M.D Bruno Romeo
email: brunoromeo@hotmail.fr

M. S Amelie Petillion

M.D Catherine Martelli

M.D Amine Benyamina

Magnetic resonance spectroscopy studies in subjects with high risk for psychosis: a meta-analysis and review

Keywords: High risk subjects, brain metabolites levels, schizophrenia, magnetic resonance spectroscopy IO/K6AXH

Introduction: Even though anomalies on brain metabolites have been found in schizophrenia, researches about subjects with high risk (HR) show heterogeneous results. Thus, this metaanalysis aims to characterize the metabolic profile of HR subjects, first, compared to controls (HC) and then compared to people with schizophrenia.

Methods: After a systematic database search, means and standard deviations were extracted to calculate standardized mean differences (SMD). Cerebral metabolites levels were compared between HR subjects and HC or patients with schizophrenia in all regions of interest investigated in included studies. Meta-regressions were performed to explore the influence of demographic and clinical variables on metabolites level's SMDs.

Results:

Thirty-nine studies were included in this meta-analysis. A higher level of glutamine + glutamate (Glx) was found in the medial prefrontal cortex (mPFC) (p < 0.01) and potentially in the basal ganglia (p = 0,05) as well as a higher level of myo-inositol (mI) in the dorsolateral prefrontal cortex (DLPFC) (p = 0.04) in HR subjects compared to HC. A higher level of choline (Cho) was found in people with schizophrenia compared to HR subjects in the DLPFC (p < 0.001) and the medial temporal lobe (p = 0.02). Meta-regression analyses showed negative associations between SMD for Cho concentration, the percentage of females or the age (p = 0.01).

Conclusions:

The present meta-analysis provides evidence that some brain metabolites concentrations are disrupted before the transition to psychosis and could be considered like a vulnerability.

Introduction

Schizophrenia is a chronic and severe mental disorder affecting more than 23 million people worldwide [START_REF] Who | World Heath Organisation[END_REF] . One of the current research challenges on schizophrenia is to provide early interventions so the functional and social impacts can be limited [START_REF] Green | Impact of cognitive and social cognitive impairment on functional outcomes in patients with schizophrenia[END_REF].

To this end, many studies focused on subjects with a high risk of psychotic transition (HR) and the notion of "ultra high risk" (UHR) subject of psychotic transition has been defined.

However, not all patients identified as UHR are in a prodromal phase of schizophrenia. Indeed, among these UHR subjects, a psychotic transition will only occur in a minority (~ 1/3 within three years) [START_REF] Fusar-Poli | The Psychosis High-Risk State: A Comprehensive State-of-the-Art Review[END_REF]. Among patients who have experienced a psychotic transition, 73% have a diagnosis of schizophrenia spectrum disorder, 16% have a diagnosis of other psychosis and 11% have a diagnosis of mood disorders with psychosis [START_REF] Fusar-Poli | At risk for schizophrenic or affective psychoses? A meta-analysis of DSM/ICD diagnostic outcomes in individuals at high clinical risk[END_REF]. It is therefore important to better understand the physiopathological mechanisms of schizophrenia and the study of HR subjects could highlight early vulnerability factors. The neurodevelopmental model is an important hypothesis regarding the physiopathology of schizophrenia. In this model, some premorbid risks have been identified like urban environment, childhood trauma or immigrant status. These premorbid risks could disturb the brain developmental processes in crucial periods of proliferation, cell migration, morphological and biochemical differentiation, and circuit formation [START_REF] Rapoport | Neurodevelopmental model of schizophrenia: update 2012[END_REF]. Some metabolic abnormalities could, then, be caused by an alteration of the brain development. Moreover, among these neurometabolic abnormalities, it is important to note that a potential link between glutamatergic and GABAergic functions and the pathoetiology of schizophrenia could possibly be related to an hypofunctioning of N-methyl D-aspartate receptors on GABAergic neurons [START_REF] Coyle | Glutamate and schizophrenia: beyond the dopamine hypothesis[END_REF]. This disturbance of glutamate concentration has also been linked with an absence of remission during follow-up in ultra-high risk of psychosis and could be linked with a psychosis risk [START_REF] Egerton | Relationship between brain glutamate levels and clinical outcome in individuals at ultra high risk of psychosis[END_REF].

Proton magnetic resonance spectroscopy (MRS) like 1 H-MRS allows the measurement of various brain metabolites concentrations like N-acetyl aspartate (NAA), a marker of neuronal integrity, Choline (Cho), considered as a potential marker of membrane phospholipid metabolism [START_REF] Yildiz-Yesiloglu | Review of 1H magnetic resonance spectroscopy findings in major depressive disorder: A meta-analysis[END_REF], myo-inositol (mI), implicated in osmoregulation in glial cells [START_REF] Frey | Myoinositol in depressive and healthy subjects determined by frontal 1H-magnetic resonance spectroscopy at 1.5 tesla[END_REF] and glutamate (Glu) or glutamate+glutamine (Glx), reflecting both intracellular and extracellular glutamate and probably involved in metabolism and neurotransmission [START_REF] Rothman | 13C MRS studies of neuroenergetics and neurotransmitter cycling in humans[END_REF]. Anomalies in these brain metabolites have been found among people with schizophrenia such as an increase of Glx concentration in prefrontal white matter and the medial prefrontal cortex (mPFC) [START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF] or a reduction in the concentration of NAA in the frontal, temporal lobe [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF] and the thalamus grey matter [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF][START_REF] Iwata | Neurometabolite levels in antipsychoticnaïve/free patients with schizophrenia: A systematic review and meta-analysis of 1 H-MRS studies[END_REF]. Another MRS acquisition is the 31 P-MRS which is a particular acquisition allowing measurement of high energy phosphate metabolite levels like phosphocreatine (PCr), adenosine triphosphate (ATP), inorganic phosphate (Pi) and phospholipid metabolites like phosphomonoesters (PME or phosphodiesters (PDE) [START_REF] Chouinard | Brain bioenergetics and redox state measured by 31 P magnetic resonance spectroscopy in unaffected siblings of patients with psychotic disorders[END_REF]. Altered levels of these brain metabolites have been shown in patients with schizophrenia which could reveal abnormalities of energy and membrane phospholipid metabolism [START_REF] Du | In vivo evidence for cerebral bioenergetic abnormalities in schizophrenia measured using 31P magnetization transfer spectroscopy[END_REF][START_REF] Yuksel | Phosphorus magnetic resonance spectroscopy studies in schizophrenia[END_REF].

To the best of our knowledge, only three 1 H-MRS meta-analyses were conducted on high risk subjects [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF][START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF][START_REF] Mondino | N-Acetyl-Aspartate Level is Decreased in the Prefrontal Cortex in Subjects At-Risk for Schizophrenia[END_REF]. Two focused exclusively on the NAA concentration [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF][START_REF] Mondino | N-Acetyl-Aspartate Level is Decreased in the Prefrontal Cortex in Subjects At-Risk for Schizophrenia[END_REF], one on glutamate concentration [START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF] and no meta-analysis was performed on 31 P-MRS. Unlike these meta-analyses, our work also examines other metabolites such as Cho, mI or Creatinine (Cr). Thus, this study aims to characterize the metabolic profile of HR subjects, by comparing them to HC, and, then, to people with schizophrenia. Based on the neurodevelopmental model of schizophrenia [START_REF] Rapoport | Neurodevelopmental model of schizophrenia: update 2012[END_REF], we hypothesize that, among individuals at risk of transition to psychotic disorders, metabolic abnormalities are present prior to the first psychotic episode.

