Mitochondrial multifaceted dysfunction in schizophrenia

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Author's Personal Copy Schizophrenia Research 187 (2017) 3–10

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Mitochondrial multifaceted dysfunction in schizophrenia; complex I as a possible pathological target Dorit Ben-Shachar ⁎ Laboratory of Psychobiology, Department of Psychiatry, Rambam Health Care Campus, B. Rappaport Faculty of Medicine, Rappaport Family Institute for Research in the Medical Sciences, Technion-IIT, Haifa, Israel

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Article history: Received 15 August 2016 Received in revised form 10 October 2016 Accepted 14 October 2016 Available online 29 October 2016 Keywords: Mitochondria Complex I Mitochondrial dynamics Mitochondrial respiration Neuronal differentiation Schizophrenia

a b s t r a c t Mitochondria are key players in various essential cellular processes beyond being the main energy supplier of the cell. Accordingly, they are involved in neuronal synaptic transmission, neuronal growth and sprouting and consequently neuronal plasticity and connectivity. In addition, mitochondria participate in the modulation of gene transcription and inflammation as well in physiological responses in health and disease. Schizophrenia is currently regarded as a neurodevelopmental disorder associated with impaired immune system, aberrant neuronal differentiation and abnormalities in various neurotransmitter systems mainly the dopaminergic, glutaminergic and GABAergic. Ample evidence has been accumulated over the last decade indicating a multifaceted dysfunction of mitochondria in schizophrenia. Indeed, mitochondrial deficit can be of relevance for the majority of the pathologies observed in this disease. In the present article, we overview specific deficits of the mitochondria in schizophrenia, with a focus on the first complex (complex I) of the mitochondrial electron transport chain (ETC). We argue that complex I, being a major factor in the regulation of mitochondrial ETC, is a possible key modulator of various functions of the mitochondria. We review biochemical, molecular, cellular and functional evidence for mitochondrial impairments and their possible convergence to impact in-vitro neuronal differentiation efficiency in schizophrenia. Mitochondrial function in schizophrenia may advance our knowledge of the disease pathophysiology and open the road for new treatment targets for the benefit of the patients. © 2016 Elsevier B.V. All rights reserved.

1. Introduction Schizophrenia is a complex and one of the most severe brain disorders, which affects main human capabilities; reality perception, emotion, cognition and social functioning (McGlashan, 1988). Schizophrenia onset is at adolescence with about 1% worldwide prevalence and is considered mostly independent of culture identity or ethnicity (Messias et al., 2007). Currently, there is no biochemical or imaging clear-cut accepted test and diagnosis of schizophrenia is entirely based on clinical assessment. Despite decades of research efforts, the etiology and pathophysiology of schizophrenia are still largely unknown. This is partly due to the longitudinal nature of schizophrenia pathophysiology, which most likely begins in-utero reaching into adulthood, the multifactorial etiology, probably involving a yet undetermined number of interacting genetic, epigenetic and environmental influences, and the inaccessibility of the living human brain for routine clinical and scientific investigations. Despite these limitation ample data accumulated over the years, point toward several possible etiological ⁎ Laboratory of Psychobiology, Department of Psychiatry, Rambam Health Care Campus and B. Rappaport Faculty of Medicine, Technion ITT, POB 9649, Haifa 31096, Israel. E-mail address: [email protected].

http://dx.doi.org/10.1016/j.schres.2016.10.022 0920-9964/© 2016 Elsevier B.V. All rights reserved.

