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VOL22.NO. 4, 1996 An In Vivo Proton Magnetic Resonance Spectroscopy Study of Schizophrenia Patients by Jeff A. Stanley, Peter C. Williamson, Dick J. Drost, R. Jane Rylett, Tom J. Carr, Ashok Malta, and R. Terry Thompson Abstract The level of the ] H metabolites in the left dorsolateral prefrontal region of schizophrenia patients at different stages of illness were measured in vivo using a short echo time spectroscopy technique. During both the early onset and chronic stages, normal A/-acetylaspartate levels were observed, which suggests that these patients had no significant neuronal cell damage and/or loss. The in vivo measurements of glutamate in thefirst-episode,drugnaive patients failed to provide convincing evidence for the involvement of the glutamatergic system in the dorsolateral prefrontal region. Significant differences in the glutamine levels were observed in the acutely medicated and chronic patients; however, the interpretation of these differences requires further study. Schizophrenia Bulletin, 22(4): 597-609,1996. Evidence from several studies has recently implicated the excitatory glutamatergic system in the pathophysiology of schizophrenia (Deutsch et al. 1989; Carlsson and Carlsson 1990; Wachtel and Turski 1990; Uras and Cotman 1993). For example, phencyclidine (PCP) and ketamine, which are specific noncompetitive antagonists of the N-methylD-aspartate (NMDA) glutamate subtype receptors, induce psychosis resembling schizophrenia in normal controls and exacerbate psychosis in schizophrenia patients (Javitt and Zukin 1991; Krystal et al. 1994). In several postmortem brain studies, evidence of glutamatergic dysfunction in schizophrenia has been reported, including abnormalities in the binding density of glutamate sub- type receptors in various brain regions (Utas and Cotman 1993); decreased release of glutamate in synaptosomes prepared from cortex tissue of schizophrenia patients (Sherman et al. 1991); and a reduction of messenger ribonucleic acid (RNA) that encodes non-NMDA glutamate receptors in hippocampal tissue observed in patients compared with controls (Harrison et al. 1991). Obtaining information in vivo on the biochemistry of patients with schizophrenia can potentially contribute further understanding of the chemical pathology that gives rise to the glutamatergic dysfunctions observed in postmortem studies. Magnetic resonance spectroscopy (MRS) is a noninvasive and nondestructive technique that can provide such information (Bottomley 1989; Kauppinen et al. 1993). MRS can assess the viability of neuronal cells by quantifying the peptide A/-acetylaspartate (NAA) (Arnold et al. 1990; De Stefano et al. 1995), reliably quantify the in vivo relative concentration of the excitatory neurotransmitter glutamate (de Graaf and Bovee 1990; Provencher 1993; Stanley et al. 1995a); and quantify additional cerebral metabolites including glutamine, phosphocreatine plus creatine (PCr + Cr), and choline-containing compound (Chot). Moreover, the localized region(s) where the in vivo metabolic information is obtained may be as small as several cm3 (Ernst et al. 1989) and can be positioned in a specific area of the brain such as the dorsolateral prefrontal region. Consequently, phosphorus (31P) and proton Reprintrequestsshould be sent to Dr. PC Williamson, Dept. of Psychiatry, University Hospital, 399 Windermere Rd., Box 5339, London, Ontario, N6A 5A5, Canada. SCHIZOPHRENIA BULLETIN 598 ('H) MRS have been applied in several studies to investigate the biochemistry in vivo of patients with schizophrenia (O'Callaghan et al. 1991; Pettegrew et al. 1991; Calabrese et al. 1992; Sharma et al. 1992; Buckley et al. 1994; Nasrallah et al. 1994; Stanley et al. 1994,1995b; Renshaw et al. 1995). In this in vivo study, 'H metabolite levels from the dorsolateral prefrontal region of first-episode, drugnaive, and acute and chronic medicated schizophrenia patients were compared to metabolite levels from controls of comparable age, gender, education, and parental education levels. It was hypothesized that differences in levels of glutamate and /or glutamine would be evident if the glutamatergic system was involved in the pathophysiology of schizophrenia. Differences in levels of NA A in patients would suggest neuronal cell damage or loss in the prefrontal region. Examination of levels of metabolites before and after medication might indicate some of the metabolic effects of these agents. Methods Subjects. Twenty-nine patients with schizophrenia ranging in age from 16 to 49 years participated in this study. There were 13 firstepisode, drug-naive schizophrenia patients, of whom 11 were classified as paranoid and 2 as undifferentiated. The length of illness (the time between the onset of positive symptoms and the MRS examination) for the drug-naive patients ranged from 1 month to 6 years. None had been exposed to any antipsychotic medication before the MRS examination. However, within the 24 hours before assessment, seven patients had received 1 to 3 mg of lorazepam. The remaining patients included 12 acute medicated schizophrenia patients (of whom 8 had been previously examined'as first-episode, drug-naive patients) and 12 chronic medicated schizophrenia patients. The acute medicated patients with a mean length of illness of 2 ± 2 years included 10 classified as paranoid and 2 as undifferentiated. The chronic medicated patients group, with a mean length of illness of 17 ± 6 years, had six diagnosed as paranoid, two as undifferentiated, and four as residuals. All medicated patients were receiving antipsychotic medication. Eighteen of these patients were also on anticholinergic medication