Method

Data sources and study selection process

The study protocol is registered at Open Science Framework, number DOI 10.17605/OSF.IO/K6AXH. We carried out a search on MEDLINE and PsycINFO databases in May 2019, without any limits on year of publication, using the key words ultrahigh risk of schizophrenia or at-risk mental state for schizophrenia or high risk for schizophrenia or prodrome of schizophrenia or genetic high risk for schizophrenia or familial high risk for schizophrenia or offspring at risk for schizophrenia or unaffected relatives of schizophrenia probands or ultra-high risk of psychosis or at-risk mental state for psychosis or high risk for psychosis or prodrome of psychosis or genetic high risk for psychosis or familial high risk for psychosis or offspring at risk for psychosis or unaffected relatives of psychosis probands; systematically associated with one of these other terms : MRS or magnetic resonance spectroscopy or glutamate or GABA or N-acetyl aspartate or brain metabolites. Studies were included if (i) they were published in English in a peer-reviewed journal, (ii) they included people with high risk for schizophrenia, (iii) they measured cerebral metabolites in vivo by MRS, (iv) they compared subjects with high risk and healthy controls or patients with schizophrenia or subjects with high risk with a transition to schizophrenia (HR-T) and HR with no transition (HR-NT). Studies that did not fulfill these three criteria were systematically excluded from the analyses. In order to obtain additional data, an email alert was created after May 2019 on MEDLINE with the same keywords to detect any publications of potential interest. The reference lists of articles identified were reviewed for additional studies. Study selection was performed by one author (BR) and verified by another (CM).

Data extraction

For each study, we obtained the means and standard deviations (SD) for all metabolites concentrations. Analysis of metabolites concentration by 1H-MRS is complicated because it could be biased by the absence of metabolites in cerebrospinal fluid. To correct this problem, in some studies, they calculated the ratio to a metabolite assumed to remain relatively constant (frequently the Cr) [START_REF] Jansen | 1 H MR spectroscopy of the brain: Absolute quantification of metabolites[END_REF]. When data were not available in these articles we contacted the corresponding authors (see Acknowledgments) of each included trial by email to improve data collection. If more than one group was available in the article, we used the method recommended by the Cochrane group (Cochrane 2011): M = (N1 M1 + N2 M2)/( N1+N2) and SD= √(( N1-1)SD1 2 +( N2-1)SD2 2 + ( N1 *N2/N1 + N1)( M1 2 + M1 2 -2 M1 M2))/( N1+ N2-1) (where N1 is the number of patients in the first group assessment, N2 is the number of patients in the second group assessment, M1 is the mean of the first group, M2 is the mean of the second group, SD1 is the standard deviation of the first group, and SD2 is the standard deviation of the second group). Articles written by a given research group were carefully scrutinized to ensure the absence of redundancy among the populations included in the trials.

A set of clinical variables was defined for the meta-regression analyses. We extracted the means and SDs for age ("age" variable), the percentages of females ("sex" variable), the Body Mass Index ("BMI" variable), the psychopathological evaluation like positive; negative; general; mood symptoms. Data extraction was performed by one author (BR) and verified by another (CM).

Data analyses

Data analyses were performed using RevMan, version 5.3 (Copenhagen, Denmark; the Nordic Cochrane Centre, Cochrane Collaboration). Effect sizes consisted in standardized mean differences (SMD) for metabolites concentrations between high risk subjects (HR) and 1) healthy controls (HC), 2) patients with schizophrenia, 3) patients with chronic schizophrenia (defined by a duration of illness over five years), 4) patients with a recent schizophrenia (defined by a duration of illness less than five years) and 5) between subjects with high risk of transition to schizophrenia (HR-T) and HR with no transition (HR-NT). Analyses were performed in all regions of interest for H 1 -MRS investigated in included studies: 1) the medial temporal cortex (MTL), 2) the medial prefrontal cortex (mPFC) including the anterior cingulate cortex, 3) the dorsolateral prefrontal cortex (DLPFC), 4) the basal ganglia, 5) the thalamus, 6) the corpus callosum, 7) the cerebellum, 8) the occipital cortex and 9) the prefrontal white matter. Analyses were performed by combining all prefrontal regions (mPFC, DLPFC) for 31 P-MRS. According to Cohen's method, SMDs were calculated as the differences between group means, divided by the pooled standard deviations. We also conducted more specific analyses including only studies with UHR subjects. A methodological investigation to evaluate the impact of field on our results was realized by performing analyses including only high field (defined by a field at 3T, 4T or 7T). In our analyses, when only one study with high field was included with other studies with low field, we excluded the high field study to assess its impact on the results. All analyses were performed with a random-effect model, which considers both between-study and within-study variability. An effect size was considered significant when the 95% confidence interval (95% CI) excluded 0 and when the p value was less than .05. Analyses were conducted when at least three studies were included. If less than three studies were available a review of different results was performed.

Study heterogeneity was estimated with the Q statistic which was calculated for all analyses and considered significant when p < .1. When a significant level of heterogeneity was reached, the I² index, an estimate of the total variation across the studies included, due to heterogeneity rather than chance was determined by the equation I² = [(Q -df)/Q] * 100% [START_REF] Higgins | Measuring inconsistency in metaanalyses[END_REF]. I² values of 25, 50, and 75 were indicative of mild, moderate, and marked heterogeneity respectively between trials. In addition, to ensure that the overall results

were not influenced by one single study, leave-one-out sensitivity analyses, performed by repeating the analyses with the consecutive exclusion of each study, were carried out for each analysis when more than three studies were included.

Funnel plots, plotting the standard error of each SMD against the SMD calculated for each study included, were drawn up when at least five individual studies contributed to an overall result, and their asymmetry was analyzed to assess the possible influence of publication and location biases [START_REF] Green | Cochrane Handbook for Systematic Reviews of Interventions 4[END_REF].

Finally, we conducted meta-regression analyses based on simple linear regression models to assess the influence of the population clinical heterogeneity on meta-analysis effect sizes.

Regression analyses were performed only when SMDs or heterogeneity were significant and when a reasonable number of data points were available, at least 5.

Results

Article identification process

The article-selection process was described in the Figure S1 and the Tables S1 andS2 according to the PRISMA guidelines [START_REF] Liberati | The PRISMA statement for reporting systematic reviews and meta-analyses of studies that evaluate health care interventions: explanation and elaboration[END_REF]. Thirty-nine studies were included in this meta-analysis. Thirty-five studies were included in 1 H-MRS analyses. Thirty-three of these studies were included in the HR vs HC analysis, thirteen studies were included in the HR vs all people with schizophrenia analysis, nine studies were included in the HR vs people with recent schizophrenia analysis, nine studies were included in the HR vs people with chronic schizophrenia analysis and five studies were included in the HR-T vs HR-NT analysis. Four studies were included in 31 P-MRS analyses. All these studies were included in the HR vs HC analysis. The main characteristics of each included study and their primary results are described in Tables 1; 2 andS3.

H 1 -MRS

MTL

HR vs HC

No difference was found for metabolites concentrations (NAA; Glx; Glu; mI; Cho; Cr) between HR and HC groups (Figure 1 andS2). The leave-one-out analysis shows that the absence of difference for the NAA concentration and the heterogeneity (I² = 55%, p = 0.02) is only due to one study [START_REF] Wood | Hippocampal pathology in individuals at ultra-high risk for psychosis: A multi-modal magnetic resonance study[END_REF]. When we excluded this study, a lower level of NAA and a decrease of the heterogeneity were found in HR subjects (n = 8 studies; HR = 243 ; HC = 223 ; test for overall effect : SMD = -0,24, 95% CI : -0,44 to -0,03 , p = 0,02; test for heterogeneity : χ² = 8,14, df = 7, p = 0,32, I² = 14%) (Figure S3). Secondary analyses including only UHR subjects were only conducted for NAA; Glx; Glu; mI; Cr and Cho levels without finding any difference between groups. We didn't observe any change on our results when only high field studies were included for NAA and Cho levels. No analyses were performed for other metabolites because only high field studies were included.