hypotheses. The first and most accepted etiology in schizophrenia is the imbalances in neurotransmitter systems, mainly in the dopaminergic (Kapur and Remington, 2001; Matthysse, 1974; Seeman, 1987), glutamatergic (Deakin et al., 1989; Kantrowitz and Javitt, 2010; Konradi and Heckers, 2003), and GABAergic (Caruncho et al., 2004; Frankel et al., 2000; Frankle et al., 2015; Lewis et al., 1999) and their disrupted interactions (Carlsson et al., 2001; Menschikov et al., 2016). Additional theories for schizophrenia pathogenesis include genetic and environmental factors (Bleuler, 1963; Fatjo-Vilas et al., 2008; McGuffin, 2004), impaired neural development and connectivity (Conrad and Scheibel, 1987; Marenco and Weinberger, 2000; McGlashan and Hoffman, 2000; White and Hilgetag, 2011), neuroinflammation (Tomasik et al., 2016; Trépanier et al., 2016) as well as abnormal bioenergetics (Ben-Shachar and Laifenfeld, 2004; Fujimoto et al., 1992; Takahashi et al., 1994; Yuksel et al., 2015). The mitochondrion, a cellular organelle involved in the regulation of a variety of complex cellular and physiological processes, is of relevance for most of the currently prevailing hypotheses in schizophrenia. Mitochondria, the cell energy source, have a crucial role in additional key cellular processes, such as keeping intracellular Ca2 + homeostasis, producing reactive oxygen species (ROS), activating the intrinsic apoptotic pathway as well as heme and steroid production, thereby driving

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biochemical and molecular processes involved in various cell functions in health and disease. The role of mitochondria in brain development and differentiation (Kasahara and Scorrano, 2014; Solá et al., 2013), neuronal activity, sprouting and plasticity (Ben-Shachar and Laifenfeld, 2004; Courchet et al., 2013; Kang et al., 2008; Li et al., 2004) has been widely reported. In addition, it has been shown that mitochondria are targets for neurotransmitters and interact with dopamine, serotonin and glutamate (Ben-Shachar et al., 2004; Brenner-Lavie et al., 2008; Chen et al., 2008; White and Reynolds, 1996). This diversity of mitochondrial functions raised the interest in mitochondrial research in the last decade across pathologies (Picard et al., 2016) including mental disorders. Numerous evidence from studies using a wide array of experimental techniques ranging from imaging studies to ultrastructural methods to genetic and molecular means, suggests a role for mitochondria in mental disorders in general and in schizophrenia in particular. The present article focuses on cellular, molecular and biochemical evidences for mitochondria dysfunction in schizophrenia, based on studies of brain postmortem specimens, somatic cells and induced pluripotent stem cells (iPSCs) differentiated into dopaminergic and glutamatergic neurons. Specific attention is paid to the first complex, (complex I), of the ETC and its pathological interaction with dopamine, as a possible main cause for mitochondrial dysfunction in schizophrenia. Additionally, we discuss the relevance of these processes to impaired brain bioenergetics, neuronal plasticity and

connectivity and thereby their consequent cognitive and behavioral anomaly, characteristic of schizophrenia. Imaging bioenergetics and metabolic pathway impairments driven by mitochondrial dysfunction as well as mitochondrial relevant genetic findings are beyond the scope of this article.

2. Mitochondrial complex I The oxidative phosphorylation system (OXPHOS) is the cellular mechanism for energy production in the form of ATP. It is comprised of four respiratory enzyme complexes (complexes I–IV) and two electron transfer shuttling proteins, coenzyme Q (CoQ) and cytochrome c, which are arranged in a specific orientation in the inner mitochondrial membrane (Fig. 1A). Reduced electron carriers such as NADH (complex I substrate) and FADH2 (complex II substrate) generated from glycolysis and the citric acid cycle, release electrons that are ultimately transferred through the respiratory chain to molecular oxygen. This process is coupled to proton translocation across the inner membrane of the mitochondria forming an electrochemical gradient with a proton motivation force of ~ 200 mV, which enables ATP synthesis by the fifth complex, ATP synthase. Each complex of the ETC consists of multiple subunits encoded either by the nuclear DNA (nDNA) (70 subunits) or by the mitochondrial DNA (mtDNA) (13 subunits).

