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Widespread Changes in the Immunoreactivity of Bioactive Peptide T14 After Manipulating the Activity of Cortical Projection Neurons
Previous studies have suggested that T14, a 14-amino-acid peptide derived from acetylcholinesterase (AChE), functions as an activity-dependent signalling molecule with key roles in brain development, and its dysregulation has been linked to neurodegeneration in Alzheimer’s disease. In this study, we examined the distribution of T14 under normal developmental conditions in the mouse forebrain, motor cortex (M1), striatum (STR), and substantia nigra (SN). T14 immunoreactivity declined from E16 to E17 and further decreased by P0, then peaked at P7 during early postnatal development before declining again by adulthood at P70. Lower T14 immunoreactivity in samples processed without Triton indicated that T14 is primarily localised intracellularly. To explore the relationship between T14 expression and neuronal activity, we used mouse models with chronic silencing (Rbp4Cre-Snap25), acute silencing (Rbp4Cre-hM4Di), and acute activation (Rbp4Cre-hM3D1). Chronic silencing altered the location and size of intracellular T14-immunoreactive particles in adult brains, while acute silencing had no observable effect. In contrast, acute activation increased T14+ density in the STR, modified T14 puncta size near Rbp4Cre cell bodies in M1 layer 5 and their projections to the STR, and enhanced co-localisation of T14 with presynaptic terminals in the SN.
ER Stress and Autophagic Perturbations Lead to Elevated Extracellular α-Synuclein in GBA-N370S Parkinson's iPSC-Derived Dopamine Neurons.
Heterozygous mutations in the glucocerebrosidase gene (GBA) represent the strongest common genetic risk factor for Parkinson's disease (PD), the second most common neurodegenerative disorder. However, the molecular mechanisms underlying this association are still poorly understood. Here, we have analyzed ten independent induced pluripotent stem cell (iPSC) lines from three controls and three unrelated PD patients heterozygous for the GBA-N370S mutation, and identified relevant disease mechanisms. After differentiation into dopaminergic neurons, we observed misprocessing of mutant glucocerebrosidase protein in the ER, associated with activation of ER stress and abnormal cellular lipid profiles. Furthermore, we observed autophagic perturbations and an enlargement of the lysosomal compartment specifically in dopamine neurons. Finally, we found increased extracellular α-synuclein in patient-derived neuronal culture medium, which was not associated with exosomes. Overall, ER stress, autophagic/lysosomal perturbations, and elevated extracellular α-synuclein likely represent critical early cellular phenotypes of PD, which might offer multiple therapeutic targets.
Transcriptomic profiling of purified patient-derived dopamine neurons identifies convergent perturbations and therapeutics for Parkinson's disease.
While induced pluripotent stem cell (iPSC) technologies enable the study of inaccessible patient cell types, cellular heterogeneity can confound the comparison of gene expression profiles between iPSC-derived cell lines. Here, we purified iPSC-derived human dopaminergic neurons (DaNs) using the intracellular marker, tyrosine hydroxylase. Once purified, the transcriptomic profiles of iPSC-derived DaNs appear remarkably similar to profiles obtained from mature post-mortem DaNs. Comparison of the profiles of purified iPSC-derived DaNs derived from Parkinson's disease (PD) patients carrying LRRK2 G2019S variants to controls identified significant functional convergence amongst differentially-expressed (DE) genes. The PD LRRK2-G2019S associated profile was positively matched with expression changes induced by the Parkinsonian neurotoxin rotenone and opposed by those induced by clioquinol, a compound with demonstrated therapeutic efficacy in multiple PD models. No functional convergence amongst DE genes was observed following a similar comparison using non-purified iPSC-derived DaN-containing populations, with cellular heterogeneity appearing a greater confound than genotypic background.
Cellular α-synuclein pathology is associated with bioenergetic dysfunction in Parkinson's iPSC-derived dopamine neurons.
Parkinson's disease (PD) is the second most common neurodegenerative disorder and a central role for α-synuclein (αSyn; SNCA) in disease aetiology has been proposed based on genetics and neuropathology. To better understand the pathological mechanisms of αSyn, we generated induced pluripotent stem cells (iPSCs) from healthy individuals and PD patients carrying the A53T SNCA mutation or a triplication of the SNCA locus and differentiated them into dopaminergic neurons (DAns). iPSC-derived DAn from PD patients carrying either mutation showed increased intracellular αSyn accumulation, and DAns from patients carrying the SNCA triplication displayed oligomeric αSyn pathology and elevated αSyn extracellular release. Transcriptomic analysis of purified DAns revealed perturbations in expression of genes linked to mitochondrial function, consistent with observed reduction in mitochondrial respiration, impairment in mitochondrial membrane potential, aberrant mitochondrial morphology and decreased levels of phosphorylated DRP1Ser616. Parkinson's iPSC-derived DAns showed increased endoplasmic reticulum stress and impairments in cholesterol and lipid homeostasis. Together, these data show a correlation between αSyn cellular pathology and deficits in metabolic and cellular bioenergetics in the pathology of PD.
