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Schizophrenia: a Case of Faulty Redox Detox?

24 October 2007. A research collaboration between Swiss and Danish researchers provides new genetic and biochemical evidence that neural damage from oxidative stress may play a role in the pathogenesis of schizophrenia. In particular, the new study, published in the October 16 issue of PNAS, implicates a trinucleotide repeat (TNR) polymorphism in a gene that is crucial for the synthesis of the antioxidant peptide glutathione, which plays a protective role in the brain by scavenging for reactive oxygen species and neutralizing them.

Reactive oxygen species are important for cell signaling (so-called redox signaling) and the immune response, but they are cytotoxic, and adequate synthesis of glutathione is essential for maintaining a balanced intracellular reducing environment that keeps destructive peroxides and free radicals in check. Gene mutations that impair glutathione synthesis have been associated with a number of serious diseases and pathologies, including Parkinson’s disease, Alzheimer’s disease, atherosclerosis, myocardial infarction, and mental retardation. Based on several previous findings of low glutathione levels in cerebrospinal fluid and postmortem tissue from patients with schizophrenia (e.g., Do et al., 2000), Kim Q. Do and colleagues at the University of Lausanne have proposed a “redox dysregulation” hypothesis of the disorder.

Anatomy of a scavenger
Glutathione is synthesized in two consecutive enzymatic reactions, the first catalyzed by glutamate cysteine ligase (GCL) and the second by glutathione synthetase (GSS). GCL, the rate-limiting enzyme in glutathione synthesis, has catalytic and modulatory subunits. A 2006 study from the Lausanne group had found that expression of both GSS and the modulatory subunit of GCL was reduced in cultured skin fibroblasts derived from schizophrenia patients, and reported an association between schizophrenia and certain alleles of the gene for GCL’s modulatory subunit.

In the new work, first author René Gysin and colleagues joined forces with Thomas Werge at Copenhagen University Hospital to more directly address the question of whether glutathione synthesis is compromised in schizophrenia.

To do so, the researchers again cultured fibroblasts obtained by skin biopsy from Swiss and Danish patients with schizophrenia and healthy controls, as assessed by Diagnostic Interview for Genetic Studies (DIGS) and DSM-IV criteria. The fibroblasts were treated with tert-butylhydroquinone (TBHQ), a phenolic compound known to induce the expression of phase 2 antioxidant genes—including those for GCL’s two subunits—and thereby increasing glutathione synthesis.

In the Swiss sample, GCL activity after TBHQ treatment was 26 percent lower in patients with schizophrenia than in controls. Protein expression of the GCL catalytic subunit (but not the modulatory subunit) was also lower in patients, by 22 percent.

A repeated problem
The GCL catalytic subunit gene, on chromosome 6p12, contains a TNR polymorphism with seven, eight, or nine guanine-adenine-guanine (GAG) repeats (Walsh et al., 2001). In both the Swiss and Danish samples, there was a marked difference in the distribution of these alleles between patients and controls. In general, the 8 and 9 alleles were far more common among the patients, whereas the 7 allele was found more often in controls. In the Danish sample, the 8/8 genotype was three times more common in patients versus controls, but the 7/7 genotype, common in controls, appeared to exert a protective effect.

Based on these findings, the researchers hypothesized that the higher number of GAG repeats in patients compromised glutathione synthesis. To explore this question, they divided the Swiss sample into “low-risk” (7/7 and 7/9 genotypes) and “high-risk” (7/8, 8/8, 8/9, and 9/9 genotypes) groups. After TBHQ treatment of fibroblasts from each group, the authors found significantly lower GCL activity, catalytic subunit expression and overall glutathione content in the high-risk group.

