Subcellular Imaging of Voltage and Calcium Signals Reveals Neural Processing In Vivo.

A mechanistic understanding of neural computation requires determining how information is processed as it passes through neurons and across synapses. However, it has been challenging to measure membrane potential changes in axons and dendrites in vivo. We use in vivo, two-photon imaging of novel genetically encoded voltage indicators, as well as calcium imaging, to measure sensory stimulus-evoked signals in the Drosophila visual system with subcellular resolution. Across synapses, we find major transformations in the kinetics, amplitude, and sign of voltage responses to light. We also describe distinct relationships between voltage and calcium signals in different neuronal compartments, a substrate for local computation. Finally, we demonstrate that ON and OFF selectivity, a key feature of visual processing across species, emerges through the transformation of membrane potential into intracellular calcium concentration. By imaging voltage and calcium signals to map information flow with subcellular resolution, we illuminate where and how critical computations arise.

Cortical preparatory activity indexes learned motor memories.

The brain's remarkable ability to learn and execute various motor behaviours harnesses the capacity of neural populations to generate a variety of activity patterns. Here we explore systematic changes in preparatory activity in motor cortex that accompany motor learning. We trained rhesus monkeys to learn an arm-reaching task1 in a curl force field that elicited new muscle forces for some, but not all, movement directions2,3. We found that in a neural subspace predictive of hand forces, changes in preparatory activity tracked the learned behavioural modifications and reassociated4 existing activity patterns with updated movements. Along a neural population dimension orthogonal to the force-predictive subspace, we discovered that preparatory activity shifted uniformly for all movement directions, including those unaltered by learning. During a washout period when the curl field was removed, preparatory activity gradually reverted in the force-predictive subspace, but the uniform shift persisted. These persistent preparatory activity patterns may retain a motor memory of the learned field5,6 and support accelerated relearning of the same curl field. When a set of distinct curl fields was learned in sequence, we observed a corresponding set of field-specific uniform shifts which separated the associated motor memories in the neural state space7-9. The precise geometry of these uniform shifts in preparatory activity could serve to index motor memories, facilitating the acquisition, retention and retrieval of a broad motor repertoire.

Brain-derived neurotrophic factor (BDNF)-induced mitochondrial motility arrest and presynaptic docking contribute to BDNF-enhanced synaptic transmission.

Appropriate mitochondrial transport and distribution are essential for neurons because of the high energy and Ca(2+) buffering requirements at synapses. Brain-derived neurotrophic factor (BDNF) plays an essential role in regulating synaptic transmission and plasticity. However, whether and how BDNF can regulate mitochondrial transport and distribution are still unclear. Here, we find that in cultured hippocampal neurons, application of BDNF for 15 min decreased the percentage of moving mitochondria in axons, a process dependent on the activation of the TrkB receptor and its downstream PI3K and phospholipase-Cγ signaling pathways. Moreover, the BDNF-induced mitochondrial stopping requires the activation of transient receptor potential canonical 3 and 6 (TRPC3 and TRPC6) channels and elevated intracellular Ca(2+) levels. The Ca(2+) sensor Miro1 plays an important role in this process. Finally, the BDNF-induced mitochondrial stopping leads to the accumulation of more mitochondria at presynaptic sites. Mutant Miro1 lacking the ability to bind Ca(2+) prevents BDNF-induced mitochondrial presynaptic accumulation and synaptic transmission, suggesting that Miro1-mediated mitochondrial motility is involved in BDNF-induced mitochondrial presynaptic docking and neurotransmission. Together, these data suggest that mitochondrial transport and distribution play essential roles in BDNF-mediated synaptic transmission.

Spyglass: a framework for reproducible and shareable neuroscience research.

Scientific progress depends on reliable and reproducible results. Progress can also be accelerated when data are shared and re-analyzed to address new questions. Current approaches to storing and analyzing neural data typically involve bespoke formats and software that make replication, as well as the subsequent reuse of data, difficult if not impossible. To address these challenges, we created Spyglass, an open-source software framework that enables reproducible analyses and sharing of data and both intermediate and final results within and across labs. Spyglass uses the Neurodata Without Borders (NWB) standard and includes pipelines for several core analyses in neuroscience, including spectral filtering, spike sorting, pose tracking, and neural decoding. It can be easily extended to apply both existing and newly developed pipelines to datasets from multiple sources. We demonstrate these features in the context of a cross-laboratory replication by applying advanced state space decoding algorithms to publicly available data. New users can try out Spyglass on a Jupyter Hub hosted by HHMI and 2i2c: https://spyglass.hhmi.2i2c.cloud/.

Dendritic calcium signals in rhesus macaque motor cortex drive an optical brain-computer interface.

