Significant outcomes
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∙ l-Theanine ameliorated positive symptoms in schizophrenia.
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∙ l-Theanine improved sleep quality in schizophrenia.
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∙ l-Theanine stabilised glutamatergic concentration in the brain.
Limitations
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∙ This was an open-label study.
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∙ The sample size was relatively small.
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∙ The observation period was 8 weeks; therefore, we do not know the long-term effect of l-theanine.
Introduction
Schizophrenia is a complex disorder characterised by symptoms such as delusions, hallucinations, disorganised communication, poor planning, reduced motivation, and blunted affect. N-methyl-d-aspartate (NMDA) receptor antagonists such as phencyclidine and ketamine induce symptoms that closely resemble those of schizophrenia, which suggests altered NMDA-glutamatergic function in schizophrenia (Reference Tsai, Van Kammen, Chen, Kelley, Grier and Coyle1). A previous 1.5-tesla (T) 1H magnetic resonance spectroscopy (MRS) study found higher levels of glutamine in the left medial prefrontal region of never-treated patients with schizophrenia compared with healthy volunteers (Reference Bartha, Williamson and Drost2). Another study using 4.0-T 1H MRS found increased levels of glutamine in the left anterior cingulate and thalamic regions of never-treated, first-episode patients with schizophrenia compared to healthy volunteers (Reference Théberge, Bartha and Drost3). In addition, one study showed increased levels of Glx (glutamine+glutamate) in the medial prefrontal cortex of unmedicated patients with schizophrenia compared to controls (Reference Kegeles, Mao and Stanford4). A review of 1H MRS studies concerning glutamate in schizophrenia showed an overall increase in the glutamine levels of patients with schizophrenia at the early phase of the disease (Reference Marsman, Van Den Heuvel, Klomp, Kahn, Luijten and Hulshoff Pol5).
In contrast, a chronic schizophrenia study showed decreased levels of glutamine and glutamate in the anterior cingulate (Reference Théberge, Al-Semaan and Williamson6), and Glx in the medial prefrontal cortex (Reference Natsubori, Inoue and Abe7). We found that patients with chronic schizophrenia with psychotic exacerbation showed increased Glx in the inferior parietal region (Reference Ota, Ishikawa and Sato8). Although 1H MRS does not selectively measure synaptic glutamate, these findings suggest that brain glutamate abnormalities may be a major neurochemical contributor to the development and exacerbation of schizophrenia.
l-Theanine (N-ethyl-l-glutamine) was originally found in green tea. l-Theanine accounts for ~50% of the total amino acids in green tea leaves. It comprises about 1–2% of the total dry weight of the green tea leaves, and the median amount of l-theanine per cup of green tea is 8–30 mg (Reference De Mejia, Ramirez-Mares and Puangpraphant9). Interestingly, l-theanine has a chemical structure that is similar to that of glutamate and affects glutamatergic neurotransmission (Reference Nathan, Lu, Gray and Oliver10–Reference Kakuda, Hinoi, Abe, Nozawa, Ogura and Yoneda12). Growing evidence suggests that l-theanine has several psychotropic effects; our previous study revealed that l-theanine attenuated K-801-induced deficits in prepulse inhibition (PPI) in rats (Reference Wakabayashi, Numakawa and Ninomiya13). l-Theanine improved the PPI in healthy humans (Reference Ota, Wakabayashi and Matsuo14), and several studies showed that l-theanine has an influence on mood (Reference Lu, Gray and Oliver15–Reference Unno, Tanida and Ishii18). It was reported that l-theanine increases sleep quality and satisfaction without increasing sleep duration or causing wake-up grogginess (Reference Lyon, Kapoor and Juneja19,Reference Ozeki, Juneja and Shirakawa20). Among them, one schizophrenia study showed the improvement of the positive and general symptoms of schizophrenia by the administration of l-theanine (Reference Ritsner, Miodownik and Ratner16). However, to the best of our knowledge, no study has assessed the in vivo effect of l-theanine on certain brain chemicals in humans.