for side effects. The mean length of time on medication for the acute medicated patients was 14 ± 10 weeks. The diagnoses of all the patients were established with the Structured Clinical Interview for DSM-1H-R (SCID; Spitzer and Williams 1985) administered by a psychiatrist. The diagnoses of the drug-naive patients were reconfirmed with the treating psychiatrist 6 months after the MRS experiments were done. Both the Scale for the Assessment of Negative Symptoms (SANS; Andreasen 1984a) and the Scale for the Assessment of Positive Symptoms (SAPS; Andreasen 1984b) were also administered by a psychiatrist without any knowledge of the 'H MRS results. Education level was rated on a 4-point scale (1 = s grade 10; 2 = grade 11-13; 3 = 1-3 years college or university; 4 = more than 3 years college or university). Parental education ratings were evaluated for the most-educated parent; however, three were evaluated for the most-educated adoptive parent. Clinical information is summarized in table 1. The handedness of each subject was defined by the hand used to write and throw a ball. All were right-handed. Twenty-four nonschizophrenia controls ranging in age from 16 to 53 were recruited by advertisement. Each control subject was evaluated with the SCID by a psychiatrist. Controls were of gender, age, education level, parental education level, and handedness (all were right-handed) comparable to the patients. Subject characteristics of the control group are shown in table 1. Patients and controls were free of any history of head injury, drug or alcohol abuse, or serious medical illness based on the information provided during the SCID interview. No gross abnormalities were detected on routine clinical magnetic resonance (MR) images that were also collected as part of the study on each subject. None of the drug-naive patients, all of the medicated patients, and 13 of the 24 controls had undergone at least one previous magnetic resonance imaging scan. J H MRS. The in vivo 1H MRS experiments were conducted using a circularly polarized head coil on a whole body MR unit (Helicon SP system, Siemens AG, Erlangen, Germany) with a static magnetic field of 1.5 Tesla. The STEAM sequence (stimulated echo acquisition mode; Frahm et al. 1989,1990) with an echo time of 20 ms provided the single voxel localization technique. The mixing time interval was 30 ms. Three Gaussian-shaped radio frequency pulses (CHESS pulses; Haase and Frahm 1985), separated by spoiler gradients, were placed at the beginning of the STEAM sequence to suppress the large contribution of the water MR signal. The 'H MR signal was obtained from 2 x 2 x 2 cm3 volume of interest (VOI) located in the left dorsolateral prefrontal region of the subjects. The VOI was positioned 599 VOL22.NO. 4, 1996 Table 1. Subject characteristics Age Group Gender (yrs) Control group Schizophrenia patient groups First-episode, drug-naive Acute medicated Chronic medicated 24M/0F 32±11 2 11 M/2F 26±7 10M/2F 26 ±7 11 M/1 F 41 ±5 Parental Education education level1 level1 Length of Illness (yrs) Length of time on medication (yrs) SANS score _ _ 37 ±13 28 ± 14 26 ±12 3.0 ± 0.92 2.5 ± 0.92 _ 2.3 ± 0.9 2.4 ± 0.7 2.6 ± 0.8 2.3 ± 0.8 2.2 ± 0.8 2.0 ± 0.9 2.0 ±1.9 — 2.5 ± 2.4 0.8 ±1.9 16 ±8.0 17 ±7.0 SAPS score _ 30+ 11 9± 16 8 ± 11 Note.—Values are mean 11 standard deviation; M = males; F = females; SANS •= Scale for the Assessment of Negative Symptoms (Andreasen 1984a); SAPS - Scale for the Assessment of Positive Symptoms (Andreasen 1984b). 'Education level: 1 « * grade 10. 2 • grade 11-13, 3 • 1-3 years college or university, 4 = more than 3 years college or university. 2 ln a two-tailed f-test, these values are not significantty different when compared with combined patient groups. with a set of sagittal and coronal 'H MR images as shown in figure 1. The dashed lines in figure 1 represent the anterior (a), posterior (b), lateral (c), and medial (d) surfaces of the three dimensional VOI (white box). The magnetic field homogeneity was maximized with a global head shim followed by a localized shim on the VOI. The interpulse repetition time was 1,500 ms, and 450 acquisitions were averaged. For each water suppressed 'H spectrum acquired, a water unsuppressed ] H spectrum was also collected (55 acquisitions) by setting the amplitudes of the CHESS pulses to zero voltage. Further details of the spectroscopy protocol are discussed by Stanley et al. (1995a). Spectral Processing. The data files were coded such that the operator had no knowledge of the subject's status. A time domain deconvolution technique, QUALITY (de Graaf et al. 1990), was first applied to the MR signal to restore the spectral lineshapes to pure Lorentzian. The time domain signal was then multiplied by a Lorentzian-to-Gaussian transformation to enhance the apparent spectral resolution (Ferrige and Lindon 1978). The MR signal was then zero filled, Fourier transformed, and phased with Oth order and 1st order (< one dwell period). No spline function was applied to the baseline. The data were processed on a personal computer using the NMR-286 software (Soft Pulse Software, Box 504, Guelph, Ontario, N1H 6K9, Canada). There is a direct relationship between the area under a spectral peak and the concentration of the metabolite associated with that peak. Therefore, Gaussian functions were fitted to each spectral peak between 1.88 and 3.45 ppm (parts per million) and the areas under the functions were used to calculate the 'H metabolite levels. To resolve the issue of quantifying complex 'H spectra with multiple peaks that overlap each other (de Graaf and Bovee 1990; Provencher 1993; Stanley et al. 1995a), a priori knowledge was incorporated into a