Misfolded protein aggregates represent a continuum with overlapping features in neurodegenerative diseases, but differences in protein components and affected brain regions. The molecular hallmark of synucleinopathies such as Parkinson's disease, dementia with Lewy bodies and multiple system atrophy are megadalton α-synuclein-rich deposits suggestive of one molecular event causing distinct disease phenotypes. Glial α-synuclein (α-SYN) filamentous deposits are prominent in multiple system atrophy and neuronal α-SYN inclusions are found in Parkinson's disease and dementia with Lewy bodies. The discovery of α-SYN assemblies with different structural characteristics or 'strains' has led to the hypothesis that strains could account for the different clinico-pathological traits within synucleinopathies. In this study we show that α-SYN strain conformation and seeding propensity lead to distinct histopathological and behavioural phenotypes. We assess the properties of structurally well-defined α-SYN assemblies (oligomers, ribbons and fibrils) after injection in rat brain. We prove that α-SYN strains amplify in vivo. Fibrils seem to be the major toxic strain, resulting in progressive motor impairment and cell death, whereas ribbons cause a distinct histopathological phenotype displaying Parkinson's disease and multiple system atrophy traits. Additionally, we show that α-SYN assemblies cross the blood-brain barrier and distribute to the central nervous system after intravenous injection. Our results demonstrate that distinct α-SYN strains display differential seeding capacities, inducing strain-specific pathology and neurotoxic phenotypes.

SummaryNeuronal function is highly sensitive to changes in oxygen levels, but how hypoxia affects dendritic spine formation and synaptogenesis is unknown. Here we report that hypoxia, chemical inhibition of the oxygen-sensing prolyl hydroxylase domain proteins (PHDs), and silencing of Phd2 induce immature filopodium-like dendritic protrusions, promote spine regression, reduce synaptic density, and decrease the frequency of spontaneous action potentials independently of HIF signaling. We identified the actin cross-linker filamin A (FLNA) as a target of PHD2 mediating these effects. In normoxia, PHD2 hydroxylates the proline residues P2309 and P2316 in FLNA, leading to von Hippel-Lindau (VHL)-mediated ubiquitination and proteasomal degradation. In hypoxia, PHD2 inactivation rapidly upregulates FLNA protein levels because of blockage of its proteasomal degradation. FLNA upregulation induces more immature spines, whereas Flna silencing rescues the immature spine phenotype induced by PHD2 inhibition.

Micro-Electrode Arrays (MEAs) have emerged as a mature technique to investigate brain (dys)functions in vivo and in in vitro animal models. Often referred to as "smart" Petri dishes, MEAs have demonstrated a great potential particularly for medium-throughput studies in vitro, both in academic and pharmaceutical industrial contexts. Enabling rapid comparison of ionic/pharmacological/genetic manipulations with control conditions, MEAs are employed to screen compounds by monitoring non-invasively the spontaneous and evoked neuronal electrical activity in longitudinal studies, with relatively inexpensive equipment. However, in order to acquire sufficient statistical significance, recordings last up to tens of minutes and generate large amount of raw data (e.g., 60 channels/MEA, 16 bits A/D conversion, 20 kHz sampling rate: approximately 8 GB/MEA,h uncompressed). Thus, when the experimental conditions to be tested are numerous, the availability of fast, standardized, and automated signal preprocessing becomes pivotal for any subsequent analysis and data archiving. To this aim, we developed an in-house cloud-computing system, named QSpike Tools, where CPU-intensive operations, required for preprocessing of each recorded channel (e.g., filtering, multi-unit activity detection, spike-sorting, etc.), are decomposed and batch-queued to a multi-core architecture or to a computers cluster. With the commercial availability of new and inexpensive high-density MEAs, we believe that disseminating QSpike Tools might facilitate its wide adoption and customization, and inspire the creation of community-supported cloud-computing facilities for MEAs users.

Cell assemblies manipulation by optogenetics is pivotal to advance neuroscience and neuroengineering. In in vivo applications, photostimulation often broadly addresses a population of cells simultaneously, leading to feed-forward and to reverberating responses in recurrent microcircuits. The former arise from direct activation of targets downstream, and are straightforward to interpret. The latter are consequence of feedback connectivity and may reflect a variety of time-scales and complex dynamical properties. We investigated wide-field photostimulation in cortical networks in vitro, employing substrate-integrated microelectrode arrays and long-term cultured neuronal networks. We characterized the effect of brief light pulses, while restricting the expression of channelrhodopsin to principal neurons. We evoked robust reverberating responses, oscillating in the physiological gamma frequency range, and found that such a frequency could be reliably manipulated varying the light pulse duration, not its intensity. By pharmacology, mathematical modelling, and intracellular recordings, we conclude that gamma oscillations likely emerge as in vivo from the excitatory-inhibitory interplay and that, unexpectedly, the light stimuli transiently facilitate excitatory synaptic transmission. Of relevance for in vitro models of (dys)functional cortical microcircuitry and in vivo manipulations of cell assemblies, we give for the first time evidence of network-level consequences of the alteration of synaptic physiology by optogenetics.

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