GAGs with serious consequences
Gysin and colleagues propose that reduced glutathione synthesis caused by the GAG TNR polymorphism in the gene for GCL’s catalytic subunit may conspire with risk factors associated with both oxidative stress and schizophrenia, causing aberrant synapse development and the perceptual, cognitive, and behavioral symptoms that characterize the schizophrenia phenotype. For example, obstetrical complications, inflammation, and viral infections, all associated with schizophrenia, are also known to cause oxidative stress. In addition, psychological stress can cause oxidative stress via the hypothalamic-pituitary-adrenal axis in the dopamine-rich brain areas affected in schizophrenia.

Summing up, the authors write that the new findings “provide evidence for a genetic source of the redox dysregulation in schizophrenia,” and add that the GAG TNR polymorphism “may serve as a marker to identify individuals at risk [and to] gather a complete picture of genetic risk factors of schizophrenia in the [glutathione] and oxidative stress associated pathways.”—Peter Farley.

Gysin R, Kraftsik R, Sandell J, Bovet P, Chappuis C, Conus P, Deppen P, Preisig M, Ruiz V, Steullet P, Tosic M, Werge T, Cuénod M, Do KQ. Impaired glutathione synthesis in schizophrenia: Convergent genetic and functional evidence. Proc Natl Acad Sci USA. 2007 Oct 16;104(42):16621-6. Abstract

Do KQ, Trabesinger AH, Kirsten-Krüger M, Lauer CJ, Dydak U, Hell D, Holsboer F, Boesiger P, Cuénod M. Schizophrenia: glutathione deficit in cerebrospinal fluid and prefrontal cortex in vivo. Eur J Neurosci. 2000 Oct;12(10):3721-8. Abstract

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Comment by:  Richard Deth
Submitted 25 November 2007
Posted 28 November 2007
  I recommend the Primary Papers

Identification of a limitation in the capacity for glutathione (GSH) synthesis by Gysin and colleagues raises several questions: "How could a redox problem (i.e., oxidative stress) lead to schizophrenia?” and "Does this finding mesh with other hypotheses?"

All cells must maintain sufficient levels of GSH to survive collateral damage from oxidative metabolism, and a number of adaptive mechanisms have evolved to meet this need. One important example is inhibition of the enzyme methionine synthase by oxidative stress. The higher the oxidative stress level, the greater the inhibition of its methylation of homocysteine to methionine, allowing the accumulating homocysteine to be diverted to GSH synthesis via the trans-sulfuration pathway. Homocysteine levels are elevated in schizophrenia, especially, but not exclusively, in first-episode males (Regland et al., 1995; Haidemenos et al., 2007), implying that methionine synthase is inhibited, perhaps by oxidative stress. Importantly, methionine synthase activity is also critical for dopamine-stimulated methylation of membrane phospholipids, a unique activity of the D4 dopamine receptor discovered by our lab in 1999 (Sharma et al., 1999). Thus, oxidative stress caused by impaired GSH synthesis will adversely affect this dopaminergic mechanism.

The physiological role of D4 receptor-mediated phospholipid methylation remains to be fully elucidated, but studies indicate a central role in attention and synchronization of neural networks, both of which are impaired in schizophrenia. The seven-repeat variant of the D4 receptor is considered to be the most important genetic risk factor for ADHD (Swanson et al., 2007), and is also associated with lower IQ, alone or in combination with dopamine transporter variants (Mill et al., 2006). MEG studies in subjects without ADHD showed stronger γ synchronized oscillatory activity during attention in subjects possessing the seven-repeat allele (Demiralp et al., 2007). We recently described a molecular mechanism by which dopamine-stimulated phospholipid methylation could tune neural networks to γ frequency during attention (Kuznetsova et al., 2007), and a restriction in GSH synthesis, as described by Gysin and colleagues, could contribute to impairments of synchronization and attention in schizophrenia.