frequency domain nonlinear least-squares-fitting algo- rithm (Marquardt 1963). The a priori knowledge included information on chemical shifts, relative amplitudes, and linewidths of the peaks for each metabolite. The quantified 1 H metabolites included NAA, glutamate, glutamine, gamma-aminobutyric acid (GABA), aspartate, Nacetylaspartylglutamate (NAAG), PCr + Cr, Chot, glucose, taurine, scy//o-inositol, and two macromolecule resonances at 2.12 and 2.9 ppm (Kauppinen et al. 1992; Behar and Ogino 1993; Behar et al. 1994; Stanley et al. 1995a). To calculate the relative metabolite levels, the integral value of the phased water peak from the unsuppressed water 'H spectrum, which represents the total MR visible water content in the VOI, was used as an internal standard to normalize the metabolite peak areas (Christiansen et al. 1993). The quantified areas of the spectral peaks were not corrected for any attenuation due to spin-spin (TJ or spin-lattice (T,) relaxation; therefore the quoted relative metabolite levels have arbitrary units. Complete details on the implementation of the a priori knowledge SCHIZOPHRENIA BULLETIN 600 Figure 1. The location of the volume of Interest (VOI) in the left dorsolateral prefrontal region ables from the multiple regression analysis. For the eight patients who were examined twice (first as drugnaive then as acute medicated patients) a two-tailed, paired f-test was also used to compare the SANS and SAPS scores and metabolite levels between pre- and on-medication measures. Pearson product moment correlations of age, length of illness, length of time on medication, equivalent dose of chlorpromazine, and SAPS and SANS scores along with levels of NAA, glutamate, glutamine, PCr + Cr, and Chot were evaluated for the controls and the combined patients. Results C) Sagittal and corona) 1H magnetic resonance Images with the 2 x 2 x 2 cm3 VOI (the white box) positioned In the left dorsolateraJ prefrontal region. The dotted lines in (a) and (b) represent position of the corona) Images In (c) and (d) while the dotted lines In (c) and (d) represent the position of the sagittal Images in (a) and (b). into the fitting routine and on testing the efficacy of quantifying in vivo 'H MR spectra has been reported by Stanley et al. (1995a) Statistical Analysis. A two-tailed ttest was performed to determine any significant differences in age, education level, and parental education level when comparing the combined patients with the control group. The quantified ] H MR parameters were modeled in a stepwise multiple regression analysis with subject group (i.e., patient and control), age, gender, education level, and parental education level as independent parameters. This method enabled us to compare differences in the NAA, glutamate, glutamine, PCr + Cr, and Cho, levels between patient and control groups while adjusting for the covariables of age, gender, education level, and parental education level. Probability values of < 0.05 were considered statistically significant and a threshold value of p = 0.2 was used to enter or remove independent vari- A typical processed in vivo TH spectrum acquired from the left dorsolateral prefrontal region of a control is shown in figure 2. The spectrum contains no dominating broad spectral resonances that underlie the baseline noise over the 1.88 to 3.45 ppm spectral region. The signal-to-noise ratio (i.e., the signal of the NAA peak at 2.02 ppm over the root mean square of the noise) is approximately 80 (figure 2), which is adequate to reliably quantify the spectrum (Stanley et al. 1995a). The quantification of this spectrum is displayed in figure 2b as a sum of all spectral peaks of the metabolites superimposed on the acquired spectrum. The difference between the two is shown as the residual plot in figure 2b. Instructions to perform specific physical or psychological tasks were not given to the subjects during the collection of the data. Therefore, in general, the observed metabolite levels reflect steady-state values at rest. The absolute difference in metabolite levels of NAA, glutamate, glutamine, PCr + Cr, and Cho, between the three patient groups and controls is shown in figure 3. In the controls, the coeffi- 601 VOL. 22, NO. 4, 1996 Figure 2. An In vivo 1H magnetic resonance (MR) spectrum collected with the STEAM sequence suppressed water peak a) /^'<'W»^oAA/V»/*-vV'>***V«^^ y\ 12.0 10.0 4.0 8.0 3.6 6.0 3.2 -2.0 2.8 2.4 2.0 1.6 -4.0 1.2 -6.0 0.8 Chemical Shift (ppm) Figure (a) contains a typical processed in vrvo 1H MR spectrum from the left dorsolateral prefrontal region acquired with the STEAM (stimulated echo acquisition mode) sequence (TE = 20 ms) and (b) shows the same spectrum with the frequency region expanded. The result of modeling the spectrum with a priori The spectral peaks are N-acetyiaspartate (NAA), glutamate (G)u), glutamlne (Qln), gamma-amino-butyric acid (GABA), NAACHJ (methyl resonance from the NAA molecule), aspartate (Asp), phosphocreatjne plus creatine (PCr + Cr), chollne-containing compounds (Cho,), glucose (Gte), myixnosrto) (myo-lns), scyflo-inositol (scyflo-lns), and taurine (Tau). ppm = parts per million. cients of variation for NAA, PCr + Cr, and Cho, were approximately 10 percent, and the coefficients of variation for glutamate and glutamine were 22 and 33 percent, respectively. Quantifying the metabolite levels of GABA, aspartate, NAAG, taurine, scy//o-inositol, and glucose was considered less reliable because their low in vivo concentration levels resulted in coefficients of variation i 30 percent (Stanley et al. 1995a), and therefore these ^H metabolites were not tested in the statistical analysis. There were no significant differences when comparing the ^H metabolite levels of the first-episode, drug-naive and acute medicated patients with the control group. The glutamate levels tended to be greater in the acute medicated patients compared with the controls; however, this difference did not reach significance (p = 0.10 where age and gender were the covariates). In the paired ttest for the eight drug-naive patients who were also examined as med- SCHIZOPHRENIA BULLETIN 602 Figure 3. Absolute metabolite level difference between schizophrenia patients and controls I— 5 r Controls ^SSI Drug-naive Acute medicated 2 Chronic medicated II \ B 0) -1 0) -2 o c CD (5 -3 Q -4 "o (/> < -5 L NAA Glu Gin' PCr+Cr Cho. NAA = AAacetytaspartate; Glu = glkitamate; Gin « glutamlne; PCr + Cr = phosphocreatine plus creatine; Cho, = choline-containing compounds; error bars are ± 1 standard deviation. 