Melbourne Veterinary School - Research Publications
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ItemPotential Molecular and Cellular Mechanism of Psychotropic Drugs(KOREAN COLL NEUROPSYCHOPHARMACOLOGY, 2014-08)Psychiatric disorders are among the most debilitating of all medical illnesses. Whilst there are drugs that can be used to treat these disorders, they give sub-optimal recovery in many people and a significant number of individuals do not respond to any treatments and remain treatment resistant. Surprisingly, the mechanism by which psychotropic drugs cause their therapeutic benefits remain unknown but likely involves the underlying molecular pathways affected by the drugs. Hence, in this review, we have focused on recent findings on the molecular mechanism affected by antipsychotic, mood stabilizing and antidepressant drugs at the levels of epigenetics, intracellular signalling cascades and microRNAs. We posit that understanding these important interactions will result in a better understanding of how these drugs act which in turn may aid in considering how to develop drugs with better efficacy or increased therapeutic reach.
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ItemValidating reference genes using minimally transformed qpcr data: findings in human cortex and outcomes in schizophrenia(BIOMED CENTRAL LTD, 2016-05-20)BACKGROUND: It is common practice, when using quantitative real time polymerase chain reaction (qPCR), to normalise levels of mRNA to reference gene mRNA which, by definition, should not vary between tissue, with any disease aetiology or after drug treatments. The complexity of human CNS means it unlikely that any gene could fulfil these criteria. METHODS: To address this issue we measured levels of mRNA for six potential reference genes (GAPDH, PPIA, SNCA, NOL9, TFB1M and SKP1) in three cortical regions (Brodmann's areas (BA) 8, 9 and 44) from 30 subjects with schizophrenia and 30 age and sex matched controls. We used a structured statistical approach to examine the characteristics of these data to determine their suitability as reference genes. We also analysed our data using reference genes selected by rank as defined using the average of the standard deviation of pair-gene ΔCt and the BestKeeper, NormFinder and geNorm algorithms to determine if they suggested the same reference genes. RESULTS: Our minimally derived data showed that levels of mRNA for all of the six genes varied between cortical regions and therefore no gene fulfilled the absolute requirements for use as reference genes. As levels of some mRNA for some genes did not vary with diagnoses within a cortical region from subjects with schizophrenia compared to controls, we normalised levels of mRNA for all the other genes to mRNA for one, two or three reference genes in each cortical region. This showed that using the geometric mean of at least two reference genes gave more reproducible results. Finally, using the reference gene ranking protocols the average of the standard deviation of pair-gene ΔCt, BestKeeper, NormFinder and geNorm we showed that these approaches ranked potential reference genes differently. We then showed that outcomes of comparing data from subjects with schizophrenia and controls varied depending on the reference genes chosen. CONCLUSIONS: Our data shows that the selection of reference genes is a significant component of qPCR study design and therefore the process by which reference genes are selected must be clearly listed as a potential confound in studying gene expression in human CNS. This should include showing that, using minimally derived qPCR data, levels of mRNA for proposed reference genes does not vary with variables such as diagnoses and CNS region.
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ItemChanges in Muscarinic M2 Receptor Levels in the Cortex of Subjects with Bipolar Disorder and Major Depressive Disorder and in Rats after Treatment with Mood Stabilisers and Antidepressants(OXFORD UNIV PRESS, 2016-04)BACKGROUND: Increasingly, data are implicating muscarinic receptors in the aetiology and treatment of mood disorders. This led us to measure levels of different muscarinic receptor-related parameters in the cortex from people with mood disorders and the CNS of rats treated with mood stabilisers and antidepressant drugs. METHODS: We measured [(3)H]AF-DX 384 binding in BA 46 and BA 24 from subjects with bipolar disorders (n = 14), major depressive disorders (n = 19), as well as age- and sex-matched controls (n = 19) and the CNS of rats treated with fluoxetine or imipramine. In addition, we used Western blots to measure levels of CHRM2 protein and oxotremorine-M stimulated [(35)S]GTPγS binding as a measure of CHRM 2 / 4 signaling. RESULTS: Compared with controls, [(3)H]AF-DX 384 binding was lower in BA 24 and BA 46 in bipolar disorders and major depressive disorders, while CHRM2 protein and oxotremorine-M stimulated [(35)S]GTPγS binding was only lower in BA 24. Compared with vehicle, treatment with mood stabilisers, antidepressant drugs for 10 days, or imipramine for 28 days resulted in higher levels of in [(3)H]AF-DX 384 binding select regions of rat CNS. CONCLUSIONS: Our data suggest that levels of CHRM2 are lower in BA 24 from subjects with mood disorders, and it is possible that signalling by that receptor is also less in this cortical region. Our data also suggest increasing levels of CHRM2 may be involved in the mechanisms of action of mood stabilisers and tricyclic antidepressants.