Oxidative stress-induced inhibition of methionine synthesis also causes accumulation of S-adenosylhomocysteine, a general inhibitor of methylation reactions, affecting more than 150 cellular methylation reactions. Lower COMT activity would augment dopamine levels, while lower activity of DNA and histone methyltransferases would alter epigenetic regulation of gene expression. Thus, a putative role for oxidative stress can be integrated with other proposed theories. Indeed, methylation defects in schizophrenia have been recognized for more than 40 years (Spiro et al., 1965), a hypothesis advanced by Seymour Kety (Kety, 1972), including the replicable finding that methionine administration provokes acute psychosis in schizophrenia subjects but is without effect in normal individuals (Cohen et al., 1974). Despite these early clues, defective sulfur metabolism has received only limited attention. Perhaps the illuminating findings of Gysin and colleagues will reinvigorate interest and encourage schizophrenia researchers to invest the time needed to understand and appreciate this important area of biochemistry.


Regland B, Johansson BV, Grenfeldt B, Hjelmgren LT, Medhus M. Homocysteinemia is a common feature of schizophrenia. J Neural Transm Gen Sect. 1995;100(2):165-9. Abstract

Haidemenos A, Kontis D, Gazi A, Kallai E, Allin M, Lucia B. Plasma homocysteine, folate and B12 in chronic schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry. 2007 Aug 15;31(6):1289-96. Abstract

Sharma A, Kramer ML, Wick PF, Liu D, Chari S, Shim S, Tan W, Ouellette D, Nagata M, DuRand CJ, Kotb M, Deth RC. D4 dopamine receptor-mediated phospholipid methylation and its implications for mental illnesses such as schizophrenia. Mol Psychiatry. 1999 May;4(3):235-46. Abstract

Swanson JM, Kinsbourne M, Nigg J, Lanphear B, Stefanatos GA, Volkow N, Taylor E, Casey BJ, Castellanos FX, Wadhwa PD. Etiologic subtypes of attention-deficit/hyperactivity disorder: brain imaging, molecular genetic and environmental factors and the dopamine hypothesis. Neuropsychol Rev. 2007 Mar;17(1):39-59. Abstract

Mill J, Caspi A, Williams BS, Craig I, Taylor A, Polo-Tomas M, Berridge CW, Poulton R, Moffitt TE. Prediction of heterogeneity in intelligence and adult prognosis by genetic polymorphisms in the dopamine system among children with attention-deficit/hyperactivity disorder: evidence from 2 birth cohorts. Arch Gen Psychiatry. 2006 Apr;63(4):462-9. Abstract

Demiralp T, Herrmann CS, Erdal ME, Ergenoglu T, Keskin YH, Ergen M, Beydagi H. DRD4 and DAT1 polymorphisms modulate human gamma band responses. Cereb Cortex. 2007 May;17(5):1007-19. Abstract

Kuznetsova AY, Deth RC. A model for modulation of neuronal synchronization by D4 dopamine receptor-mediated phospholipid methylation. J Comput Neurosci. 2007 Oct 11; [Epub ahead of print] Abstract

Spiro HR, Schimke RN, Welch JP. Schizophrenia in a patient with a defect in methionine metabolism. J Nerv Ment Dis. 1965 Sep;141(3):285-90. Abstract

Kety SS. Toward hypotheses for a biochemical component in the vulnerability to schizophrenia. Semin Psychiatry. 1972 Aug;4(3):233-8. Abstract

Cohen SM, Nichols A, Wyatt R, Pollin W. The administration of methionine to chronic schizophrenic patients: a review of ten studies. Biol Psychiatry. 1974 Apr;8(2):209-25. Abstract

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Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  John Krystal
Submitted 6 December 2007
Posted 9 December 2007

The paper by Behrens and colleagues provides exciting new data to suggest that NADPH oxidase plays an important role in the impact of the NMDA receptor antagonist, ketamine, upon parvalbumin-containing (PVC) fast-spiking GABA interneurons. The authors show that ketamine causes an activation of NADPH oxidase, resulting in increases in superoxide production. The elevation in free radicals, presumably toxic to these neurons, is associated with reduction in the expression of parvalbumin and GAD67. These effects of ketamine could be prevented by inhibition of NADPH oxidase.