Meta-regression analyses showed a positive association between SMD for mI concentration and percentage of females (β = 0,06; Adjusted R-squared = 0,89; p = 0,01) which can be interpreted as a sex by diagnosis interaction (Figure S4).

HR vs patients with schizophrenia

When we included all types of people with schizophrenia, only analyses for NAA, Glu and Cho concentrations could be performed. A lower level of Cho was found in HR subjects (n = 4 studies; HR = 84 ; HC = 109 ; test for overall effect : SMD = -0,43, 95% CI : -0,73 to -0,13, p = 0,0005 ; test for heterogeneity : χ² = 0,81, df = 3, p = 0,85, I² = 0%) (Figure S5). No difference was found for other metabolites (NAA or Glu). Secondary analyses for UHR subjects could only be conducted for NAA and Cho concentrations without changing the results. We didn't observe any change on our results when only high field studies were included for NAA and Cho levels. When we exclusively included patients with a recent schizophrenia, we were able to perform analyses of NAA and Cho concentrations. A lower level of Cho was found (p=0,01) in HR (Figure S6) and no difference was found for NAA concentration (idem when we only included UHR subjects). No analysis could be performed between HR and people with chronic schizophrenia. Two studies investigated the difference of Glu concentrations among HR compared to first episode of schizophrenia in the first study [START_REF] Nenadic | Associations of hippocampal metabolism and regional brain grey matter in neuroleptic-naïve ultra-high-risk subjects and first-episode schizophrenia[END_REF] and compared to chronic schizophrenia in the second [START_REF] Lutkenhoff | Proton MRS in twin pairs discordant for schizophrenia[END_REF]. No difference was found in these two studies. Only one study investigated mI concentration between HR and chronic schizophrenia [START_REF] Lutkenhoff | Proton MRS in twin pairs discordant for schizophrenia[END_REF] but no difference was found.

HR-T vs HR-NT

Three studies investigated differences between HR-T and HR-NT [START_REF] Bossong | Association of hippocampal glutamate levels with adverse outcomes in individuals at clinical high risk for psychosis[END_REF][START_REF] Wood | Hippocampal pathology in individuals at ultra-high risk for psychosis: A multi-modal magnetic resonance study[END_REF][START_REF] Wood | Proton magnetic resonance spectroscopy in first episode psychosis and ultra highrisk individuals[END_REF]. No difference was found for NAA concentration between HR-T and HR-NT (Figure S7). Significant higher Glu, mI and Cr concentrations in HR-T were found in one study [START_REF] Bossong | Association of hippocampal glutamate levels with adverse outcomes in individuals at clinical high risk for psychosis[END_REF]. No difference was found for Cho and Glx concentrations in these three studies, even though one study showed a statistical increasing trend of the NAA /Cho ratio [START_REF] Wood | Proton magnetic resonance spectroscopy in first episode psychosis and ultra highrisk individuals[END_REF].

mPFC

HR vs HC

A higher level of Glx was found in HR subjects (n = 12 studies ; HR = 314 ; HC = 297 ; test for overall effect : SMD = 0,25, 95% CI : 0,07 to 0,43, p = 0,007 ; test for heterogeneity : χ² = 13,06, df = 11, p = 0,29, I² = 16%). No difference was found for all other metabolites (NAA; Glu; GABA; mI; Cho and Cr) (Figure 2 andS8). Leave-one-out analyses showed that the absence of difference for Cho concentration and the heterogeneity (I² = 34%; p = 0.1) were only due to one study [START_REF] Egerton | Relationship between brain glutamate levels and clinical outcome in individuals at ultra high risk of psychosis[END_REF]. When we excluded this study, analyses showed a higher level of Cho in HR subjects and a decrease of the heterogeneity (n = 13 studies; HR = 255 ; HC = 281 ; test for overall effect: SMD = 0,21, 95% CI : 0,04 to 0,39, p = 0,02 ; test for heterogeneity : χ² = 12,15, df = 12, p = 0,43, I² = 1%) (Figure S9). Secondary analyses including only UHR subjects did not show any difference between the two groups for NAA; Glx; mI; Cho; Cr and GABA concentrations. Glutamine concentration was investigated by one study [START_REF] Stone | Glutamate Dysfunction in People with Prodromal Symptoms of Psychosis: Relationship to Gray Matter Volume[END_REF] that revealed a higher level among UHR subjects. When we included only high field studies, the results remained the same for NAA, Glx, mI, Cho, and Cr levels excepting a heterogeneity decrease from 22% to 0% for the NAA level. No analyses were performed for Glu and GABA levels as all included studies used a high field.

Meta-regression analyses showed a negative association between SMD for Cho concentration and percentage of females (β = -0,01) and age of HR (β = -0,02) (Adjusted R-squared = 0,46; p = 0,01) (Figure S10).

HR vs patients with schizophrenia

Analyses were performed for NAA and Cho concentrations (because we didn't identify more than two studies for other metabolites) without finding any difference between groups when we included all people with schizophrenia (Figure S11). This result did not change by including only UHR subjects. No analyses could be performed between HR subjects and people with recent schizophrenia. When we included only high field studies, the results remained the same for NAA and Cho levels except that we found a heterogeneity decrease from 44% to 0% for the Cho level. Only one study investigated the difference between HR subjects and patients with a first episode of schizophrenia [START_REF] Uhl | 1H-MR spectroscopy in ultra-high risk and first episode stages of schizophrenia[END_REF]. They did not find any difference in NAA and Cho concentrations between the two groups. When we compared HR subjects to people with chronic schizophrenia, no difference was found for NAA and Cho concentrations (Figure S12). Three other studies investigated other metabolites concentrations like Glx [START_REF] Natsubori | Reduced Frontal Glutamate + Glutamine and N-Acetylaspartate Levels in Patients With Chronic Schizophrenia but not in Those at Clinical High Risk for Psychosis or With First-Episode Schizophrenia[END_REF], Glu [START_REF] Lutkenhoff | Proton MRS in twin pairs discordant for schizophrenia[END_REF] or GABA [START_REF] Marenco | Prefrontal GABA Levels Measured With Magnetic Resonance Spectroscopy in Patients With Psychosis and Unaffected Siblings[END_REF]. Compared to HR subjects, two studies found a lower level of Glx [START_REF] Natsubori | Reduced Frontal Glutamate + Glutamine and N-Acetylaspartate Levels in Patients With Chronic Schizophrenia but not in Those at Clinical High Risk for Psychosis or With First-Episode Schizophrenia[END_REF] or Glu [START_REF] Lutkenhoff | Proton MRS in twin pairs discordant for schizophrenia[END_REF] in patients with chronic schizophrenia but not in patients with a first episode of schizophrenia [START_REF] Natsubori | Reduced Frontal Glutamate + Glutamine and N-Acetylaspartate Levels in Patients With Chronic Schizophrenia but not in Those at Clinical High Risk for Psychosis or With First-Episode Schizophrenia[END_REF]. The third study [START_REF] Marenco | Prefrontal GABA Levels Measured With Magnetic Resonance Spectroscopy in Patients With Psychosis and Unaffected Siblings[END_REF] found a lower level of GABA concentration in HR compared to patients with treated psychosis.

