# Ultra-fast high-resolution imaging of whole mouse brain for the study of drug addiction

> **NIH NIH R21** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2021 · $249,919

## Abstract

PROJECT SUMMARY
Opioid and cocaine abuse prevalence has skyrocketed in the United States, fueling the current epidemic of
overdose deaths. Despite the public health impact of opioids and cocaine, we still lack a fundamental
understanding of the mechanisms by which these drugs work, particularly across cellular and circuit levels.
Further understanding of the neuroanatomy of the neural circuitry underlying opioid and cocaine reward is a
critical initial step in targeting and elucidating their mechanisms. However, comprehensively visualizing relevant
circuits in drug reward has been limited by approaches to contextualize these circuits and their response to drugs
of abuse in the whole brain. We developed an approach to rapidly image the whole brain in three-dimensional
(3D) space using ultra-fast high-resolution ribbon-scanning confocal microscopy. Our ribbon-scanning confocal
imaging approach can image and visualize an entire rodent brain in less than 24 hours, where more conventional
approaches (e.g., light-sheet) currently require days or even weeks. Furthermore, our ribbon-scanning confocal
approach reaches diffraction-limited resolutions (~200-300nm), enabling us to visualize individual cells in the
brain and their ultrastructure. We can apply these unique tools to begin solving the fundamental questions: 1)
What is the precise circuitry that defines drug reward? And 2) What are the differential effects of cocaine and
opioids on this circuitry? Like many drugs of abuse, cocaine and opioids rely on neurotransmission from
dopamine (DA) neurons in the ventral tegmental area (VTA). However, until recently, parsing the connectivity of
unique subpopulations of DA neurons and their potential roles in drug reward has been difficult. We developed
a suite of intersectional genetic tools to definitively dissect the anatomical and functional properties of these
different subpopulations within the same brain. We will integrate our 3D ribbon-scanning confocal imaging of DA
neuron subpopulations with immunolabeling of neuronal activity markers to visualize precisely which DA neurons
are activated in response to cocaine and opioids. Using whole brain immunolabeling and imaging, we will also
visualize and map drug-dependent neuronal activity changes in the whole brain with the potential to reveal new
populations of neurons differentially response to cocaine and opioids. Our overall objectives are to:
Comprehensively map the distribution of DA neuron subpopulations including DA/glutamate co-transmitting cells
relative to the overall DA system within whole brain (Aim 1); and to determine how cocaine and opioids
differentially affect the activity of these DA neuron subpopulations (Aim 2). We will generate a comprehensive
3D brain atlas to identify the roles of unique subpopulations of DA neurons highly relevant to cocaine and opioids,
which will serve as a proof of principle for the implementation of our ultra-fast high-resolution 3D ribbon-scanning
confocal micr...

## Key facts

- **NIH application ID:** 10101820
- **Project number:** 1R21DA052419-01
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** ZACHARY FREYBERG
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $249,919
- **Award type:** 1
- **Project period:** 2021-03-01 → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10101820

## Citation

> US National Institutes of Health, RePORTER application 10101820, Ultra-fast high-resolution imaging of whole mouse brain for the study of drug addiction (1R21DA052419-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10101820. Licensed CC0.

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