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Millimeter field-of-view miniature two-photon microscopy for brain imaging in freely moving mice
Abstract:
Development of miniature two-photon microscopy (m2PM) has made it possible to observe fine structure and activity of neurons in the brain of freely moving animals. However, the imaging field-of-view of existing m2PM is still significantly smaller than that of miniature single-photon microscopy. Here we report that, through the design of low-magnification objective, large field-of-view scan lens and small tilt angle microscanner, a 2.5-g m2PM achieved a field-of-view of 1000 × 788 µm2, comparable to that of a typical single-photon miniscope. We demonstrated its capability by imaging neurons, dendrites and spines in the millimeter field-of-view, and simultaneous recording calcium activities, through a gradient-index lens, of approximately 400 neurons in the dorsal hippocampal CA1 in a freely moving mouse. Integrated with a detachable 1.2-g fast z-scanning module, it enables a 1000 × 788 × 500 µm3 volumetric neuronal imaging in the cerebral cortex. Thus, millimeter FOV m2PM provides a powerful tool for deciphering neuronal population dynamics in experimental paradigms allowing for animal's free movement.
Insights
A new miniature two-photon microscopy (m2PM) offers a millimeter field-of-view for observing neural structures and activity in freely moving animals. This breakthrough enables broader insights into neuronal population dynamics.
Area of Science:
- Neuroscience
- Biomedical Engineering
- Microscopy Technology
Background:
- Miniature two-photon microscopy (m2PM) allows neural observation in freely moving animals.
- Current m2PM systems have limited field-of-view (FOV) compared to single-photon methods.
Purpose of the Study:
- To develop an m2PM with a significantly enlarged FOV for comprehensive neural imaging.
- To enable simultaneous recording of neuronal activity across large populations in behaving animals.
Main Methods:
- Designed a low-magnification objective, large FOV scan lens, and microscanner for m2PM.
- Achieved a 1000 × 788 µm² FOV with a 2.5-g device.
- Integrated a fast z-scanning module for volumetric imaging.
Main Results:
- Demonstrated millimeter-scale imaging of neurons, dendrites, and spines.
- Simultaneously recorded calcium activity from ~400 neurons in mouse hippocampus.
- Achieved 1000 × 788 × 500 µm³ volumetric imaging in the cortex.
Conclusions:
- The millimeter FOV m2PM significantly expands imaging capabilities in neuroscience.
- This technology is a powerful tool for studying neuronal population dynamics in freely moving subjects.
- Facilitates research in naturalistic behavioral paradigms.
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