Francesca Tozzi

The locomotor activity (LMA) test is widely used to study spontaneous activity in rodents and the effects of drug treatments. However, standard automated metrics such as “distance traveled” often fail to capture the true complexity of behavior in this test, while manual annotations are labor intensive and prone to variability. Although recent advances in computational ethology, both unsupervised and supervised, have pushed forward analysis of spontaneous activity, challenges remain in providing interpretability and in temporal modeling. In this work, we adapt ActionFormer, a transformer-based architecture for temporal action localization, to analyze spontaneous activity of laboratory mice. ActionFormer significantly outperforms prior state-of-the-art methods in Average Precision (AP) across 7 common behaviors, and can capture characteristic drug-induced effects. By introducing behavior signatures, we showcase ActionFormer’s ability to uncover group differences and dose-response relationships in preclinical studies. Our findings demonstrate the potential of ActionFormer to further advance the frontier of computational ethology for laboratory rodents.

The beam walk is widely used to study coordination and balance in rodents. While the task has ethological validity, the main endpoints of ‘foot slip counts’ and ‘time to cross’ are prone to human-rater variability and offer limited sensitivity and specificity. We asked if machine learning-based methods could reveal previously hidden, but biologically relevant, insights from the task. Marker-less pose estimation, using DeepLabCut, was deployed to label 13 anatomical points on mice traversing the beam. Next, we automated classical endpoint detection, including foot slips, with high recall (>90%) and precision (>80%). A total of 395 features were engineered and a random-forest classifier deployed that, together with skeletal visualizations, could test for group differences and identify determinant features. This workflow, named Forestwalk, uncovered pharmacological treatment effects in C57BL/6J mice, revealed phenotypes in transgenic mice used to study Angelman syndrome and SLC6A1-related neurodevelopmental disorder, and will facilitate a deeper understanding of how the brain controls balance in health and disease.

Background: Synaptic dysfunction is an early mechanism in Alzheimer’s disease (AD) which involves progressively larger areas of the brain over time. However, how it starts and propagates is unknown. We hypothesised that large extracellular vesicles (EVs) released by microglia may be responsible for these early events in AD.Method: Large EVs carrying Aβ peptides (Aβ‐EVs) were obtained from primary microglia exposed to Aβ_42. Optical manipulation was combined with time lapse imaging to study EV‐neuron interaction. Then, EVs were stereotaxically injected into naïve mice entorhinal cortex (EC), a vulnerable region in AD, to evaluate: i) long‐term potentiation (LTP) in EC and its main target region, the dentate gyrus of the hippocampus (DG), through field potential extracellular recordings in slices; ii) possible alterations in cortico‐hippocampal network activity by chronic EEG recordings; iii) progressive memory impairment in behavioural tasks. Finally, taking advantage of high‐resolution accurate‐mass spectrometry SWATH™‐MS, we investigated the molecular differences between Aβ‐ and ctrl‐EVs (from cells not exposed to Aβ). The influence of mesenchymal stem cell (MSC) indirect co‐colture with microglia primed with Aβ on cell phenotype, EVs and functions is also being explored.Result: Aβ‐EVs were able to move along the axonal surface more efficiently than ctrl‐EVs and predominantly in an anterograde direction, altering dendritic spine morphology in vitro. LTP was impaired in the EC 1h after Aβ‐EV (but not ctrl‐EV) local injection, while 24h later LTP was impaired also in the DG, indicating a spreading of synaptic dysfunction between connected regions. This translated into network alterations and progressive memory deficits. Interestingly, Aβ₄₂ alone was unable to spread alterations. Aβ‐EVs limited in their motion could neither propagate impairment nor cause network abnormalities, thus implicating Aβ‐EV extracellular motion in these processes. Proteomic analysis provided insights into the molecular mechanisms underneath Aβ‐EV motion at the neuronal surface.Conclusion: Our data provide evidence of the involvement of microglial EVs in early synaptic dysfunction in AD, through a new mechanism ‐ extracellular motion at the neuronal surface ‐ not implying EV internalization, paving the way for novel therapeutic strategies. Reference: Gabrielli et al., Brain, 2022 https://doi.org/10.1093/brain/awac083 Fundings: 2018‐AARF‐588984, Horizon2020#874721PREMSTEM

Perineuronal nets (PNNs) surround specific neurons in the brain and are involved in various forms of plasticity and clinical conditions. However, our understanding of the PNN role in these phenomena is limited by the lack of highly quantitative maps of PNN distribution and association with specific cell types. Here, we present the first comprehensive atlas of PNN distribution (in Allen Brain Atlas coordinates) and colocalization with parvalbumin (PV) cells for over 600 regions of the adult mouse brain. Data analysis showed that PV expression is a good predictor of PNN aggregation. In the cortex, PNNs are dramatically enriched in layer 4 of all primary sensory areas in correlation with thalamocortical input density, and their distribution mirrors intracortical connectivity patterns. Gene expression analysis identified many PNN correlated genes. Strikingly, PNN anticorrelated transcripts were enriched in synaptic plasticity genes, generalizing PNN role as circuit stability factors. Overall, this atlas offers novel resources for understanding the organizational principles of the brain extracellular matrix.

Alzheimer’s disease (AD) is considered by many to be a synaptic failure. Synaptic function is in fact deeply affected in the very early disease phases and recognized as the main cause of AD-related cognitive impairment. While the reciprocal involvement of amyloid beta (Aβ) and tau peptides in these processes is under intense investigation, the crucial role of extracellular vesicles (EVs) released by different brain cells as vehicles for these molecules and as mediators of early synaptic alterations is gaining more and more ground in the field. In this review, we will summarize the current literature on the contribution of EVs derived from distinct brain cells to neuronal alterations and build a working model for EV-mediated propagation of synaptic dysfunction in early AD. A deeper understanding of EV–neuron interaction will provide useful targets for the development of novel therapeutic approaches aimed at hampering AD progression.

Many studies have revealed a central role of p38 MAPK in neuronal plasticity and the regulation of long-term changes in synaptic efficacy, such as long-term potentiation (LTP) and long-term depression (LTD). However, p38 MAPK is classically known as a responsive element to stress stimuli, including neuroinflammation. Specific to the pathophysiology of Alzheimer’s disease (AD), several studies have shown that the p38 MAPK cascade is activated either in response to the Aβ peptide or in the presence of tauopathies. Here, we describe the role of p38 MAPK in the regulation of synaptic plasticity and its implication in an animal model of neurodegeneration. In particular, recent evidence suggests the p38 MAPK α isoform as a potential neurotherapeutic target, and specific inhibitors have been developed and have proven to be effective in ameliorating synaptic and memory deficits in AD mouse models.