Florey Department of Neuroscience and Mental Health - Research Publications
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ItemNo Preview AvailableEvaluating retinal biomarkers in a mouse model of Parkinson's disease(Association for Research in Vision and Ophthalmology, 2019-07-01)Purpose : The retina, an accessible outpouching of the central nervous system, may manifest cortical changes that occur with Parkinson’s disease (PD), lending itself as a potential biomarker. PD is characterised by reduced dopamine levels, a neurotransmitter found in amacrine cells. Human PD patients have also shown structural changes in the outer retina. This work aims to determine if retinal function and structure are altered in a murine model of PD and whether deficits can be ameliorated with L-DOPA treatment. Methods : A PD model was induced in adult C57BL6/J mice using MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, 4x i.p. injections, 20mg/kg) and vehicle control and examined at day 21 and 45. Another MPTP group was administered L-DOPA (L-3,4-dihydroxyphenylalanine 0.2 mg/ml) or control in their drinking water and assessed at day 45 (n=12–15/group). In ketamine:xylazine anaesthetised (80:10mg/kg) mice full-field dark- and light-adapted electroretinography (ERG) was assessed to target dopamine-related responses. Optical coherence tomography (OCT) was used to quantify thickness of retinal layers. Retinal and cortical tissue were collected for immunohistochemical assessment of changes in tyrosine hydroxylase (TH)and imaged using confocal microscopy. Data (mean±SEM) were compared using unpaired ANOVA and t-tests as appropriate. Results : At day 21 no retinal changes were found. At day 45 dark and light adapted ERGs showed slower amacrine cell responses (oscillatory potential, p<0.05), a finding which reversed with L-DOPA treatment (p<0.05). Other components of the ERG were unchanged. TH staining showed a trend towards decreased retinal levels in MPTP mice but this did not reach significance (p=0.10). Reduced levels of TH were found in the ventral hippocampus of MPTP mice compared with control (p<0.05). OCT revealed thinning of the outer plexiform layer at day 45, and the L-DOPA group exhibited a thinning of the outer nuclear layer (p<0.05). Conclusions : This study shows for the first time that the MPTP model recapitulates key dopaminergic changes previously reported in humans. In particular, electroretinographic changes that correspond with dopaminergic retinal cells occur in the Parkinson’s model and reverse with therapeutic treatment. Structural thinning of the outer retinal layers also occur, which parallels some human findings. This work paves the way for retinal measures as preclinical screening tools in drug development.
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ItemL-3,4-dihydroxyphenylalanine (L-DOPA) modulates brain iron, dopaminergic neurodegeneration and motor dysfunction in iron overload and mutant alpha-synuclein mouse models of Parkinson's disease(WILEY, 2019-07)Treatment with the dopamine (DA) precursor l-3,4-dihydroxyphenylalanine (l-DOPA) provides symptomatic relief arising from DA denervation in Parkinson's disease. Mounting evidence that DA autooxidation to neurotoxic quinones is involved in Parkinson's disease pathogenesis has raised concern about potentiation of oxidative stress by l-DOPA. The rate of DA quinone formation increases in the presence of excess redox-active iron (Fe), which is a pathological hallmark of Parkinson's disease. Conversely, l-DOPA has pH-dependent Fe-chelating properties, and may act to 'redox silence' Fe and partially allay DA autoxidation. We examined the effects of l-DOPA in three murine models of parkinsonian neurodegeneration: early-life Fe overexposure in wild-type mice, transgenic human (h)A53T mutant α-synuclein (α-syn) over-expression, and a combined 'multi-hit' model of Fe-overload in hA53T mice. We found that l-DOPA was neuroprotective and prevented age-related Fe accumulation in the substantia nigra pars compacta (SNc), similar to the mild-affinity Fe chelator clioquinol. Chronic l-DOPA treatment showed no evidence of increased oxidative stress in wild-type midbrain and normalized motor performance, when excess Fe was present. Similarly, l-DOPA also did not exacerbate protein oxidation levels in hA53T mice, with or without excess nigral Fe, and showed evidence of neuroprotection. The effects of l-DOPA in Fe-fed hA53T mice were somewhat muted, suggesting that Fe-chelation alone is insufficient to attenuate neuron loss in an animal model also recapitulating altered DA metabolism. In summary, we found no evidence in any of our model systems that l-DOPA treatment accentuated neurodegeneration, suggesting DA replacement therapy does not contribute to oxidative stress in the Parkinson's disease brain.