HR-T vs HR-NT

No analysis could be performed because only two studies have been realized for this comparison [START_REF] Jessen | Proton magnetic resonance spectroscopy in subjects at risk for schizophrenia[END_REF]Modinos et al., 2018). [START_REF] Jessen | Proton magnetic resonance spectroscopy in subjects at risk for schizophrenia[END_REF] found a higher Cho/Cr ratio and a lower NAA/Cho ratio in HR-T. Modinos et al (2018) found no difference for GABA concentration between the two groups.

DLPFC

HR vs HC

A higher level of mI was found in HR subjects (n = 3 studies; HR = 77 ; HC = 61 ; test for overall effect : SMD = 0,36, 95% CI: 0.01 to 0,70, p = 0,04 ; test for heterogeneity: χ² = 0,27, df = 2, p = 0,87, I² = 0%). No difference was found for other metabolites (NAA; Glx and Cho) (Figure 3 andS13). Leave-one-out analyses did not change these results. A secondary analysis has been performed only for NAA and Cho concentrations without any difference found between UHR and controls. When we included only low field studies, the results remained the same for NAA level. No analyses were performed for Glx, mI, and Cho levels because all studies included used a low field.

Meta-regression analyses did not show significant association between SMD and sex or age for NAA and Cho concentrations.

HR vs patients with schizophrenia

Analyses were only performed for NAA and Cho concentrations between HR and all patients with schizophrenia. No difference was found for Cho and NAA concentrations (Figure S14).

A secondary analysis was performed only for NAA concentration without finding any difference. When we included only low field studies, the results remained the same for NAA level. No analyses were performed for Cho level as all included studies used a low field. Only two studies investigated the difference between HR and people with recent schizophrenia [START_REF] Liemburg | Prefrontal NAA and Glx Levels in Different Stages of Psychotic Disorders: a 3T 1H-MRS Study[END_REF][START_REF] Wood | Proton magnetic resonance spectroscopy in first episode psychosis and ultra highrisk individuals[END_REF]. No difference was found for NAA [START_REF] Liemburg | Prefrontal NAA and Glx Levels in Different Stages of Psychotic Disorders: a 3T 1H-MRS Study[END_REF][START_REF] Wood | Proton magnetic resonance spectroscopy in first episode psychosis and ultra highrisk individuals[END_REF], Cho [START_REF] Wood | Proton magnetic resonance spectroscopy in first episode psychosis and ultra highrisk individuals[END_REF], and Glx [START_REF] Liemburg | Prefrontal NAA and Glx Levels in Different Stages of Psychotic Disorders: a 3T 1H-MRS Study[END_REF] concentrations between the two groups. Compared to patients with chronic schizophrenia, a lower level of Cho was found in HR (n = 3 studies ; HR = 68 ; HC = 59 ; test for overall effect : SMD = -0,81, 95% CI : -1,19 to -0,44, p < 0,001 ; test for heterogeneity : χ² = 0,79, df = 2, p = 0,67, I² = 0%) (Figure S15).

HR-T vs HR-NT

Only two studies investigated this comparison [START_REF] Jessen | Proton magnetic resonance spectroscopy in subjects at risk for schizophrenia[END_REF][START_REF] Wood | Proton magnetic resonance spectroscopy in first episode psychosis and ultra highrisk individuals[END_REF] and did not find any difference between HR-T and HR-NT for NAA and Cho concentrations.

Basal ganglia

HR vs HC

Analyses were performed for NAA, Glx and Cho concentrations. A statistical trend to a higher level of Glx was found in HR subjects (n = 5 studies; HR = 122 ; HC = 131 ; test for overall effect: SMD = 0,4, 95% CI : 0,00 to 0,8, p = 0,05 ; test for heterogeneity : χ² = 9,51, df = 4, p = 0,05, I² = 58%) (Figure 5 andS16). The leave-one out analyses found a significant higher level of Glx when we excluded [START_REF] Block | Decreased frontal lobe ratio of N-acetyl aspartate to choline in familial schizophrenia: a proton magnetic resonance spectroscopy study[END_REF] (p = 0,04) or [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF] (p = 0,005) with a heterogeneity decrease (I² = 41%, p=0,16) (Figure S17). No difference was found for NAA and Cho concentrations and we couldn't perform a secondary analysis. When we included only high field studies, the results remained the same for NAA, Glx and Cho levels. Regarding other metabolites, contradictory results were found for glutamate and GABA levels [START_REF] De | Higher levels of glutamate in the associative-striatum of subjects with prodromal symptoms of schizophrenia and patients with first-episode psychosis[END_REF][START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF]. No difference was found for Glx, Gln [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF] and mI concentrations [START_REF] Block | Decreased frontal lobe ratio of N-acetyl aspartate to choline in familial schizophrenia: a proton magnetic resonance spectroscopy study[END_REF].

HR vs patients with schizophrenia

Analyses were only performed for Glx concentration between HR and all patients with schizophrenia and no difference was found (Figure S18). Regarding other metabolites, no difference was found for NAA [START_REF] Block | Decreased frontal lobe ratio of N-acetyl aspartate to choline in familial schizophrenia: a proton magnetic resonance spectroscopy study[END_REF][START_REF] De | Higher levels of glutamate in the associative-striatum of subjects with prodromal symptoms of schizophrenia and patients with first-episode psychosis[END_REF], Glu [START_REF] De | Higher levels of glutamate in the associative-striatum of subjects with prodromal symptoms of schizophrenia and patients with first-episode psychosis[END_REF][START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF], Gln [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF], GABA , [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF], Cho [START_REF] Block | Decreased frontal lobe ratio of N-acetyl aspartate to choline in familial schizophrenia: a proton magnetic resonance spectroscopy study[END_REF] and mI [START_REF] Block | Decreased frontal lobe ratio of N-acetyl aspartate to choline in familial schizophrenia: a proton magnetic resonance spectroscopy study[END_REF] concentrations.

When we included only high field studies, the results remained the same for Glx level.

HR-T vs HR-NT

Only one study (de la Fuente-Sandoval et al., 2013) investigated this topic and found a higher level of Glu in UHR-T but no difference for other metabolites like GABA, NAA and Glx.

Thalamus

No difference was found between HR and HC for NAA, Glx, mI, Cr and Cho concentrations (Figure 5). Only one study [START_REF] Egerton | Relationship between brain glutamate levels and clinical outcome in individuals at ultra high risk of psychosis[END_REF] investigated Glu concentration and found a lower level in UHR subjects. No study has investigated the difference between HR subjects and people with schizophrenia or HR-T and HR-NT. When we included only low field studies, the results remained the same for NAA, Glx, mI, Cho and Cr levels excepting a heterogeneity decrease from 92% to 66% for the mI level.

Corpus Callosum

Only one study investigated this region of interest [START_REF] Aydin | Altered Metabolic Integrity of Corpus Callosum Among Individuals at Ultra High Risk of Schizophrenia and First-Episode Patients[END_REF] and found a lower level of NAA in UHR subjects compared to HC and a decrease of NAA concentration among patients with a first episode of schizophrenia compared to UHR subjects. No difference was found between UHR subjects and HC or first episode of schizophrenia for Cho concentration.

Cerebellum

Only one study investigated this region of interest [START_REF] De | Higher levels of glutamate in the associative-striatum of subjects with prodromal symptoms of schizophrenia and patients with first-episode psychosis[END_REF] and found no difference between UHR and HC for Cho, mI, Glu, Glx and NAA concentrations.

