizophrenia, have significantly lower activation of the right PFC compared to subjects with the Gln/Gln genotype (p = 0.013). These data suggest that the SIGMAR1 polymorphism is associated with an increased risk of schizophrenia and differential activation of the PFC.     

The kynurenine pathway in adolescent depression: Preliminary findings from a proton MR spectroscopy study

Background

Cytokine induction of the enzyme indoleamine 2,3-dioxygenase (IDO) has been implicated in the development of major depressive disorder (MDD). IDO metabolizes tryptophan (TRP) into kynurenine (KYN), thereby decreasing TRP availability to the brain. KYN is further metabolized into several neurotoxins. The aims of this pilot were to examine possible relationships between plasma TRP, KYN, and 3-hydroxyanthranilic acid (3-HAA, neurotoxic metabolite) and striatal total choline (tCho, cell membrane turnover biomarker) in adolescents with MDD. We hypothesized that MDD adolescents would exhibit: i) positive correlations between KYN and 3-HAA and striatal tCho and a negative correlation between TRP and striatal tCho; and, ii) the anticipated correlations would be more pronounced in the melancholic subtype group.

Methods

Fourteen adolescents with MDD (seven with melancholic features) and six healthy controls were enrolled. Minimums of 6 weeks MDD duration and a severity score of 40 on the Children's Depression Rating Scale-Revised were required. All were scanned at 3 T with MRI, multi-voxel 3-dimensional, high, 0.75 cm³, spatial resolution proton magnetic resonance spectroscopic imaging. Striatal tCho concentrations were assessed using phantom replacement. Spearman correlation coefficients were Bonferroni-corrected.

Results

Positive correlations were found only in the melancholic group, between KYN and 3-HAA and tCho in the right caudate (r = 0.93, p = 0.03) and the left putamen (r = 0.96, p = .006), respectively.

Conclusions

These preliminary findings suggest a possible role of the KYN pathway in adolescent melancholic MDD. Larger studies should follow.     

Association analysis of Group II metabotropic glutamate receptor genes (GRM2 and GRM3) with mood disorders and fluvoxamine response in a Japanese population

Background

Several lines of evidence implicate abnormalities in glutamate neural transmission in the pathophysiology of mood disorders, including major depressive disorder (MDD) and bipolar disorder (BP). Preclinical antidepressant effects were also reported for group II metabotropic glutamate receptor (Group II mGluRs) antagonists show dose-dependent antidepressant-like effects in murine models of depression. Also, it has been suggested that abnormalities in the hypothalamic-pituitary-adrenal axis and serotonergic neural transmission are important mechanisms in the pathophysiology of mood disorders. Group II mGluRs play an important role in regulating the function of these mechanisms. From these results, it has been suggested that abnormalities in Group II mGluRs might be involved in the pathophysiology of mood disorders, including MDD) and BP, and may influence the clinical response to treatment with SSRIs in MDD. Therefore, we studied the association between Group II mGluR genes (GRM2 and GRM3) and mood disorders and the efficacy of fluvoxamine treatment in Japanese MDD patients.

Materials and methods

Using three tagging SNPs in GRM2 and an SNP (rs6465084) reported functional variant in GRM3, we conducted a genetic association analysis of case-control samples (325 MDD patients, 155 BP patients and 802 controls) in the Japanese population. In addition, we performed an association analysis of GRM2 and GRM3 and the efficacy of fluvoxamine treatment in 117 Japanese patients with MDD. The MDD patients in this study had scores of 12 or higher on the 17 items of the Structured Interview Guide for Hamilton Rating Scale for Depression (SIGH-D). We defined a clinical response as a decrease of more than 50% in baseline SIGH-D within 8 weeks, and clinical remission as an SIGH-D score of less than 7 at 8 weeks.

Results

We found an association between rs6465084 in GRM3 and MDD in the allele-wise analysis after Bonferroni's correction (P-value = 0.0371). However, we did not find any association between GRM3 and BP or the fluvoxamine therapeutic response in MDD in the allele/genotype-wise analysis. We also did not detect any association between GRM2 and MDD, BP or the fluvoxamine therapeutic response in MDD in the allele/genotype-wise or haplotype-wise analysis.

Discussion

We detected an association between only one marker (rs6465084) in GRM3 and Japanese MDD patients. However, because we did not perform an association analysis based on LD and a mutation scan of GRM3, a replication study using a larger sample and based on LD may be required for conclusive results.     

Heart rate variability during motor and cognitive tasks in females with major depressive disorder

Research indicates that major depressive disorder (MDD) is associated with alterations in autonomic control, particularly cardiac control as measured by heart rate variability (HRV). In this preliminary study, we investigated the neural correlates of autonomic control by measuring both HRV and associated brain activity during the performance of mildly stressful tasks. Medically healthy female subjects with MDD (N = 10) and healthy controls (N = 7) underwent H₂¹⁵O-positron emission tomography (PET) and electrocardiographic ECG recording while performing a handgrip motor task and an n-back task. Indices of HRV were calculated and correlated with regional cerebral blood flow (rCBF). Differences in the rCBF and HRV correlations between depressed and healthy subjects were evident in both the medial and lateral orbital cortices. In addition, these areas appeared to be involved in different facets of autonomic control with regard to sympathetic or parasympathetic dominance of cardiac control. These results are consistent with the known roles of networks within the orbital cortex in both autonomic control and the pathophysiology of MDD.     