These data were interpreted by the authors to help explain the schizophrenia-like effects of ketamine in healthy humans. I think that these data provide important insights into the impact of reductions in NMDA receptor function, and they may be relevant to schizophrenia. First, the data amplify the implications of the work of Kinney, Cunningham, and others who have shown that PVC interneurons express the NR2A subunit of the NMDA receptor and that deficits in NMDA receptor function may contribute to reduced GAD expression by these neurons. Since PVC deficits in GAD expression have been described in postmortem cortical tissue from people diagnosed with schizophrenia, the current data suggest that some of these findings may be attributable to activation of NADPH oxidase. It would be interesting to know whether there is an interaction between this consequence of deficits in NMDA receptor function, a feature associated with schizophrenia, and reductions in the cortical levels of glutathione, also associated with this disorder. Glutathione is a free radical scavenger. In other words, the emergence of GABA neuronal deficits may be an unfortunate consequence of the convergence of a disturbance in glutamatergic neurotransmission and a heritable abnormality in neural metabolism. These data highlight the potential importance of some very preliminary new data that suggest that N-acetyl-cysteine (NAC) may augment antipsychotic effects in treating schizophrenia. NAC raises intracellular glutathione and might be a treatment that targets the cellular process described by Behrens and colleagues.

The Behrens paper also highlights the importance of research studies exploring ketamine effects from a systems and cognitive neuroscience perspective. For example, it does not explain why ketamine effects produce symptoms and cognitive impairments associated with schizophrenia. It is likely that the work of scientists including H. Grunze, R. Greene, B. Moghaddam, R. Dingeldine, M. Cunningham, and others is important to consider. These investigators have shown that NMDA receptor antagonists reduce the recruitment of PVC interneurons in feed-forward inhibition pathways resulting in increased glutamatergic output. When NMDA receptors are blocked, the activity of these neurons produces dysfunctional effects, in that neural activity seems chaotic and the organized oscillatory activity of networks is disrupted. These disturbances in network function are paralleled by abnormal behaviors and cognitive impairments in animals and "schizophrenia-like" symptoms and cognitive deficits in humans. One potential solution to this problem would be to reduce glutamate release, a paradoxical suggestion for a disorder commonly thought of as "hypoglutamatergic" based on loss of cortical connectivity. Yet, in animals and humans, drugs that reduce glutamate release (lamotrigine, group II metabotropic glutamate receptor agonists) reduce the physiologic and behavioral effects of NMDA glutamate receptor antagonists. Further, there are now some encouraging clinical data that lamotrigine and, particularly, group II mGluR agonists, might have clinical efficacy in treating schizophrenia.

Overall, we seem to be working in a period where a wide variety of data from many sources is rapidly converging to capitalize on the insight that NMDA receptor antagonists, when administered to healthy people, transiently produce effects that resemble schizophrenia.

View all comments by John Krystal

Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  Steven Siegel (Disclosure)
Submitted 6 December 2007
Posted 9 December 2007

The article by Behrens and colleagues provides evidence for a mechanistic link between NADPH oxidase and disruption of normal protein expression in some interneurons following the drug ketamine. Data presented demonstrate that addition of an NADPH oxidase inhibitor, given in the animal’s drinking water, blocked the effects of ketamine on a specific class of interneurons that contains parvalbumin. Several lines of research suggest that this population of cells is disrupted in schizophrenia, and that reductions of NMDA-type glutamate receptor activity may lead to that impairment. The important iterative advance in the current study links the reduction in NMDA receptor-mediated glutamate transmission to a specific intracellular mechanism and molecular pathway. Furthermore, the authors demonstrate that they can effectively block the cellular changes by inhibiting that pathway, suggesting a novel therapeutic target.

