An investigation of the factors that regulate muscarinic receptor expression in schizophrenia

Abstract

We previously identified a group of subjects with schizophrenia who, on average, have a 75% decrease in cholinergic receptor, muscarinic 1 (CHRM1) in Brodmann's area (BA) 9. To extend this finding, we determined i) if the decrease in CHRM1 was present in another functionally related CNS region (BA6), ii) whether the marked decrease in CHRM1 was accompanied by changes in levels of other CHRMs and iii) potential factors responsible for the decreased CHRM1 expression. We measured CHRM1 and CHRM3 using in situ radioligand binding with [(3)H]pirenzepine and [(3)H]4-DAMP respectively in BA6 from 20 subjects with schizophrenia who had low levels of CHRM1 in BA9 (SzLow[(3)H]PZP), 18 subjects with schizophrenia whose levels of CHRM1 were similar to controls (SzNormal[(3)H]PZP) and 20 control subjects. Levels of CHRM1, 3 and 4 mRNA were measured using qPCR and levels of the transcription factors, SP1 and SP3, were determined using Western blots. In BA6, the density of [(3)H]pirenzepine binding was decreased in subjects with SzLow[(3)H]PZP (p<0.001) compared to controls. The density of [(3)H]4-DAMP binding, levels of CHRM1, 3 and 4 mRNA and levels of SP1 and SP3 was not significantly different between the three groups. This study shows that the previously identified decrease in CHRM1 expression is not confined to the dorsolateral prefrontal cortex but is present in other cortical areas. The effect shows some specificity to CHRM1, with no change in levels of binding to CHRM3. Furthermore, this decrease in CHRM1 does not appear to be associated with low levels of CHRM1 mRNA or to simply be regulated by the transcription factors, SP1 and SP3, suggesting that other mechanisms are responsible for the decreased CHRM1 in these subjects.

Evidence for impaired glucose metabolism in the striatum, obtained postmortem, from some subjects with schizophrenia

“… 20 We have shown that it is possible to distinguish a subset of individuals within the disorder based on a marked loss of cortical muscarinic receptor binding; we have termed this subset as muscarinic receptor deficit schizophrenia (MRDS). Subjects with MRDS have been shown to have a widespread loss of cortical muscarinic receptors 21 , 22 similar to what was reported in some individuals with the disorder in a neuroimaging study of muscarinic receptors in schizophrenia. 23 Recently we have reported that subjects with MRDS also have decreased radioligand binding to striatal muscarinic receptors and higher levels of post-synaptic density protein 95, 24 showing changes in the molecular cytoarchitecture of the CNS in subjects with MRDS extends beyond the cortex.…”

“…We have previously reported that subjects with MRDS have been shown to have a widespread loss of cortical muscarinic receptors, 21 , 22 have disruptions in muscarinic M1 receptor signaling, 48 have a distinct pattern of muscarinic M1 receptor gene promoter methylation and higher levels of a microRNA that acts to reduce receptor expression, 49 and have a reduced response to a muscarinic M1-receptor-positive allosteric modulator. 50 In addition, we have reported that subjects with MRDS also have decreased radioligand binding to striatal muscarinic receptors and higher levels of post-synaptic density protein 95, 24 showing changes in the molecular cytoarchitecture of the CNS in subjects with MRDS extends beyond the cortex.…”

Changes in BQCA Allosteric Modulation of [3H]NMS Binding to Human Cortex within Schizophrenia and by Divalent Cations

“…Finally, to allow us to better understand the potential impact of differing methods used for [ 3 H]NMS and [ 3 H]pirenzepine binding, we investigated whether the binding of these radioligands was ion dependent. Our studies were completed using Brodmann's area (BA) 6 because we confirmed that the marked decrease in [ 3 H]pirenzepine binding in subjects with MRDS, which was first identified in BA 9, was detectable in BA 6 (Seo et al, 2014) and because we had shown that this region of the cortex had decreased levels of neuregulin 1 (NRG1) (Barakat et al, 2010). As changes in NRG1 gene sequence has been associated with an increased risk of schizophrenia (Stefansson et al, 2002), we postulated molecular changes in BA 6 may reflect the impact of a biology dictated by an increased genetic predisposition for the disorder.…”

“…Following approval from the Tissue Access Committee of the Victorian Brain Bank Network, tissue was obtained from BA 6 (a cortical region that includes the anterior part of the paracentral lobule and the adjacent superior gyrus, the dorsal bank of the callosomarginal sulcus (minus its posterior third), and the bases of superior and middle frontal gyrus and pre-central gyrus not included in BA 4 (Garey, 1994)) from five subjects for methodological optimization. In addition, tissue was obtained from BA 6 from 40 subjects with schizophrenia and 20 age/sex-matched controls (demographics summarized in Table 1, complete detail in Supplementary Table 1) who had essentially been used in a previous study of [ 3 H]pirenzepine binding in that cortical region (Seo et al, 2014). The subjects with schizophrenia comprised 20 MRDS and 20 non-MRDS cases; the 20 subjects with MRDS were selected from the cases that we have previously shown separated into a subgroup within the syndrome of schizophrenia because of a marked loss of [ 3 H]pirenzepine binding in BA 9 ).…”

“…The lack of change in density of [ 3 H]NMS binding was significant because the selection of subjects with schizophrenia for this study was dictated by their levels of cortical [ 3 H]pirenzepine binding in BA 6 (Seo et al, 2014) and thus, compared with control, [ 3 H]pirenzepine binding was lower in subjects with schizophrenia (mean ± SEM: schizophrenia = 47 ± 5.9 vs Controls = 98 ± 4.0 fmol/mg ETE, po0.0001; Cohen's d = − 1.631). Levels of [ 3 H]pirenzepine were lowest in MRDS (mean ± SEM: 18 ± 4.1 fmol/mg ETE, po0.0001; Cohen's d = − 4.444) but binding in non-MRDS (mean ± SEM: 79 ± 5.2 fmol/mg ETE, po0.01; Cohen's d = − 0.945) was lower than controls.…”