Genetic Labeling of Synapses

Abstract Detailing the ways drugs of abuse physically alter dopaminergic circuits would provide new mechanisms for explaining addictive behaviors, future targets for therapeutic intervention, and insights into the nature of synaptic plasticity. We combine recent advances in genetic labeling with large volume serial electron microscopy to detail how normal dopaminergic (DA) axons interact with putative targets in the Nucleus Accumbens (NAc) and how those interactions change in mice briefly exposed to cocaine. We find that while most DA axonal boutons are devoid of obvious signs of synapses (i.e. synaptic vesicles or synaptic densities), many DA boutons physically interdigitate with dendrites or excitatory and inhibitory axons. A brief exposure to cocaine results in large-scale remodeling: extensive DA axonal branching and frequent occurrences of axonal blind-ended “bulbs”, filled with mitochondria and reminiscent of axonal retraction in the developing and damaged brain. The number of physical interdigitations and vesicle filled boutons in DA axons scales linearly with the length of axon in both controls and cocaine exposed animals and the size or the type of interaction (i.e. axo-axonic or axo-dendritic) do not change. Finally, we find in cocaine exposed animals, mitochondrial lengths are increased ~2.5 times relative to control. Mitochondrial elongation is cell type specific: primarily in DA neurons and downstream spiny dendrites, and localized to DA axons and not DA soma or dendrites. We show for the first time the effects of cocaine on remodeling of dopamine axon morphology and mitochondria and reveal new details on how dopamine neurons physically associate with downstream targets.

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Light microscopy based approach for mapping connectivity with molecular specificity

10.1101/2020.02.24.963538 ◽

2020 ◽

Cited By ~ 1

Author(s):

Fred Y. Shen ◽

Margaret M. Harrington ◽

Logan A. Walker ◽

Hon Pong Jimmy Cheng ◽

Edward S. Boyden ◽

...

Keyword(s):

Light Microscopy ◽

Brain Function ◽

Inhibitory Neuron ◽

Cell Types ◽

Synaptic Connections ◽

Connectivity Analysis ◽

Brain Section ◽

Molecular Specificity ◽

Molecular Information ◽

Genetic Labeling

AbstractMapping neuroanatomy is a foundational goal towards understanding brain function. Electron microscopy (EM) has been the gold standard for connectivity analysis because nanoscale resolution is necessary to unambiguously resolve chemical and electrical synapses. However, molecular information that specifies cell types is often lost in EM reconstructions. To address this, we devised a light microscopy approach for connectivity analysis of defined cell types called spectral connectomics. We combined multicolor genetic labeling (Brainbow) of neurons with a multi-round immunostaining Expansion Microscopy (miriEx) strategy to simultaneously interrogate morphology, molecular markers, and connectivity in the same brain section. We applied our multimodal profiling strategy to directly link inhibitory neuron cell types with their network morphologies. Furthermore, we showed that correlative Brainbow and endogenous synaptic machinery immunostaining can be used to define putative synaptic connections between spectrally unique neurons, as well as map putative inhibitory and excitatory inputs. We envision that spectral connectomics can be applied routinely in neurobiology labs to gain insights into normal and pathophysiological neuroanatomy across multiple animals and time points.

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Integration of signals from different cortical areas in higher order thalamic neurons

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2104137118 ◽

2021 ◽

Vol 118 (30) ◽

pp. e2104137118

Author(s):

Vandana Sampathkumar ◽

Andrew Miller-Hansen ◽

S. Murray Sherman ◽

Narayanan Kasthuri

Keyword(s):

Higher Order ◽

Cortical Layer ◽

Thalamic Neurons ◽

Cortical Areas ◽

Medial Nucleus ◽

Afferent Information ◽

Posterior Medial Nucleus ◽

Depth And Complexity ◽

Genetic Labeling

Higher order thalamic neurons receive driving inputs from cortical layer 5 and project back to the cortex, reflecting a transthalamic route for corticocortical communication. To determine whether or not individual neurons integrate signals from different cortical populations, we combined electron microscopy “connectomics” in mice with genetic labeling to disambiguate layer 5 synapses from somatosensory and motor cortices to the higher order thalamic posterior medial nucleus. A significant convergence of these inputs was found on 19 of 33 reconstructed thalamic cells, and as a population, the layer 5 synapses were larger and located more proximally on dendrites than were unlabeled synapses. Thus, many or most of these thalamic neurons do not simply relay afferent information but instead integrate signals as disparate in this case as those emanating from sensory and motor cortices. These findings add further depth and complexity to the role of the higher order thalamus in overall cortical functioning.

