Short Communication

Decreased Hippocampal (CA3) NMDAReceptors in Schizophrenia

BRIAN DEAN, ELIZABETH SCARR, ROBYN BRADBURY, AND DAVID COPOLOVMolecular Schizophrenia Division, The Mental Health Research Institute,

Locked Bag 11, Parkville, Victoria, 3052, Australia

Changed interactions between dopaminergic and glu-tamatergic neurons in the central nervous system(CNS) may be important in the pathogenesis of schizo-phrenia (Carlsson et al., 1997), particularly in themedial temporal lobe (MTL), a region thought to beimportant in the pathology of the illness (Arnold, 1997).There have been separate studies of dopaminergic(Goldsmith et al., 1997; Lahti et al., 1996) and glutama-tergic (Kerwin et al., 1990; Kornhuber et al., 1989;Simpson et al., 1991) markers in the MTL of schizo-phrenic subjects but no study has attempted to examinethe interaction between these two neurotransmittersystems. To begin to address this issue, we have mea-sured the density of dopamine (DA) receptors, thedopamine transporter (DAT) and the N-methyl-D-aspartate (NMDA) receptor in MTL in schizophrenia.The NMDA receptor was investigated because thisreceptor is critical in dopaminergic/glutamatergic inter-actions in human CNS (Starr, 1995).

Dopaminergic and glutamatergic markers were mea-sured in the left MTL collected at autopsy from 25schizophrenic and 25 control subjects (Table I). Limita-tions on available MTL meant that all binding param-eters were only available on 15 matched schizophrenicand control subjects. Where death was witnessed,postmortem interval (PMI) was the time between deathand autopsy. If death was not witnessed, tissue wastaken from individuals seen alive within 5 hours ofbeing found dead. The PMI was taken as the intervalhalfway between the donor being found and being lastseen alive.

Radioligand binding to frozen sections (220°C: 5 3 20µm frozen tissue per radioligand) of MTL was mea-sured at a single concentration, approximately threetimes that of the reported Kd for each radioligand at thebinding site of interest. The density of [3H] SCH 23390(3 nM) binding to DA-D1 receptors was taken as bindingin the absence (3 sections: total binding) minus bindingin the presence (2 sections: non-specific binding) of1026M cis-flupenthixol (Pimoule et al., 1985). Similarly,differences in the binding of [3H]YM-09151-2 (4 nM) 61026M (1)-butaclamol was taken as a measure ofDA-D2 receptors (Lahti et al., 1996), [3H]TCP (20 nM) 6

MK-801 (1026M) as a measure of NMDA receptors(Simpson et al., 1991) and [3H]mazindol (15 nM) in thepresence of 0.3 µM desmethylimipramine (DMI) 61026M mazindol as a measure of DAT (Donnan et al.,1991). After incubating with radioligand, all sectionswere washed twice in ice cold assay buffer, dipped intoice cold distilled water, thoroughly dried and apposed toHyperfilm-3Ht along with [3H]Micro-scales until animage of appropriate optical density was obtained.Images were then analyzed with an MCID imageanalysis system and results expressed as fmol/mgestimated tissue equivalents (wet weight) (ETE). Com-paring data from schizophrenic and control subjects,relationships between radioligand binding within MTL,subject age and tissue PMI were examined using theMann-Whitney U-test. The analysis of radioligand bind-ing across laminae was carried out using a one-wayANOVA. Relationships between age, PMI, and finalrecorded drug dose with radioligand binding wereassessed using the Pearson product-moment correla-tion coefficients calculated using an assumed straightline fit. Donor age, tissue PMI, and final recordedantipsychotic drug were not confounding factors in thisstudy.

Overall, there was one significant difference in theneurochemical markers studied in the MTL from theschizophrenic subjects; this was a decrease in NMDAreceptors in the CA3 region of the hippocampal forma-tion (CA 3) (Table II). However, there was a trend tolower levels of NMDA receptor throughout the hippo-campus. This could be why a previous study, usingparticulate membrane, showed an overall decrease inNMDA receptors in the hippocampus from schizo-phrenic subjects (Simpson et al., 1991). Another studyusing autoradiography reported a 59% decrease in thedensity of NMDA receptors in the CA3 from the lefthemisphere of 7 schizophrenic subjects compared tothat in 8 control subjects (Kerwin et al., 1990). This

*Correspondence to: Brian Dean, Molecular Schizophrenia Division, TheMental Health Research Institute, Locked Bog 11, Parkville, Victoria, 3052,Australia.

Received 18 May 1998; accepted in revised form 13 October 1998

SYNAPSE 32:67–69 (1999)

r 1999 WILEY-LISS, INC.

apparently large decrease in the density of NMDAreceptor did not reach significance but tends to agreewith the findings in this study. This study has alsoshown a trend towards an increase in DAT in the CA2from schizophrenic subjects (Table II). However, in theMTL from the 30 subjects in which DAT and NMDAwere measured, there was there no relationship be-tween these markers (CA2 r 5 0.14, CA3 r 5 0.06).This study cannot resolve whether the tendency towardincreased DAT in the CA2 from the schizophrenicsubjects was due to increased number of DAT contain-ing neurons or an increased expression of DAT on astable number of neurons.