This leads to two major questions: 1) Could NADPH oxidase inhibitors, or similar mechanisms be used to avert the onset of schizophrenia if administered during a prodromal period? 2) Is the process of reduced parvalbumin expression reversible? Some studies have shown that drugs like ketamine, which reduce activity at NMDA receptors, actually lead to cell death, suggesting that only prevention would be possible. Alternatively, there is evidence that the parvalbumin-containing cells in schizophrenia may not be dead and gone, but rather have impaired function and loss of this particular protein. In this latter scenario, it is possible that the effects of the illness could be reversible. Given that ketamine also causes a variety of functional changes in animals, including electrical brain activity and behavior, the current work lays the groundwork for future studies to determine if co-administration of NADPH oxidase inhibitors can block the functional consequences of ketamine and, by extension, reduce NMDA receptor activity in general.

View all comments by Steven Siegel

Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  Dan Javitt, SRF Advisor
Submitted 7 December 2007
Posted 10 December 2007

The study by Behrens and colleagues is an excellent illustration of how breaking with traditional paradigms can lead to identification of novel potential targets for intervention in schizophrenia. As detailed on the pages of Schizophrenia Research Forum (e.g., Interview with D. Lewis) and the cited articles from F. Benes, one of the most consistent findings in schizophrenia is the downregulation of PV and GAD67 expression in PV+ GABAergic interneurons. Dysfunction of these neurons, in turn, may be responsible for frontal neurocognitive and dopaminergic deficits. The underlying cause of the GABAergic interneuron changes, however, has only intermittently been investigated.

One of the leading potential mechanisms underlying reduced PV and GAD67 expression in brain in schizophrenia has always been NMDA dysfunction, given the strong expression of NMDA receptors on GABA interneurons, as described by Behrens and colleagues, and the well-known ability of NMDA antagonists to induce both symptoms and neurocognitive deficits closely resembling those of schizophrenia. Last year, Kinney and colleagues demonstrated that exposure to the NMDA antagonist ketamine reduced PV and GAD67 expression in GABAergic interneurons in vitro (Kinney et al., 2006). The present study builds upon this finding and demonstrates a similar phenomenon in vivo. Moreover, it builds upon this finding to demonstrate that these changes can be reversed by antagonists of NADPH oxidase, suggesting a potential target for intervention.

This study thus adds reduced GAD67 and PV expression in PV+ GABAergic interneurons to the long list of findings in schizophrenia that can be viewed as “downstream” of a more proximal deficit in NMDA-mediated neurotransmission, and supports interventions aimed specifically at frontal GABAergic interneurons, as well as more generally at reduced NMDA activity throughout brain. This preparation, moreover, may be appropriate to the testing of novel glutamatergic agents.

Behrens and colleagues' article, however, also leaves many questions unanswered. For example, loss of PV and GAD67 in schizophrenia is not confined to prefrontal cortex. It would be of interest to know, therefore, whether histological changes induced by ketamine are or are not confined to this region. As with all proposed new drug targets, it will also be important to know what other processes NADPH oxidase is involved with both inside and outside brain before proposing it too seriously as a drug target. It is one thing to reverse a specific deficit in a short-term treatment model, another to contemplate long-term treatment. At first glance, NADPH oxidase would seem to be a very general enzyme, which is being targeted to treat a very specific condition. Nevertheless, if NADPH oxidase activity can safely be blocked throughout the body long term, the present findings may point the way for new treatments for schizophrenia.

View all comments by Dan Javitt

Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  Julie MarkhamJames I. Koenig
Submitted 10 December 2007
Posted 10 December 2007

The role of reactive oxygen species in the pathogenesis of schizophrenia is currently unclear. Several lines of evidence support a greater production of these reactive molecules in schizophrenia because of reduced levels of important buffers for superoxides, such as glutathione. Other research, however, suggests that antipsychotic drugs themselves increase the production of oxygen radicals. In this week’s issue of Science, Behrens and colleagues present data supporting the involvement of reactive oxygen species in the pathophysiology of schizophrenia. The authors have previously shown that administration of an NMDA receptor antagonist to primary cultures of cortical neurons results in the loss of GAD67 and parvalbumin (PV; a calcium-binding protein) from PV positive GABAergic interneurons (Kinney et al., 2006), similar to what has been observed in studies using postmortem tissue from patients with schizophrenia (e.g., Volk et al., 2000; Hashimoto et al., 2003). In this study, administration of the NMDA receptor antagonist ketamine was found to increase production of reactive molecules both in vitro (following bath application of the drug to cultured neurons) and in vivo (following two injections of the drug to mice). Moreover, inhibition of the enzyme NADPH oxidase prevented the reduction of both PV and GAD67 expression. The authors suggest that inhibition of NADPH oxidase may represent a novel treatment for both ketamine-induced psychosis and schizophrenia.

