Altered NMDA receptor activity and glutamate signalling might underlie the pathogenesis of both schizophrenia and depression in subgroups of patients. In schizophrenia, pharmacological modelling, post‐mortem and imaging data suggest reduced NMDA signalling. In contrast, recent clinical trials demonstrating the efficacy of the NMDA antagonist ketamine in severely depressed patients suggest increased NMDA receptor signalling. We conducted a proof‐of‐concept study to assess whether there is any in vivo evidence for an inverse association in depression and schizophrenia with respect to the NMDA receptor function. For this purpose, we used a translational approach, based on findings from animal studies that NMDA receptor is a key mediator of arginine vasopressin (AVP) release into the bloodstream. Using hypertonic saline to increase plasma osmolality ( P Osm ) and thereby induce AVP release, as done in animal studies, we found that in depressed patients the NMDA receptor‐mediated AVP release induced by hypertonic saline infusion was significantly increased [0.24 (0.15) pg ml −1 mosmol −1 , P < 0.05] compared with schizophrenia patients [0.07 (0.07) pg ml −1 mosmol −1 ]. Slopes for healthy control subjects were 0.11 (0.09) pg ml −1 mosmol −1 which was less than the depressed group. These findings are consistent with implicated NMDA receptor‐related abnormalities in depression and schizophrenia in subgroups of patients and provide the first in vivo evidence of this dichotomy.

In response to hyperosmotic challenge, depressed subjects showed increased P [AVP] response compared with healthy control and schizophrenic subjects. However, schizophrenic subjects were not different from healthy control subjects in this small sample. The ‘ P [AVP] response to P Osm ’ is a suitable biomarker to distinguish depressed versus schizophrenic patients when used with psychiatric screening. This is the first objective physiological measure for schizophrenia or depression.

Can the change in plasma arginine vasopressin concentration ( P [AVP] ) in response to osmotic stimulation ( P Osm ) serve as a biomarker for NMDA receptor signalling in schizophrenia and depression and thereby distinguish between these mental illnesses?

Introduction The NMDA receptor is a major subtype of glutamate receptor that mediates fast synaptic transmission in the CNS. Reduced NMDA receptor (NMDAR) signalling has long been implicated in the pathogenesis of schizophrenia, as reviewed in detail elsewhere (Olney & Farber, 1995; Krystal et al. 2002; Arnsten, 2011), and thought to underlie both the cognitive dysfunction and symptom profile observed in this disorder. Of note, NMDAR antagonists provide a pharmacological animal model of schizophrenia for drug development (Jentsch & Roth, 1999) and were found to increase symptom severity when administered to patients (Lahti et al. 1995, 2001; Malhotra et al. 1997). In contrast to schizophrenia, some depressed patients show a remarkable improvement following NMDA antagonist administration (Berman et al. 2000; Zarate et al. 2006), raising the possibility of increased NMDA receptor signalling in depression. In the pathogenesis of depression, the recognition of a glutamate‐mediated ‘neuroplasticity hypothesis’ is relatively recent (Pittenger & Duman, 2008; Sanacora et al. 2012) and supported by several lines of evidence, including post‐mortem (Ongur et al. 1998; Rajkowska et al. 1999; Miguel‐Hidalgo et al. 2000, 2002; Cotter et al. 2001; Bowley et al. 2002; Hamidi et al. 2004; Khundakar et al. 2011), imaging (Yuksel & Ongur, 2010) and clinical trials data (Berman et al. 2000; Zarate et al. 2006; DiazGranados et al. 2010). As both schizophrenia and depression are heterogeneous, it is highly likely that the suspected abnormal NMDAR signalling in these disorders represent subsets of patients. For example, the administration of the NMDAR antagonist ketamine affects some schizophrenic patients more than others, as evidenced by high variability of the response pattern among patients (Lahti et al. 2001). Likewise, the antidepressant effect of ketamine is also variable, because one‐third of treatment‐resistant patients with depression do not respond to ketamine (Moaddel et al. 2015), and ketamine may have a preferential positive effect