'Values are relative to that of the controls (i.e., mean metabolite level of patient - mean metabolite level of controls). 'Significantly different when comparing the chronic medicated patients and the controls (p » 0.013). icated patients, glutamine levels (p = 0.020) and SAPS scores (p = 0.006) were both significantly reduced in the on-medication measures compared with the premedication measures. When comparing the metabolite levels of the chronic medicated patients and the control group, the only significant difference was the increase in glutamine in the chronic medicated patients (p = 0.013 where age, gender, education level, and parental education level were the covariates). Age, education levels, and parental education levels of the patients with schizophrenia were not significantly different from the controls. These results are summarized in table 1 and figures 3 and 4. Combining the schizophrenia patients together, the length of illness, length of time on medication, equivalent dose of chlorpromazine, and SANS and SAPS scores were not significantly correlated (after applying a Bonferroni correction for multiple comparisons) with the levels of NAA, glutamate, glutamine, PCr + Cr, or ChOj, except for a significant positive correlation between the glu- VOL. 22, NO. 4, 1996 603 Figure 4. Palrwise comparison of the glutamine levels between the premedication and on-medication measurements of the eight schizophrenia patients 9 r § I -2. iE ca 2 2 O 33 CD > jg 2 <D 1 Pre-medication measurement On-medication measurement1 A solid line connects the premedteatlon and on-medlcatlon measurements of the same patient. 'On-medlcation measurements are significantly different than the premedication measurements (p = 0.020). famine level and the length of illness (r = 0.53, p = 0.0007, figure 5). The glutamine level and age correlation was not significant in the controls. Discussion Assessing Neuronal Damage and Loss. NAA levels in the left dorsolateral prefrontal region did not differ between any of the patient groups and the controls. In terms of the total concentration of free amino acids in mammals, NAA is second only to glutamate (Tallan 1957). This is reflected on the dominant spectral feature of NAA (figure 2) that gives rise to a reliable measure (Stanley et al. 1995a). While the function of NAA in the central nervous system (CNS) has not been elucidated (Birken and Oldendorf 1989), it has been established that NAA is found exclusively in mature neurons and neuronal processes (Matalon et al. 1988; Birken and Oldendorf 1989; Urenjak et al. 1993). Decreased NAA levels have been observed by MRS in numerous cerebral pathologies involving neuronal cell damage and loss (Arnold et al. 1990; Menon et al. 1990; Graham et al. 1992; Klunk et al. 1992; Cendes et al. 1994). Recently, recovery of NAA levels has been reported in patients with acute CNS damage (De Stefano et al. 1995). The absence of a difference in NAA levels in the left dorsolateral prefrontal region between schizophrenia patients and controls suggests there is no cell damage or loss in this part of the brain. A previous localized in vivo 'H MRS study of the left frontal region (Buckley et al. 1994) also observed no significant differences in the percent of NAA signal between schizophrenia patients (who included first- episode, drug-naive and medicated patients) and controls. However, when only male patients were tested, a 23 percent decrease in the NAA was observed compared with the male controls. The subjects used in this study were mostly males, so a gender effect could not be completely assessed. These findings are in contrast to other MRS findings in the temporal lobes, which show marked reductions in NAA in schizophrenia patients compared with controls (Buckley et al. 1994; Nasrallah et al. 1994; Renshaw et al. 1995). MRI volumetric studies have shown that deficits in gray matter have been less striking in the frontal lobes than in the temporal lobes (Breier et al. 1992; Zipursky et al. 1992; Buchanan et al. 1993). Involement of the Glutamatergic System in Schizophrenia. Based on in vitro studies, the glutamate concentration in the frontal cortex is approximately 9.0 mmol/kg wet weight (w. wt.) and the concentration of glutamine is approximately twofold to threefold smaller (Perry et al. 1971; Erecinska and Silver 1990). Using the identical acquisition and processing protocol as this study, our group has reported glutamate and glutamine concentration levels in the left dorsolateral prefrontal region of approximately 9.4 and 4.8 mmol/kg w. wt., respectively (Stanley et al. 1995a). This would suggest that the bulk of the in vivo concentration of SCHIZOPHRENIA BULLETIN 604 Figure 5. 12 Glutamine level versus length of illness r 10 c/T 'c • • o o o • 13 I _ o 8 - o CO • * * 1 CD _ l CD I* o 0 o iS * O o ' o 1 0 ^ 1 1 I 1 I . 0 . . 