Fig. 1. A schematic presentation of mitochondrial oxidative phosphorylation system (OXPHOS) (A) and complex I (CoI) (B). Through the (OXPHOS) the electrons flow from the electron donors NADH (CoI) and FADH2 (CoII) to oxygen reducing it to water. This reaction is coupled with the proton pumping from the matrix through the inner mitochondrial membrane, which generates an electrochemical gradient used for ATP synthesis by CoV. Complex I has an L-shape structure with 14 core subunits, out of its 44–45 subunits, well conserved among species in mammals. Seven subunits are encoded by the mitochondrial DNA and are embedded in the inner membrane of the mitochondria. In SZ reduced CoI activity associated with mRNA and protein levels of NDUFV1, NDUFV2 and NDUFS1 has been reported. Less consistent changes were reported for its other subunits and for the other complexes of the OXPHOS.

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Complex I (NADH:ubiquinone oxidoreductase) is the first and largest complex of the respiratory chain and a cornerstone of mitochondrial function (Fig. 1B). It exerts a high level of control over OXPHOS activity, as it contributes to about 40% of the proton motive force for ATP synthesis (Galkin et al., 2006; Telford et al., 2009) and is a major site for ROS production (Kudin et al., 2008; Murphy, 2009). Eukaryotic complex I consists of 14 conserved subunits, which are homologous to the bacterial subunits, and more than 26 accessory subunits. In mammals, complex I consists of 44 or 45 subunits, which must be assembled correctly to form the properly functioning mature complex. For further complex I structural-functional information see the following reviews (Hirst, 2013; Hunte et al., 2010; Vinothkumar et al., 2014; Wirth et al., 2016). Complex I deficits have been associated with various disorders including mitochondrial disorders and neuropsychiatric disorders. In mitochondrial disorders, complex I deficiency is the most common enzyme defect, mostly due to a total of 36 mutations that have been identified in its nuclear DNA encoded subunits. Additional mutations have been observed in genes involved in complex I assembly and in its 7 mtDNA encoded subunits (Swalwell et al., 2011). Diseases associated with these mutations include lethal infantile mitochondrial disease, Leigh's syndrome, mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) and mitochondrial myopathy (Calvo et al., 2010; Kirby, 2004; McFarland et al., 2004). These disorders are frequently associated with psychiatric symptoms including psychosis (Anglin et al., 2012; Fattal et al., 2006; Mancuso et al., 2013). The connection of complex I to neuropsychiatric disorders is less direct, however decreased activity and SNP polymorphism have been observed in disorders such as Parkinson's disease, dystonia, Leber's hereditary optic neuropathy, schizophrenia, and bipolar disorder (Akarsu et al., 2015; Andreazza et al., 2010; Ben-Shachar, 2009; Konradi et al., 2004; Schapira, 1998; Washizuka et al., 2009).

3. Complex I as a target for dopamine and antipsychotic drugs An additional link between complex I and schizophrenia is its interaction with dopamine, a major pathological substrate in the disease, and with antipsychotic drugs. Dopamine is a catechol neurotransmitter involved in movement and cognitive functions through its interaction with its pre- and post-synaptic receptors. In addition, due to its high redox reduction potential it can be auto-oxidized in aqueous solution to highly reactive dopamine-quinones and ultimately to melanine (Das et al., 1978; Zhang and Dryhurst, 1993). Thus, depending on the cellular state, dopamine can exert oxidative stress and thereby neurodegeneration by two different mechanisms, its enzymatic metabolism by monoamine oxidase, which results in the formation of H2O2, and its auto-oxidation to quinones, which can bind to cysteine moiety of different proteins or enzymes to produce 5-S-cysteinyldopamine (Zhang and Dryhurst, 1993). It has been shown that dopamine inhibits mitochondrial respiration by products obtained by both mechanisms (Barros-Miñones et al., 2015; Berman et al., 1999; Kroll and Czyzyk-Krzeska, 1998; Mytilineou et al., 1993). We have suggested an alternative mechanism by which dopamine inhibits complex I activity via a direct interaction, without the induction of reactive oxygen species (Ben-Shachar et al., 1995). In this context, it is important to note that mitochondria are target organelles for dopamine, as monoamine oxidase is located on their outer membrane. We have shown that dopamine can be taken up by mitochondria (Brenner-Lavie et al., 2008) and through its iron chelating catechol moiety (Ben-Shachar et al., 1995; Rajan et al., 1976), directly interact with complex I, probably with an iron-sulfur cluster between the complex I binding site for NADH and its N1 cluster (Ben-Shachar et al., 2004). This inhibition of complex I activity is specific as dopamine does not affect complexes II–V of the ETC. In addition, dopamine inhibition of complex I is associated with reduced ATP production both in-vivo and in neuronal cell cultures, dissipates mitochondrial membrane potential and impairs