The overall absence of change in [3H]YM-09151-2 inthe MTL from schizophrenic subjects is consistent witha previous study using this radioligand (Lahti et al.,1996). However, as both DA-D2 receptors and s recep-

tors display (1)-butaclamol sensitive [3H]YM-09151-2binding this finding must be interpreted with caution.Another study (Goldsmith et al., 1997) has reporteddecreased [125I]epidepride binding to DA-D2 receptorsin laminae I and II and increased binding in lamina IVof the perirhinal cortex and temporal isocortex of theMTL from schizophrenic subjects. Significantly, only[3H]YM-09151-2 binding appeared to show any varia-tion across laminae with a tendency to higher levels ofradioligand binding in lamina I in the parahippocampalgyrus from the control subjects (laminae I 5 22 6 4.7,II 5 12 6 3.7, III 5 11 6 3.4, IV 5 10 6 3.6, V 5 8.7 63.1, and VI 5 8.1 6 2.8 fmol/mg ETE: mean 6 SEM),which was not apparent in the schizophrenics (laminaeI 5 6.3 6 3.3, II 5 8.7 6 3.6, III 5 8.3 6 3.6, IV 5 7.8 63.1, V 5 7.3 6 2.9, and VI 5 6.3 6 2.9 fmol/mg ETE).However, in neither group was there a significant

TABLE I. Demographic and Pharmacological Data on the 25 Schizophrenic Subjects Diagnosed According to DSM-III-R Criteria (AmericanPsychiatric Association, 1995) Following Extensive Case History Review and the 25 Age/Sex-Matched Control Subjects1

Schizophrenics Controls

ID Sex Age PMI Antipsychotic drugsLastdose* Cause of death ID Sex Age PMI Cause of death

1 M 27 22 ChlorpromazinePimozide

1,200 Burning 1 M 44 44 Acute myocardialinfarction

2 M 23 43 Haloperidol 1,750 Hanging 2 M 23 36 Asthma attack3 M 25 49 Trifluoperazine

Stelazine200 Drug overdose 3 M 62 63 Thoracic aorta disorder

42 M 53 37 FluphenazineChlorpromazine

1,700 IntestinalIschaemic

42 M 65 41 Ischaemic heart disease

5 M 63 46 Not available Ischaemic heart disease 52 M 65 21 Acute myocardialinfarction

62 M 63 73 TrifluoperazineChlorpromazine

No drug7 yr.

Chronic cardiac failure 6 M 35 27 Acute myocardialinfarction

72 M 67 21 Fluphenazine 75 Pneumonia 72 M 25 50 Exsanguination8 M 44 32 Thioridazine 600 Ischaemic heart disease 82 M 53 45 Acute myocardial

infarction92 M 35 81 Fluphenazine 400 Perforated gastric ulcer 92 F 80 55 Exsanguination

102 M 47 42 ChlorpromazineHaloperidol

1,400 Multiple injuries 10 M 27 31 Asthma

112 F 81 25 TrifluoperazineChlorpromazine

100 Aspiration, food 112 M 46 47 Acute myocardialinfarction

122 M 71 48 Thioridazine 150 Aspiration, food 122 M 22 62 Iatrogenic haemorrhage132 M 32 17 Haloperidol 670 Carbon monoxide poi-

soning132 M 63 38 Acute myocardial

infarction142 M 53 43 Chlorpromazine No drug

6 yr.Aspiration, food 142 M 69 38 Acute myocardial

infarction152 M 69 45 Trifluoperazine 100 Ischaemic heart disease 152 M 72 45 Pyelonephritis162 F 68 42 Trifluoperazine

Chlorpromazine75 Ischaemic heart disease 162 M 35 55 Coronary artery athero-

sclerosis172 M 61 38 Fluphenazine No drug

2 yr.Ischaemic heart disease 172 F 66 43 Acute myocardial

infarction182 F 34 42 Risperidone 1,200 Drowning 182 M 48 52 Ischaemic heart disease192 M 22 37 Pimozide 200 Drug overdose 192 F 33 43 Multiple injuries202 M 42 47 Haloperidol 128 Hanging 202 M 43 43 Coronary artery athero-

sclerosis21 M 35 47 Fluphenazine 550 Poisoning, welding solu-

tion21 M 28 22 Hanging

22 M 71 48 Chlorpromazine 600 Hanging 22 M 38 51 Ischaemic heart disease23 M 47 42 Trifluoperazine

Flupenthixol450 Pericarditis 23 M 45 18 Hanging

24 F 34 42 FlupenthixolChlorpromazine

610 Drowning 24 F 29 26 Stab wound

25 M 53 53 Clozapine 50 Meningo encepha-lopathy

25 M 35 31 Ischaemic heart disease

1Dopamine subjects 5 1–20; NMDA subjects 5 6–25.2Dopamine and NMDA subjects.*Expressed as chlorpromazine equivalent (mg).