in depressed patients with high anxiety levels (Ionescu et al. 2014). Therefore, it would be simplistic to propose that NMDAR abnormality solely would explain the entire pathophysiology of these disorders. As yet, in vivo assessment of the NMDAR in humans is not possible. Thus, the need for development of a blood‐based biomarker that represents the NMDAR signalling in schizophrenia and depression is essential. Such a biomarker would enable us to evaluate the vulnerability of individuals towards schizophrenia versus depression, especially in early illness when there is significant overlap in the symptoms and consequently diagnostic uncertainty. Schizophrenia most often manifests during adolescence. Initial signs and symptoms are typically non‐specific, ranging from subtle changes in behaviour to depressive symptoms, sometimes including normal adolescent behaviour, such as academic decline and social withdrawal. In addition, a biomarker could be used to identify patients with abnormal NMDAR signalling among those with established diagnoses in order to offer targeted treatments using glutamatergic agents. One approach to a blood‐based biomarker would be to examine hormones that are regulated by the NMDAR in the CNS. As hormones are directly released into the bloodstream, assessment of such hormones in peripheral blood could serve as a biomarker of the CNS disorders that are associated with NMDAR signalling. Arginine vasopressin (AVP) is one such hormone whose release is regulated by the NMDAR. Arginine vasopressin is synthesized in the cell bodies of magnocellular neurosecretary cells, which are large neurons located in the hypothalamic nuclei that extend to the posterior pituitary and terminate on the pituitary portal system. Magnocellular neurosecretory cells express functional NMDA receptors and release AVP in response to glutamatergic activation (McKinley et al. 1992; Richard & Bourque, 1995; Bourque, 1998; Swenson et al. 1998; Leng et al. 2001; Sladek, 2004). Glutamatergic synapses that terminate on the magnocellular neurosecretary cells contain NMDA, α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) and metabotropic glutamate receptors (Panatier, 2009). Physiological studies have demonstrated that the plasma AVP (P [AVP] ) response to plasma osmolality (P Osm ) is sensitive to the function of the NMDA receptor. When rats are infused with hypertonic saline, P Osm increases linearly during the infusion, as in humans, producing a linear P [AVP] response to P Osm . However, when rats are pretreated with the non‐competitive NMDA receptor antagonist MK‐801 and then administered hypertonic saline, the slope of the P [AVP] response to P Osm decreases (Onaka & Yagi, 2001), suggesting that the NMDA receptor signalling modulates the release of AVP. Therefore, the purpose of the present study was to determine whether the P [AVP] response to P Osm could serve as a biomarker for the NMDA receptor signalling in schizophrenia and depression. As mentioned earlier, although resting P [AVP] is low, AVP release is sensitive to increases in P Osm as slight as 5 mosmol kg−1 (2–3%). Moreover, above this threshold for osmotic AVP stimulation, the P [AVP] –P Osm relationship is tight and linear (Calzone et al. 2001; Stachenfeld et al. 2001). We have safely increased the P Osm by administration of hypertonic saline infusion (HSI) in a number of populations (Stachenfeld et al. 1996, 1998, 2001; Calzone et al. 2001). We therefore propose that because the release of AVP is NMDA receptor dependent, we can determine the extent of NMDA receptor activity in humans by measuring the osmotically controlled release of AVP using HSI. Specifically, we would expect the magnitude of the linear slope of the P [AVP] –P Osm relationship to serve as an indicator of the NMDA receptor activity. We hypothesized that the lower NMDA receptor signalling in schizophrenics would produce a flatter HSI‐induced P [AVP] –P Osm slope compared with healthy individuals without mental illness consistent with disrupted NMDAR function. In contrast, we hypothesized that the greater NMDA receptor signalling in depression would produce a steeper HSI‐induced P [AVP] –P Osm slope compared with healthy control subjects without mental illness, and steeper than in schizophrenics.