Early deficits in an in vitro striatal microcircuit model carrying the Parkinson's GBA-N370S mutation.
Understanding medium spiny neuron (MSN) physiology is essential to understand motor impairments in Parkinson's disease (PD) given the architecture of the basal ganglia. Here, we developed a custom three-chambered microfluidic platform and established a cortico-striato-nigral microcircuit partially recapitulating the striatal presynaptic landscape in vitro using induced pluripotent stem cell (iPSC)-derived neurons. We found that, cortical glutamatergic projections facilitated MSN synaptic activity, and dopaminergic transmission enhanced maturation of MSNs in vitro. Replacement of wild-type iPSC-derived dopamine neurons (iPSC-DaNs) in the striatal microcircuit with those carrying the PD-related GBA-N370S mutation led to a depolarisation of resting membrane potential and an increase in rheobase in iPSC-MSNs, as well as a reduction in both voltage-gated sodium and potassium currents. Such deficits were resolved in late microcircuit cultures, and could be reversed in younger cultures with antagonism of protein kinase A activity in iPSC-MSNs. Taken together, our results highlight the unique utility of modelling striatal neurons in a modular physiological circuit to reveal mechanistic insights into GBA1 mutations in PD.
αSynuclein and Mitochondrial Dysfunction: A Pathogenic Partnership in Parkinson's Disease?
Parkinson's Disease (PD) is a complex, chronic, progressive, and debilitating neurodegenerative disorder. Neither a cure nor effective long-term therapy exist and the lack of knowledge of the molecular mechanisms responsible for PD development is a major impediment to therapeutic advances. The protein αSynuclein is a central component in PD pathogenesis yet its cellular targets and mechanism of toxicity remains unknown. Mitochondrial dysfunction is also a common theme in PD patients and this review explores the strong possibility that αSynuclein and mitochondrial dysfunction have an inter-relationship responsible for underlying the disease pathology. Amplifying cycles of mitochondrial dysfunction and αSynuclein toxicity can be envisaged, with either being the disease-initiating factor yet acting together during disease progression. Multiple potential mechanisms exist in which mitochondrial dysfunction and αSynuclein could interact to exacerbate their neurodegenerative properties. Candidates discussed within this review include autophagy, mitophagy, mitochondrial dynamics/fusion/fission, oxidative stress and reactive oxygen species, endoplasmic reticulum stress, calcium, nitrosative stress and αSynuclein Oligomerization.
Adaptive microscope methods with embedded intelligence
Adaptive optics (AO) was shown to improve microscope imaging quality. However, issues like prolonged sample exposures are often associated with sensorless AO implementation. We present an AO solution with embedded intelligence to achieve outstanding performance.
Pupil Correlates of Decision Variables in Mice Playing a Competitive Mixed-Strategy Game.
In a competitive game involving an animal and an opponent, the outcome is contingent on the choices of both players. To succeed, the animal must continually adapt to competitive pressure, or else risk being exploited and lose out on rewards. In this study, we demonstrate that head-fixed male mice can be trained to play the iterative competitive game "matching pennies" against a virtual computer opponent. We find that the animals' performance is well described by a hybrid computational model that includes Q-learning and choice kernels. Comparing between matching pennies and a non-competitive two-armed bandit task, we show that the tasks encourage animals to operate at different regimes of reinforcement learning. To understand the involvement of neuromodulatory mechanisms, we measure fluctuations in pupil size and use multiple linear regression to relate the trial-by-trial transient pupil responses to decision-related variables. The analysis reveals that pupil responses are modulated by observable variables, including choice and outcome, as well as latent variables for value updating, but not action selection. Collectively, these results establish a paradigm for studying competitive decision-making in head-fixed mice and provide insights into the role of arousal-linked neuromodulation in the decision process.
Training enhances the ability of listeners to exploit visual information for auditory scene analysis.
