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

https://doi.org/10.1016/j.pnpbp.2011.05.010Get rights and content

Abstract

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.

Research highlights

► We used postmortem cortical tissue to examine mTOR signaling in depression. ► Reductions in mTOR, p70S6K, eIF4B and p-eIF4B were identified. ► No differences were seen in eIF4E, p-eIF4E or actin levels. ► Specific dysregulation of mTOR/p70S6K/eIF4B signaling is evident in depression. ► Dysfunction of mTOR signaling may underlie synaptic deficits in depression.

Introduction

A major limitation of established antidepressants is the delayed onset of therapeutic response, resulting in non-compliance and dramatically increased risk for suicidal behavior. Of particular relevance is the demonstration that a single dose of ketamine, a glutamate N-methyl-d-aspartate (NMDA) receptor antagonist, induced a rapid (within hours), long lasting (up to 1 week), and robust antidepressant effect in treatment-resistant cases of MDD (Berman et al., 2000, Zarate et al., 2006). Recent animal studies indicate that the fast antidepressant response to NMDA receptor antagonists (ketamine and Ro 25–6981) is mediated by rapid activation of the mammalian target of rapamycin (mTOR) pathway leading to an increase in synaptic signaling proteins and increased number and function of new spine synapses in the prefrontal cortex (PFC) of rats (Li et al., 2010). Moreover, it has been demonstrated that a single dose of these antagonists rapidly reversed the chronic stress-induced behavioral and synaptic deficits in an mTOR-dependent manner (Li et al., 2011). Our recent postmortem studies show significant reductions in the expression of prominent postsynaptic proteins involved in glutamate neurotransmission, including NMDA receptor subunits (NR2A and NR2B), metabotropic glutamate receptor subtype 5 (mGluR5) and postsynaptic density 95 kDa (PSD-95) in the PFC from depressed subjects (Deschwanden et al., 2011, Feyissa et al., 2009). These studies may indicate an association between marked deficits in synaptic proteins and dysregulation of mTOR signaling in MDD (Karolewicz et al., 2011).

Traditionally, it was thought that the change in the proteome is caused by transcriptional activity. Now it is known that regulation of translation is another way of altering protein production (Nilsson et al., 2004). Protein synthesis is a highly regulated process that can be separated into three general phases: initiation, elongation and termination (Hoeffer and Klann, 2010, Klann et al., 2004). The rate-limiting step in the process of protein synthesis is translation initiation (Hoeffer and Klann, 2010, Holz et al., 2005). The activity of mTOR, an ubiquitously expressed serine/threonine kinase, is central to the regulation of translation initiation and, consequently, protein synthesis required for long-term potentiation and new synaptic connections (Hashimoto, 2011, Hoeffer and Klann, 2010, Klann et al., 2004, Tang et al., 2002, Tang and Schuman, 2002).

It has been reported that neuronal mTOR function is influenced by the activity of growth factors, insulin, cytokines, as well as glutamate activity via NMDA receptors and metabotropic glutamate receptors (mGluR) (Antion et al., 2008, Gong et al., 2006, Hay and Sonenberg, 2004, Hoeffer and Klann, 2010) (Fig. 1). Activated mTOR phosphorylates p70-kDa ribosomal protein S6 kinase (p70S6K) followed by p70S6K-induced phosphorylation of eukaryotic initiation factor 4B (eIF4B) which promotes the initiation of protein translation (Raught et al., 2004). mTOR also phosphorylates and inactivates eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), reducing its affinity for eIF4E and releasing eIF4E to facilitate translation initiation. Activated translation initiation factors, particularly eIF4E and eIF4B, are responsible for ribosome recruitment to the 5′ end of mRNA. The 5′ end of all nuclear-transcribed mRNAs possess a cap structure (m7GpppN, in which “m” represents a methyl group and “N”, any nucleotide) that is specifically recognized by eIF4E. Thus, eIF4E guides the ribosome to an mRNA 5′ end and facilitates its binding. On the other hand, eIF4B potentiates ribosome recruitment by stimulating the helicase activity of the eukaryotic initiation factor 4A (eIF4A), to unwind mRNA secondary structure for efficient translation (Gingras et al., 1999, Hay and Sonenberg, 2004, Holz et al., 2005, Rogers et al., 2002). Thus, mTOR controls the efficiency of protein translation within cells via its critical downstream targets (Fig. 1).

There is abundant evidence linking mTOR signaling to synaptic plasticity, memory, neurological disorders, and cancer (Gong et al., 2006, Hay and Sonenberg, 2004, Hoeffer and Klann, 2010). To date there are no studies that implicate the mTOR signaling pathway in the pathology of MDD. We hypothesize that deficits in the mTOR-dependent translation initiation pathway contribute to the molecular pathology seen in the PFC in MDD. Therefore, the goal of this study is to examine MDD-related changes in the protein level of mTOR and its downstream signaling targets: p70S6K, eIF4E, eIF4B in cortical tissue (PFC BA10) from the same MDD subjects as those used in our previous postmortem studies (Deschwanden et al., 2011, Feyissa et al., 2009). Additionally, levels of eIF4E phosphorylated at serine 209 (p-eIF4E Ser209) and eIF4B phosphorylated at serine 504 (p-eIF4B Ser504) were examined.

Section snippets

Human subjects

Postmortem brain samples were collected at autopsy at the Cuyahoga County Coroner's Office in Cleveland, OH. Informed written consent was obtained from the legal next-of-kin of all subjects. Next-of-kin were interviewed and retrospective psychiatric assessments were conducted in accordance with Institutional Review Board policies at Case Western Reserve University and the University of Mississippi Medical Center. A trained interviewer administered the Schedule for Affective Disorders and

Results

Amounts of mTOR, p70S6K, eIF4E, eIF4B, p-eIF4E (Ser209), and p-eIF4B (Ser504) were analyzed in the PFC BA10 from 12 pairs of subjects with MDD and matched healthy controls. Fig. 2 shows representative immunoblots from 3 pairs of subjects used in the analysis. The amount of mTOR immunoreactivity from depressed subjects (0.42 ± 0.06) was significantly lower compared to controls (0.616 ± 0.055; t = 4.13 df = 11, p = 0.0017, Fig. 3). Similarly, there was a robust reduction in the level of p70S6K in depressed

Discussion

The present study is the first to analyze levels of the mTOR-dependent translation initiation factors in the PFC from subjects diagnosed with MDD. Significant reductions in the expression of mTOR, p70S6K, eIF4B, and p-eIF4B were observed in MDD subjects as compared to psychiatrically healthy controls. In contrast, levels of eIF4E and p-eIF4E were unchanged in depressed subjects. Previously, we reported marked deficits in prominent postsynaptic proteins involved in glutamate neurotransmission

Acknowledgements

We thank Dr. Feyza Aricioglu for comments on the manuscript. The authors thank Drs. Dorota Maciag and Warren May for assistance in statistical analysis. We gratefully acknowledge the assistance of Drs James C Overholser, George Jurjus, Herbert Y Meltzer, and Ginny Dilley and Lisa Konick, MA, in the establishment of retrospective psychiatric diagnoses. The excellent assistance of the Cuyahoga County Coroner's Office, Cleveland, OH, is greatly appreciated. We thank the next-of-kin for their

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    Mrs. Jernigan and Dr. Goswami contributed equally to this article.

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