Methods Ethical approval All subjects gave written informed consent to participate in the study, which conformed to the guidelines contained in the Declaration of Helsinki and received prior approval by the Human Investigation Committee of Yale School of Medicine (approval no. 0910005875) and the Human Subjects Subcommittee of the VA Connecticut Healthcare System (Project No: HGB0006). Upon establishment of eligibility, subjects were scheduled for a single test day at the John Pierce Laboratory, New Haven, CT, USA. Overview We recruited men and women with a Diagnostic and Statistical Manual of Mental Disorders, 4th Edition (DSM‐IV) diagnosis of schizophrenia or unipolar depression and healthy control subjects via referrals from the outpatient clinics of Yale School of Medicine, VA Medical Center and the local clinics in the New Haven area. All subjects received a brief telephone screening for basic eligibility, such as age, sex and medical history, after which they were brought in for the consenting process and screening procedures. These procedures included a history and physical examination, laboratory assessment, ECG and a structured interview. Trained research personnel conducted The Structured Clinical Interview (SCID) using the DSM‐IV, patient and non‐patient versions for diagnostic assessment. Trained research assistants determined subjects’ diagnoses by conducting structured interviews, which is standard of practice in psychiatric diagnoses, and also by the American Board of Psychiatry and Neurology certified psychiatrist (prinicipal investigator of the study). Protocol Subjects In order to reduce individual variability in hydration levels, the subjects were instructed to eat only a prescribed low‐fat breakfast, drink 5 ml kg−1 water and refrain from alcohol and caffeine for 12 h before testing. Subjects arrived at the laboratory at 0800 h. Hydration state was assessed by urine specific gravity. If specific gravity was >1.02, the subjects were given another 5 ml kg−1 of water to drink, followed by a 60 min control period. The subjects were seated in a comfortable chair for catheter placement and took the semi‐recumbent position that was maintained throughout the experiment (ambient temperature = 28°C). During a 60 min resting period, the subjects were instrumented for the measurement of plasma variables (using a 22 gauge Teflon i.v. cannula), heart rate and arterial blood pressure (Colin Medical Instruments, Komaki, Japan) and blood oxygenation using a pulse oximeter. Testing procedures We infused hypertonic saline (3.0% NaCl) at 0.1 ml kg−1 min−1 for 120 min to examine the osmotic regulation of AVP in the subjects. We obtained blood samples at 15, 25, 35, 45, 60, 75, 90, 105 and 120 min during the infusion, along with measurements of blood pressure, heart rate and thirst sensation. After the infusion, we collected a second urine sample and, following a 30 min stabilization period, the subjects drank 15 ml kg−1 of water over 30 min. After water ingestion, the subjects rested quietly for 60 min, at the end of which a third urine sample was collected. During this recovery period, blood samples were obtained at 30 min intervals, and blood pressure and heart rate were monitored at 10 min intervals. Haematocrit, haemoglobin, osmolality and concentrations of AVP and electrolytes were determined in all blood samples. Volume, osmolality and electrolyte concentrations were determined in all urine samples for a comprehensive characterization of the water regulation abnormalities in schizophrenia that are dependent on the P [AVP] response to P Osm . Blood analysis Blood samples were separated immediately into aliquots and first analysed for haemoglobin and haematocrit. A second aliquot was transferred to a heparinized tube, and all other aliquots were placed in prechilled tubes containing EDTA. The tubes were centrifuged and the plasma was taken off the heparinized samples and analysed for sodium, potassium and osmolality. The EDTA samples were analysed for concentrations of AVP. Plasma and urine sodium and potassium were measured by flame photometry and P Osm by freezing point depression. The P [AVP] was determined after extraction from plasma on octadecylsilane cartridges (SEP‐PAK C18; Waters Corporation, Milford, MA, USA). Extracted samples were assayed using a disequilibrium assay, with the extracts incubated with the antiserum at 4°C for 72 h, followed by the addition of 125I‐labelled AVP (New England Nuclear, Boston, MA, USA). Bovine albumin‐coated charcoal was used for separation of free and antibody‐bound labelled AVP. This assay is highly specific for AVP, with the antiserum prepared against a lysine vasopressin–thyroglobin conjugate, and has a sensitivity of 0.6 pg ml−1. Extraction recovery of AVP was determined using plasma spiked with a known concentration of AVP (Peninusula Laboratories, Belmont, CA, USA). The recovery sample was extracted and analysed along with the subjects’ samples. The extraction recovery was 89%. Intra‐ and interassay coefficients of variation for the midrange standards (2.3 pg ml−1) were 9.2 and 4.6%, respectively. Thirst ratings We assessed thirst by asking the subject to make a mark on a line rating scale in response to the question, ‘How thirsty do you feel now?’ The line is 175 mm in length and is marked ‘not at all’ on one end and ‘extremely thirsty’ at the 125 mm point. We told subjects that they could mark beyond the extremely thirsty point if they wished and might even extend the line if they felt it necessary. This method was developed by Marks et al. (1988) and has been used with great success in the evaluation of several sensory systems. We have found an extraordinarily good relationship between the perception of thirst and plasma osmolality during HSI and dehydration in young volunteers.