Calcium imaging is a powerful tool for recording from large populations of neurons in vivo. Imaging in rhesus macaque motor cortex can enable the discovery of fundamental principles of motor cortical function and can inform the design of next generation brain-computer interfaces (BCIs). Surface two-photon imaging, however, cannot presently access somatic calcium signals of neurons from all layers of macaque motor cortex due to photon scattering. Here, we demonstrate an implant and imaging system capable of chronic, motion-stabilized two-photon imaging of neuronal calcium signals from macaques engaged in a motor task. By imaging apical dendrites, we achieved optical access to large populations of deep and superficial cortical neurons across dorsal premotor (PMd) and gyral primary motor (M1) cortices. Dendritic signals from individual neurons displayed tuning for different directions of arm movement. Combining several technical advances, we developed an optical BCI (oBCI) driven by these dendritic signalswhich successfully decoded movement direction online. By fusing two-photon functional imaging with CLARITY volumetric imaging, we verified that many imaged dendrites which contributed to oBCI decoding originated from layer 5 output neurons, including a putative Betz cell. This approach establishes new opportunities for studying motor control and designing BCIs via two photon imaging.

Accurate Estimation of Neural Population Dynamics without Spike Sorting.

A central goal of systems neuroscience is to relate an organism's neural activity to behavior. Neural population analyses often reduce the data dimensionality to focus on relevant activity patterns. A major hurdle to data analysis is spike sorting, and this problem is growing as the number of recorded neurons increases. Here, we investigate whether spike sorting is necessary to estimate neural population dynamics. The theory of random projections suggests that we can accurately estimate the geometry of low-dimensional manifolds from a small number of linear projections of the data. We recorded data using Neuropixels probes in motor cortex of nonhuman primates and reanalyzed data from three previous studies and found that neural dynamics and scientific conclusions are quite similar using multiunit threshold crossings rather than sorted neurons. This finding unlocks existing data for new analyses and informs the design and use of new electrode arrays for laboratory and clinical use.

Blockage of GSK3ß-mediated Drp1 phosphorylation provides neuroprotection in neuronal and mouse models of Alzheimer's disease.

It is well established that mitochondrial fragmentation plays a key role in the pathogenesis of Alzheimer's disease (AD). Mitochondrial fission is mediated by dynamin-related protein 1 (Drp1), which is highly expressed in nervous system and regulated by various posttranslational modifications including phosphorylation. We identified glycogen synthase kinase (GSK)3ß-dependent Drp1 phosphorylation at Ser(40) and Ser(44), which increases Drp1 GTPase activity and its mitochondrial distribution and could induce mitochondrial fragmentation. Moreover, neurons transfected with Ser(40)Ser(44) phosphomimic Drp1 showed increased mitochondria fragmentation and were more vulnerable to amyloid-ß (Aß)-induced apoptosis. Therefore, blocking GSK3ß-induced Drp1 phosphorylation may be an effective way to protect neurons from Aß toxicity. To address this, we designed and synthesized an artificial polypeptide named TAT-Drp1-SpS, which could specifically block GSK3ß-induced Drp1 phosphorylation. Our results demonstrated that TAT-Drp1-SpS treatment could significantly reduce Aß-induced neuronal apoptosis in cultured neurons. Notably, TAT-Drp1-SpS administration in hippocampus Cornu Ammonis 1 (CA1) region significantly reduced Aß burden and rescued the memory deficits in AD transgenic mice. Although Aß has multiple targets to exert its neurotoxicity, our findings suggested that GSK3ß-induced mitochondrial fragmentation was, at least partially, mediated by Aß toxicity and contribute to the pathogenesis of AD. Taken together, GSK3ß-induced Drp1 phosphorylation provides a novel mechanism for mitochondrial fragmentation in AD, and our findings suggested a novel therapeutic strategy for AD.

Dynamic changes in xylanases and ß-1,4-endoglucanases secreted by Aspergillus niger An-76 in response to hydrolysates of lignocellulose polysaccharide.

Aspergillus niger is an effective secretor of glycoside hydrolases that facilitate the saprophytic lifestyle of the fungus by degrading plant cell wall polysaccharides. In the present study, a series of dynamic zymography assays were applied to quantify the secreted glycoside hydrolases of A. niger cultured in media containing different carbon sources. Differences in the diversity and concentrations of polysaccharide hydrolysates dynamically regulated the secretion of glycoside hydrolases. The secretion of ß-1,4-endoglucanase isozymes was observed to lag at least 24 h behind, rather than coincide with, the secretion of xylanase isozymes. Low concentrations of xylose could induce many endoxylanases (such as Xyn1/XynA, Xyn2, and Xyn3/XynB). High concentrations of xylose could sustain the induction of Xyn2 and Xyn3/XynB but repress Xyn1/XynA (GH10 endoxylanase), which has a broad substrate specificity, and also triggers the low-level secretion of Egl3/EglA, which also has a broad substrate specificity. Mixed polysaccharide hydrolysates sustained the induction of Egl1, whereas the other ß-1,4-endoglucanases were sustainably induced by the specific polysaccharide hydrolysates released during the hydrolysis process (such as Egl2 and Egl4). These results indicate that the secretion of glycoside hydrolases may be specifically regulated by the production of polysaccharide hydrolysates released during the process of biomass degradation.