1 . . . . 1 . . . . 1 10 15 20 Length of Illness (years) . . . . 1 25 . . . . 1 30 Scatter plot of the glutamine levels of the drug-naive (*), acute medicated (O), and chronic medicated (0) patients. The solid Hne represents the regression Bne ( r » 0.53, p = 0.0007). glutamate and glutamine is observable with this short echo MRS technique. This observation is consistent with the 'H MRS study by Kauppinen and Williams (1991), in which 79 percent of the total glutamate concentration was estimated as MR-visible. Approximately 80 percent of the total glutamate concentration is found in glutamatergic neurons (large compartment) and approximately 2 to 20 percent in glial cells (small compartment) (Erecinska and Silver 1990). The larger glutamate compartment has been described as a slow turnover pool of glutamate (i.e., the metabolic pool) derived from glucose precursors, and the smaller glutamate compartment as a raster turnover pool of glutamate (i.e., the neurotransmitter pool) that serves as a substrate to glutamine (Erecinska and Silver 1990). Glutamine plays an important role in the recycling of the neurotransmitter glutamate in the smaller compartment (Nicholls and Attwell 1990). For instance, following the release of glutamate by calciumdependent exocytosis, excess glutamate is transported into glia and is subsequently converted to glutamine by glutamine synthetase. Following release from glial cells, glutamine may then enter the presynaptic neuron and serve as a precursor for glutamate by mitochondrial glutaminase. Overall, the localization of glutamine is found predominantly in glial cells (Erecinska and Silver 1990; 605 VOL. 22, NO. 4, 1996 Urenjak et al. 1993). In this study, the in vivo glutamate and glutamine levels observed in the left dorsolateral prefrontal region did not differ between the first-episode, drug-naive schizophrenia patients and controls. This finding suggests that there are no abnormalities in the metabolic and (with less confidence) the neurotransmirter pool of glutamate at the early-onset stage of schizophrenia before treatment. Additionally, this does not provide support for the involvement of the prefrontal glutamatergic system in schizophrenia. However, this study would have found significant differences for our sample size only if those differences had been greater than 20 percent because of the precision in quantifying the in vivo level of glutamate. In the chronic medicated patients, glutamine levels were, however, increased compared with controls. Since glutamine is predominantly in glial cells, increased glial cell volume in the chronic medicated patients is one possible interpretation of our data. However, the failure to find any differences on NAA levels would suggest that there is no change in the proportion of neurons to glial cells. Glutamine is directly associated with the recycling of the neurotransmirter glutamate (Nicholls and Attwell 1990). Recently, Pellerin and Magistretti (1994) have shown that the glucose uptake is directly dependent on the glutamate uptake into glia. It is unclear which metabolic process or processes are responsible for the increase observed in the steady-state level of glutamine. This alteration could be in keeping with the decreased glucose uptake observed in the prefrontal lobe of schizophrenia patients from positron emission tomography studies (Buchsbaum 1990) if glutamine was not being converted to glutamate in the neurons. Nevertheless, further investigation is required to determine the physiological significance of the observed increase in glutamine in these patients. Levels of glutamine in the combined patient group were correlated positively with the length of illness, while there was no age glutamine correlation from the controls. There also were no significant correlations between the equivalent dose of chlorpromazine or the length of time on medication. Considering only the 12 chronic medicated patients (i.e., the patient subgroup with the greatest range in length of illness), a stronger relationship between glutamine and length of illness was observed (r = 0.61, p = 0.035). This would suggest that the abnormal levels of glutamine have a stronger dependency on the progression of the illness than on the medication treatment. The decreased glutamate and increased glutamine levels that were reported in the first preliminary in vivo 'H MRS study on first-episode, drug-naive patients by Stanley et al. (1992) were not reproduced in this study. The sampling of a larger subject population and advancements made in the spectral quantification procedure may account for these discrepancies. In this study, a priori knowledge on the spectral peak arrangement for each metabolite was incorporated into the quantification fitting algorithm, increasing the precision and accuracy of our results (de Graaf and Bovee 1990; Provencher 1993; Stanley et al. 1995a). Effects of Treatment The 1H metabolites observed in the firstepisode, drug-naive patients and the acute medicated patients were not significantly different from the controls. Glutamate levels tended to be higher in the acute medicated patients, but this finding was not sta- tistically significant. However, of the eight first-episode, drug-naive patients who also participated as acute medicated patients, the glutamine levels were significantly decreased in the on-medication measurements compared with the premedication measurements. Additionally, during this acute on-medication period (i.e., with a mean length of time on medication of 14 ± 8 weeks), the SAPS scores also were significantly decreased. This finding would suggest that during the initial period of neuroleptic treatment, the antipsychotic