Statistical analysis All data were first summarized descriptively, and distributional assumptions for continuous variables were tested using Kolmogorov–Smirnov tests and normal probability plots. When normality was not satisfied, transformations were considered, and if it was not possible to normalize the data, non‐parametric methods were used. Demographics were compared between groups using ANOVA for continuous measures and Fisher's exact tests for categorical variables. Mixed effects models were used to test for differences by group and time in all repeatedly measured continuous outcomes. Group [healthy control (HC), depression (Dep) and Schizophrenia (Sch)], time (available time points) and the interaction between group and time were the predictors in the models. Subject was the clustering factor. The best‐fitting variance–covariance matrix for each model was selected based on the Schwarz Bayesian criterion. Tests of simple effects and pairwise comparisons were performed to explain significant interactions in the models. Residual plots were used to evaluate the fit of the model to the data. Sensitivity analyses with sex as a fixed covariate were also performed. The relationship between P [AVP] and plasma osmolality was assessed using a mixed model, with AVP as the dependent variable, group (HC, Dep and Sch), plasma osmolality and the interaction between group and plasma osmolality as predictors. This analysis was restricted to the time points during the infusion. For this analysis, a model with random intercept and slope for osmolality fitted the best. This model allows the between‐subject heterogeneity in slopes to be taken into account and accounts for the correlations of repeated measures within individual. Pairwise slope comparisons between the different groups were performed to explain interaction effects. As a sensitivity analysis, a general linear model and a non‐parametric Kruskal–Wallis test were also used to compare the slopes for the relationship between P [AVP] and P Osm between groups. The dependent variables in these models were individual slopes calculated using ordinary linear regression based on all available P [AVP] and P Osm data for each individual during the infusion.

Results There were more depressed females and more schizophrenic males, but the difference was not statistically significant (Fisher's exact test, P = 0.19; Table 1) There were no significant differences in ethnicity (Fisher's exact test, P = 0.37) or age [F(2,22) = 2.80, P = 0.08] across groups. Baseline values of physiological measures are given in Table 2. All subjects tolerated the procedures well, with no adverse events. Table 1. Demographic characteristics of the subjects Characteristic Healthy control group (n = 10) Schizophrenia group (n = 7) Depression group (n = 8) Age [years; mean (SD)] 31 (9) 39 (13) 42 (8) Sex (male; %) 70 88 43 Ethnicity (%) Caucasian 40 38 71.4 African American 30 63 14.3 Hispanic 20 0 14.3 Asian 10 0 0 Table 2. Baseline physiological measures Parameter Healthy control group Schizophrenia group Depression group Height (cm) 173.5 (8) 177.8 (12) 166.6 (12) Weight (kg) 76 (11) 102 (28) 77 (22) P [AVP] (pg ml−1) 0.98 (0.32) 1.08 (0.35) 1.09 (0.15) P Osm [mosmol kg−1] 283 (4) 282 (4) 281 (5) U Osm [mosmol kg−1] 417 (347) 668 (337) 355 (229) Mixed model analysis of P [AVP] over time (up to 240 min) showed only a significant time effect [F(13,253) = 5.70, P < 0001]. The interaction between group and time was not significant [F(26,253) = 1.16, P = 0.27], indicating that in all groups P [AVP] increased significantly during the infusion and then decreased following the infusion. Next, we examined whether there were any significant differences in P Osm by group and time. There was only a significant time effect [F(13,262) = 79.22, P < 0001]. The interaction between group and time was not significant [F(26,262) = 1.29, P = 0.17]. Plasma osmolality increased during the infusion and then decreased following the infusion in all groups. Restricting this analysis to the 120 min, likewise, we found only a significant time effect [F(9,186) = 139.18, P < 0001]. The interaction between group and time was not significant [F(18,186) = 1.16, P = 0.30]. Plasma osmolality increased during the infusion for all groups (Fig. 1). Figure 1. Increase in plasma osmolality over time as stimulated by hypertonic saline infusion Open in figure viewer