They found a higher level of NAA concentration in FEP patients compared to UHR subjects but no difference for Glu, Glx, Cho and mI concentrations.

Occipital

Two studies investigated this region of interest (Grent-'t-Jong et al., 2018;[START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF]. One found an higher level of Glx concentration (Grent-'t-Jong et al., 2018) and the other found a decrease [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF] in HR subjects vs controls. A lower level of Glu concentration [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF] in HR vs HC was found. No difference was found for GABA concentration (Grent-'t-Jong et al., 2018;[START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF] and Gln concentration [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF]. Compared to patients with schizophrenia, a lower level of GABA concentration was found in HR and no difference was found for Gln, Glx, Glu concentrations [START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF].

Prefrontal white matter

Only one study investigated this region of interest [START_REF] Lutkenhoff | Proton MRS in twin pairs discordant for schizophrenia[END_REF] and found no difference for NAA, Glu, Cho and mI concentrations between HR and HC or people with schizophrenia.

31 P-MRS

We did not find any difference in the prefrontal cortex between HR subjects and HC for PME, PDE, PCR and Pi. We found a statistical trend to a decrease of intracellular pH in HR subjects (p = 0.06) (Figure S19). Only one study compared [START_REF] Chouinard | Brain bioenergetics and redox state measured by 31 P magnetic resonance spectroscopy in unaffected siblings of patients with psychotic disorders[END_REF] UHR subjects and patients with a first episode of psychosis without finding any difference for intracellular pH, phosphocreatine/β-ATP; inorganic phosphate/ β-ATP, phosphodiester/ β-ATP, phosphomonoester/ β-ATP.

Discussion

A better understanding of the schizophrenia pathophysiology could allow a change in the medical intervention provided to these patients. By studying subjects with a risk of developing this pathology, we could, on the one hand, determine potential early vulnerability factors and, on the other hand, to consider earlier care so the functional impact could be limited. This meta-analysis shows a higher level of Glx in the mPFC cortex and a potential higher level of Glx in the basal ganglia as well as a higher level of mI in the DLPFC in subjects at risk compared to control subjects. A potential lower level of NAA concentration was also found in HR subjects in the medial temporal cortex. It also shows a higher level of Cho in subjects with schizophrenia compared to subjects at risk in the DLPFC and the hippocampus.

Some results from this meta-analysis are common to previous ones [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF][START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF][START_REF] Mondino | N-Acetyl-Aspartate Level is Decreased in the Prefrontal Cortex in Subjects At-Risk for Schizophrenia[END_REF]) as a higher level of Glx in the mPFC cortex [START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF]. Nevertheless, we found a higher level of Glx concentration in the basal ganglia that has not been described yet. Compared to the meta-analysis of Merritt et al. ( 2016) [START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF], this result could be explained by a difference in the included studies.

Indeed, we could not include the study by Keshevan et al. (2009) due to the impossibility of obtaining the data. However, we included two recent studies (de la [START_REF] De La Fuente-Sandoval | Cortico-Striatal GABAergic and Glutamatergic Dysregulations in Subjects at Ultra-High Risk for Psychosis Investigated with Proton Magnetic Resonance Spectroscopy[END_REF][START_REF] Thakkar | 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings[END_REF] that were not included in the meta-analysis by [START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF]. It is important to note that the lack of difference found in the meta-analysis of [START_REF] Merritt | Nature of Glutamate Alterations in Schizophrenia: A Meta-analysis of Proton Magnetic Resonance Spectroscopy Studies[END_REF] has a more important heterogenity (I 2 = 73.7%), than our analysis (I 2 = 58%), and this heterogeneity is mainly due to the [START_REF] Keshavan | Striatal metabolic alterations in non-psychotic adolescent offspring at risk for schizophrenia: A 1H spectroscopy study[END_REF] study, which found an SMD at -0,34. Other methodological differences are important to note compared to previous metaanalyses. First, as numerous studies were recently published on the subject, we were able to include a higher number of studies and thereby increase the statistical power, and so, we were able to perform leave-one-out studies to investigate the heterogeneity of our analyses. For example, [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF] included 4 studies in their analyses between HR and HC in the temporal lobe for NAA concentration whereas our meta-analysis included 9 studies.

Moreover, our leave-one-out study showed that the absence of difference and the heterogeneity were only due to one study [START_REF] Wood | Hippocampal pathology in individuals at ultra-high risk for psychosis: A multi-modal magnetic resonance study[END_REF] that was included in the Brugger et al. ( 2011) meta-analysis. Thus, we showed that the level of NAA was potentially lower in the MTL among HR subjects unlike [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF]. A second difference is the comparison between HR subjects and patients with schizophrenia that, to our knowledge, has never been performed. A third is that, contrary to three previous meta-analyses, we investigated other brain metabolites such as Cho, mI, Cr or GABA, which allowed us to show a higher level of mI in the DLPFC in HR subjects as well as a higher level of Cho in people with schizophrenia compared to HR subjects. Unlike [START_REF] Brugger | Proton Magnetic Resonance Spectroscopy and Illness Stage in Schizophrenia-A Systematic Review and Meta-Analysis[END_REF] and [START_REF] Mondino | N-Acetyl-Aspartate Level is Decreased in the Prefrontal Cortex in Subjects At-Risk for Schizophrenia[END_REF], we chose to fragment the prefrontal cortex into the mPFC and the DLPFC for 1 H-MRS analyses which could explain some difference in the results.

Our meta-analysis shows a higher level of Glx in subjects at risk compared to controls, particularly in the mPFC and potentially in the basal ganglia. Indeed, in basal ganglia we only found a statistical trend to an increase of Glx level (p = 0.05). The result became clearly significant (p = 0.005) and a decrease of the heterogeneity was obtained when we excluded the study from Takkhar et al. ( 2017) confirming that the statistical trend was principally driven by this study. Also, we didn't find any difference regarding Glx level between HR and patients with schizophrenia in basal ganglia. These results might be consistent with an increase in the glutamatergic system activity in people with schizophrenia possibly due to NMDA receptors hypofunctioning [START_REF] Moghaddam | Bringing order to the glutamate chaos in schizophrenia[END_REF][START_REF] Moghaddam | Activation of glutamatergic neurotransmission by ketamine: a novel step in the pathway from NMDA receptor blockade to dopaminergic and cognitive disruptions associated with the prefrontal cortex[END_REF][START_REF] Moghaddam | Glutamatergic animal models of schizophrenia[END_REF]. Similar results have been described in animals, as well as in healthy subjects, after taking ketamine, which is an NMDA receptor antagonist [START_REF] Moghaddam | Activation of glutamatergic neurotransmission by ketamine: a novel step in the pathway from NMDA receptor blockade to dopaminergic and cognitive disruptions associated with the prefrontal cortex[END_REF]. The increase in Glx concentration in the basal ganglia may have a role in modulating the dopaminergic system via NMDA receptors [START_REF] Lodge | Hippocampal dysregulation of dopamine system function and the pathophysiology of schizophrenia[END_REF]. It seems important to note that hyperactivation of the glutamatergic system in subjects at risk, in the basal ganglia, has also been described as a potential predictor of transition to psychosis (de la [START_REF] De La Fuente-Sandoval | Striatal glutamate and the conversion to psychosis: a prospective 1H-MRS imaging study[END_REF]. Regarding the higher level of Glx in the mPFC, relatively similar results have been shown in patients presenting a first episode without psychotropic treatment [START_REF] Bartha | Measurement of glutamate and glutamine in the medial prefrontal cortex of never-treated schizophrenic patients and healthy controls by proton magnetic resonance spectroscopy[END_REF][START_REF] Théberge | Longitudinal grey-matter and glutamatergic losses in first-episode schizophrenia[END_REF][START_REF] Théberge | Glutamate and glutamine measured with 4.0 T proton MRS in never-treated patients with schizophrenia and healthy volunteers[END_REF]. These results could point to a vulnerability even before the disease onset It is important to note that we found a dissociation in our results with a higher Glx level in mPFC in HR but not in UHR subjects compared to HC subjects. Most of the time, HR subjects were defined as subjects with familial antecedent of psychosis which are closely to the UHR sub-classes genetic and deterioration syndrome defined by the presence of familial antecedent of psychosis or schizotypical personality and a functioning deterioration. A possible explanation of this result is that HR subjects, as UHR with genetic risk and deterioration syndrome, could have a genetic predisposition associated with a modification of cerebral metabolites that could be part of an innate vulnerability. This genetic vulnerability could be less important in other sub-classes of UHR, which explains the dissociation of our results. Another explanation of this difference is a statistical hypothesis.