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cellular respiration (Ben-Shachar et al., 2004; Brenner-Lavie et al., 2009; Chan et al., 1994; Przedborski et al., 1993). Complex I is also a target for typical and atypical antipsychotic drugs as these drugs specifically inhibit complex I driven, yet not complex II driven, mitochondrial respiration in isolated mitochondria, in-vivo in animal models and in human tissue (Balijepalli et al., 1999, 2001; Burkhardt et al., 1993; Maurer and Moller, 1997; Modica-Napolitano et al., 2003; Prince et al., 1997). In stable schizophrenic patients treated with antipsychotic drugs a decreased activity of complex I in peripheral blood mononuclear cells has been reported (Casademont et al., 2007). The mechanism by which antipsychotic drugs inhibit complex I is still unclear, and can be due to a direct interaction with the drug or its metabolites, due to an indirect mechanism, or both. Our findings in rat brain isolated mitochondrial homogenate, in which antipsychotic drugs inhibited complex I activity, favor a direct interaction mechanism (Rosenfeld et al., 2011). Regardless, the findings that complex I is a mutual target for DA and antipsychotic drugs accentuate complex I relevance to the pathology of schizophrenia. The findings that dopamine inhibits complex I activity and ATP production in an ADP dependent reversible manner (Brenner-Lavie et al., 2009) suggest that this is a natural mechanism by which the self compensates for increased activity of the dopaminergic neurons. We suggest that antipsychotic drugs mimic dopamine inhibitory action, which may not be sufficient during psychosis, reduce neuronal activity and dopamine release and thereby contribute to the relief of psychosis. Interestingly, in schizophrenia-derived cells such as platelets, lymphocytes, Epstein Barr Virus (EBV) transformed B lymphocytes (lymphoblasts), keratinocytes and iPSCs, dopamine inhibition of complex I activity and complex I driven respiration was two times greater than in cells derived from healthy subjects, patients with bipolar disorder and patients with major depression (Ben-Shachar et al., 1999; Robicsek et al., 2013; Rosenfeld et al., 2011). Antipsychotic drugs however, inhibit complex I to the same extent in schizophrenia and the control groups. This difference between antipsychotic drugs and dopamine is probably due to the different site of interaction of the antipsychotic drugs, which is downstream to the N1 cluster of complex I (Fig. 1B) (Rosenfeld et al., 2011) and suggests a conformational change at its site of interaction with dopamine in schizophrenia. 4. Complex I in schizophrenia 4.1. Complex I activity Mitochondrial OXPHOS dysfunction has been repeatedly observed in mental disorders, specifically in schizophrenia and bipolar disorder, in brain and peripheral cells (Andreazza et al., 2010; Bergman and Ben-Shachar, 2016; Cavelier et al., 1995; Prince et al., 2000). Early studies focused on complex IV activity in schizophrenia brains, as it was regarded an endogenous metabolic marker for neuronal activity (Wong-Riley, 1989). Yet, controversial findings were obtained (Cavelier et al., 1995; Maurer et al., 2001; Prince et al., 2000; Rice et al., 2014) similar to those obtained for brain complex I activity (Andreazza et al., 2010; Maurer et al., 2001; Whatley et al., 1996). The reliability of measuring mitochondrial enzyme activity in postmortem brain specimen is questionable and can lead to inconsistent findings. Thus, it has been shown that OXPHOS enzymes' activity is particularly sensitive to postmortem delay (Mizuno et al., 1990; Prince et al., 1998). In addition, assay accuracy in whole tissue compared to isolated mitochondria is questionable as mitochondrial contribution to the measured activity is often difficult to distinguish from whole tissue activity. Finally, analyzing different brain regions can contribute to the controversial findings in OXPHOS complexes' activity. In isolated mitochondria from peripheral cells however, we and others observed consistent disease-state dependent alterations in complex I activity in schizophrenia. No change was observed in patients with bipolar disorder or major depression (Ben-Shachar et al., 1999; Gubert et al., 2013). These changes