The ability to use temporal relationships between cross-modal cues facilitates perception and behavior. Previously we observed that temporally correlated changes in the size of a visual stimulus and the intensity in an auditory stimulus influenced the ability of listeners to perform an auditory selective attention task (Maddox, Atilgan, Bizley, & Lee, 2015). Participants detected timbral changes in a target sound while ignoring those in a simultaneously presented masker. When the visual stimulus was temporally coherent with the target sound, performance was significantly better than when the visual stimulus was temporally coherent with the masker, despite the visual stimulus conveying no task-relevant information. Here, we trained observers to detect audiovisual temporal coherence and asked whether this changed the way in which they were able to exploit visual information in the auditory selective attention task. We observed that after training, participants were able to benefit from temporal coherence between the visual stimulus and both the target and masker streams, relative to the condition in which the visual stimulus was coherent with neither sound. However, we did not observe such changes in a second group that were trained to discriminate modulation rate differences between temporally coherent audiovisual streams, although they did show an improvement in their overall performance. A control group did not change their performance between pretest and post-test and did not change how they exploited visual information. These results provide insights into how crossmodal experience may optimize multisensory integration.
Ketosis regulates K+ ion channels, strengthening brain-wide signaling disrupted by age.
Aging is associated with impaired signaling between brain regions when measured using resting-state fMRI. This age-related destabilization and desynchronization of brain networks reverses itself when the brain switches from metabolizing glucose to ketones. Here, we probe the mechanistic basis for these effects. First, we confirmed their robustness across measurement modalities using two datasets acquired from resting-state EEG (Lifespan: standard diet, 20-80 years, N = 201; Metabolic: individually weight-dosed and calorically-matched glucose and ketone ester challenge, μ a g e = 26.9 ± 11.2 years , N = 36). Then, using a multiscale conductance-based neural mass model, we identified the unique set of mechanistic parameters consistent with our clinical data. Together, our results implicate potassium (K+) gradient dysregulation as a mechanism for age-related neural desynchronization and its reversal with ketosis, the latter finding of which is consistent with direct measurement of ion channels. As such, the approach facilitates the connection between macroscopic brain activity and cellular-level mechanisms.
Physiological function of cyclic nucleotide phosphodiesterases in atrial myocytes and their potential as therapeutic targets for atrial fibrillation.
Cyclic nucleotide hydrolyzing phosphodiesterases (PDEs) are key regulators of cyclic nucleotide (e.g., cAMP and cGMP) signaling. Here, we examine the role of PDEs in the physiology of atrial myocytes (AMs), the pathogenesis of atrial fibrillation (AF), and the potential of PDEs as therapeutic targets for AF. PDE1-5 and 8 are present and functional in AMs. PDE2-4 are important regulators of AM contraction but their role beyond atrial contractility is unclear. The role of PDE1,5 and 8 in healthy AMs is unknown but of interest because of their roles in ventricular myocytes. We propose that PDE2-5 and PDE8 are potential targets to prevent the triggering of AF considering their effects on Ca2+ handling and/or electrical activity. PDE1-5 are possible targets to treat patients with paroxysmal or persistent AF caused by pulmonary vein automaticity. PDE8B2 is a possible target for patients with persistent AF due to its altered expression. Research should aim to identify the presence, localization, and function of specific PDE isoforms in human atria. Ultimately, the paucity of PDE isoform-specific small molecule modulators and the difficulty of delivering PDE-targeted medications or therapies to particular cell types limit current research and its application.