Aims of the study
The aims of the present study were two-fold: to examine the possible effect of l-theanine on symptoms and sleep quality in chronic schizophrenia patients and to observed any changes in chemical mediators, including glutamate + glutamine, in the brain by using 1H magnetic resonance spectroscopy.
Material and methods
Subjects
Data were collected between December 2011 and July 2013. Outpatients who were treated by the authors and those who voluntarily responded to our poster announcement in the Hospital were recruited. The subjects were 17 chronic schizophrenia outpatients who fulfilled the Diagnostic and Statistical Manual of Mental Disorders (4th edition) (DSM-IV) (21) criteria for schizophrenia. Additionally, 22 age- and sex-matched healthy subjects (male/female 11/11, mean age=41.9±14.9 years) were recruited from the community through local magazine advertisements and our website announcement for a study of MRS metabolites.
Research psychiatrists with board certification (M.O., H.H., and T.T.) made the diagnoses and rated the symptom severity using the Positive and Negative Syndrome Scale (PANSS) (Reference Kay, Opler and Fiszbein22). Healthy subjects were interviewed for enrollment by research psychiatrists using the Japanese version of the Mini-International Neuropsychiatric Interview (Reference Otsubo, Tanaka and Koda23,Reference Sheehan, Lecrubier and Sheehan24). Sleep quality was assessed by the Pittsburgh Sleep Quality Index (PSQI) (Reference Buysse, Reynolds, Monk, Berman and Kupfer25). Participants were excluded if they had a history of central nervous system disease or severe head injury.
After the study was explained to the subjects, written informed consent was obtained for participation in the study from every subject. This study was approved by the Ethics Committee of the National Center of Neurology and Psychiatry, Japan.
Drug treatment and test schedule
The baseline clinical assessment (PANSS and PSQI) and MRS (scan 1) were conducted before the l-theanine treatment. Then 250 mg/day of l-theanine (Suntheanine, Taiyo Kagaku Co. Ltd, Yokkaichi, Japan) was added to ongoing antipsychotic medication for 8 weeks. The second clinical assessment and MRS (scan 2) were performed immediately after the completion of the 8-week trial. Other medications were essentially kept unchanged during the trial period.
MRS data acquisition
MR imaging was performed on a Magnetom Symphony 1.5-T (Siemens, Erlangen, Germany). Single-voxel 1H spectra, recommended to use by the widespread analysis tool ‘LCModel’ (Reference Provencher26), were acquired with a point-resolved pulse sequence (‘PRESS’, repetition time (TR) =1500 ms, echo time (TE)=30 ms, 1024 points, 1000 Hz spectral width, 160 averages water-suppressed and 30 averages without water suppression) from a 1.5×1.5×2.5 cm voxel placed in the left middle frontal white matter region and a 1.5×2.5×1.5 cm voxel in the left inferior parietal white matter region because these are regarded as the areas associated with the psychopathology of schizophrenia (Reference Rowland, Spieker, Francis, Barker, Carpenter and Buchanan27), and because there are sufficient white matter volumes at these sites to put regions of interest (ROIs) large enough to get adequate signal-to-noise ratios. The voxels were positioned by the scanner operator based on anatomical landmarks. The scan took ~10 min per region. Voxels were placed to maximise the white matter volume and minimise grey matter and cerebrospinal fluid. Global and local shimming were performed before the 1H-MRS sequence.
MRS analysis
Water-suppressed spectra were analysed using ‘LCModel’ (Reference Provencher26), a fully automated, commercially available curve-fitting software program that uses a least-squares analysis method for estimating metabolite concentrations in the millimolar range. The quantification model included the following metabolites: N-acetylaspartate (NAA), Glx, inositol, and glycerophosphocholine+phosphocholine. For each spectrum, the area under each peak was normalised to the unsuppressed water peak (corrected for water T1 relaxation) (Reference Barker, Soher, Blackband, Chatham, Mathews and Bryan28), yielding metabolite concentrations. The fitting quality of each spectrum is shown as the per cent standard deviation (SD), and those with SD values over 20% were excluded from analysis.