were disease state dependent, as complex I activity was increased in patients with positive symptomatology, whether medicated or un-medicated, and reduced in patients with residual schizophrenia. A high and significant correlation was observed between complex I activity and PANSS scores of patients (Dror et al., 2002). Interestingly, cerebral glucose metabolism in regions implicated in schizophrenia showed significant correlation with platelet complex I activity in patients but not in healthy subjects, suggesting that the existence of such a brain/peripheral correlation is a pathological factor in the disorder (Ben-Shachar et al., 2007). 4.2. Complex I subunit expression Alterations in complex I activity may stem from changes in the expression of genes encoding for its subunits. Review of the literature of microarray studies of complex I genes in postmortem brain samples, mostly from areas in the frontal cortex, revealed both increases and decreases in the expression of 6 genes in schizophrenia (Scola et al., 2013). We argue that due to a low effect size and heterogeneity of patients, whole genome analyses may skip genes that may be revealed by candidate gene analyses. Indeed, we and others have shown impaired expression of complex I nuclear encoded labile subunits NDUFV1, NDUFV2 and NDUFS1, which form the first N module of complex I and are located at its site of interaction with dopamine (Fig. 1B) (Akarsu et al., 2014; Dror et al., 2002; Karry et al., 2004; Mehler-Wex et al., 2006; Taurines et al., 2010; Washizuka et al., 2009). Furthermore, protein levels of these genes were also abnormal in both different postmortem brain areas and somatic cells. Changes in these subunits have been also observed in brains of patients with bipolar disorders and major depression, however the neuroanatomical distribution of the impairment showed a disease specific pattern (Akarsu et al., 2015; Karry et al., 2004). Interestingly, nuclear encoded subunits of mitochondrial complex I including NDUFV1, NDUFV2 and NDUFS1, show high average expression in the CA3/CA1 hippocampal GABAergic parvalbumin interneurons, which have been implicated in cognitive and behavioral dysfunction in schizophrenia (Kann et al., 2011; Wirtz and Schuelke, 2011). Genome-wide association studies (GWAS) failed to identify genetic risk factors in these genes. To the best of our knowledge one study reported single nucleotide polymorphisms (SNPs) in 3′-UTR regions of NDUFV2 associated with schizophrenia (Washizuka et al., 2006). This sparse data may suggest that the altered expression of these genes is not due to a consistent difference in DNA sequence. A possible factor that can modulate gene expression is a transcription factor. We have previously shown that the ubiquitous transcription factor Sp1 is regulating the expression of NDUFV1, NDUFV2 and NDUFS1, and that its levels are impaired in four different brain areas and blood cells of schizophrenia patients. We further showed a parallel pattern of change in Sp1 and these genes in controls and in patients, while a highly significant correlation between them in healthy subjects, which was distorted in schizophrenia (Ben-Shachar and Karry, 2007). Others have also shown alterations in Sp1 and Sp4 in the frontal cortex, hippocampus and blood cells in schizophrenia (Fusté et al., 2013; Pinacho et al., 2014). Notably, dysregulation of Sp1 has been observed in additional neuropsychiatric disorders including autism, Alzheimer's and Huntington diseases (Citron et al., 2008; Dunah et al., 2002; Thanseem et al., 2012). These findings do not exclude post-transcriptional and posttranslational modifications of these genes, which are more intensively subjected to intracellular and external stimuli. 5. Mitochondrial dysfunction The hitherto described impairments in complex I driven cellular respiration, enzymatic activity in isolated mitochondria and expression of candidate genes in postmortem brain tissue and in cells, are at the range of 30% of change. One may wonder whether such a change with no consistent change in the activity of the other complexes of the

