Decreased synaptic and mitochondrial density in the postmortem anterior cingulate cortex in schizophrenia

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Abstract

Schizophrenia (SZ) is a mental illness characterized by psychosis, negative symptoms, and cognitive deficits. The anterior cingulate cortex (ACC), a structurally and functionally diverse region, is one of several brain regions that is abnormal in SZ. The present study compared synaptic organization and mitochondrial number and morphology in postmortem ACC in SZ versus normal control (NC). Total synaptic density in the combined ACC was decreased in SZ, to 72% of normal controls (NCs), due to selective decreases in axospinous synapses, both asymmetric (excitatory) and symmetric (inhibitory). These changes were present in layers 3 and 5/6. The density of mitochondria in all axon terminals combined in SZ was decreased to 64% of NC. In layer 3, mitochondrial density was decreased only in terminals forming asymmetric synapses with spines, while in layers 5/6 mitochondrial density was decreased in terminals forming symmetric synapses with spines and dendrites. The proportion of terminals making symmetric synapses that contained mitochondria was significantly lower in SZ than in NCs, especially for symmetric axospinous synapses. The number of mitochondria per neuronal somata was decreased in the ACC in SZ compared to NCs; this finding was present in layers 5–6. The size of mitochondria in neuronal somata and throughout the neuropil was similar in SZ and NCs. Our results, though preliminary, are well supported by the literature, and support an anatomical substrate for some of the altered executive functions found in SZ.

Introduction

The anterior cingulate cortex (ACC) is part of prefrontal cortex and its neuroanatomy has been well described (DeFelipe et al., 2002, Jones, 1998, Lewis et al., 2002, Peters, 2002). The ACC is composed of distinct anatomical subregions, each with different functional properties (Peterson et al., 1999, Palomero-Gallagher et al., 2008, Vogt et al., 1992). At the microscopic level, each layer of the ACC has characteristic types of neurons, which project to particular targets; each layer is the recipient of inputs from specific regions (DeFelipe et al., 2002, Lewis et al., 2002). Human imaging studies indicate that the dorsal ACC is involved in mediating attention and executive functions, such as task difficulty, remote memory (Koski and Petrides, 2001), conflict (Barch et al., 2001, Kerns et al., 2004), response inhibition and error commission (Braver et al., 2001, Mathalon et al., 2002). The subcallosal ACC is involved in emotional processing (Bush et al., 2000) or internal states (Greicius et al., 2003), the dorsal ACC is involved in cognitive function, and the rostral ACC, located between these two subdivisions, plays an important role in the integration of these functions (Vogt et al., 1992).

The ACC is one of several brain regions that are abnormal in schizophrenia (SZ), as shown in both in vivo imaging and postmortem studies (Fornito et al., 2009). In vivo imaging of people with SZ has shown abnormalities in multiple transmitter systems (Egerton et al., 2012, Kraguljac et al., 2012, Rowland et al., 2013, Théberge et al., 2003), blood flow (Holcomb et al, 2000) and metabolism (Nordahl et al., 1996, Tamminga et al., 1992). Functional impairments in cognitive interference (Heckers et al., 2004), error or conflict monitoring (Alain et al., 2002, Carter et al., 1997, Carter et al., 2001) and response monitoring (Kopp and Rist, 1999, Mathalon et al., 2002) have also been demonstrated.

Postmortem studies have shown multiple defects in the ACC in SZ (Eastwood and Harrison, 2001, Fornito et al., 2009). Neurochemical and molecular changes include altered distribution or modulation of dopamine (Benes et al., 1997), abnormalities in multiple aspects of the glutamate system (Barksdale et al., 2014, Bauer et al., 2008, Bauer et al., 2010, Drummond et al., 2013, Katsel et al., 2011, Oni-Orisan et al., 2008, Woo et al., 2004, Woo et al., 2007) and intracellular signaling abnormalities (Funk et al., 2012, Funk et al., 2014). Mitochondrial pathology has been implicated repeatedly in schizophrenia (Anglin et al., 2012, Manji et al., 2012) and in the ACC, oxidative stress, a result of mitochondrial production of reactive oxygen species, is increased (Wang et al., 2009). Anatomical abnormalities include layer specific alterations in neuronal distribution and density (Benes and Bird, 1987, Brune et al., 2010, Todtenkopf et al., 2005), and increased numbers of glutamatergic (Benes et al., 1987, Benes et al., 1992) and parvalbumin axons (Kalus et al., 1997, Kalus et al., 1999). Of note, previous electron microscopic studies in SZ have shown layer specific abnormalities such as fewer synapses in the ACC (Aganova and Uranova, 1992) and alterations in synapses, mitochondria and oligodendrocytes in other areas of prefrontal cortex (Uranova et al., 2004, Uranova et al., 2007, Uranova et al., 2011).

The purpose of the present study is to compare the synaptic organization and mitochondrial number and morphology in SZ versus normal control postmortem ACC. Several synaptic features were quantified, including morphological features which identify excitatory and inhibitory synapses. This work has been presented in preliminary form (Barksdale et al., 2012a, Barksdale et al., 2012b, Roberts et al., 2013).

Section snippets

Methods

Human brain tissue was obtained with IRB approved protocols from the Maryland Brain Collection and the Alabama Brain Collection. Diagnostic criteria have been described previously (McCollum et al., in press, Roberts et al., 2008). Demographics are presented in Table 1.

Coronal blocks from the dorsal ACC were preserved in fixative and processed for electron microscopy as previously described (McCollum et al., in press). One series of sections was stained for Kluver-Barrera stain as previously

Demographics

Demographics for the groups were similar for age, PMI, sex and race (Table 1).

Synapses

There was no apparent difference in the integrity of the tissue between the NC (Fig. 3) and SZ groups (Fig. 4). As expected, the majority of synapses in both groups were asymmetric axospinous synapses (Figs. 3A,C,D, 4A,D). Asymmetric axodendritic synapses (Figs. 3B, 4A), symmetric axospinous synapses (Figs. 3C, 4B), symmetric axodendritic synapses (Figs. 3E, 4C) were also present in both groups.

In the ACC (layers 3, 5

Discussion

Our main results show reduced overall synaptic density in the ACC in SZ compared to NCs. The synaptic loss was confined to axospinous synapses and was present in layers 3 and 5/6. Both asymmetric and symmetric axospinous synapses were affected, indicating abnormalities in both excitatory and inhibitory transmission, respectively. Overall decreases in synaptic density in the SZ group are consistent with the reduced neuropil hypothesis, which posits fewer cortical synaptic connections (Selemon

Role of funding source

NIMH supported ACL and RCR for time, travel and supplies. NIMH funded JKR for time. The MERIT scholarship funded KAB for time spent on the project.

Contributions

Dr. Roberts designed the experiment, analyzed data and wrote the manuscript. Dr. Barksdale collected data on the electron microscope and analyzed data. Ms. Roche processed the tissue for electron microscopy, did all of the thin sectioning and collected data on the electron microscope. Dr. Lahti designed the experiment and provided input on the meaning and interpretation of the results. All authors contributed to and have approved the final manuscript.

Conflicts of interest

The authors have no conflicts of interest.

Acknowledgments

This research was supported by NIH grant MH081014 (ACL), and a 5K12GM088010-02 MERIT Scholarship (KAB). We thank the staff of the Maryland and Alabama Brain Collections and Racheal McKinney for formatting the references.

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