medication, whose function is to block dopamine receptors, has altered directly or indirectly the steady-state level of glutamine in the prefrontal region. It may also suggest that the antipsychotic medication has influenced the glutamatergic system since glutamine is directly associated with the recycling of the neurotransmitter glutamate. There is little support in the literature for this interpretation. Antipsychotic drugs have been reported to affect the dopaminergic activity in the nigrostriatal and limbic structures but not the glutamatergic activity in the prefrontal region after acute and chronic administration in animals (Yamamoto and Cooperman 1994). However, Pehek and colleagues (1991) and Daly and Moghaddam (1993) found increased extracellular concentrations of glutamate in the prefrontal cortex following acute administration of antipsychotic drugs. Further in vivo MRS studies on larger patient populations are required to confirm the involvement of the glutamatergic system in the prefrontal region during acute administration of antipsychotic drugs. Limitations. The sagittal and coronal images in figure 1 suggest that within the dimension of the VOI, the SCHIZOPHRENIA BULLETIN 606 observed ] H MR signal is dominated by white matter compared with gray matter. The identical spectroscopy protocol was used in the repeated measures study, and the volume of gray matter for the 8 cm3 VOI was estimated at approximately 30 percent (Stanley et al. 1995a). If metabolite differences are only present in the gray matter of schizophrenia patients, then the observed intensity of the difference is reduced because of sampling relatively less gray matter volume. However, acquiring ] H spectra with a smaller VOI to reduce the partial white matter volume decreases the signal-to-noise ratio, which reduces the reliability of the quantification. To account for the ] H signal from macromolecules (i.e., signal from mobile proteins and polypeptides), two macromolecule resonances were incorporated in the fitting routine (Stanley et al. 1995a). These two resonances are dominant spectral peaks that are observed in spectra of macromolecules in vivo (Behar et al. 1994). In the controls, the two macromolecule resonances combined accounted for 3 ± 2 percent of the total signal compared with 28 ± 2 percent for NAA. We do acknowledge that there are inaccuracies associated with this approach that may result in overestimating the levels of certain metabolites (Stanley et al. 1995a). The in vivo T, (spin-lattice relaxation time) of glutamine is approximately 2,100 msec (Hanicke et al. 1993), which is greater than the interpulse repetition time used and implies that the observed glutamine signal is partially saturated. A decrease in T, of glutamine could account for the observed increase; however, the T, would have to decrease significantly by about 34 percent to observe the 30 percent increase in glutamine level. Differences in the T2 values of the metabolites between patients and controls could not be detected because the relative linewidths (i.e., T2 values) of the metabolites were held constant in the quantification technique (Stanley et al. 1995a). References Andreasen, N. Scale for the Assessment of Negative Symptoms (SANS). Iowa City, IA: The University of Iowa, 1984a. Andreasen, N. Scale for the Assessment of Positive Symptoms (SAPS). Iowa City, IA: The University of Iowa, 1984b. Arnold, D.L.; Matthews, P.M.; Francis, G.; and Antel, J. Proton magnetic resonance spectroscopy of human brain in vivo in the evaluation of multiple sclerosis: Assessment of the load of disease. Magnetic Resonance in Medicine, 14:154-159,1990. Behar, K.L., and Ogino, T. Characterization of macromolecule resonances in the 'H NMR spectrum of rat brain. Magnetic Resonance in Medicine, 30:38-44,1993. Behar, K.L.; Rothman, D.L.; Spencer, D.D.; and Petroff, O.A.C. Analysis of macromolecule resonances in H NMR spectra of human brain. Magnetic Resonance in Medicine, 32:294- 302,1994. Birken, D.L., and Oldendorf, W.H. N-acetyl-L-aspartic acid: A literature review of a compound prominent in ^-NMR spectroscopic studies of brain. Neuroscience and Biobehavioral Reviews, 13:23-31,1989. Bottomley, P.A. Human in vivo NMR spectroscopy in diagnostic medicine: Clinical tool or research probe? Radiology, 170:1-15,1989. Breier, A.; Buchanan, R.W.; Elkashef, A.; Munson, R.C.; Kirkpatrick, B.; and Gellad, F. Brain morphology and schizophrenia: A magnetic resonance imaging study of limbic, prefrontal cortex, and caudate structures. Archives of General Psychiatry, 49:921-926,1992. Buchanan, R.W.; Breier, A.; Kirkpatrick, B.; Elkashef, A.; Munson, R.C.; Gellad, F.; and Carpenter, W.T., Jr. Structural abnormalities in deficit and nondeficit schizophrenia. American Journal of Psychiatry, 150:59-66, 1993. Buckley, P.F.; Moore, C; Long, H.; Larkin, C; Thompson, P.; Mulvany, F.; Stack, J.P.; Ennis, J.T.; and Waddington, J.L. H-l magnetic resonance spectroscopy of the left temporal and frontal lobes in schizophrenia: Clinical, neurodevelopmental, and cognitive correlates. Biological Psychiatry, 36:792-800,1994. Buchsbaum, M.S. The frontal lobes, basal ganglia, and temporal lobes as sites for schizophrenia. Schizophrenia Bulletin, 16(3):379-389,1990. Calabrese, G.; Deicken, R.F.; Fein, G.; Merrin, E.L.; Schoenfeld, F.; and Weiner, M.W. 31-Phosphorus magnetic resonance spectroscopy of the temporal lobes in schizophrenia. Biological Psychiatry, 32:26-32,1992. Carlsson, M., and Carlsson, A. Schizophrenia: A subcortical neurorransmitter imbalance syndrome? Schizophrenia Bulletin, 16(3):425-432,1990. Cendes, F.; Andermann, F.; Preul, M.C.; and Arnold, D.L. Lateralization of temporal lobe epilepsy based on regional metabolic abnormalities in proton magnetic resonance spectroscopic images. Annals of Neurology, 35:211-216,1994. Christiansen, P.; Henriksen, O.; Stubgaard, M.; Gideon, P.; and Larsson, H.B.W. In vivo quantification of brain metabolites by 'H-MRS using water as an internal standard. Magnetic Resonance Imaging, 11:107-118,1993. 