PowerPoint Abbreviations: Dep, depression; HC, healthy control; and Sch, schizophrenic. The interaction between P Osm and group was statistically significant [F(2,162) = 4.94, P = 0.008]. There was also a significant main effect of P Osm [F(1,22) = 47.94, P < 0001]. As hypothesized, the slope for subjects with schizophrenia was the smallest [fixed slope estimate = 0.07, 95% confidence interval (CI): (0.00, 0.14)] and for the depressed subjects, the largest [slope = 0.22, 95% CI: (0.15, 0.30)]. The slope for normal control subjects was in between [slope = 0.11, 95% CI: (0.05, 0.17)]. See Fig. 2 for a healthy control representative subject and Fig. 3 for all data. The slope for the depressed group was significantly steeper than the slopes for healthy control subjects and for schizophrenics [slope difference = 0.11, 95% CI: (0.02, 0.21), P = 0.02 and slope difference = 0.15, 95% CI: (0.02, 0.21), P = 0.0025, respectively]. The slopes for healthy control subjects and schizophrenics were not significantly different [slope difference = 0.04, 95% CI: (−0.05, 0.13), P = 0.35]. The lowest slope in the healthy control group (Fig. 3) was outside of the range of what is expected for healthy control subjects. This subject was also unusual in his response to the thirst rating, reporting no thirst at baseline and no change throughout the experiment. We kept this subject in the database because we could not rule out possibilities such as anatomical defects or pituitary lesions. Low slope values such as this could also be caused by central diabetes insipidus, although patients with central diabetes insipidus have intact thirst sensation, unlike this subject (Verbalis, 2003). Slopes and calculated threshold values are given in Table 3. Figure 2. Response of plasma concentration of arginine vasopressin (P [AVP] ) to increasing plasma osmolality (P Osm ) over time as stimulated by hypertonic saline infusion in a healthy individual in our laboratory Open in figure viewer PowerPoint The slope of the P [AVP] response to P Osm is calculated by the slope of the regression line of the P [AVP] –P Osm relationship. The calculated osmotic threshold for P [AVP] represents the maximal P Osm value at which P [AVP] could be 0.0 pg ml−1. Here, this subject's calculated threshold was 275 mosmol kg−1. Figure 3. Individual P [AVP] –P Osm slopes for all three groups Open in figure viewer PowerPoint Table 3. Slope and threshold values in the groups Healthy control group (n = 10) Schizophrenia group (n = 8) Depression group (n = 7) Slope 0.11 (0.09) 0.07 (0.07) 0.24 (0.15)*† Threshold 258.7 (55.6) 238.7 (99.3) 279.6 (6.6) Urine volume over time showed a statistically significant time effect [F(3,66) = 12.77, P < 0001], but there were no group‐related differences. For urine osmolality, both the group‐by‐time interaction [F(6,66) = 2.33, P = 0.04] and the time effect were statistically significant [F(3,66) = 4.15, P = 0.009]. There were differences among the three groups only at 1 h postinfusion (P = 0.01). Healthy control subjects had higher urine osmolality than depressed and schizophrenic subjects at the 180 min time point. No significant between‐group differences were observed at the other time points. Only for the thirst ratings, we used non‐parametric analysis because of their non‐normal distribution (Brunner & Langer, 2002). Only the time effect was statistically significant [ANOVA type statistic (4.19) = 17.86, P < 0001]. No group‐related differences over time were observed [ANOVA type statistic (8.26) = 1.47, P = 0.16]. As would be expected, there was a significant main effect of time [F(5,106) = 6.31, P < 0001] for the heart rate measures. No group‐related differences were observed [F(10,106) = 1,19, P = 0.30]. We observed a significant main effect of time [F(5,107) = 6.81, P < 0001] and a significant main effect of group [F(2,22) = 4.32, P = 0.03] for systolic blood pressure. Depressed subjects had lower systolic blood pressure than healthy control subjects and schizophrenics; however, the group‐by‐time interaction was not significant [F(10,107) = 1.48, P = 0.16]. Controlling for sex differences, for haematocrit values there was a significant main effect of time [F(2,36) = 152.61, P < 0001] and a significant interaction between group and time [F(4,36) = 2.88, P = 0.04]. However, none of the between‐group comparisons by time point was statistically significant (P > 0.17).

