Uncovering the role of the nucleus accumbens in schizophrenia: A postmortem analysis of tyrosine hydroxylase and vesicular glutamate transporters

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Abstract

The nucleus accumbens (NAcc) is often implicated in schizophrenia (SZ) pathology, but with little evidence to support its role. This study examined postmortem human tissue to determine if abnormalities are present in the dopaminergic or glutamatergic systems in the NAcc in SZ. We compared the protein levels of tyrosine hydroxylase (TH) and vesicular glutamate transporters vGLUT1 and vGLUT2 in control (n = 7) and schizophrenia (n = 13) subjects using Western blot analysis. The SZ subjects were further divided by treatment status: SZ on-drug (SZ-ON, n = 6) and SZ off-drug (SZ-OFF, n = 7), to assess the effects of antipsychotic treatment. TH protein levels were similar between control and SZ subjects, and there was no difference between SZ-ON and SZ-OFF subjects. Protein levels of vGLUT1 were similar in control and SZ subjects, and there was no difference in vGLUT1 protein levels between SZ-ON and SZ-OFF subjects. In contrast, vGLUT2 protein levels were significantly elevated in the SZ group (25% increase). Protein levels of vGLUT2 did not differ between SZ-ON and SZ-OFF subjects. Similar levels of TH suggest the presynaptic DA pathway may be normal in the NAcc in SZ. The elevated vGLUT2 protein levels, but not vGLUT1, suggest the NAcc receives increased glutamatergic input in SZ, possibly from thalamic or other subcortical origins. The similarity between SZ-ON and SZ-OFF subjects suggests that the results are not caused by APD treatment. These findings provide further insight into the role of the NAcc in SZ.

Introduction

The cause of schizophrenia (SZ) remains elusive, however decades of research have revealed common pathologies, some of which are now hallmarks of the disorder. One of these hallmarks is abnormalities in the dopamine (DA) system in the striatum of patients with SZ (Miyake et al., 2011). Of special interest is a subregion of the ventral striatum, the nucleus accumbens (NAcc), which has been assumed to be a prime location for the elevated DA levels in SZ, based on its functional properties and evidence of antipsychotic drug (APD) action here (Deutch and Cameron, 1992, Deutch et al., 1992, Robertson and Fibiger, 1992Merchant and Dorsa, 1993). Until recently, imaging studies have had to analyze the striatum as a single region, without the ability to distinguish between key functional and anatomical areas. Thus, not only has the role of the NAcc has never been confirmed, but recent studies with improved imaging techniques suggest that this subregion may not be implicated as previously thought (Howes et al., 2009, Howes et al., 2011, Kegeles et al., 2010). Additionally, conclusions surrounding the DA abnormalities in the striatum lack solid support from postmortem studies, which offer the ability to study these subregions individually. Previous studies in postmortem NAcc report conflicting results, typically finding no change (Crow et al., 1979, Farley et al., 1977, Owen et al., 1978, Toru et al., 1982) or increases (Bird et al., 1979, Mackay et al., 1982) in DA measures.

Another, more recent hypothesis of SZ is a causal role of glutamate abnormalities. Within the NAcc, a hypothesis emerged that pathology of the glutamate system could drive DA dysfunction (Lodge and Grace, 2007, Lodge and Grace, 2011). NMDA antagonists have been shown to enhance DA efflux in the striatum in both preclinical studies (Miller and Abercrombie, 1996) and imaging studies in healthy volunteers (Kegeles et al., 2000). Further, stimulation of glutamatergic input to the NAcc in rats causes elevated DA release in the region (Blaha et al., 1997, Legault and Wise, 1999). It is not known however, if glutamatergic abnormalities are actually present in the NAcc in SZ.

The purpose of this study was to determine if neurochemical abnormalities are present in the NAcc of postmortem SZ. To study the dopaminergic system, we analyzed tyrosine hydroxylase (TH), the rate-limiting synthesizing enzyme of DA. To study the glutamatergic system, we analyzed the vesicular glutamate transporters vGLUT1 and vGLUT2, which are essential for the uptake and storage of glutamate into synaptic vesicles (Bellocchio et al., 2000, Takamori et al., 2000). They have complementary localization patterns in cortical and subcortical structures, respectively (Hisano et al., 2000, Fremeau et al., 2001, Herzog et al., 2001) thus studying both markers provides a comprehensive analysis of glutamatergic input to the NAcc. The Western blot analyses were performed both in SZ subjects that were on medication at the time of death, and in SZ subjects that were off medication at the time of death to assess APD effects.

Section snippets

Postmortem tissue

Postmortem human brain tissue was obtained from the Maryland Brain Collection. DSM-IV diagnosis of SZ was confirmed by two psychiatrists based on patient medical records, family interviews, autopsy reports, and neuropathologic assessments. Schizophrenia subjects were considered off-drug if they had been untreated with APDs for at least 6 months prior to death. Control cases had no history of psychiatric or neurological disease. Cases were chosen based on the best match of age, race, sex,

Actin protein levels in human NAcc

Protein levels of actin were measured in SZ and control subjects. Western blotting yielded an intense band at the correct molecular weight of ~ 42 kDa. Un-normalized actin levels did not differ between the NC and SZ groups. The mean un-normalized actin levels for the blots used in the TH analysis were 0.250 ± 0.007 (NC) and 0.247 ± 0.012 (SZ); p = 0.563. The mean un-normalized actin levels for the blots used in the vGLUT analysis were 0.251 ± 0.034 (NC) and 0.292 ± 0.046 (SZ); p = 0.053.

Tyrosine hydroxylase protein levels in human NAcc

Protein levels of TH

Discussion

In this study we found that TH protein levels did not differ between control and SZ groups in the NAcc. Protein levels of vGLUT2 were significantly increased in SZ subjects compared to controls, whereas levels of vGLUT1 were similar. There were no differences in TH, vGLUT1, or vGLUT2 protein measures between the SZ-ON and SZ-OFF subjects. To our knowledge, this is the first study to analyze TH and vGLUT protein levels in postmortem NAcc in on-drug and off-drug SZ subjects.

Role of funding sources

This research was supported by the National Institute of Mental HealthF31MH098566 (LAM) and RO1MH066123 (RCR). The funding source had no role in study design, data collection, analysis and interpretation of data, writing the manuscript, or in the decision to submit the paper for publication.

Contributors

Drs. Roberts and McCollum conceived the idea and methodology of this study. Dr. McCollum conducted the data collection and statistical analyses. Both authors contributed to and have approved the final manuscript.

Conflict of interest

The authors have no conflicts of interest to declare.

Acknowledgments

We would like to thank the staff of the Maryland Brain Collection for their assistance with collection of the human tissue used in this study, and Courtney Walker for her technical assistance.

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