While the authors’ findings are undoubtedly exciting, some limitations of their approach need to be addressed before over-enthusiasm regarding NADPH oxidase inhibition as a treatment for schizophrenia is generated. Although the title advertises a “loss of phenotype of fast-spiking interneurons,” the reduction in PV and GAD67 expression from neurons that remain PV positive does not represent a loss of phenotype, and the ketamine-induced increase in superoxide production was not specific to interneurons (only 5-10 percent of primary cortical neuron cultures are PV positive, yet the effect was observed throughout sampled cells). Also, although their findings bear similarity to those observed in schizophrenia, there are notable differences. For instance, whereas the level of PV expression per cell is reduced in schizophrenia (Hashimoto et al., 2003), the level of GAD67 mRNA expression per cell does not differ between individuals with schizophrenia and controls; rather, it appears to be a reduction in the density of neurons that express the transcript. In contrast, Behrens and colleagues report a reduction in the expression per cell for both PV and GAD67. While this difference may simply be due to the fact that Behrens and colleagues examined levels of the proteins, the potential discrepancy should be recognized.

Perhaps the most important limitation to the work is the absence of a functional measure to determine whether the reduction in PV and GAD67 in cortical interneurons observed following ketamine administration results in any of the schizophrenia-associated endophenotypes which can be modeled in rodents. Animal models of schizophrenia employing developmental strategies have been very successful in this regard (reviewed in Carpenter and Koenig, in press), and it is unclear how functional outcomes from the acute pharmacological challenge in mature animals used in the present study might compare. Although the data as they stand are promising, they would be much more compelling if a functional deficit as a result of the treatment was observed and the authors could demonstrate that inhibition of NADPH oxidase prevented this deficit. Unfortunately, such a deficit is unlikely to be found following such a limited ketamine exposure. This is actually quite fortunate since ketamine is a popular general anesthetic in both human and veterinary medicine. Additionally, countless biomedical investigators routinely use ketamine as an anesthetic for survival surgeries; even in cases where the experimental design calls for multiple anesthetizations over the course of the study, no major functional disturbances in experimental animals have been reported. Our conclusion is that, while exposure to ketamine may induce features of neuropathology that bear some similarity to those observed in schizophrenia, the excitement about a treatment for ketamine-induced superoxide production should be tempered until it can be demonstrated that the treatment reverses a functional deficit that is relevant to schizophrenia.


Carpenter WT, Koenig JI. The evolution of drug development in schizophrenia: past issues and future opportunities. Neuropsychopharmacology. (In press, 2007)

Hashimoto T, Volk DW, Eggan SM, Mirnics K, Pierri JN, Sun Z, Sampson AR, Lewis DA. Gene expression deficits in a subclass of GABA neurons in the prefrontal cortex of subjects with schizophrenia. J Neurosci. 2003 Jul 16;23(15):6315-26. Abstract

Kinney JW, Davis CN, Tabarean I, Conti B, Bartfai T, Behrens MM. A specific role for NR2A-containing NMDA receptors in the maintenance of parvalbumin and GAD67 immunoreactivity in cultured interneurons. J Neurosci. 2006 Feb 1;26(5):1604-15. Abstract

Volk DW, Austin MC, Pierri JN, Sampson AR, Lewis DA. Decreased glutamic acid decarboxylase67 messenger RNA expression in a subset of prefrontal cortical gamma-aminobutyric acid neurons in subjects with schizophrenia. Arch Gen Psychiatry. 2000 Mar;57(3):237-45. Abstract