OXPHOS, could have functional manifestations at the level of mitochondrial function and thereby cellular oxygen consumption. This is of utmost importance as the OXPHOS regulates ROS production, calcium homeostasis, neuronal activity, and eventually, neuronal plasticity and connectivity, which can modulate neuronal survival and ultimately the organism behaviors. The first evidence for a change in mitochondrial function in schizophrenia was the observation of a reduced oxygen uptake in brain biopsies from schizophrenia patients (Takahashi, 1954). We have shown reduced cellular oxygen consumption (mitochondrial respiration) and its increased inhibition by dopamine, in schizophrenia derived lymphoblasts and keratinocytes compared with cells derived from healthy subjects. In schizophrenia hair follicle derived iPSCs basal respiration was normal, while its interaction with dopamine was pathological, similar to our findings in other cell types (Robicsek et al., 2013; Rosenfeld et al., 2011). Complex I driven respiration, but not that through complex II or III was impaired in schizophrenia lymphoblasts. In addition, we have shown a dissipation of mitochondrial membrane potential (ΔѰm) in these cells and also in glutamatergic and dopaminergic neurons differentiated from schizophrenia derived iPSCs. ΔѰm is the driving force for ATP production by complex V and is formed by protons, which are pumped out against the concentration gradient by complexes I, III and IV, during the flow of electrons through respiratory chain. Complex I contributes 40% of this driving force. ΔѰm is regulated by uncoupling proteins (UCPs), which are mitochondrial transporter proteins involved in proton conductance across inner mitochondrial membrane. Protons leak through UCPs into the mitochondria produce a motive force (Δp) which is dissipated as heat. Interestingly, two studies have reported a significant association between schizophrenia and UCP2 and UCP4, which have predominant expression in the brain (Mouaffak et al., 2011; Yasuno et al., 2007). An additional study showed reduced mRNA levels of UCP2, yet not of its protein in postmortem prefrontal cortex of patients with schizophrenia (Gigante et al., 2011). The interaction between complex I and UCPs in the modulation of ΔѰm is not known and warrants further studies under normal and impaired activity of complex I. An additional feature of the mitochondria is their ability to exist in two interconverting forms, extended filaments, networks, or clusters and spherical or ellipsoid isolated particles. In a network mitochondria are electrically united and can share their content including their respiratory chain complexes and DNA to facilitate energy delivery between different parts of the cell (Bakeeva et al., 1978, 1983; Gottlieb and Bernstein, 2016; Muster et al., 2010; Skulachev, 2001; Westermann, 2008). As isolated particles, mitochondria can be trafficked along microtubules in cells and even transfer between cells (Csordas, 2006; MacAskill et al., 2010). Mitochondrial structure dynamics is tightly connected to cellular metabolic demand and is regulated by two continuous processes fission and fusion. These two processes are also important for inheritance and maintenance of mtDNA and together with mitophagy, form a quality control pathway. The main proteins involved in mitochondrial network dynamics are fission 1 protein (Fis1) and mitochondrial fission factor (Mff), located on the outer mitochondrial membrane, which recruit the cytosolic factor dynamin-related protein 1 (Drp1) mediating fission, as well as mitofusins 1 and 2 (Mfn1 and 2), located on the outer mitochondrial membrane, and optic atrophy-1 (OPA1) located in the inner mitochondrial membrane, which mediate the fusion of mitochondria (Gottlieb and Bernstein, 2016; Shirihai et al., 2015; Westermann, 2008). In schizophrenia impairments in mitochondrial cellular distribution and connectivity have been observed in lymphoblasts, keratinocytes and dopaminergic neurons differentiated from iPSCs. As mitochondria were more aggregated in cells and less connected we suggest that aggregation is a compensation for reduced connectivity. Interestingly, in iPSCs and in differentiated glutamatergic cells, cellular distribution was abnormal but connectivity was unimpaired in line with the reduced severity of deficits in basal respiration and differentiation efficiency, respectively observed in these cells. Reduced connectivity was associated

