An Algorithm Based on a Cable-Nernst Planck Model Predicting Synaptic Activity throughout the Dendritic Arbor with Micron Specificity.

Recent technological advances have enabled the recording of neurons in intact circuits with a high spatial and temporal resolution, creating the need for modeling with the same precision. In particular, the development of ultra-fast two-photon microscopy combined with fluorescence-based genetically-encoded Ca2+-indicators allows capture of full-dendritic arbor and somatic responses associated with synaptic input and action potential output. The complexity of dendritic arbor structures and distributed patterns of activity over time results in the generation of incredibly rich 4D datasets that are challenging to analyze (Sakaki et al. in Frontiers in Neural Circuits 14:33, 2020). Interpreting neural activity from fluorescence-based Ca2+ biosensors is challenging due to non-linear interactions between several factors influencing intracellular calcium ion concentration and its binding to sensors, including the ionic dynamics driven by diffusion, electrical gradients and voltage-gated conductances. To investigate those dynamics, we designed a model based on a Cable-like equation coupled to the Nernst-Planck equations for ionic fluxes in electrolytes. We employ this model to simulate signal propagation and ionic electrodiffusion across a dendritic arbor. Using these simulation results, we then designed an algorithm to detect synapses from Ca2+ imaging datasets. We finally apply this algorithm to experimental Ca2+-indicator datasets from neurons expressing jGCaMP7s (Dana et al. in Nature Methods 16:649-657, 2019), using full-dendritic arbor sampling in vivo in the Xenopus laevis optic tectum using fast random-access two-photon microscopy. Our model reproduces the dynamics of visual stimulus-evoked jGCaMP7s-mediated calcium signals observed experimentally, and the resulting algorithm allows prediction of the location of synapses across the dendritic arbor. Our study provides a way to predict synaptic activity and location on dendritic arbors, from fluorescence data in the full dendritic arbor of a neuron recorded in the intact and awake developing vertebrate brain.

Quantifying neuronal structural changes over time using dynamic morphometrics.

Brain circuit development involves tremendous structural formation and rearrangement of dendrites, axons, and the synaptic connections between them. Direct studies of neuronal morphogenesis are now possible through recent developments in multiple technologies, including single-neuron labeling, time-lapse imaging in intact tissues, and 4D rendering software capable of tracking neural growth over periods spanning minutes to days. These methods allow detailed quantification of structural changes of neurons over time, called dynamic morphometrics, providing new insights into fundamental growth patterns, underlying molecular mechanisms, and the intertwined influences of external factors, including neural activity, and intrinsic genetic programs. Here, we review the methods of dynamic morphometrics sampling and analyses, focusing on their applications to studies of activity-driven dendritogenesis in vertebrate systems.

Comprehensive Imaging of Sensory-Evoked Activity of Entire Neurons Within the Awake Developing Brain Using Ultrafast AOD-Based Random-Access Two-Photon Microscopy.

Determining how neurons transform synaptic input and encode information in action potential (AP) firing output is required for understanding dendritic integration, neural transforms and encoding. Limitations in the speed of imaging 3D volumes of brain encompassing complex dendritic arbors in vivo using conventional galvanometer mirror-based laser-scanning microscopy has hampered fully capturing fluorescent sensors of activity throughout an individual neuron's entire complement of synaptic inputs and somatic APs. To address this problem, we have developed a two-photon microscope that achieves high-speed scanning by employing inertia-free acousto-optic deflectors (AODs) for laser beam positioning, enabling random-access sampling of hundreds to thousands of points-of-interest restricted to a predetermined neuronal structure, avoiding wasted scanning of surrounding extracellular tissue. This system is capable of comprehensive imaging of the activity of single neurons within the intact and awake vertebrate brain. Here, we demonstrate imaging of tectal neurons within the brains of albino Xenopus laevis tadpoles labeled using single-cell electroporation for expression of a red space-filling fluorophore to determine dendritic arbor morphology, and either the calcium sensor jGCaMP7s or the glutamate sensor iGluSnFR as indicators of neural activity. Using discrete, point-of-interest scanning we achieve sampling rates of 3 Hz for saturation sampling of entire arbors at 2 µm resolution, 6 Hz for sequentially sampling 3 volumes encompassing the dendritic arbor and soma, and 200-250 Hz for scanning individual planes through the dendritic arbor. This system allows investigations of sensory-evoked information input-output relationships of neurons within the intact and awake brain.