When we included only UHR subjects in the analyses, we lost approximately one third of HR which could cause an important loss of statistical power. It also seems that antipsychotic treatments would tend to correct this hyperactivation [START_REF] Kegeles | Elevated prefrontal cortex γ-aminobutyric acid and glutamate-glutamine levels in schizophrenia measured in vivo with proton magnetic resonance spectroscopy[END_REF] or that an initial hyperactivation would be a factor of good prognosis for the response to antipsychotic treatments [START_REF] Poels | Glutamatergic abnormalities in schizophrenia: A review of proton MRS findings[END_REF]. However, considering the medication status seems to be insufficient to explain all the differences between the phases of the illness. For example, Natsubori et al. ( 2014) found a higher level of Glx in FES compared to chronic schizophrenia whereas the two groups were sufficiently treated by antipsychotics (respectively 558.1 and 839.8 mg/day chlorpromazine equivalents) but found no difference between UHR and FES even though FES received more treatment (respectively 117.6 and 558.1 mg/day chlorpromazine equivalents). Other confounding factors may partly explain the difference of Glx levels at the different stages of illness such as the absence of remission [START_REF] Egerton | Anterior cingulate glutamate levels related to clinical status following treatment in first-episode schizophrenia[END_REF]; or a possible association with the progressive brain volume reduction in the mPFC [START_REF] Natsubori | Reduced Frontal Glutamate + Glutamine and N-Acetylaspartate Levels in Patients With Chronic Schizophrenia but not in Those at Clinical High Risk for Psychosis or With First-Episode Schizophrenia[END_REF] that are associated with longer durations of schizophrenia [START_REF] Koo | A cross-sectional and longitudinal magnetic resonance imaging study of cingulate gyrus gray matter volume abnormalities in first-episode schizophrenia and first-episode affective psychosis[END_REF][START_REF] Premkumar | Association between a longer duration of illness, age and lower frontal lobe grey matter volume in schizophrenia[END_REF][START_REF] Velakoulis | Increased duration of illness is associated with reduced volume in right medial temporal/anterior cingulate grey matter in patients with chronic schizophrenia[END_REF]. Moreover, it is important to note that the interpretation of the higher level of Glx needs to be tempered as the link between MRSdetected levels of Glx and glutamatergic activity is currently not known.

In our meta-analysis, we found a possible decrease of NAA concentration in the MTL. It is interesting to note that a similar result has been described in first psychotic episodes and in chronic schizophrenia in one meta-analysis and a statistical decreasing trend was also found in another [START_REF] Iwata | Neurometabolite levels in antipsychoticnaïve/free patients with schizophrenia: A systematic review and meta-analysis of 1 H-MRS studies[END_REF]. This result could also be considered as a vulnerability marker of schizophrenia. The role of brain NAA metabolites is various [START_REF] Moffett | N-Acetylaspartate in the CNS: From neurodiagnostics to neurobiology[END_REF]. The NAA have a role in neurotransmission with some interactions with glutamatergic [START_REF] Ohrmann | Evidence for glutamatergic neuronal dysfunction in the prefrontal cortex in chronic but not in first-episode patients with schizophrenia: a proton magnetic resonance spectroscopy study[END_REF] and dopaminergic [START_REF] Bertolino | The relationship between dorsolateral prefrontal neuronal N-acetylaspartate and evoked release of striatal dopamine in schizophrenia[END_REF] systems. To illustrate these interactions it is interesting to note that the modulation of the GRM3 allele (gene encoding for the 2/3 type metabotropic glutamate receptor) could decrease the NAA levels in the right prefrontal cortex [START_REF] Marenco | Effect of metabotropic glutamate receptor 3 genotype on N-acetylaspartate measures in the dorsolateral prefrontal cortex[END_REF]. The relation with the dopaminergic system could be illustrated by the association found in people with schizophrenia between a decrease of NAA concentrations in the prefrontal cortex and a dopaminergic release dysregulation in the striatum [START_REF] Bertolino | The relationship between dorsolateral prefrontal neuronal N-acetylaspartate and evoked release of striatal dopamine in schizophrenia[END_REF]. A possible explanation of these different interactions is that the N-acetylaspartylglutamate (NAAG) is directly synthesized, by a NAAG synthetase, from NAA and glutamate [START_REF] Moffett | N-Acetylaspartate in the CNS: From neurodiagnostics to neurobiology[END_REF]. This NAAG could regulate the glutamate and the dopamine release probably via the activation of presynaptic 2/3 type metabotropic receptors [START_REF] Moffett | N-Acetylaspartate in the CNS: From neurodiagnostics to neurobiology[END_REF].

In schizophrenia, an altered membrane metabolism has been described [START_REF] Keshavan | Magnetic resonance spectroscopy in schizophrenia: methodological issues and findings--part II[END_REF]. This could be explained by anomalies in the membrane phospholipid synthesis and a phospholipid breakdown [START_REF] Keshavan | Magnetic resonance spectroscopy in schizophrenia: methodological issues and findings--part II[END_REF][START_REF] Stanley | Magnetic resonance spectroscopy in schizophrenia: methodological issues and findings--part I[END_REF]. This dysregulation of the phospholipid metabolism could be related to stress, a viral infection or hormonal influences [START_REF] Horrobin | Schizophrenia: the illness that made us human[END_REF]. In our meta-analysis, we have investigated this metabolism with two markers. The first is the Cho concentration obtained by 1 H-MRS (Yildiz-Yesiloglu and Ankerst, 2006) and others like PME or PDE by 31 P-MRS. We did not find any difference between HR and HC concerning PME, PDE, and PCR concentrations but we found a statistical trend to a decrease of intracellular pH in HR subjects in the frontal cortex. These results were similar to results found in patients with schizophrenia [START_REF] Du | In vivo evidence for cerebral bioenergetic abnormalities in schizophrenia measured using 31P magnetization transfer spectroscopy[END_REF]) and could be interpreted as an increase in the contribution of glycolysis to ATP synthesis with no impact on levels of substrates as PME or PCR. We found also an increase of Cho concentration was found in people with schizophrenia vs HR in the DLPFC and the MTL. This Cho concentration increase could be considered as an increase of phospholipid membrane turnover leading to an alteration of phospholipid metabolism [START_REF] Ende | The hippocampus in patients treated with electroconvulsive therapy: a proton magnetic resonance spectroscopic imaging study[END_REF]. A hypothesis to explain this result is that these anomalies are not present before the onset of the pathology and could be a marker of the transition to schizophrenia. Unfortunately, we could not confirm this hypothesis with a direct comparison between HR-T and HR-NT because no sufficient studies have been published.