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Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  Gavin Reynolds
Submitted 10 December 2007
Posted 10 December 2007

For two decades, following the work by Benes and her colleagues, it has been increasingly apparent that there is a deficit in cortical GABAergic neurons in schizophrenia. Ten years ago we found that the parvalbumin (PV)-containing, but not calretinin-containing, subgroup of these neurons was selectively affected, and recently this specific deficit has been seen in animal models of the disease. Repeated administration of non-competitive NMDA receptor antagonists such as PCP, MK801, and ketamine can induce in rats some behaviors reminiscent of schizophrenia, as well as enduring deficits in PV expression.

Behrens and colleagues have identified some of the molecular mechanisms underlying this specific neurotoxicity of ketamine and, probably, other NMDA antagonists. That the effects of ketamine involve generation of reactive oxygen species (ROS) is not surprising, given the ubiquity of oxidative free radical production in neurotoxic processes. However, identifying the role of NADPH oxidase in producing ROS in response to ketamine, and demonstrating that this process determines the consequent toxic effects of ketamine on PV-containing and other neurons, are potentially important developments.

The importance of these findings to schizophrenia relies on the assumption that repeated administration of ketamine and, presumably, other NMDA antagonists not only models (some of) the pathophysiology of schizophrenia, it also mimics the process leading to this neuronal pathology. This is far from proven, although the NMDA receptor hypofunction hypothesis of Olney and Farber provides a useful model mechanism for this pathogenesis.

A useful proof of concept would be to move away from pharmacological approaches to other animal models of the disease. One such is the isolation rearing paradigm; in this model, induction of abnormal “schizophrenia-like” behaviors is also paralleled by a deficit in PV-containing neurons (Harte et al., 2007). A simple but very informative study here would be to determine whether inhibition of NADPH oxidase might protect against the development of these deficits. Of course, how the NMDA receptor-mediated deficits relate temporally to the natural history of schizophrenia is unclear; we do not know when the PV deficits occur in schizophrenia. There may be some hope for targeted treatment with, e.g., NADPH oxidase inhibitors if the neuronal pathology parallels a neurotoxic process that underlies the progressive cognitive disturbances as implied by Olney and Farber, but not if the PV deficits relate to an early and primary pathology of the disease.


Harte M, Powell S, Swerdlow N, Geyer M, Reynolds GP (2007) Deficits in Parvalbumin and Calbindin Immunoreactive cells in the Hippocampus of Isolation Reared Rats. J Neural Transm 114, 893-898. Abstract

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Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  Kenneth Johnson
Submitted 18 December 2007
Posted 18 December 2007

The recent study by Behrens and colleagues provides in vitro evidence that blockade of NMDA receptors by ketamine leads to a selective reduction in PV and GAD67 that appears to be due to the toxic effects of superoxide anion arising subsequent to the activation of NADPH oxidase. Blockade of the sublethal, toxic effects of ketamine in neuronal culture is consistent with our report demonstrating that the apoptotic effect of phencyclidine (PCP) on cortical neurons in vivo also could be prevented by the superoxide dismutase mimetic, M40403 (Wang et al., 2003). Though seemingly non-specific, superoxide dismutase mimetics may prove to be useful in the treatment of ketamine or PCP-induced psychosis because of the relative sparseness of critical life-promoting processes that require superoxide anion. Perhaps more importantly, a better understanding of the mechanisms underlying ketamine-induced loss of PV/GAD67 may lead to novel treatment modalities for schizophrenia.

While the primary focus of the report by Behrens and colleagues is on PV-expressing GABAergic interneurons, Fig. 1 clearly demonstrates that ketamine also affects a large population of non-PV neurons. This is consistent with our recent in vivo experiments in developing rats demonstrating that PCP administration on PN7 induces apoptosis of cortical PV-containing interneurons as well as principal neurons in layers II-IV of the cortex (Wang et al., 2007). Early postnatal administration of PCP also results in neuronal apoptosis in the hippocampus, striatum, and thalamus (Wang and Johnson, 2007. Thus, it may be premature to focus solely on this population of interneurons.

