# Open-source miniaturized two-photon microscopes for large field-of-view and volumetric imaging

> **NIH NIH U01** · UNIVERSITY OF CALIFORNIA LOS ANGELES · 2023 · $982,172

## Abstract

Abstract:
Single-photon (1P) epifluorescence miniaturized microscopy coupled with genetically encoded calcium sensors
has allowed investigators to record the activity of large populations of identified neurons over days to weeks in
freely behaving animals, answering fundamental questions in neuroscience. Our group's efforts with the UCLA
Miniscope Project have allowed over 600 labs to build and use over 2500 open-source miniaturized microscopes
with expanded capabilities at a small fraction of the cost of those offered by commercial versions, thus
democratizing access. Yet, 1P miniscopes lack the lateral and axial resolution to image activity in fine structures
such as dendrites and axons. In addition, 1P imaging is limited to superficial structures or requires removal of
overlying tissue for imaging of deeper neurons. Two-photon (2P) microscopy has exquisite lateral and axial
resolution and bypasses all of these obstacles. Recent advances in technology have made the construction of
two-photon miniaturized microscopes for mice possible. However, the field of view (FOV) is still limited, and
these microscopes require custom-built optics and cost several hundred thousand dollars to acquire
commercially. We have designed and built a two-photon miniaturized microscope for mice, including a custom-
made objective lens, that allows 2P imaging of an 800 micrometer FOV nearly quadrupling the FOV from the
latest published 2P miniaturized microscope (Mini2P-V1). In this proposal, we will optimize this microscope and
test it in freely behaving mice for axonal, dendritic and deep somatic imaging. This microscope will be tested in
three labs. The Golshani Lab will test the scope with calcium imaging of thalamic axons in anterior cingulate
cortex during social interaction. The Silva Lab will test the scope by performing dendritic calcium and glutamate
imaging in retrosplenial cortex during memory linking. The Shtrahman Lab will test deep imaging capability by
imaging dentate granule neurons through an intact CA1. We will also build a larger miniaturized microscope
suitable for rats and non-human primates with expanded capabilities, including a higher numerical aperture (NA),
large FOV and temporal multiplexing capability to allow volumetric imaging at high frame rates (MiniMux2P).
This microscope will be tested by the Blair Lab to dissect the role of superficial and deep CA1 neurons of rats in
navigation. It will also be tested in the Churchland Lab to image rat posterior parietal cortical neurons during
decision-making tasks. Finally, we will disseminate the technology using our open-source wiki that has already
disseminated miniscope technology to thousands of users. We will provide parts-lists, optical designs and
methods for obtaining custom lens elements. As we have done before, we will educate users through online
videos and hands-on workshops where imaging basics, surgical techniques and analysis tools are demonstrated.
We hope these cutting edge, novel...

## Key facts

- **NIH application ID:** 10675751
- **Project number:** 5U01NS128664-02
- **Recipient organization:** UNIVERSITY OF CALIFORNIA LOS ANGELES
- **Principal Investigator:** Daniel Aharoni
- **Activity code:** U01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $982,172
- **Award type:** 5
- **Project period:** 2022-08-15 → 2026-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10675751

## Citation

> US National Institutes of Health, RePORTER application 10675751, Open-source miniaturized two-photon microscopes for large field-of-view and volumetric imaging (5U01NS128664-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10675751. Licensed CC0.

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