We also found an association between the concentration of Cho in the mPFC, the percentage of females and the age. These associations could explain, in part, our analyses' heterogeneity (I² = 34%; p = 0.1). Some studies showed sex differences [START_REF] Chang | Effects of age and sex on brain glutamate and other metabolites[END_REF][START_REF] Hädel | Effects of age and sex on the concentrations of glutamate and glutamine in the human brain: Brain Glutamate and Glutamine With Age and Sex[END_REF][START_REF] Pfleiderer | N-acetylaspartate levels of left frontal cortex are associated with verbal intelligence in women but not in men: a proton magnetic resonance spectroscopy study[END_REF] with a decrease of Cho concentration in ACC in women but not all studies [START_REF] Endres | On the Effect of Sex on Prefrontal and Cerebellar Neurometabolites in Healthy Adults: An MRS Study[END_REF]. Among the included studies, it is important to note that some studies have not matched for sex their HR subjects and their controls. We found a growing difference with controls as the number of women among HR subjects increases, which is consistent with the literature. A hypothesis to explain this sex-difference is an effect of menstruation cycle on cortical concentrations of Cho and in particular an impact of estrogen which could regulate the acetyltransferase activity which converts choline in acetylcholine [START_REF] Bohacek | Long-Term Ovarian Hormone Deprivation Alters the Ability of Subsequent Oestradiol Replacement to Regulate Choline Acetyltransferase Protein Levels in the Hippocampus and Prefrontal Cortex of Middle-Aged Rats[END_REF][START_REF] Hädel | Effects of age and sex on the concentrations of glutamate and glutamine in the human brain: Brain Glutamate and Glutamine With Age and Sex[END_REF][START_REF] Rasgon | Menstrual cycle-related brain metabolite changes using 1H magnetic resonance spectroscopy in premenopausal women: a pilot study[END_REF]. The link between choline concentration and age is more harder to explain. To our knowledge, few studies have investigated this association and no link was found between these two variables [START_REF] Pfleiderer | N-acetylaspartate levels of left frontal cortex are associated with verbal intelligence in women but not in men: a proton magnetic resonance spectroscopy study[END_REF]Saunders et al., 1999).

It is important to note that more than 70% of HR subjects have a non-psychotic disorder [START_REF] Fusar-Poli | Comorbid Depressive and Anxiety Disorders in 509 Individuals With an At-Risk Mental State: Impact on Psychopathology and Transition to Psychosis[END_REF] with an important prevalence of depressive disorders, anxiety disorders, obsessive-compulsive disorder and substance use disorders [START_REF] Albert | Prevalence of non-psychotic disorders in ultrahigh risk individuals and transition to psychosis: A systematic review[END_REF][START_REF] Fusar-Poli | Comorbid Depressive and Anxiety Disorders in 509 Individuals With an At-Risk Mental State: Impact on Psychopathology and Transition to Psychosis[END_REF]. These non-psychotic disorders could also impact results of MRS studies in our meta-analysis. However, 72% of included studies had excluded substance abuse or dependence so the impact of this comorbidity is limited. For other comorbidities, some studies had excluded or included very few HR subjects with comorbid Axis I diagnosis (de la [START_REF] De La Fuente-Sandoval | Cortico-Striatal GABAergic and Glutamatergic Dysregulations in Subjects at Ultra-High Risk for Psychosis Investigated with Proton Magnetic Resonance Spectroscopy[END_REF], 2013;[START_REF] De | Higher levels of glutamate in the associative-striatum of subjects with prodromal symptoms of schizophrenia and patients with first-episode psychosis[END_REF][START_REF] Wobrock | Reduction of the Internal Capsule in Families Affected with Schizophrenia[END_REF], or HR subjects with bipolar disorder [START_REF] Aydin | Altered Metabolic Integrity of Corpus Callosum Among Individuals at Ultra High Risk of Schizophrenia and First-Episode Patients[END_REF][START_REF] Chouinard | Brain bioenergetics and redox state measured by 31 P magnetic resonance spectroscopy in unaffected siblings of patients with psychotic disorders[END_REF][START_REF] Keshavan | Prefrontal membrane phospholipid metabolism of child and adolescent offspring at risk for schizophrenia or schizoaffective disorder: an in vivo 31P MRS study[END_REF][START_REF] Klemm | Cerebral phosphate metabolism in first-degree relatives of patients with schizophrenia[END_REF] or HR subjects with major depressive disorder [START_REF] Keshavan | Prefrontal membrane phospholipid metabolism of child and adolescent offspring at risk for schizophrenia or schizoaffective disorder: an in vivo 31P MRS study[END_REF][START_REF] Klemm | Cerebral phosphate metabolism in first-degree relatives of patients with schizophrenia[END_REF][START_REF] Natsubori | Reduced Frontal Glutamate + Glutamine and N-Acetylaspartate Levels in Patients With Chronic Schizophrenia but not in Those at Clinical High Risk for Psychosis or With First-Episode Schizophrenia[END_REF] which could decrease the impact of these non-psychotic disorders in results of MRS. In MDD, results of MRS metaanalyses showed a lower level of Glx in frontal lobe and notably in anterior cingular cortex [START_REF] Luykx | Region and state specific glutamate downregulation in major depressive disorder: A metaanalysis of 1H-MRS findings[END_REF][START_REF] Yildiz-Yesiloglu | Review of 1H magnetic resonance spectroscopy findings in major depressive disorder: A meta-analysis[END_REF]. These results could strengthen our result showing a higher level of Glx in HR subjects in mPFC. In fact, despite the inclusion of HR subjects with MDD (who could have a lower level of Glx) we found the same result, that is, a higher level among HR subjects.

This meta-analysis has several limitations. First, we can mention a limited number of included studies that restricts the number of data available to perform some analyses and therefore the statistical power. Indeed, we couldn't perform analyses comparing HR subjects to people with schizophrenia and HR-T to HR-NT, as less than three studies were available. A second limit is a possible heterogeneity in ours populations. First, the HR subjects group in which HR subjects could be symptomatic or not. Indeed, some studies included subjects defined as HR because of a family history, while others included subjects defined as UHR (using CAARMS criteria). We tried to temper this limit by performing secondary analyses, which included only the UHR subjects, but, unfortunately, many analyses could not be reproduced due to a lack of available data and, when they were realized, their interpretations must be made with great caution because of a possible lack of statistical power. Second, the heterogeneity is also present when we performed the comparison between HR subjects and patients with schizophrenia with a possible impact of age. Indeed, patients with schizophrenia are older than HR subjects and this difference could disturb the cerebral metabolites concentrations independently. A third limitation is that MRS cannot be used as a diagnostic tool to evaluate a subject's vulnerability. Indeed, the associations and differences are too affected by scatter to be useful for individual diagnosis, even though they are statistically significant. Thus, our results are a research tool. A fourth limitation is the difference between the MRS acquisition (notably field strength, voxel size and placement, echo time), which could impact the results.

The heterogeneity found in some nalyses (as for the NAA level analyze between HR and HC in mPFC) was explained by these differences in the field strength. A final limitation is that we could not determine the influence of neurometabolite reference quantification (e.g. Crratio). However, we did not find any difference between HR subjects and controls for Cr level in mPFC and thalamus.

