@article {46862, title = {From a meso- to micro-scale connectome: array tomography and mGRASP.}, journal = {Frontiers in Neuroanatomy}, volume = {9}, year = {2015}, month = {2015 Jun 04}, pages = {78}, abstract = {

Mapping mammalian synaptic connectivity has long been an important goal of neuroscience because knowing how neurons and brain areas are connected underpins an understanding of brain function. Meeting this goal requires advanced techniques with single synapse resolution and large-scale capacity, especially at multiple scales tethering the meso- and micro-scale connectome. Among several advanced LM-based connectome technologies, Array Tomography (AT) and mammalian GFP-Reconstitution Across Synaptic Partners (mGRASP) can provide relatively high-throughput mapping synaptic connectivity at multiple scales. AT- and mGRASP-assisted circuit mapping (ATing and mGRASPing), combined with techniques such as retrograde virus, brain clearing techniques, and activity indicators will help unlock the secrets of complex neural circuits. Here, we discuss these useful new tools to enable mapping of brain circuits at multiple scales, some functional implications of spatial synaptic distribution, and future challenges and directions of these endeavors.

}, issn = {1662-5129}, doi = {10.3389/fnana.2015.00078}, author = {Rah, Jong-Cheol and Feng, Linqing and Druckmann, Shaul and Lee, Hojin and Kim, Jinhyun} } @article {45852, title = {Live imaging of endogenous PSD-95 using ENABLED: a conditional strategy to fluorescently label endogenous proteins.}, journal = {Journal of Neuroscience}, volume = {34}, year = {2014}, month = {2014 Dec 10}, pages = {16698-712}, abstract = {

Stoichiometric labeling of endogenous synaptic proteins for high-contrast live-cell imaging in brain tissue remains challenging. Here, we describe a conditional mouse genetic strategy termed endogenous labeling via exon duplication (ENABLED), which can be used to fluorescently label endogenous proteins with near ideal properties in all neurons, a sparse subset of neurons, or specific neuronal subtypes. We used this method to label the postsynaptic density protein PSD-95 with mVenus without overexpression side effects. We demonstrated that mVenus-tagged PSD-95 is functionally equivalent to wild-type PSD-95 and that PSD-95 is present in nearly all dendritic spines in CA1 neurons. Within spines, while PSD-95 exhibited low mobility under basal conditions, its levels could be regulated by chronic changes in neuronal activity. Notably, labeled PSD-95 also allowed us to visualize and unambiguously examine otherwise-unidentifiable excitatory shaft synapses in aspiny neurons, such as parvalbumin-positive interneurons and dopaminergic neurons. Our results demonstrate that the ENABLED strategy provides a valuable new approach to study the dynamics of endogenous synaptic proteins in vivo.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3888-14.2014}, author = {Fortin, Dale A and Tillo, Shane E and Yang, Guang and Rah, Jong-Cheol and Melander, Joshua B and Bai, Suxia and Soler-Cede{\~n}o, Omar and Qin, Maozhen and Zemelman, Boris V and Guo, Caiying and Mao, Tianyi and Zhong, Haining} } @article {6674, title = {Thalamocortical input onto layer 5 pyramidal neurons measured using quantitative large-scale array tomography.}, journal = {Frontiers in Neural Circuits}, volume = {7}, year = {2013}, month = {2013}, pages = {177}, abstract = {

The subcellular locations of synapses on pyramidal neurons strongly influences dendritic integration and synaptic plasticity. Despite this, there is little quantitative data on spatial distributions of specific types of synaptic input. Here we use array tomography (AT), a high-resolution optical microscopy method, to examine thalamocortical (TC) input onto layer 5 pyramidal neurons. We first verified the ability of AT to identify synapses using parallel electron microscopic analysis of TC synapses in layer 4. We then use large-scale array tomography (LSAT) to measure TC synapse distribution on L5 pyramidal neurons in a 1.00 {\texttimes} 0.83 {\texttimes} 0.21 mm(3) volume of mouse somatosensory cortex. We found that TC synapses primarily target basal dendrites in layer 5, but also make a considerable input to proximal apical dendrites in L4, consistent with previous work. Our analysis further suggests that TC inputs are biased toward certain branches and, within branches, synapses show significant clustering with an excess of TC synapse nearest neighbors within 5-15 μm compared to a random distribution. Thus, we show that AT is a sensitive and quantitative method to map specific types of synaptic input on the dendrites of entire neurons. We anticipate that this technique will be of wide utility for mapping functionally-relevant anatomical connectivity in neural circuits.

}, issn = {1662-5110}, doi = {10.3389/fncir.2013.00177}, author = {Rah, Jong-Cheol and Bas, Erhan and Colonell, Jennifer and Mishchenko, Yuriy and Karsh, Bill and Fetter, Richard D and Myers, Eugene W and Chklovskii, Dmitri B and Svoboda, Karel and Harris, Timothy D and Isaac, John T R} }