High resolution Imaging: 3-D EM reconstrucfion and STORM microscopy

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PROJECT SUMMARY (See instructions): The present impediments to detailed connetomic data of brain circuitry are largely technical. In this core we will set up two powerful imaging modalities that provide detailed information about the number and type of synapses that innervate a class of cortical neurons and in addition, provide the full listing of all the targets cells a neuron innervates. One of the two imaging modalities is a super-resolution fluorescence imaging method termed STochasfic Optical Reconstruction Microscopy (STORM) invented by one of the core leaders. This method offers nanometer-scale imaging resolution with molecular specificity needed to identify specific neuronal and synapse types. In the core, we will develop staining and imaging capabilities to allow visualization of neurons and associated synapses with sub-diffraction-limit resolution. We further aim at automating the imaging process to allow the entire synapse distribution of a neuron, in particular its presynaptic input, to be routinely reconstructed. The second imaging modality is serial electron microscopy. A new approach to electron microscopy will be supported in the core that allows high resolution reconstruction of neural circuits in large volumes of brain tissue. The approach uses a novel automated microtome that puts ultrathin sections of brain on tape and an automated secondary-electron-detecting scanning electron microscope to image these sections. Complementary to the STORM facility, this serial EM facility will aim at reconstruction of the entire postsynaptic target field of specific neurons. Staff members in the core will not only make these imaging facilities available to researchers in the three Pi's lab, but also work closely with researchers in these labs to develop specific imaging capabilities needed for the project goals