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ItemEarly existence and biochemical evolution characterise acutely synaptotoxic PrPSc(PUBLIC LIBRARY SCIENCE, 2019-04)Although considerable evidence supports that misfolded prion protein (PrPSc) is the principal component of "prions", underpinning both transmissibility and neurotoxicity, clear consensus around a number of fundamental aspects of pathogenesis has not been achieved, including the time of appearance of neurotoxic species during disease evolution. Utilizing a recently reported electrophysiology paradigm, we assessed the acute synaptotoxicity of ex vivo PrPSc prepared as crude homogenates from brains of M1000 infected wild-type mice (cM1000) harvested at time-points representing 30%, 50%, 70% and 100% of the terminal stage of disease (TSD). Acute synaptotoxicity was assessed by measuring the capacity of cM1000 to impair hippocampal CA1 region long-term potentiation (LTP) and post-tetanic potentiation (PTP) in explant slices. Of particular note, cM1000 from 30% of the TSD was able to cause significant impairment of LTP and PTP, with the induced failure of LTP increasing over subsequent time-points while the capacity of cM1000 to induce PTP failure appeared maximal even at this early stage of disease progression. Evidence that the synaptotoxicity directly related to PrP species was demonstrated by the significant rescue of LTP dysfunction at each time-point through immuno-depletion of >50% of total PrP species from cM1000 preparations. Moreover, similar to our previous observations at the terminal stage of M1000 prion disease, size fractionation chromatography revealed that capacity for acute synpatotoxicity correlated with predominance of oligomeric PrP species in infected brains across all time points, with the profile appearing maximised by 50% of the TSD. Using enhanced sensitivity western blotting, modestly proteinase K (PK)-resistant PrPSc was detectable at very low levels in cM1000 at 30% of the TSD, becoming robustly detectable by 70% of the TSD at which time substantial levels of highly PK-resistant PrPSc was also evident. Further illustrating the biochemical evolution of acutely synaptotoxic species the synaptotoxicity of cM1000 from 30%, 50% and 70% of the TSD, but not at 100% TSD, was abolished by digestion of immuno-captured PrP species with mild PK treatment (5μg/ml for an hour at 37°C), demonstrating that the predominant synaptotoxic PrPSc species up to and including 70% of the TSD were proteinase-sensitive. Overall, these findings in combination with our previous assessments of transmitting prions support that synaptotoxic and infectious M1000 PrPSc species co-exist from at least 30% of the TSD, simultaneously increasing thereafter, albeit with eventual plateauing of transmitting conformers.
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ItemMigration and Differentiation of Neural Stem Cells Diverted From the Subventricular Zone by an Injectable Self-Assembling β-Peptide Hydrogel(FRONTIERS MEDIA SA, 2019-11-08)Neural stem cells, which are confined in localised niches are unable to repair large brain lesions because of an inability to migrate long distances and engraft. To overcome these problems, previous research has demonstrated the use of biomaterial implants to redirect increased numbers of endogenous neural stem cell populations. However, the fate of the diverted neural stem cells and their progeny remains unknown. Here we show that neural stem cells originating from the subventricular zone can migrate to the cortex with the aid of a long-lasting injectable hydrogel within a mouse brain. Specifically, large numbers of neuroblasts were diverted to the cortex through a self-assembling β-peptide hydrogel that acted as a tract from the subventricular zone to the cortex of transgenic mice (NestinCreERT2:R26eYFP) in which neuroblasts and their progeny are permanently fluorescently labelled. Moreover, neuroblasts differentiated into neurons and astrocytes 35 days post implantation, and the neuroblast-derived neurons were Syn1 positive suggesting integration into existing neural circuitry. In addition, astrocytes co-localised with neuroblasts along the hydrogel tract, suggesting that they assisted migration and simulated pathways similar to the native rostral migratory stream. Lower levels of astrocytes were found at the boundary of hydrogels with encapsulated brain-derived neurotrophic factor, comparing with hydrogel implants alone.
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ItemThe Down Syndrome-Associated Protein, Regulator of Calcineurin-1, is Altered in Alzheimer's Disease and Dementia with Lewy Bodies(OMICS International, 2019)There is a known relationship between Alzheimer's disease (AD) and Down syndrome (DS), with the latter typically developing AD-like neuropathology in mid-life. In order to further understand this relationship we examined intersectin-1 (ITSN1) and the regulator of calcineurin-1 (RCAN1), proteins involved in endosomal and lysosomal trafficking that are over-expressed in DS. We examined RCAN1 and ITSN1 levels (both long (-L) and short (-S) isoforms) and the level of endogenous metals in White Blood Cells (WBCs) collected from AD patients who were enrolled in the Australian Imaging, Biomarker and Lifestyle Study on Ageing (AIBL). We also examined RCAN1 and ITSN1-S and -L in post-mortem brain tissue in a separate cohort of patients with AD or other types of dementia including Dementia with Lewy Bodies (DLB) and non-Alzheimer's disease dementia. We found that RCAN1 was significantly elevated in AD and DLB brain compared with controls, but there was no difference in the level of RCAN1 in WBCs of AD patients. There were no differences in the levels of ITSN1-L and -S between AD and the control, nor between other types of dementia and the control. We found that there were no differences in the levels of metals between AD and the control WBCs. In conclusion, our data demonstrate that RCAN1 is differentially regulated between the peripheral and central compartments in AD and should be further investigated to understand its potential role in dementia of AD and DLB.