This should enable neuroscientists to obtain more rapid insights into the roles of specific neural circuits in the brains of healthy animals, and to identify cases where this wiring goes awry and leads to disease. This pipeline of steps reduces the time required to map the shapes and connectivity of neurons with electron microscopy by some two orders of magnitude. Lastly, an algorithm pieces together the individual images to produce a reconstruction of the cell. Small areas of that neuron are then re-imaged at higher resolution to map the connections between neurons.
Next, a labeled target neuron is imaged at relatively low resolution to reveal its overall structure. Then, a tissue sample is collected and treated with a chemical that enhances the electron density of the stained neurons, without disrupting the tissue’s structure. First, gene technology is used to selectively stain specific types of neurons in mice and flies. ARTEMIS makes use of genetic engineering, serial-scanning electron microscopy, an enhanced chemical staining procedure and a new image processing approach. have now developed a new procedure – named ARTEMIS – that uses a combination of multiple techniques to speed up the mapping of neurons and their connections. These are imaged individually, and the images are then pieced together to reconstruct the sample. In an approach called serial-section electron microscopy, a tissue sample is first cut into extremely thin sections. Labeled cells scatter more electrons, which increases the contrast of the images. Stains are used to make specific neurons less permeable to electrons, or more “electron dense”. However, this technique is so time-consuming that thousands of hours of work are typically required to image even the smallest of tissue samples.Įlectron microscopes fire beams of electrons at tissue samples, and detect the scattering of the electrons. The only way to do this precisely enough is by using electron microscopy. Mapping these connections to obtain a so-called wiring diagram is an essential step in learning how the brain works. Neurons connect with each other to form complex circuits that underlie mental activities. This pipeline reduces imaging and reconstruction times by two orders of magnitude, facilitating directed inquiry of circuit motifs. The high contrast of the marked neurons enabled two innovations that speed data acquisition: targeted high-resolution reimaging of regions selected from rapidly-acquired lower resolution reconstruction, and an unsupervised segmentation algorithm. We used viral vectors to deliver peroxidase derivatives, which catalyze production of an electron-dense tracer, to genetically identify neurons, and developed a protocol that enhances the electron-density of the labeled cells while retaining the quality of the ultrastructure. Here, we present an alternative strategy, targeted reconstruction of specific neuronal types. However, complete circuit reconstruction is prohibitively slow and may not be necessary for many purposes such as comparing neuronal structure and connectivity among multiple animals.
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Resolving patterns of synaptic connectivity in neural circuits currently requires serial section electron microscopy.