607 VOL. 22, NO. 4, 1996 Daly, D.A., and Moghaddam, B. Actions of dozapine and haloperidol on extracellular levels of excitatory amino acids in the prefrontal cortex and striatum of conscious rats. Neuroscience Letters, 152:61-64,1993. de Graaf, A. A., and Bovee, W.M.M.J. Improved quantification of in vivo ^H NMR spectra by optimization of signal acquisition and processing and by incorporation of prior knowledge into the spectral fitting. Magnetic Resonance in Medicine, 15:305-319,1990. de Graaf, A.A.; van Dijk, J.E.; and Bovee, W.M.M.J. QUALITY: Quantification improvement by converting lineshapes to the Lorentzian type. Magnetic Resonance in Medicine, 13:343-357,1990. De Stefano, N.; Matthews, P.M.; and Arnold, D.L. Reversible decreases in N-acetylaspartate after acute brain injury. Magnetic Resonance in Medicine, 34:721-727,1995. Deutsch, S.I.; Mastropaolo, J.; Schwartz, B.L.; Rosse, R.B.; and Morihisa, J.M. A glutamatergic hypothesis of schizophrenia: Rationale for pharmacotherapy with glycine. Clinical Neuropharmacology, 12:1-13,1989. Erecinska, M., and Silver, I.A. Metabolism and role of glutamate in mammalian brain. Progress in Neurobiology, 35:245-2%, 1990. Ernst, T.; Hennig, J.; Ott, D.; and Friedburg, H. The importance of the voxel size in clinical 'H spectroscopy of the human brain. NMR in Biomedicine, 2:216-224,1989. Ferrige, A.G., and Lindon, J.C. Resolution enhancement in FT NMR through the use of a double exponential function, journal of Magnetic Resonance, 31:337-340,1978. Frahm, J.; Bruhn, H.; Gyngell, M.L.; Merboldt, K.D.; Hanicke, W.; and Sauter, R. Localized high-resolution proton NMR spectroscopy using stimulated echoes: Initial applications to human brain in vivo. Magnetic Resonance in Medicine, 9:79-93,1989. Frahm, J.; Michaelis, T; Merboldt, K.D.; Bruhn, H.; Gyngell, M.L.; and Hanicke, W. Improvements in localized proton NMR spectroscopy of human brain and water suppression, short echo times, and 1 ml resolution. Journal of Magnetic Resonance, 90:464- 473,1990. Graham, G.D.; Blamire, A.M.; Howseman, A.M.; Rothman, D.L.; Fayad, P.B.; Brass, L.M.; Petroff, O.A.; Shulman, R.G.; and Prichard, J.W. Proton magnetic resonance spectroscopy of cerebral lactate and other metabolites in stroke patients. Stroke, 23:333-340,1992. Haase, A., and Frahm, J. Multiple chemical-shift-selective NMR imaging using stimulated echoes. Journal of Magnetic Resonance, 64:94-102,1985. Hanicke, W.; Michaelis, T; Merboldt, K.D.; and Frahm, J. On the use of a fully automated data analysis method for in vivo MRS: Metabolite concentrations and relaxation times from proton spectra of human brain. In: Proceedings of the 12th Annual Meeting of the Society of Magnetic Reso- nance in Medicine. Berkeley Springs, CA: Society of Magnetic Resonance in Medicine, 1993. p. 977. Harrison, P.J.; McLaughlin, D.; and Kerwin, R.W. Decreased hippocampal expression of a glutamate receptor gene in schizophrenia. Lancet, 337:450-152,1991. Javitt, D.C., and Zukin, S.R. Recent advances in the phencyclidine model of schizophrenia. American Journal of Psychiatry, 148:1301-1308,1991. Kauppinen, R.A.; Kokko, H.; and Williams, S.R. Detection of mobile proteins by proton nuclear magnetic resonance spectroscopy in the guinea pig brain ex vivo and their partial purification. Journal of Neurochemistry, 58:967-974,1992. Kauppinen, R.A., and Williams, S.R. Nondestructive detection of glutamate by ] H nuclear magnetic resonance spectroscopy in cortical brain slices from the guinea pig: Evidence for changes in detectability during severe anoxic insults. Journal of Neurochemistry, 57:1136-1144,1991. Kauppinen, R.A.; Williams, S.R.; Busza, A.L.; and van Bruggen, N. Applications of magnetic resonance spectroscopy and diffusion-weighted imaging to the study of brain biochemistry and pathology. Trends in Neuroscience, 16:88-95,1993. Klunk, W.E.; Panchalingam, K.; Mossy, J.; McClure, R.J.; and Pettegrew, J.W. W-acetyl-L-aspartate and other amino acid metabolites in Alzheimer's disease brain: A preliminary proton nuclear magnetic resonance study. Neurology, 42:1578-1585, 1992. Krystal, J.H.; Karper, L.P.; Seibyl, J.P.; Freeman, G.K.; Delaney, R.; Bremner, J.D.; Heninger, G.R.; Bowers, M.B.; and Charney, D.S. Subanesthetic effects of the noncompetitive NMDA antagonist, ketamine, in humans. Archives of General Psychiatry, 51:199-214,1994. Marquardt, D.W. An algorithm for least-squares estimation of non-linear parameters. Society of Industrial and Applied Mathematics Journal, 11:431441,1963. Matalon, R.; Michals, K.; Sebesta, D.; Deanching, M.; Gashkoff, P.; and Casanova, J. Aspartoacylase deficiency and N-acetylaspartic aciduria in patients with Canavan disease. American Journal of Medical Genetics, 29:463-471,1988. Menon, D.K.; Baudouin, C.J.; Tomlinson, D.; and Hoyle, C. Proton MR spectroscopy and imaging of the SCHIZOPHRENIA BULLETIN 608 brain in AIDS: Evidence of neuronal loss in regions that appear normal with imaging. Journal of Computer Assisted Tomography, 14:882-885,1990. Nasrallah, H.A.; Skinner, T.E.; Schmalbrock, P.; and Robitaille, P.M. Proton magnetic resonance spectroscopy (H-l MRS) of the hippocampal formation in schizophrenia: A pilot study. British Journal of Psychiatry, 165:481-485,1994. Nicholls, D., and Attwell, D. The release and uptake of excitatory amino acids. Trends in Pharmacological Sciences, 