As a conclusion, this meta-analysis revealed a higher level of Glx in the mPFC, potentially in the basal ganglia, and a potential lower level of NAA in the medial temporal cortex in HR.

These results provide evidence that some brain metabolites concentrations could be disturbed before the transition to psychosis and could be considered as vulnerability factors. Future studies should continue to investigate this topic to provide greater understanding of the physiopathology of schizophrenia and also develop markers that could change the clinical practice.

Figures and tables

Figure

  Figure 1. Results of meta-analysis for medial temporal lobe between high risk subjects

Table 1

 1 CrIncrease GABA and Glx concentration in UHR in NC and mPFC Nb : number, ROI : region of interest ; UHR : ultra-high risk ; HR: high risk; HC : healthy controls ; SCZ chro : chronic schizophrenia ; SCZ recent : recent schizophrenia ; FES : first episode of schizophrenia ; FEP : first episode of psychosis ; UHR-T: ultra-high risk with a transition to schizophrenia; UHR-NT: ultra-high risk with no transition; MTL : medial temporal lobe ; mPFC : medial prefrontal cortex ; DLPFC : dorsolateral prefrontal cortex; WMPFC : white matter prefrontal cortex ; NC : caudate nucleus ; NAA : N-acetyl aspartate ; GABA : acide γ amino-butirique ; Cr : creatine ; Cho : choline ; mI : myo-inositol ; Glx : glutamine+glutamate ; Glu : Glutamate ; Gln : Glutamine.

	: 1 H-MRS included studies
	Study	Population (nb)	Controls (nb)	Female (%)	Age	ROI (Voxels cm 3 )	Metabolites		Main results
	Wood et al. 2003	UHR (30)	HC (21) ; FEP (56)	43	19.5	MTL (3,38) ; DLPFC (3,38)	NAA/Cr ; Cho/Cr ; NAA/Cho		DLPFC NAA/Cr and Cho/Cr ratio increase in UHR
	Jessen et al. 2006	UHR (9)	HC (31) ; SCZ (21)	55.6	28.7	MTL (17,5) ; mPFC (29) ; DLPFC (30)	NAA/Cr ; NAA/Cho ; Cho/Cr		mPFC and DLPFC NAA/Cr ratio decrease in UHR compare to HC
	Aydin et al. 2008	UHR (17)	HC (15) ; FES (14)	17.6	19.6	Corpus callosum (NA)	NAA ; Cr ; Cho ;		Decrease NAA concentration in UHR and FES compare to HC. Decrease NAA concentration in UHR compare to FES.
	Buyn et al. 2009	UHR (20)	HC (20)	45	21.8	mPFC (8) ; DLPFC (6) ; Thalamus (6)	NAA ; Cr ; Cho ; mI ; Glx		No difference between groups
	Stone et al. 2009	UHR (27)	HC (27)	52	25	mPFC (8); MTL (6); Thalamus (6)	Glu ; Glx ; Gln ; NAA ; Cho ; mI, Cr	Decrease thalamus Glu concentration and increase mPFC Gln in UHR
	Wood et al. 2010	UHR (66)	HC (29)	62	19.2	MTL (NA)	NAA ; Glx ; mI, Cr+PCr ; TMA	No analysis between UHR and HC. No difference between UHR-T and UHR-NT.
	Uhl et al. 2011	UHR (30)	HC (31) ; FES (24)	36.7	25.6	mPFC (8) ; MTL (8)	NAA/Cr ; Cho/Cr		No difference between groups
	Fuente-Sandoval et al. 2011	UHR (18)	HC (40) ; FEP (18)	22.2	19.6	NC (8) ; cerebellum (8)	Glu ; Glx ; NAA ; Cho ; mI ; Cr		Increase NC Glu concentration in UHR and FEP compare to HC.
	Bloemen et al. 2011	UHR (11)	HC (11)	NA	NA	MTL (8)	Glu		Decrease Glu concentration in UHR
	Fuente-Sandoval et al. 2013	UHR (19)	HC (26)	26.3	19.7	NC (8)	Glu		Increase Glu concentration in UHR-T compare to UHR-NT or HC
	Egerton et al. 2014	UHR (75)	HC (56)	45.3	23.3	mPFC (8) ; Thalamus (6)	Glu ; Glx ; NAA ; Cr ; mI ; Cho		Decrease thalamus Glu concentration in UHR
	Natsubori et al. 2014	UHR (24)	HC (26) ; SCZ chro (25)	50	21.7	mPFC (8)	NAA ; Glx ; mI ; Cho ; Cre	Decrease mPFC NAA and Glx concentrations in SCZ chro compare to UHR or HC
	Nenadic et al. 2015	UHR (31)	HC (42) ; FES (18)	45.2	23.7	MTL (3)	Glu ; NAA		Decrease MTL NAA in UHR and FES compare to HC
	Liemburg et al. 2016	UHR (16)	HC (36) ; SCZ chro (60) ; SCZ recent (31)	44	22.9	DLPFC (8)	Glx ; NAA		Decrease Glx concentration in SCZ chro compare to ohers groups.
	Menchikov et al. 2016	UHR (20)	HC (26)	0	20,2	mPFC (30)	GABA/Cr ; Glx/Cr ; GABA/NAA ; Glx/NAA ; Cr/NAA	mPFC GABA/Cr and GABA/Glx ration decrease in UHR.
	Fuente-Sandoval et al. 2016	UHR (23)	HC (24)	34.8	20.7	mPFC (18.8) ; NC (22.5)	GABA ; Glx ; NAA ; Cho ;

Acknowledgments

We would like to thank all the following authors for their kind collaboration: Dr Edith Liemburg (Department of Neuroscience, and BCN Neuroimaging Center, University of Groningen, University Medical Center Groningen, Groningen, the Netherlands) for providing seminal data for the study "Prefrontal NAA and Glx Levels in Different Stages of Psychotic Disorders: a 3T 1 H-MRS Study "; Dr Petr Menschikov (Emanuel Institute of biochemical Physics, Russian academy of sciences, ul. Kosygina 4, Moscow, 117977 Russia) for providing data for the studies " 1 H MRS and MEGA PRESS Pulse Sequence in the Studyof Balance of Inhibitory and Excitatory Neurotransmittersin the Human Brain of Ultra High Risk of Schizophrenia Patients ".

Funding

None

Declaration of interest

Bruno Romeo, Amelie Petillion and Catherine Martelli have no conflict of interest. Amine Benyamina has given talk for Lundbeck, Mylan, Merck-Serono and Bristol-Myers Squibb and is a member of Indivior board. GABA ; Glu ; Gln ; Glx No difference in basal ganglia concentration. Decrease Glu in first degree relatives and in SCZ vs HC in occipital cortex. Decrease GABA in SCZ in occipital cortex Nb : number, ROI : region of interest ; UHR : ultra-high risk ; HR: high risk; HC : healthy controls ; SCZ chro : chronic schizophrenia ; SCZ recent : recent schizophrenia ; FES : first episode of schizophrenia ; FEP : first episode of psychosis ; UHR-T: ultra-high risk with a transition to schizophrenia; UHR-NT: ultra-high risk with no transition; MTL : medial temporal lobe ; mPFC : medial prefrontal cortex ; DLPFC : dorsolateral prefrontal cortex; WMPFC : white matter prefrontal cortex ; NC : caudate nucleus ; NAA : N-acetyl aspartate ; GABA : acide γ amino-butirique ; Cr : creatine ; Cho : choline ; mI : myo-inositol ; Glx : glutamine+glutamate ; Glu : Glutamate ; Gln : Glutamine