Diagnostic and prognostic value of α-synuclein seed amplification assay kinetic measures in Parkinson's disease: a longitudinal cohort study
Background: α-synuclein seed amplification assay (SAA) positivity has been proposed as a diagnostic biomarker for Parkinson's disease. However, studies of the prognostic value of this biomarker have been limited to small, single-centre studies over short follow-up periods. We aimed to assess the diagnostic and prognostic value of quantitative CSF α-synuclein SAA kinetic measures in Parkinson's disease. Methods: In this longitudinal cohort study, we collected and analysed data from participants with Parkinson's disease, progressive supranuclear palsy, and healthy controls enrolled in three cohorts: the UK parkinsonism cohort, the Parkinson's Progression Markers Initiative (PPMI) international observational study, and the Tübingen Parkinson's disease cohort. Baseline CSF α-synuclein SAA data and longitudinal clinical data were collected between Jan 1, 2005, and Nov 1, 2023. The following seeding kinetic measures were calculated from the α-synuclein SAA curve for each SAA-positive sample: time to threshold (TTT) for a positive SAA result; maximum Thioflavin T fluorescence during the reaction time (MaxThT); and area under the fluorescence curve during the reaction time (AUC). We compared seeding kinetic measures between sporadic Parkinson's disease and progressive supranuclear palsy, and between sporadic Parkinson's disease and monogenic Parkinson's disease. We used time-to-event analyses to assess the ability of α-synuclein SAA kinetic measures to predict an unfavourable outcome in Parkinson's disease, adjusting for sex, age, and disease duration at SAA testing. Findings: We analysed data from 1631 participants: newly generated data from the UK parkinsonism cohort (Parkinson's disease, n=66; progressive supranuclear palsy, n=52; controls, n=9) and previously generated data from the PPMI (Parkinson's disease, n=1036; controls, n=239) and Tübingen (Parkinson's disease, n=229) cohorts. In the UK parkinsonism cohort, α-synuclein SAA was positive in 63 (96%) of 66 Parkinson's disease samples and eight (15%) of 52 progressive supranuclear palsy samples, with six (75%) of eight positive progressive supranuclear palsy samples having distinct low and slow seeding kinetics (low MaxThT and high TTT) as a marker of Lewy body co-pathology. TTT was faster in GBA1-associated Parkinson's disease compared with sporadic Parkinson's disease in both the PPMI (p=0·04) and Tübingen (p=0·01) cohorts. In the PPMI cohort, after excluding individuals who had an unfavourable outcome at the time of baseline SAA testing, an unfavourable outcome was observed in 593 (73%) of 810 participants with α-synuclein SAA-positive Parkinson's disease during a median follow-up period of 4·5 years (IQR 2–9). TTT at baseline predicted only cognitive decline (Montreal Cognitive Assessment score ≤21) as a component of an unfavourable outcome in Parkinson's disease in both the PPMI (n=824, hazard ratio [HR] 2·36 [95% CI 1·60–3·46], p=0·001) and Tübingen (n=135, 2·17 [1·07–4·41], p=0·03) cohorts. TTT also predicted cognitive decline in a subgroup of participants with Parkinson's disease in the PPMI cohort who were Alzheimer's disease biomarker negative (n=355, HR 1·80 [95% CI 1·03–3·18], p=0·04). Interpretation: Assessing α-synuclein SAA kinetic measures might aid in the diagnostic differentiation of Parkinson's disease from progressive supranuclear palsy with Lewy body co-pathology. Furthermore, faster seeding kinetics are found in GBA1-Parkinson's disease and predict cognitive decline in Parkinson's disease independently of Alzheimer's disease co-pathology. Funding: Medical Research Council, PSP Association. Copyright: © 2025 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license.
A compilation of reported alterations in the cerebrospinal fluid proteome in Alzheimer's disease.
Alzheimer's disease is a multifaceted neurodegenerative disorder, with diverse underlying pathophysiological processes extending beyond amyloid-β and tau accumulation. The heterogeneity of Alzheimer's disease necessitates the identification of a broad array of biomarkers that capture the diverse mechanisms contributing to disease onset and progression. In this study, we systematically compiled and analysed cerebrospinal fluid proteomics data from omics studies utilizing mass spectrometry, Olink, or SomaScan platforms. Systematic literature searches for each platform revealed a total of 264 studies. From this, a set of 18 studies were selected based on sample size, number of markers analysed, and open data availability. We found a total of 1,448 differentially expressed proteins between Alzheimer's disease and amyloid negative controls across these datasets, with 635 being found in more than one study. A 'top' set of 61 differentially expressed proteins were consistently reported in at least six studies. Clustering and functional enrichment analysis of the top differentially expressed proteins indicated involvement in metabolic regulation, glutathione metabolism and proteins of the 14-3-3 family, reflecting importance of reactive oxygen species (ROS) response. Synaptic signalling processes were found to generally be downregulated. We further integrated the top differentially expressed proteins with results from a study on familial Alzheimer's disease cerebrospinal fluid to assess at which stage of disease progression these proteins change, highlighting markers shared between sporadic and familial Alzheimer's disease datasets. Lastly, we examine the overlap of the top differentially expressed proteins between cerebrospinal fluid and brain tissue using a publicly available database. This analysis provides a comprehensive overview of the Alzheimer's disease cerebrospinal fluid proteomic landscape, indicating changes in key pathways and cellular processes associated with Alzheimer's disease pathology. By integrating data from different platforms, we highlight reproducible protein changes that may serve as promising candidates for further biomarker research aimed at improving patient stratification, tracking disease progression, and assessing therapeutic interventions.