Molecular Psychiatry (1997) 2, 495–500 1997 Stockton Press All rights reserved 1359–4184/97 $12.00 ORIGINAL RESEARCH ARTICLE consistent with their specific distribution in the human striatum.11,12 Within the range studied, postmortem Prodynorphin mRNA interval (PMI) was not found to significantly influence the levels of prodynorphin, proenkephalin, or D1 and D2 receptor mRNAs expressed throughout the striatum expression is increased in (eg Spearman correlation values between PMI and pro- the patch vs matrix dynorphin mRNA levels measured in the caudate, putamen, and nucleus accumbens, were −0.0099, compartment of the −0.0474, and 0.0442, respectively). No significant cor- relations were found between proenkephalin, D1,orD2 caudate nucleus in suicide mRNA expression and age, sex, race, blood ethanol level, or freezer storage time of the brain tissue subjects samples. The same applied for the expression of levels of prodynorphin mRNA and these variables, except for YL Hurd1, MM Herman2, TM Hyde2,LB 2 2 2 age, where there was a positive correlation in the Bigelow , DR Weinberger and JE Kleinman nucleus accumbens (Spearman correlation: 0.7592; = 1Karolinska Institute, Dept of Clinical Neuroscience, P 0.0004; with Bonferroni inequality the significance Psychiatry Section, S-171 76 Stockholm, Sweden; 2Clinical level was 0.0017). To consider any possible contri- Brain Disorders Branch, NIMH, Neuroscience Center at St bution of age, ANOVA statistical evaluations were also carried out with covariance for age. Elizabeths Hospital, Washington DC 20032, USA As presented in Table 1, no significant group differ- ences were seen in the mRNA expression levels of pro- Keywords: opioid neuropeptides; limbic; striatum; enkephalin or D1 and D2 dopamine receptors measured depression; human; in situ hybridization; enkephalin; dopa- within the caudate, putamen, or nucleus accumbens. mine receptors Also, no significant group differences were apparent Experimental and clinical studies suggest an involve- for the prodynorphin hybridization signals measured ment of the opioid neuropeptide system in psychiatric within the total caudate, putamen, and nucleus accum- disorders. Notably, opioid peptide immunoreactivity is bens. Of the markers tested, however, the mRNA altered in the cerebrospinal fluid of chronic schizo- expression of prodynorphin in the human striatum phrenics and manic-depressive subjects.1–3 Despite showed a pronounced heterogeneous distribution pat- these clinical findings, few postmortem investigations4,5 tern, with cell clusters of high mRNA expression sur- have examined the association of endogenous opioid rounded by regions of low expression (Figure 1), con- neuropeptides with schizophrenia and suicide. Anatom- sistent with its patch/matrix striatal organization.9 ically, a tight interaction exists within the neostriatum ANOVA with age as a covariate revealed a significant between the opioid peptide (dynorphin and enkephalin) group difference (P = 0.027) of the prodynorphin system and classical neurotransmitters such as dopam- mRNA expression in the caudate nucleus patch com- ine6 which has been implicated in both the psychotic partment: suicide subjects expressed higher levels of symptoms and the cognitive deficits that characterize 7 prodynorphin mRNA than that measured in schizo- schizophrenia (see review). The neostriatum is differen- # tially organized into patch and matrix neurochemical phrenics (P 0.05; Tukey-Kramer), and a similar trend mosaic compartments anatomically connected to limb- (P # 0.1; Tukey-Kramer) when compared to normal ic- and sensorimotor-related brain regions, respect- controls (Table 1 and Figure 1). Prodynorphin mRNA ively.6,8 Moreover, the human neostriatum is charac- expression in the caudate nucleus patch compartment terized by a heterogenous expression of the of schizophrenic subjects was not significantly differ- prodynorphin opioid gene: high in the patch, but low in ent from normal controls. No significant group differ- the matrix compartment.9,10 The present results show ences were found for prodynorphin mRNA expression for the first time a differential alteration of prodynorphin levels measured in the matrix compartment of the cau- within distinct striatal compartments in postmortem date nucleus. Thus overall, the same group difference tissue from nonschizophrenic suicide subjects. The pro- (P = 0.018) was apparent for the prodynorphin dynorphin patch/matrix mRNA expression was elevated in the caudate nucleus of suicide subjects as compared patch/matrix ratio in the caudate nucleus: high to normal controls and schizophrenics in which no alter- patch/matrix prodynorphin mRNA expression in the suicide subjects vs the control (P # 0.05) and schizo- ations in opioid peptides or D1 and D2 mRNA expression were apparent. Altogether the findings suggest that