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with reduced level of the fusion protein OPA1 in the frontal cortex, hippocampus and the different cell types mentioned above in schizophrenia (Engmann et al., 2011; Petit et al., 2012; Robicsek et al., 2013; Rosenfeld et al., 2011). Notably, it was reported that complex I inhibition leads to an oxidative-dependent disruption of OPA1 oligomeric complexes (Ramonet et al., 2012). Taken together, the accumulating data suggest that mitochondrial dysfunction in schizophrenia stems, at least in part from abnormalities in complex I. Having a major role in the regulation of the OXPHOS activity it is not surprising that a mild impairment in complex I can be responsible for different aspects of mitochondrial activities. 6. Mitochondrial dysfunction and neuronal function By the virtue of being the main source of ATP, having an essential role in the regulation of cellular calcium homeostasis and ROS production, mitochondria are fundamental for neuronal activity and plasticity. For their activity, neurons have to maintain the intracellular ion concentration against the concentration gradient, via Na+/K+- and Ca2 +ATPase pumps situated in the plasma and the endoplasmic reticulum membranes, a process that require a high-energy supply. Indeed, it has been shown that mitochondria are transported along axons and recruited to high-activity zones in neurons (Saxton and Hollenbeck, 2012). Thus, mitochondria are recruited to axon terminals during neurotransmitter release in response to an increase in synaptic activity, while their loss results in defective neurotransmitter release (Brodin et al., 1999; Ly and Verstreken, 2006). An example for the role mitochondria play in neuronal function comes from studies of GABAergic parvalbumin interneurons, which have been shown to have a pathogenic role in schizophrenia (Nakazawa et al., 2012). It has been shown that mitochondria dysfunction affects the fast spiking properties of these neurons, which generate synchronous oscillatory output, mainly gamma oscillation that are associated with extremely high utilization of mitochondrial oxidative capacity, are highly dependent on energy, regulate neuronal circuits in the cortex and are essential for the balance of excitation and inhibition (Inan et al., 2016; Kann et al., 2014; Whittaker et al., 2011). An interference with these neurons in the frontal cortex of animals as well as dysfunction of their mitochondria has been shown to be associated with behavioral abnormalities that are hallmarks of schizophrenia and autism spectrum disorder such as abnormal sociability, sensory gating and cognitive functions including working memory (Inan et al., 2016; Kann, 2016). Interestingly, it was shown that gamma oscillations are highly dependent on complex I activity and can be completely blocked by complex I inhibitor, rotenone in rat CA3 slice cultures (Kann et al., 2011). Mitochondria have been also shown to accumulate in the vicinity of active growth cones of developing neurons and to have an essential role in neuronal development and sprouting. In line with the latter, a link between presynaptic mitochondrial immobilization and axon branching has been observed (Courchet et al., 2013). Mitochondrial dynamics (fission fusion and mitophagy), which has been shown to influence complex signaling pathways and gene expression, also contributes to delineate cell differentiation, for reviews see Kasahara and Scorrano (2014) and Mattson et al. (2008). Finally, it was demonstrated that reduced content of mitochondria in dendrites leads to loss of synapse and plasticity, whereas increasing dendritic mitochondrial content or activity enhances the number and plasticity of spines and synapses (Li et al., 2004). Based on epidemiological, clinical, brain-imaging and genetic studies, schizophrenia is currently conceptualized as a neurodevelopmental disorder, with impaired neuronal plasticity and connectivity (Balu and Coyle, 2011; Bhandari et al., 2016; Rapoport et al., 2012; Stephan et al., 2006). Schizophrenia derived iPSC and their differentiation into neurons provide us the opportunity to study the relationship between mitochondrial function and neuronal differentiation in the disease. We and others have shown that iPSCs derived from schizophrenia patients