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ItemAberrant expression of microRNAs as biomarker for schizophrenia: from acute state to partial remission, and from peripheral blood to cortical tissue(SPRINGERNATURE, 2016-01-19)Based on our previous finding of a seven-miRNA (hsa-miR-34a, miR-449a, miR-564, miR-432, miR-548d, miR-572 and miR-652) signature as a potential biomarker for schizophrenia, this study aimed to examine if hospitalization could affect expressions of these miRNAs. We compared their expression levels between acute state and partial remission state in people with schizophrenia (n=48) using quantitative PCR method. Further, to examine whether the blood and brain show similar expression patterns, the expressions of two miRNAs (hsa-miR-34a and hsa-miR-548d) were examined in the postmortem brain tissue of people with schizophrenia (n=25) and controls (n=27). The expression level of the seven miRNAs did not alter after ~2 months of hospitalization with significant improvement in clinical symptoms, suggesting the miRNAs could be traits rather than state-dependent markers. The aberrant expression seen in the blood of hsa-miR-34a and hsa-miR-548d were not present in the brain samples, but this does not discount the possibility that the peripheral miRNAs could be clinically useful biomarkers for schizophrenia. Unexpectedly, we found an age-dependent increase in hsa-miR-34a expressions in human cortical (Brodmann area 46 (BA46)) but not subcortical region (caudate putamen). The correlation between hsa-miR-34a expression level in BA46 and age was much stronger in the controls than in the cases, and the corresponding correlation in the blood was only seen in the cases. The association between the miRNA dysregulations, the disease predisposition and aging warrants further investigation. Taken together, this study provides further insight on the candidate peripheral miRNAs as stable biomarkers for the diagnostics of schizophrenia.
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ItemAutophagy has a key role in the pathophysiology of schizophrenia(NATURE PUBLISHING GROUP, 2015-02)Autophagy is a process preserving the balance between synthesis, degradation and recycling of cellular components and is therefore essential for neuronal survival and function. Several key proteins govern the autophagy pathway including beclin1 and microtubule associated protein 1 light chain 3 (LC3). Here, we show a brain-specific reduction in beclin1 expression in postmortem hippocampus of schizophrenia patients, not detected in peripheral lymphocytes. This is in contrast with activity-dependent neuroprotective protein (ADNP) and ADNP2, which we have previously found to be deregulated in postmortem hippocampal samples from schizophrenia patients, but that now showed a significantly increased expression in lymphocytes from related patients, similar to increases in the anti-apoptotic, beclin1-interacting, Bcl2. The increase in ADNP was associated with the initial stages of the disease, possibly reflecting a compensatory effect. The increase in ADNP2 might be a consequence of neuroleptic treatment, as seen in rats subjected to clozapine treatment. ADNP haploinsufficiency in mice, which results in age-related neuronal death, cognitive and social dysfunction, exhibited reduced hippocampal beclin1 and increased Bcl2 expression (mimicking schizophrenia and normal human aging). At the protein level, ADNP co-immunoprecipitated with LC3B suggesting a direct association with the autophagy process and paving the path to novel targets for drug design.
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ItemSELENBP1 expression in the prefrontal cortex of subjects with schizophrenia(SPRINGERNATURE, 2015-08-04)Selenium binding protein 1 (SELENBP1) messenger RNA (mRNA) has previously been shown to be upregulated in the brain and blood from subjects with schizophrenia. We aimed to validate these findings in a new cohort using real-time PCR in Brodmann's Area (BA) 9, and to determine the disease specificity of increased SELENBP1 expression by measuring SELENBP1 mRNA in subjects with major depressive disorder and bipolar disorder. We then extended the study to include other cortical regions such as BA8 and BA44. SELENBP1 mRNA was higher in BA9 (P = 0.001), BA8 (P = 0.003) and BA44 (P = 0.0007) from subjects with schizophrenia. Conversely, in affective disorders, there was no significant difference in SELENBP1 mRNA in BA9 (P = 0.67), suggesting that the upregulation may be diagnosis specific. Measurement of SELENBP1 protein levels showed that changes in mRNA did not translate to changes in protein. In addition, chronic treatment of rats with antipsychotics did not significantly affect the expression of Selenbp1 in the cortex (P = 0.24). Our data show that elevated SELENBP1 transcript expression is widespread throughout the prefrontal cortex in schizophrenia, and confirm that this change is a consistent feature of schizophrenia and not a simple drug effect.
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ItemBiomarkers for Psychiatry: The Journey from Fantasy to Fact, a Report of the 2013 CINP Think Tank(OXFORD UNIV PRESS, 2015-09)BACKGROUND: A think tank sponsored by the Collegium Internationale Neuropsychopharmacologium (CINP) debated the status and prospects of biological markers for psychiatric disorders, focusing on schizophrenia and major depressive disorder. METHODS: Discussions covered markers defining and predicting specific disorders or domains of dysfunction, as well as predicting and monitoring medication efficacy. Deliberations included clinically useful and viable biomarkers, why suitable markers are not available, and the need for tightly-controlled sample collection. RESULTS: Different types of biomarkers, appropriate sensitivity, specificity, and broad-based exploitability were discussed. Whilst a number of candidates are in the discovery phases, all will require replication in larger, real-life cohorts. Clinical cost-effectiveness also needs to be established. CONCLUSIONS: Since a single measure is unlikely to suffice, multi-modal strategies look more promising, although they bring greater technical and implementation complexities. Identifying reproducible, robust biomarkers will probably require pre-competitive consortia to provide the resources needed to identify, validate, and develop the relevant clinical tests.