Metabotropic Glutamate Receptor 2 and 3 Gene Expression in The Human Prefrontal Cortex and Mesencephalon in Schizophrenia

Group II metabotropic glutamate receptors (mGluR2 and mGluR3) are implicated in schizophrenia. We characterized mGluR2 and 3 mRNA in the human prefrontal cortex (PFC) and mesencephalon, and then compared cases with schizophrenia to matched controls. In the human brain, both receptors were expressed in the PFC and, unlike the rodent, in dopaminergic (DA) cell groups. In schizophrenia, we found significantly higher levels of mGluR2 mRNA in the PFC white matter. The expression of mGluR2, 3 in DA cells provide a mechanism for glutamate to modulate dopamine release in the human brain and this species-specific difference may be critical to understanding rodent models in schizophrenia.     

Metabotropic glutamate receptor 2 and 3 gene expression in the human prefrontal cortex and mesencephalon in schizophrenia

Group II metabotropic glutamate receptors (mGluR2 and mGluR3) are implicated in schizophrenia. We characterized mGluR2 and 3 mRNA in the human prefrontal cortex (PFC) and mesencephalon, and then compared cases with schizophrenia to matched controls. In the human brain, both receptors were expressed in the PFC and, unlike the rodent, in dopaminergic (DA) cell groups. In schizophrenia, we found significantly higher levels of mGluR2 mRNA in the PFC white matter. The expression of mGluR2, 3 in DA cells provide a mechanism for glutamate to modulate dopamine release in the human brain and this species-specific difference may be critical to understanding rodent models in schizophrenia.     

Reduced platelet BDNF level in patients with major depression

Serum and plasma brain-derived neurotrophic factor (BDNF) levels as well as brain BDNF have previously been shown to be decreased in patients with major depressive disorder (MDD). We explored whether platelet BDNF levels, circulating stored BDNF, would be lower in MDD patients than in normal controls. BDNF levels were examined in platelet-rich plasma (PRP) and platelet-poor plasma (PPP) in 20 hospitalized non-suicidal MDD patients, 20 recent-suicidal MDD patients, and 20 normal controls. Platelet BDNF content was calculated by subtracting PPP BDNF level from PRP BDNF level, and dividing the result by the total platelet count, and it was expressed as pg/10⁶ platelets. Individuals were evaluated using a Structured Clinical Interview for DSM-IV and a Hamilton Depression Rating Scale. Platelet BDNF contents were significantly lower in non-suicidal patients (3.09 ± 2.53 pg/10⁶ platelets) and recent-suicidal MDD patients (3.16 ± 1.99 pg/10⁶ platelets) than in healthy controls (6.17 ± 2.64 pg/10⁶ platelets) (p < 0.01). However, platelet BDNF contents had no significant differences between non-suicidal and recent-suicidal patients. PRP BDNF levels were also significantly lower in non-suicidal and suicidal MDD patients than in healthy controls (p = 0.029), while PPP BDNF had no significant difference between 3 groups (p = 0.971). Our findings suggest that there is a decrease in the platelet BDNF of patients with major depression. Reduced platelet BDNF contents as circulating stored BDNF could be associated with lower serum BDNF level in patients with major depression.     

Repeated electroconvulsive seizure treatment in rats reduces inducibility of early growth response genes and hyperactivity in response to cocaine administration

Regulated expression of immediate early genes (IEGs) in the brain reflects neuronal activity in response to various stimuli and recruits specific gene programs involved in long-term neuronal modification and behavioral alterations. Repeated electroconvulsive seizure (ECS) treatment reduces the expression level of several IEGs, such as c-fos, which play important roles in psychostimulant-induced behavioral changes. In this study, we investigated the effects of repeated ECS treatment on the basal expression level of IEGs and its effects on cocaine-induced activation of IEGs and locomotor activity in rats. Repeated ECS treatment for 10 days (E10×) reduced Egr1, Egr2, Egr3, and c-fos mRNA and protein levels in the rat frontal cortex at 24 h after the last ECS treatment, and these changes were evident in the neuronal cells of the prefrontal cortex. In particular, downregulation of Egr1 and c-fos was evident until 5 days after the last ECS treatment. Moreover, E10× pretreatment attenuated the cocaine-induced increase in Egr1, Egr2, and c-fos expression in the rat frontal cortex, whereas phosphorylation of ERK1/2, one of the representative upstream activators of these genes, increased significantly following cocaine treatment. Additionally, E10× pretreatment attenuated the increase in locomotor activity in response to a cocaine injection. In conclusion, repeated ECS treatment reduced the expression and inducibility of Egrs and c-fos, which could attenuate the response of the brain to psychostimulants.