In thinking about the mechanism underlying the selective loss of PV interneurons following PCP, it is important to note that PV is not yet expressed on PN7, which is when PCP was administered in our paradigm (Wang et al., 2007). (The loss of PV-containing interneurons was measured at PN56, well after the time of PV expression on about PN10.) Interestingly, interneurons expressing calretinin and calbindin at the time of PCP administration were spared. These neurons showed no colocalization with cellular markers of apoptosis (terminal dUTP nick-end labeling [TUNEL] of broken DNA or cleaved caspase-3), indicating that calretinin- and calbindin-containing neurons were protected from the toxic effect of PCP and survived into adulthood (Wang et al., 2007). The mechanism underlying this selectivity for cortical PV-containing interneurons is unknown, but as Behrens and colleagues suggest, it could be because these neurons are dependent on a relatively large glutamatergic input for survival. It is also possible that the differing calcium buffering capacities of these interneurons play a role in the selective neurotoxic effect of NMDA receptor blockade. That is, since calcium binding proteins could also act to buffer decreases in intracellular Ca2+ levels caused by ketamine-induced blockade of NMDA receptors, it is possible that the lack of PV in these vulnerable interneurons reduces the ability of these cells to adequately buffer the ketamine-induced decrease in intracellular calcium. This is consistent with the lack of effect on other interneurons that express the calcium binding proteins calretinin and calbindin at the time of PCP administration. This suggests NMDA receptor blockade could cause the deletion of PV neurons because of a specific effect at a critical stage of development. However, cleaved caspase-3 (a hallmark of apoptosis) showed no colocalization with BrdU, a specific marker of S-phase proliferation (Wang et al., 2007). These data suggest that the loss of PV-containing neurons in this paradigm was not due to an effect of PCP on proliferating neurons, but rather an effect on postmitotic neurons.

We have reported recently that PCP in cortical neuronal culture causes neuronal apoptosis by interfering with the Akt-GSK-3β cascade, which is necessary for neuronal survival during development (Lei et al., 2007). Moreover, increasing synaptic strength by various means such as increasing calcium current via activation of L-type calcium channels completely blocks PCP-induced cell death by increasing Akt phosphorylation. It would be of great interest to determine whether PV-containing interneurons respond in a similar fashion.

In order to fully appreciate the significance of ketamine-induced loss of PV-containing neurons, it will be necessary to carefully compare the in vivo dose-related effects of ketamine or PCP that are truly selective for PV/GAD67-containing interneurons to those cortically mediated behaviors that have relevance to schizophrenia.


Wang C, McInnis J, West JB, Bao J, Anastasio N, Guidry JA, Ye Y, Salvemini D, Johnson KM. Blockade of phencyclidine-induced cortical apoptosis and deficits in prepulse inhibition by M40403, a superoxide dismutase mimetic. J Pharmacol Exp Ther. 2003 Jan 1;304(1):266-71. Abstract

Wang, C.Z., Yang, S.F., Xia, Y. and Johnson, K.M. Induction of a selective cortical deficit of parvalbumin-containing interneurons by phencyclidine administration during postnatal brain development. Neuropsychopharmacology (In press, 2007).

Wang CZ, Johnson KM. The role of caspase-3 activation in phencyclidine-induced neuronal death in postnatal rats. Neuropsychopharmacology. 2007 May 1;32(5):1178-94. Abstract

Lei, G., Xia, Y. and Johnson, K.M. The role of Akt-GSK-3β signaling and synaptic strength in phencyclidine-induced neurodegeneration. Neuropsychopharmacology (In press, 2007).

View all comments by Kenneth Johnson

Related News: Does Oxidative Stress Link NMDA and GABA Hypotheses of Schizophrenia?

Comment by:  Patricia Estani
Submitted 11 January 2008
Posted 13 January 2008
  I recommend the Primary Papers