Statistical analysis
The differences in PANSS and PSQI scores between before and after the l-theanine treatment were compared using the paired t-test. We evaluated the differences in the metabolic concentrations of NAA, Glx, inositol and glycerophosphocholine+phosphocholine in the left frontal and inferior parietal white matter in schizophrenic patients before and after the l-theanine treatment using paired t-tests. With respect to Glx, we calculated the % ratio of the Glx levels before and after the l-theanine treatment, then analysed the effects of the l-theanine treatment by partial correlation using the age and sex as covariates. We calculated the % ratio by the formula:
$$\,\%\,{\rm ratio}={\rm Glx \ in\ scan\ 2}/{\rm Glx\ in\ scan\ 1}.$$
We also evaluated the relationships between the Glx levels and the psychiatric symptoms and the antipsychotic medication by generalised estimating equations using the Glx as dependent variable, and the dose of antipsychotic drug and PANSS score as covariates.
Statistical analyses were performed using the SPSS Statistics for Windows 22.0 software program (SPSS Japan, Tokyo, Japan).
Results
Nine male and eight female outpatients with chronic schizophrenia participated in this study. Their mean age was 40.6±12.3 years and their mean education years were 13.4±2.1. The mean chlorpromazine equivalent dose of antipsychotic medication was 958.5±516.6 mg/day (29,Reference Inagaki and Inada30). The results of the clinical assessments, MRS data before and after l-theanine treatment and the MRS data of the healthy subjects are shown in Table 1. There were significant differences in the PANSS total score (t=2.38, p=0.030) and the PSQI total score (t=3.01, p=0.008) between pre- and post-treatment with l-theanine. With respect to the PANSS scores, we found a nominal improvement in the positive score after the 8-week administration of l-theanine (t=2.47, p=0.025; Table 1).
Clinical characteristics of the schizophrenic patients
MRS, magnetic resonance spectroscopy; NAA, N-acetyl-aspartate; PANSS, Positive and Negative Syndrome Scale; WM, white matter.
* Significant difference between pre- and post-treatment after multiple comparisons.
As for MRS data, there were no significant differences in any metabolite between scans 1 and 2 (Fig. 1; Table 1). The relationships between Glx in scan 1 and the % ratio for Glx after the l-theanine treatment are shown in Fig. 2. There were significant negative correlations between Glx at baseline and the % ratio for Glx at the frontal (p=0.003, partial correlation coefficient=−0.72) and inferior parietal (p=0.010, partial correlation coefficient=−0.64) regions. We found no relationships between the Glx in the frontal and parietal regions and the clinical characteristics (Table 2).
Relation between the glutamate + glutamine (Glx) in scans 1 and 2. There were no significant differences in Glx between scans 1 and 2 in the frontal region (a) or the inferior parietal region (b).
Relations between glutamate + glutamine (Glx) in scan 1 and the % ratio for Glx after 8-week l-theanine treatment. There were significant negative correlations between Glx at the baseline (scan 1) and the % ratio in Glx following the l-theanine treatment in the frontal (a) and inferior parietal (b) regions.
Estimated effects of clinical characteristics on glutamate + glutamine
PANSS, the Positive and Negative Syndrome Scale.
* Chlorpromazine equivalent dose.
Discussion
We found that the 8-week add-on l-theanine treatment significantly reduced the PANSS scores in schizophrenia patients who were under treatment with antipsychotic medication. MRS revealed that l-theanine affected the concentration of Glx in the frontal and inferior parietal regions. Interestingly, there were significant negative correlations between Glx at baseline and the % ratio in Glx at the frontal and inferior parietal regions. To our knowledge, this is the first study that obtained evidence of the effect of l-theanine on Glx, which is related to glutamatergic neurotransmission in humans.