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Efficient transient genetic labeling of human CD34+progenitor cells forin vivoapplication

Regenerative Medicine ◽

10.2217/17460751.1.2.223 ◽

2006 ◽

Vol 1 (2) ◽

pp. 223-234 ◽

Cited By ~ 7

Author(s):

Juliane MI Wiehe ◽

Carola Niesler ◽

Jan Torzewski ◽

Oliver Zimmermann ◽

Markus Wiesneth ◽

...

Keyword(s):

Progenitor Cells ◽

Human Cd34 ◽

Genetic Labeling

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Tumor-selective, adenoviral-mediated GFP genetic labeling of human cancer in the live mouse reports future recurrence after resection

Cell Cycle ◽

10.4161/cc.10.16.16756 ◽

2011 ◽

Vol 10 (16) ◽

pp. 2737-2741 ◽

Cited By ~ 58

Author(s):

Hiroyuki Kishimoto ◽

Ryoichi Aki ◽

Yasuo Urata ◽

Michael Bouvet ◽

Masashi Momiyama ◽

...

Keyword(s):

Human Cancer ◽

Genetic Labeling ◽

Tumor Selective ◽

Live Mouse

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Isolation by genetic labeling of a new mycobacterial plasmid, pJAZ38, from Mycobacterium fortuitum.

Journal of Bacteriology ◽

10.1128/jb.179.13.4115-4122.1997 ◽

1997 ◽

Vol 179 (13) ◽

pp. 4115-4122 ◽

Cited By ~ 27

Author(s):

J A Gavigan ◽

J A Aínsa ◽

E Pérez ◽

I Otal ◽

C Martín

Keyword(s):

Mycobacterium Fortuitum ◽

Genetic Labeling

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Cortical distribution of GABAergic interneurons is determined by migration time and brain size

Development ◽

10.1242/dev.185033 ◽

2020 ◽

Vol 147 (14) ◽

pp. dev185033

Author(s):

Pietro Fazzari ◽

Niall Mortimer ◽

Odessa Yabut ◽

Daniel Vogt ◽

Ramon Pla

Keyword(s):

Molecular Mechanisms ◽

Brain Size ◽

Migration Time ◽

Cortical Interneurons ◽

Rostrocaudal Axis ◽

Genetic Labeling ◽

Cortical Distribution ◽

Transplantation Experiments ◽

Host Brain

ABSTRACTCortical interneurons (CINs) originate in the ganglionic eminences (GEs) and migrate tangentially to the cortex guided by different attractive and repulsive cues. Once inside the cortex, the cellular and molecular mechanisms determining the migration of CINs along the rostrocaudal axis are less well understood. Here, we investigated the cortical distribution of CINs originating in the medial and caudal GEs at different time points. Using molecular and genetic labeling, we showed that, in the mouse, early- and late-born CINs (E12 versus E15) are differentially distributed along the rostrocaudal axis. Specifically, late-born CINs are preferentially enriched in cortical areas closer to their respective sites of origin in the medial or caudal GE. Surprisingly, our in vitro experiments failed to show a preferential migration pattern along the rostrocaudal axis for medial- or caudal-born CINs. Moreover, in utero transplantation experiments suggested that the rostrocaudal dispersion of CINs depends on the developmental stage of the host brain and is limited by the migration time and the increasing size of the developing brain. These data suggest that the embryonic expansion of the cortex contributes to the rostrocaudal distribution of CINs.

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