Improved methods for marking active neuron populations

Benjamien Moeyaert; Graham Holt; Rajtarun Madangopal; Alberto Perez-Alvarez; Brenna C Fearey; Nicholas F Trojanowski; Julia Ledderose; Timothy A Zolnik; Aniruddha Das; Davina Patel; Timothy A Brown; Robert N S Sachdev; Britta J Eickholt; Matthew E Larkum; Gina G. Turrigiano; Hod Dana; Christine E Gee; Thomas G Oertner; Bruce T Hope; Eric R Schreiter

doi:10.1038/s41467-018-06935-2

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Improved methods for marking active neuron populations

Journal article

Open access

Peer reviewed

Improved methods for marking active neuron populations

Benjamien Moeyaert, Graham Holt, Rajtarun Madangopal, Alberto Perez-Alvarez, Brenna C Fearey, Nicholas F Trojanowski, Julia Ledderose, Timothy A Zolnik, Aniruddha Das, Davina Patel, …

Nature communications, Vol.9(1), p.4440

10/25/2018

DOI: https://doi.org/10.1038/s41467-018-06935-2

PMCID: PMC6202339

PMID: 30361563

Abstract

Antibodies - metabolism

Protein Engineering - methods

Rats, Wistar

Humans

Neurons - cytology

Animals

Light

Zebrafish - metabolism

Mice

Neurons - metabolism

HeLa Cells

Visual Cortex - metabolism

Luminescent Proteins - metabolism

Fluorescence

Neuroanatomy

Neurobiology

Neurons

Neuroscience

Visual Perception

Marking functionally distinct neuronal ensembles with high spatiotemporal resolution is a key challenge in systems neuroscience. We recently introduced CaMPARI, an engineered fluorescent protein whose green-to-red photoconversion depends on simultaneous light exposure and elevated calcium, which enabled marking active neuronal populations with single-cell and subsecond resolution. However, CaMPARI (CaMPARI1) has several drawbacks, including background photoconversion in low calcium, slow kinetics and reduced fluorescence after chemical fixation. In this work, we develop CaMPARI2, an improved sensor with brighter green and red fluorescence, faster calcium unbinding kinetics and decreased photoconversion in low calcium conditions. We demonstrate the improved performance of CaMPARI2 in mammalian neurons and in vivo in larval zebrafish brain and mouse visual cortex. Additionally, we herein develop an immunohistochemical detection method for specific labeling of the photoconverted red form of CaMPARI. The anti-CaMPARI-red antibody provides strong labeling that is selective for photoconverted CaMPARI in activated neurons in rodent brain tissue.

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Title: Improved methods for marking active neuron populations
Creators: Benjamien Moeyaert - Howard Hughes Medical Institute, Janelia Research Campus, 19700 Helix Drive, Ashburn, VA, 20147, USA
Graham Holt - Program in Computational Biology and Bioinformatics, Duke University, Durham, NC, 27708, USA
Rajtarun Madangopal - Neuronal Ensembles in Addiction Section, Behavioral Neuroscience Research Branch, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Baltimore, MD, 21224, USA
Alberto Perez-Alvarez - Institute for Synaptic Physiology, University Medical Center Hamburg-Eppendorf, 20251, Hamburg, Germany
Brenna C Fearey - Institute for Synaptic Physiology, University Medical Center Hamburg-Eppendorf, 20251, Hamburg, Germany
Nicholas F Trojanowski - Department of Biology, Brandeis University, Waltham, MA, 02454, USA
Julia Ledderose - Institute of Biochemistry, Charité - Universitätsmedizin Berlin, 10117, Berlin, Germany
Timothy A Zolnik - NeuroCure Cluster of Excellence, Department of Biology, Humboldt University, 10117, Berlin, Germany
Aniruddha Das - Department of Neurosciences, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH, 44195, USA
Davina Patel - Department of Neurosciences, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH, 44195, USA
Timothy A Brown - Howard Hughes Medical Institute, Janelia Research Campus, 19700 Helix Drive, Ashburn, VA, 20147, USA
Robert N S Sachdev - NeuroCure Cluster of Excellence, Department of Biology, Humboldt University, 10117, Berlin, Germany
Britta J Eickholt - NeuroCure Cluster of Excellence, Department of Biology, Humboldt University, 10117, Berlin, Germany
Matthew E Larkum - NeuroCure Cluster of Excellence, Department of Biology, Humboldt University, 10117, Berlin, Germany
Gina G. Turrigiano - Brandeis University, Department of Biology
Hod Dana - Department of Neurosciences, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH, 44195, USA
Christine E Gee - Institute for Synaptic Physiology, University Medical Center Hamburg-Eppendorf, 20251, Hamburg, Germany
Thomas G Oertner - Institute for Synaptic Physiology, University Medical Center Hamburg-Eppendorf, 20251, Hamburg, Germany
Bruce T Hope - Neuronal Ensembles in Addiction Section, Behavioral Neuroscience Research Branch, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Baltimore, MD, 21224, USA
Eric R Schreiter - Howard Hughes Medical Institute, Janelia Research Campus, 19700 Helix Drive, Ashburn, VA, 20147, USA. schreitere@janelia.hhmi.org
Publication Details: Nature communications, Vol.9(1), p.4440
Publisher: Springer Nature; England
Grant note: R37 NS092635 / NINDS NIH HHS F32 NS101832 / NINDS NIH HHS
Identifiers: 9923906479901921
Academic Unit: Benjamin and Mae Volen National Center for Complex Systems; Department of Biology; Interdepartmental Program in Neuroscience
Language: English
Resource Type: Journal article

Improved methods for marking active neuron populations

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