Gene expression profiling in Brodmann's area 46 from subjects with schizophrenia.

OBJECTIVE: To identify altered gene expression in the dorsolateral prefrontal cortex obtained after death from subjects with schizophrenia. METHOD: Restriction fragment differential display (RFDD) was used to measure levels of mRNA in Brodmann area (BA) 46 from schizophrenia and control subjects. Findings on specific mRNA identified with RFDD were further investigated using real-time polymerase chain reaction (real-time PCR), PCR and western blotting. RESULTS: Levels of mRNA for 63 of approximately 12,500 genes differed in BA 46 in schizophrenia. Subsequent real-time PCR has shown that mRNA for muscleblind protein 1 (MBNL1) and protocadherin 17 (PCDH17) are increased in BA 46 from subjects with schizophrenia of short, but not long, duration. Altered levels of mRNA for neither gene were present in BA 9 from subjects with schizophrenia or in either cortical area from subjects with bipolar 1 disorder. By contrast, both RFDD and real-time PCR failed to show altered expression of the schizophrenia candidate gene disrupted in schizophrenia 1 (DISC1) BA46 from any diagnostic cohort. CONCLUSION: The present study has identified genes that are differentially expressed in BA 46 in schizophrenia. Initial studies have shown that there is a need for a careful validation of genes shown to be affected in schizophrenia using high-throughput technologies. In addition the present study has shown that gene expression may vary considerably depending on the duration of schizophrenia. This raises the hypothesis that changing gene expression may be underlying the change in symptom profile that occurs with disease progression in some subjects with schizophrenia.

Altered hippocampal muscarinic M4, but not M1, receptor expression from subjects with schizophrenia.

BACKGROUND: Having shown a decrease in [3H]pirenzepine binding in the hippocampus from subjects with schizophrenia, we wished to determine whether such a change in radioligand binding was associated with changes in hippocampal mRNA for the muscarinic1 (M1) and muscarinic4 (M4) receptors in tissue from different cohorts of subjects. METHOD: The [3H]pirenzepine binding using autoradiography and in situ hybridization with oligonucleotides specific for muscarinic M1 and M4 receptors were completed using hippocampal tissue obtained postmortem from 20 control subjects and 20 subjects with schizophrenia. RESULTS: The [3H]pirenzepine binding was decreased in the dentate gyrus (p < .05), CA3 (p < .01), CA2 (p < .05), and CA1 (p < .01) regions of the hippocampus from subjects with schizophrenia. Levels of M4 mRNA varied with the diagnosis of schizophrenia (p = .01), but significant region-specific changes were not apparent. Changes in levels of mRNA for the muscarinic M1 receptor were not detected with diagnosis. CONCLUSIONS: This study suggests that decreases in hippocampal [3H]pirenzepine binding in subjects with schizophrenia are most likely associated with widespread changes in expression levels of the M4 receptor. These data further implicate the hippocampal formation in the pathology of schizophrenia.

No change in cortical muscarinic M2, M3 receptors or [35S]GTPgammaS binding in schizophrenia.

Muscarinic M1, but not M4, receptors have been shown to be decreased in Brodmann's area (BA) 9 obtained postmortem from subjects with schizophrenia. This study extends that data by measuring levels of muscarinic M2 and M3 receptor protein and mRNAs in BA 9 and BA 40 from the same cohorts of subjects used in the study of M1 and M4 receptors. In addition, the ability of carbachol to stimulate muscarinic receptors that signal through the Gi/o G-proteins was measured in BA 9 from the same cohorts of subjects. There were no changes in levels of muscarinic M2 or M3 protein or M3 mRNA with diagnosis in either CNS region. M2 receptor mRNA could not be detected in BA 9 or BA 40. Finally, carbachol-stimulated GTPgammaS binding did not differ between the diagnostic cohorts in BA 9 (p = 0.64). These data add considerable weight to the argument that the muscarinic M1 receptor is the muscarinic receptor predominantly affected in BA 9 by the pathology of schizophrenia. Given the widespread changes in muscarinic receptors identified in the CNS of subjects of schizophrenia using functional neuroimaging it remains possible that receptors other than the M1 receptor may be altered in different CNS regions.

Differential changes in apolipoprotein E in schizophrenia and bipolar I disorder.

BACKGROUND: This study extends an initial finding of increased levels of apoE in Brodmann's area (BA) 9 from subjects with schizophrenia to determine if apoE is altered in other brain regions and in brains from subjects with bipolar I disorder (BID). METHODS: ApoE was quantified apoE in BA 9, 10, 40, 46 and caudate putamen from control (n = 18), schizophrenic (n = 19) and BID (n = 8) subjects using Western blotting. RESULTS: In schizophrenia, there was increased apoE in BA9 (mean +/- SEM: schizophrenia 3.8 +/- .18 vs. control 3.2 +/- .19) and BA46 (schizophrenia 2.7 +/- .26 vs. control 1.6 +/- .20). In BID, increased levels of the apolipoprotein were detected in the caudate putamen (BID 3.3 +/- .44 vs. control 2.4 +/- .19) and BA9 (BID 4.0 +/- .27 vs. control 3.2 +/- .19) with a decrease in apoE being measured in BA10 (BID 1.6 +/- .16 vs. control 3.9 +/- .53). CONCLUSIONS: This study has shown disease specific, regionally discrete changes in levels of apoE in brain obtained post mortem from schizophrenic and BID subjects. Our data adds weight to the hypothesis that changes in the levels of apolipoproteins may be involved in the pathologies of schizophrenia and bipolar disorder.

Cloning of a differentially expressed tropomyosin isoform from cultured rabbit aortic smooth muscle cells.

The four known tropomyosin genes have highly conserved DNA and amino acid sequences, and at least 18 isoforms are generated by alternative RNA splicing in muscle and non-muscle cells. No rabbit tropomyosin nucleotide sequences are known, although protein sequences for alpha- and beta-tropomyosin expressed by rabbit skeletal muscle have been described. Subtractive hybridisation was used to select for genes differentially expressed in rabbit aortic smooth muscle cells (SMC), during the change in cell phenotype in primary culture that is characterised by a loss of cytoskeletal filaments and contractile proteins. This led to the cloning of a tropomyosin gene predominantly expressed in rabbit SMC during this change. The full-length cDNA clone, designated "rabbit TM-beta", contains an open reading frame of 284 amino acids, 5' untranslated region (UTR) of 117 base pairs and 3' UTR of 79 base pairs. It is closely related to the beta-gene isoforms in other species, with the highest homology in DNA and protein sequences to the human fibroblast isoform TM-1 (91.7% identity in 1035 bp and 93.3% identity in the entire 284 amino acid sequence of the protein). It differs from rabbit skeletal muscle beta-tropomyosin (81.7% homology at the protein level) mainly in two regions at amino acids 189-213 and 258-283 suggesting alternative splicing of exons 6a for 6b and 9d for 9a. Since this TM-beta gene was the only gene strongly enough expressed in SMC changing phenotype to be observed by the subtractive hybridisation screen, it likely plays a significant role in this process.