68 B. DEAN ET AL.

difference in the mean density of [3H]YM-09151-2 acrosslaminae (P 5 0.11 and P 5 0.99, respectively). Therewas a significant decrease in the density of [3H]YM-09151-2 binding in lamina I from the schizophrenicsubjects compared to controls (P , 0.01) that was notdetectable in the other laminae. Thus, there wouldseem to be laminae specific changes in DA-D2 receptorsin specific regions of the MTL from schizophrenicsubjects. We are not aware of any previous studies ofDA-D1 receptors in the MTL from schizophrenic subjects.

This study has shown a decrease in NMDA receptorsin CA 3 from schizophrenic subjects but does notprovide evidence that this change is related to changesin dopaminergic markers in the MTL.

REFERENCES

American Psychiatric Association. 1995. Diagnostic criteria fromDSM-IV. Washington, DC: American Psychiatric Association.

Arnold SE. 1997. The medial temporal lobe in schizophrenia. JNeuropsychiatr Clin Neurosci 9:460–470.

Carlsson A, Hansson LO, Waters N, Carlsson ML. 1997. Neurotrans-mitter aberrations in schizophrenia: New perspectives and therapeu-tic implications. Life Sci 61:75–94.

Donnan GA, Kaczmarczyk SJ, Paxinos G, Chilco PJ, Kalnins RM,Woodhouse DG, Mendelsohn FA. 1991. Distribution of catechol-amine uptake sites in human brain as determined by quantitative[3H] mazindol autoradiography. J Comp Neurol 304:419–434.

Goldsmith SK, Shapiro RM, Joyce JN. 1997. Disrupted pattern of D2dopamine receptors in the temporal lobe in schizophrenia.Apostmor-tem study. Arch Gen Psychiatr 54:649–658.

Kerwin RW, Patel S, Meldrum B. 1990. Quantitative autoradiographicanalysis of glutamate binding sites in the hippocampal formation innormal and schizophrenic brain postmortem. Neuroscience 39:25–32.

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Lahti RA, Roberts RC, Conley RR, Cochrane EV, Mutin A, TammingaCA. 1996. D2-type dopamine receptors in postmortem human brainsections from normal and schizophrenic subjects. Neuroreport7:1945–1948.

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TABLE II. Binding of [3H]SCH 23390, [3H]YM-19151-2, [3H]NMDA, and [3H]Mazindol to Regions of MTL From Schizophrenicand Control Subjects as Well as the Age of the Tissue Donors and Tissue Postmortem Interval

Radioactive ligand

Dopamine studies Glutamate studies

Schizophrenic Control P* Schizophrenic Control P*

Age (year) 49 6 4.1 49 6 4.06 NS 52 6 3.7 48 6 2.7 NSPMI (hour) 42 6 3.5 44 6 2.39 NS 43 6 3.3 65 6 7.6 NS[3H]SCH 23390 binding†

Parahippocampal gyrus 7.26 6 1.67 7.23 6 1.27 NSSubiciculum 5.76 6 1.63 4.87 6 1.13 NSCA1 11.4 6 2.0 8.96 6 1.37 NSCA2 3.1 6 1.12 2.84 6 1.17 NSCA3 5.37 6 1.68 3.03 6 1.07 NSDentate gyrus 3.14 6 0.53 2.74 6 0.90 NS

[3H] YM-09151-2 binding†Parahippocampal gyrus 6.50 6 1.97 13.5 6 3.85 NSSubiciculum 8.25 6 2.39 10.4 6 3.24 NSCA1 13.2 6 2.65 17.02 6 5.10 NSCA2 8.67 6 2.81 11.6 6 3.69 NSCA3 4.75 6 1.57 8.30 6 2.08 NSDentate gyrus 4.81 6 1.75 11.7 6 3.00 NS

[3H]Mazindol binding†Parahippocampal gyrus 18.4 6 4.90 17.5 6 6.01 NSSubiciculum 18.9 6 4.80 12.3 6 3.61 NSCA1 14.8 6 5.10 10.6 6 3.95 NSCA2 32.9 6 8.14 14.1 6 5.00 0.059CA3 23.4 6 6.19 14.3 6 4.57 NSDentate gyrus 25.5 6 6.62 12.3 6 4.74 NS

[3H]TCP bindingParahippocampal gyrus 58.4 6 6.6 65.5 6 7.85 NSSubiciculum 50.9 6 6.9 59.3 6 7.64 NSCA1 103 6 10.2 121 6 9.35 NSCA2 59.4 6 7.0 76.8 6 6.3 0.09CA3 78.1 6 12.0 110 6 13.1 ,0.02Dentate gyrus 92.2 6 6.0 109 6 7.00 NS

*NS 5 not significant.†fmol/mg Estimated Tissue Equivalents.

69DECREASED NMDA RECEPTORS IN SCHIZOPHRENIA