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show impaired efficiency to differentiate into dopaminergic and glutamatergic neurons associated with impaired neurotransmitter release, reduced ability to form synapse or neuronal contacts, and abnormal expression of genes implicated in this disease (Brennand et al., 2011; Hook et al., 2014; Pedrosa et al., 2011; Robicsek et al., 2013; Wen et al., 2014). In our hands, dopaminergic neurons almost failed to differentiate and most cells remained in their progenitor stage, while glutamatergic neurons failed to maturate. The deficits in neuronal differentiation of schizophrenia derived cells were associated with mitochondrial dysfunction, expressed by impaired cellular oxygen consumption in iPSCs, abnormal ΔѰm, mitochondrial dynamics and aberrant expression of complex I subunits NDUFV1, NDUFV2, and NDUFS1 and of the fission protein OPA1 along all stages of differentiation (Robicsek et al., 2013). These studies suggest that there is a link between dysfunctional mitochondria and neuronal differentiation in schizophrenia. Thus, one may extrapolate from mitochondrial role in neuronal plasticity and connectivity and reason that mitochondrial deficits are involved in brain processes leading to the abnormal cognition and behaviors observed in the disease. 7. Conclusion The accumulating evidence reviewed above, together with morphological, genetic and imaging data that have not been addressed in this article, delineates mitochondrial multifaceted dysfunction as a pathological factor in schizophrenia. Mitochondria have been implicated in additional mental and general disorders, since beyond being the “power house” of the cell, they are involved in the regulation of many essential cellular and physiological functions including transcription, brain function, inflammation and immune function (Fig. 2). However, a review of the literature regarding schizophrenia, shows that a disease specific profile can be identified as long as an array of functional parameters of the mitochondria is scanned and performed in various tissues. We have discussed in this article several such factors including complex I activity, cellular oxygen consumption, mitochondrial membrane potential and their vulnerability to dopamine, parameters of mitochondrial network dynamics and the expression of proteins and genes involved in mitochondrial dysfunction. Using various parameters related to mitochondrial function may help to overcome the controversial data observed upon using a single parameter, due to intrinsic heterogeneity of patients, tissue samples or methodology. Establishing a disease specific profile may turn mitochondria or rather an array of interrelated mitochondrial factors into a diagnostic, treatment efficacy and follow-up biomarker for the disease. Advanced knowledge of mitochondrial various functions and their disease related failure could unravel new pathological processes in schizophrenia and open up new opportunities for therapy. Mitochondria are intertwined with the cellular network, being remodeled by and remodeling various signaling pathways, transcription and structural effectors. In addition, mitochondria are a two-edge sword involved in cellular processes leading either to cell death or supporting its survival and sprouting. This multitasking of the mitochondria turns them into an intricate and challenging target for interference, similar to the complexity faced with cortisol aimed therapies in major depression. Still, focusing on a specific target in the mitochondria, such as complex I or his subunits, may lead to the delineation of a novel target, which if transiently modulated, may be of benefit for patients with schizophrenia. Conflict of interest The author has no conflict of interest. Role of funding source No funding source has funded this review. Contributors Dorit Ben-Shachar wrote this review paper.



Fig. 2. Mitochondrial multifaceted dysfunction and its consequential pathologies in schizophrenia.

Acknowledgment None.

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