Discussion Longitudinal studies demonstrated that only 35% of individuals with early symptoms convert to schizophrenia over 2.5 years (Cannon et al. 2008). A similar percentage of patients develop mood disorders, and some may grow out of these early symptoms. This presents major ethical challenges for early intervention in both disorders because of false‐positive and false‐negative diagnoses by current interview‐based assessments. Thus, to distinguish schizophrenia from depression and even normal adolescent behaviour, an objective assessment of the NMDA receptor signalling is crucial for diagnostic guidance and early intervention. We conducted this proof‐of‐concept study to assess the central NMDA receptor signalling in healthy control subjects, subjects with schizophrenia and subjects with depression. Based on animal findings, we stimulated the activity of the hypothalamic NMDA receptor by increasing plasma osmolality via i.v. hypertonic saline and measured the concentration of AVP in the peripheral blood over time. Consistent with our hypothesis, we demonstrated that in response to the hyperosmotic challenge, subjects with depression showed a significantly increased AVP response compared with both healthy control subjects and schizophrenic subjects. However, although subjects with schizophrenia showed a decreased AVP response compared with the depressed group, they did not differ significantly from the healthy controls. Nonetheless, the lowest values were observed in the schizophrenia group, which may represent a subgroup of patients with reduced NMDAR signalling. Hence, our findings support the use of ‘P [AVP] response to P Osm ’ as a potential biomarker to distinguish depressed versus schizophrenic patients when used together with psychiatric screening when there is diagnostic uncertainty. A potential caveat is that there was some overlap across the groups, so although this measure is a potential biomarker, it requires further testing and should be used along with an overall diagnostic battery. Nonetheless, our data will certainly contribute to our understanding of the pathophysiology of both schizophrenia and depression. It is well recognized that both schizophrenia and depression are heterogeneous conditions, and there may be different underlying scientific underpinnings among patients in each disorder (Andreasen & Carpenter, 1993; Sullivan et al. 2000; Kennedy et al. 2003; Hasler et al. 2004). Both schizophrenia and depression also show differences in course, treatment response, functionality and prognosis. In this context, our method, for the first time, identifies subgroups of patients within the schizophrenia group with indication of low NMDAR activity and subgroups of patients within the depression group high NMDAR activity. Therefore, this method has potential to identify a particular subtype of patients in each disorder, and thus is a crucial step forward. Previously, HSIs have been administered to schizophrenic patients in order to study the underlying physiological abnormalities in a subset of schizophrenic patients with polydipsia (Goldman et al. 1996). In this previous study, polydipsic patients demonstrated the lowest AVP response to hypertonic stimulation. To our knowledge, this protocol has not been used in depressed patients previously. Importantly, no prior studies have examine the P [AVP] response to P Osm in these groups compared with each other or compared with a healthy control group. In the present study, our schizophrenic subjects were not polydipsic, as assessed by clinical interview and laboratory values. In addition, their urine osmolality were somewhat higher than that the other groups at baseline. NMDA antagonists are used to provide pharmacological animal models of schizophrenia in drug development. Rodents pretreated with NMDA antagonists also show a decreased AVP response to hypertonic stimulation (Onaka & Yagi, 2001; Yamaguchi & Watanabe, 2002). These observations are consistent with reduced NMDAR activity in schizophrenia, at least in some patients. It could be argued that the subunit composition of the hypothalamic NMDA receptor may be significantly different from that in those brain regions implicated in the disorder, such as the prefrontal cortex. However, the composition of the hypothalamic NMDA receptor includes all four subunits of the NMDA receptor heteromer (NR2A, NR2B, NR2C and NR2D), in addition to the obligatory NR1 (Doherty & Sladek, 2011). The similarity of the hypothalamic NMDA receptor composition to most central NMDA receptors provides reassurance that our approach is representative of the implicated NMDA receptor dysfunction in schizophrenia. Finally, to our knowledge, there are a handful of hormones that may be regulated by stimulation of the NMDA receptor in addition to AVP, such as gonadotrophin‐releasing hormone (Yin et al. 2007), prolactin (D'Aniello et al. 2000) and leptin (Carbone et al. 2005). No direct examinations have been made between these neuroendocrine systems in depression and schizophrenia with respect to the NMDA receptors, but this might be an interesting area for future research. We found a significantly greater P [AVP] response to a change in P Osm in the depressed group compared with both healthy control subjects and subjects with schizophrenia. To the extent that the response to the NMDA antagonist ketamine in depression suggests overactive NMDAR signalling, our finding is consistent with this interpretation in a subgroup of subjects, as three of the seven depressed subjects showed the highest responses. Also of interest is the spread of these slopes within the depressed group (Fig. 3). Based on our understanding of the NMDAR, we suspect that the members of this group in the upper range represent patients with refractory depression, although this hypothesis requires further testing. Another limitation of our results is that we had a small sample size; hence, the chance of false‐positive or false‐negative findings cannot be overlooked. Replication of our results in a larger sample is needed. Potential medication effects also deserve comment, because all of our patients were treated with antipsychotics in the schizophrenia group and antidepressants in the depressed group, and this constitutes a major limitation in our study. Water regulation abnormalities in schizophrenia have been documented before the availability of antipsychotics (Hoskins, 1933; Sleeper, 1935) and are also observed in antipsychotic‐free patients (Hariprasad et al. 1980). In 1933, before the discovery of antipsychotics, Hoskins intensively studied 54 schizophrenic patients and found that that the average 24 h urine volume (obtained by catheter from all subjects) in the patients (2602 ± 120 ml) was twice the normal amount in healthy subjects (1328 ± 83 ml). In some patients, urine volume was 3–8 l day−1. In the other study, in 92 patients the mean urine volume was similarly increased, at 2532 ± 172 ml (Hariprasad et al. 1980). More detailed studies showed that patients’ ability to concentrate or dilute urine was within the normal range and that restricted water intake did not lead to a craving for water for schizophrenic subjects, in sharp contrast to what is observed in diabetes insipidus. Animal studies have shown either no effect of antipsychotics (Forsling et al. 1988; Hirayama et al. 2001) or an increased P [AVP] response to P Osm (Wells & Forsling, 1992). Collectively, these data suggest that antipsychotic treatments are not causal in water regulation abnormalities in schizophrenia. To our knowledge, the P [AVP] response to P Osm has not been studied in patients with depression or in animal models of depression, but in rats the P [AVP] response to P Osm increases with acute treatment and decreases with chronic (21 day) treatment with fluoxetine (selective serotonin reuptake inhibitor, a commonly used antidepressant; Faull et al. 1993). However, SSRIs may induce hyponatraemia from inappropriate antidiuretic hormone secretion, although this occurs primarily in the elderly and rarely in the age group studied here. Nonetheless, these types of medications remain a potential a contributory factor in the few subjects taking them in our investigation. Based on this finding, although clearly more research needs to be done, our finding of an increased AVP response in depression may even be an underestimation. There is also evidence suggesting that the AVP response to hyperosmotic stimulation is genetically regulated. The variability among dizygotic twins (r = 0.24) and high correlation in monozygotic twins (r = 0.94) in the general population indicates that the robust genetic mechanism that governs this water regulation response is crucial for survival (Zerbe et al. 1991). The stability of the AVP response to hypertonic stimulation in the same individuals over time (r = 0.95) also provides reassurance that that this approach is well suited for the assessment of differences among individuals. Conclusions Our findings may represent the first in vivo evidence that in depression the central NMDA receptor signalling is enhanced and that in schizophrenia it may be decreased, at least in comparison to depression. Once replicated in larger samples and fully developed, our approach might be used to detect vulnerability towards depression and schizophrenia in individuals early in the disease process and enable early intervention in a subgroup of patients. In addition, this approach might be useful in quantifying the central NMDA receptor responsiveness in individuals and guide clinicians in targeted drug treatment using novel glutamatergic agents.

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