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ItemChanged gene expression in subjects with schizophrenia and low cortical muscarinic M1 receptors predicts disrupted upstream pathways interacting with that receptor(NATURE PUBLISHING GROUP, 2018-02)We tested the hypothesis that, compared with subjects with no history of psychiatric illness (controls), changes in gene expression in the dorsolateral prefrontal cortex from two subgroups of subjects with schizophrenia, one with a marked deficit in muscarinic M1 receptors (muscarinic receptor-deficit schizophrenia (MRDS)), would identify different biochemical pathways that would be affected by their aetiologies. Hence, we measured levels of cortical (Brodmann area 9) mRNA in 15 MRDS subjects, 15 subjects with schizophrenia but without a deficit in muscarinic M1 receptors (non-MRDS) and 15 controls using Affymetrix Exon 1.0 ST arrays. Levels of mRNA for 65 genes were significantly different in the cortex of subjects with MRDS and predicted changes in pathways involved in cellular movement and cell-to-cell signalling. Levels of mRNA for 45 genes were significantly different in non-MRDS and predicted changes in pathways involved in cellular growth and proliferation as well as cellular function and maintenance. Changes in gene expression also predicted effects on pathways involved in amino acid metabolism, molecular transport and small-molecule biochemistry in both MRDS and non-MRDS. Overall, our data argue a prominent role for glial function in MRDS and neurodevelopment in non-MRDS. Finally, the interactions of gene with altered levels of mRNA in the cortex of subjects with MRDS suggest many of their affects will be upstream of the muscarinic M1 receptor. Our study gives new insight into the molecular pathways affected in the cortex of subjects with MRDS and supports the notion that studying subgroups within the syndrome of schizophrenia is worthwhile.
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ItemThe Role of Muscarinic Receptors in the Pathophysiology of Mood Disorders: A Potential Novel Treatment?(BENTHAM SCIENCE PUBL LTD, 2015)The central cholinergic system has been implicated in the pathophysiology of mood disorders. An imbalance in central cholinergic neurotransmitter activity has been proposed to contribute to the manic and depressive episodes typical of these disorders. Neuropharmacological studies into the effects of cholinergic agonists and antagonists on mood state have provided considerable support for this hypothesis. Furthermore, recent clinical studies have shown that the pan-CHRM antagonist, scopolamine, produces rapid-acting antidepressant effects in individuals with either major depressive disorder (MDD) or bipolar disorder (BPD), such as bipolar depression, contrasting the delayed therapeutic response of conventional mood stabilisers and antidepressants. This review presents recent data from neuroimaging, post-mortem and genetic studies supporting the involvement of muscarinic cholinergic receptors (CHRMs), particularly CHRM2, in the pathophysiology of MDD and BPD. Thus, novel drugs that selectively target CHRMs with negligible effects in the peripheral nervous system might produce more rapid and robust clinical improvement in patients with BPD and MDD.
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ItemChanged frontal pole gene expression suggest altered interplay between neurotransmitter, developmental, and inflammatory pathways in schizophrenia(NATURE PUBLISHING GROUP, 2018-02-20)Schizophrenia (Sz) probably occurs after genetically susceptible individuals encounter a deleterious environmental factor that triggers epigenetic mechanisms to change CNS gene expression. To determine if omnibus changes in CNS gene expression are present in Sz, we compared mRNA levels in the frontal pole (Brodmann's area (BA) 10), the dorsolateral prefrontal cortex (BA 9) and cingulate cortex (BA 33) from 15 subjects with Sz and 15 controls using the Affymetrix™ Human Exon 1.0 ST Array. Differences in mRNA levels (±≥20%; p < 0.01) were identified (JMP Genomics 5.1) and used to predict pathways and gene x gene interactions that would be affected by the changes in gene expression using Ingenuity Pathway Analysis. There was significant variation in mRNA levels with diagnoses for 566 genes in BA 10, 65 genes in BA 9 and 40 genes in BA 33. In Sz, there was an over-representation of genes with changed expression involved in inflammation and development in BA 10, cell morphology in BA 9 and amino acid metabolism and small molecule biochemistry in BA 33. Using 94 genes with altered levels of expression in BA 10 from subjects with Sz, it was possible to construct an interactome of proven direct gene x gene interactions that was enriched for genes in inflammatory, developmental, oestrogen, serotonergic, cholinergic and NRG1 regulated pathways. Our data shows complex, regionally specific changes in cortical gene expression in Sz that are predicted to affect homeostasis between biochemical pathways already proposed to be important in the pathophysiology of the disorder.