11:462-468,1990. O'Callaghan, E.; Redmond, O.; Ennis, R.; Stack, ].; Kinsella, A.; Ennis, J.T.; and Waddington, J.L. Initial investigation of the left temporoparietal region in schizophrenia by 31P magnetic resonance spectroscopy. Biological Psychiatry, 29:1149-1152,1991. Pehek, E.A.; Yamamoto, B.K.; and Meltzer, H.Y. The effects of clozapine on dopamine, 5-HT, and glutamate release in the rat medial prefrontal cortex. Schizophrenia Research, 4:323, 1991. Pellerin, L., and Magistretti, P.J. Glutamate uptake into astrocytes stimulates aerobic glycolysis—A mechanism coupling neuronal activity to glucose utilization. Proceedings of tlie National Academy of Sciences of the United States of America, 91:1062510629,1994. Perry, T.L.; Hansen, S.; Berry, K.; Mok, C; and Lesk, D. Free amino acids and related compounds in biopsies of human brain. Journal of Ncumdiemistry, 18:521-528, 1971. Pettegrew, J.W.; Keshavan, M.S.; Panchalingam, K.; Strychor, S.; Kaplan, D.B.; Tretta, M.G.; and Allen, M. Alterations in brain high-energy phosphate and membrane phospholipid metabolism in first-episode, drug-naive schizophrenics: A pilot study of the dorsolateral prefrontal cortex by in vivo phosphorus 31 nuclear magnetic resonance spectroscopy. Archives of General Psychiatry, 48:563-568,1991. Provencher, S.W. Estimation of metabolite concentrations from localized in vivo proton NMR spectra. Magnetic Resonance in Medicine, 30:672-679,1993. Renshaw, PR; Yurgelun-Todd, D.A.; Tohen, M.; Gruber, S.; and Cohen, B.M. Temporal lobe proton magnetic resonance spectroscopy of patients with first-episode psychosis. American Journal of Psychiatry, 152:444—446, 1995. Sharma, R.; Venkatasubramanian, P.N.; Barany, M.; and Davis, J.M. Proton magnetic resonance spectroscopy of the brain in schizophrenic and affective patients. Schizophrenia Research, 8:43-49,1992. Sherman, A.D.; Davidson, A.T.; Baruah, S.; Hegwood, T.S.; and Waziri, R. Evidence of glutamatergic deficiency in schizophrenia. Neuroscience Letters, 121:77-80,1991. Spitzer, R., and Williams, J. Structured Clinical Interview for DSM-III-R. New York, NY: New York Psychiatric Institute, 1985. Stanley, J.A.; Drost, D.J.; Williamson, PC.; and Thompson, R.T. The use of a priori knowledge to quantify short echo in vivo 'H MR spectra. Magnetic Resonance in Medicine, 34:17-24, 1995a. Stanley, J.A.; Williamson, PC; Drost, D.J.; Canr, T.J.; Rylett, R.J.; Malla, A.; and Thompson, R.T. An in vivo study of the prefrontal cortex of schizophrenic patients at different stages of illness via phosphorus magnetic resonance spectroscopy. Archives of General Psychiatry, 52:399-406,1995b. Stanley, J.A.; Williamson, PC; Drost, D.J.; Carr, T.J.; Rylett, R.J.; and Merskey, H. "In Vivo Proton MRS in Never Treated Schizophrenics." Presented at the Annual Meeting of the American Psychiatric Association, Washington, DC, May 1992. Stanley, J.A.; Williamson, PC; Drost, D.J.; Carr, T.J.; Rylett, R.J.; and Thompson, R.T. Membrane phospholipid metabolism and schizophrenia: An in vivo P-31-MR spectroscopy study. Schizophrenia Research, 13:209215,1994. Tallan, H.H. Studies on the distribution of N-acetyl-L-aspartic acid in brain. Journal of Biological Chemistry, 224:41^5,1957. Uras, J., and Cotman, C.W. Excitatory amino acid receptors in schizophrenia. Schizophrenia Bulletin, 19(l):105-117,1993. Urenjak, J.; Williams, S.R.; Godown, D.G.; and Noble, M. Proton nuclear magnetic resonance spectroscopy unambiguously identifies different neural cell types. Journal ofNeurosciencc, 13:981-989,1993. Wachtel, H., and Turski, L. Glutamate: A new target in schizophrenia? Trends in Pharmacological Sciences, 11:219-220,1990. Yamamoto, B.K., and Cooperman, M. A. Differential effects of chronic antipsychotic drug treatment on extracellular glutamate and dopamine concentrations. Journal of Ncuroscience, 14:4159-4166,1994. Zipursky, R.B.; Lim, K.O.; Sullivan, E.V.; Brown, B.W.; and Pfefferbaum, A. Widespread cerebral gray matter volume deficits in schizophrenia. Archives of General Psydiiatry, 49:195205,1992. Acknowledgments This study was supported by grant MH-50768 from the National Institute of Mental Health and grant MT-12078 609 VOL. 22, NO. 4, 1996 from the Ontario Mental Health Foundation, Medical Research Council of Canada. The authors thank Mr. John Potwarka for computer software development assistance. The Authors Jeff A. Stanley, Ph.D., is a Senior Announcement Research Fellow, Laboratory of Neurophysics, University of Pittsburgh Medical Center, Pittsburgh, PA. Peter C. Williamson, M.D., is Associate Professor of Psychiatry; Dick J. Drost, Ph.D., is Associate Professor of Medical Biophysics; R. Jane Rylett, Ph.D., is Professor of Physiology; Tom J. Carr, M.D., is Clinical Assistant Pro- fessor of Diagnostic Radiology and Nuclear Medicine; Ashok Malla, M.D., is Associate Professor of Psychiatry; and R. Terry Thompson, Ph.D., is Associate Professor of Medical Biophysics, Nuclear Medicine and Magnetic Resonance, St. Joseph's Health Centre, University of Western Ontario, London, Ontario, Canada. The Pacific Clinics and the Alliance for the Mentally 111 will sponsor a conference entitled Integrating Approaches to Intervention With Persons Who Have Schizophrenia to be held in Pasadena, California, January 30 and 31 and February 1,1997. This meeting focuses on exercises, demonstrations, and role plays to teach usable skills from the neurobiological, recovery, and family psychoeducational approaches to schizophrenia. A model for integrating the interventions into a unified view of person, illness, and family is presented. For more information about the conference, please contact: Sarah Shatford Pacific Clinics 909 South Fair Oaks Pasadena, CA 91105 Telephone: 818-577-6697