dis- phrenic (P # 0.03) groups, and no change from control crete dysfunction of the endogenous opioid dynorphin for the schizophrenics. It was noted that although all system might contribute to depression and the risk of suicide victims showed elevated patch/matrix prody- suicide in nonschizophrenic subjects. norphin mRNA expression, one subject was found to The mRNA expression of prodynorphin, proenke- express very low prodynorphin mRNA throughout the phalin, and D1 and D2 receptors was assessed using in entire striatum, which could not be explained by any situ hybridization histochemistry. The hybridization documented clinical information. In the putamen, no signals obtained using selective oligonucleotides or statistically significant group difference was evident in riboprobes complementary to these mRNA transcripts either the patch or matrix compartment. showed unique hybridization patterns and intensities The failure to detect any significant differences in Prodynorphin mRNA expression in the striatum YL Hurd et al 496 Table 1 DPM mg−1 values (mean ± s.e.m.) in the striatum of schizophrenic, suicide, and normal control subjects Caudate Putamen Nucleus accumbens PDYN (total area) Normal 88.9 ± 3.0 84.6 ± 2.6 93.2 ± 6.0 Suicide 93.8 ± 2.8 90.9 ± 4.1 108.8 ± 10.7 Schizophrenic 84.6 ± 5.3 80.3 ± 5.5 101.3 ± 9.9 PDYN patch Normal 105.6 ± 2.7 97.2 ± 5.5 n.d. Suicide 120.5 ± 6.5b 103.9 ± 9.8 n.d. Schizophrenic 97.6 ± 6.1 80.9 ± 5.4 n.d. PDYN matrix Normal 54.1 ± 2.2 48.3 ± 7.2 n.d. Suicide 48.1 ± 5.4 43.9 ± 5.6 n.d. Schizophrenic 52.2 ± 4.9 39.8 ± 6.4 n.d. PDYN patch/matrix Normal 1.9 ± 0.10 2.2 ± 0.20 n.d. Suicide 2.6 ± 0.19a,b 2.4 ± 0.15 n.d. Schizophrenic 1.9 ± 0.17 2.2 ± 0.24 n.d. PENK Normal 125.8 ± 9.2 133.0 ± 8.8 129.1 ± 8.7 Suicide 133.3 ± 7.0 136.4 ± 6.0 139.7 ± 4.1 Schizophrenic 138.7 ± 6.4 141.9 ± 6.7 141.1 ± 6.5 D1 dopamine receptor Normal 65.3 ± 2.9 65.7 ± 3.1 65.2 ± 2.9 Suicide 66.7 ± 2.2 65.9 ± 2.3 67.2 ± 2.3 Schizophrenic 65.2 ± 0.7 65.2 ± 1.1 65.6 ± 0.8 D2 dopamine receptor Normal 85.4 ± 2.2 85.6 ± 1.6 80.5 ± 2.5 Suicide 83.9 ± 1.2 85.6 ± 1.6 84.5 ± 2.6 Schizophrenic 87.1 ± 3.3 86.8 ± 4.4 82.0 ± 2.5 PDYN, prodynorphin; PENK, proenkephalin; n.d., not determined. aP , 0.05, vs control; bP , 0.05 vs schizophrenic; Tukey-Kramer. 17 the mRNA levels of dopamine D1 and D2 receptors typically inducing dysphoric experiences in humans. expressed in the caudate, putamen, or nucleus accum- While the present study cannot directly address the bens of drug-free schizophrenics is also matched at the state of depression, if any, at the time of death, the fact protein level; negative results were also obtained for remains that depression is the single most important 18 D1 and D2 receptor sites using radioligand binding on risk factor for suicide. Moreover, the prodynorphin the same postmortem brain specimen currently exam- elevations were predominantly localized in the patch 13 ined. The lack of D2 receptor alterations found at compartment of the caudate nucleus which has sig- postmortem fits with in vivo imaging studies that have nificant implications for affect, since it is the patch stri- failed to detect any alterations of dopamine D2 receptor atal compartment that receives a greater input from density in drug-free schizophrenic patients.14 How- limbic brain regions such as the amygdala8 and allo- 6 ever, D1 and D2 receptor and proenkephalin mRNA cortices, in comparison to the matrix which is more expression were also found to be normal in schizo- closely linked to sensorimotor brain areas. phrenic subjects who had chronic neuroleptic treat- Although striatal prodynorphin neurons preferen- 15 19 ment prior to death. It is feasible that although the D1 tially express D1 dopamine receptors, no changes and D2 receptor sites are major targets of antipsychotic were detected in mRNA expression for this dopamine drugs, other dopamine receptor subtypes, eg D3 and D4, receptor subtype or for D2 mRNA expression in the that are more tightly linked to the limbic system, are suicide group. Again, the lack of any alteration of the 16 affected to a greater extent in schizophrenic subjects. mRNA expression of D1 and D2 receptors is consistent In contrast to the failure to detect any mRNA abnor- with the negative results observed for D1 and D2 recep- malities at the level of striatal organization currently tor-binding sites in these subjects13 and in other sui- studied in schizophrenics, the increased expression of cide victims free of antidepressants prior to death.20 prodynorphin mRNA in the patch compartment of the In addition to expressing high levels of prodynor- caudate nucleus provides the first postmortem evi- phin mRNA, the human patch compartment is charac- dence of mRNA abnormalities in the striatal opioid terized by high m receptor-binding sites.12 Recently, m peptide system in suicide victims.