The therapeutic effects of l-theanine against the schizophrenic symptoms observed in the present study are consistent with those of previous studies (Reference Ritsner, Miodownik and Ratner16,Reference Miodownik, Maayan and Ratner17). In addition, our finding of a positive effect of l-theanine on sleep supports the results of preceding studies (Reference Lyon, Kapoor and Juneja19,Reference Ozeki, Juneja and Shirakawa20).
l-Theanine, which has a chemical structure that is similar to that of glutamate, affects glutamatergic neurotransmission (Reference Nathan, Lu, Gray and Oliver10–Reference Kakuda, Hinoi, Abe, Nozawa, Ogura and Yoneda12). The effects of l-theanine on schizophrenia symptoms and Glx concentrations in the brain observed in the present study might be attributable to its chemical structure. It is known that l-theanine has weak affinities for kainate, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid, and NMDA receptors (Reference Nathan, Lu, Gray and Oliver10,Reference Yokogoshi and Terashima31). On the other hand, l-theanine was reported to be able to suppress the excitotoxic release of glutamate derived from the Gln/glutamate cycle through the inhibition of Gln incorporation in glutamatergic neurons in a particular situation (Reference Kakuda, Hinoi, Abe, Nozawa, Ogura and Yoneda12).
In a previous study, we focused on the effect of l-theanine on PPI, a measure of sensorimotor gating that is known to be impaired in schizophrenia (Reference Braff and Geyer32,Reference Kunugi, Tanaka, Hori, Hashimoto, Saitoh and Hironaka33), and we observed the enhancement of PPI at particular doses of l-theanine (Reference Ota, Wakabayashi and Matsuo14). It is possible that l-theanine exerts its effects, at least in part, through a partial agonistic-like action on the glutamatergic system. Another study evaluated the dopamine synthesis capacities at resting condition and after the oral administration of a single dose of the partial agonist antipsychotic aripiprazole, revealing a significant negative correlation between the baseline and aripiprazole-induced changes in dopamine synthesis capacities (Reference Ito, Takano and Arakawa34). These findings suggested that aripiprazole may have a stabilising effect on the dopamine synthesis capacity. Similarly, l-theanine may have a stabilising effect on the glutamatergic neurotransmission.
Several studies examined glutamatergic changes in first-episode schizophrenia (Reference Bartha, Williamson and Drost2,Reference Théberge, Bartha and Drost3,Reference Théberge, Williamson and Aoyama35), chronic schizophrenia (Reference Théberge, Al-Semaan and Williamson6–Reference Ota, Ishikawa and Sato8,Reference Van Elst, Valerius and Büchert36,Reference Pakkenberg, Scheel-Kruger and Kristiansen37), and individuals at high risk for schizophrenia (Reference Tibbo, Hanstock, Valiakalayil and Allen38). The results obtained at each clinical stage and the clinical severity indicate that the glutamatergic metabolites of schizophrenia change in a course-dependent manner. l-Theanine was shown to be a safe and well-tolerated medication (Reference Ritsner, Miodownik and Ratner16), and the stabilising effect of l-theanine on glutamatergic neurotransmission could be of significant benefit in clinical practice.
The present study has several limitations. First, this was an open-label study and thus subject to bias in clinical assessments, although the MRS findings are free from such bias. Second, the sample size was relatively small, and thus subject to type II errors. Our results reached statistical significance, however, future studies with larger numbers of subjects are necessary to verify the present findings. Thirdly, the observation period was only 8 weeks and we therefore do not yet know the long-term effects of l-theanine.
Acknowledgement
This research was funded by an unrestricted research grant provided by the Taiyo Life Insurance Himawari Foundation, Tokyo, Japan. Contributors: M. Ota designed the study and wrote the first draft of the manuscript. H. Hori, K. Hattori and T. Teraishi collected the data. C. Wakabayashi, N. Sato, H. Ozawa, T. Okubo and H. Kunugi managed the analyses. All authors contributed to and have approved the final manuscript.
Conflicts of Interest
This research was funded by an unrestricted research grant provided by the Taiyo Life Insurance Himawari Foundation, Tokyo, Japan. H.O. and T.O. were the employees of Taiyo Kagaku Co. Ltd.; however, they only contributed to inform the other authors about l-theanine and to offer the supplement tablets. They did not analyse the clinical data or manage the analysis. This clinical study was registered in the UMIN Clinical Trials Registry (registration date 12/26/2011, registration no. 20111226-234711).
Supplementary material
To view supplementary material for this article, please visit http://dx.doi.org/10.1017/neu.2015.22
