Ultrawidefield microscope for high-speed fluorescence imaging and targeted optogenetic stimulation

Area of Science:

• Cellular and Molecular Neuroscience
• Optical Engineering
• Biomedical Imaging

Background:

• Advances in fluorescent reporters and optogenetic tools necessitate improved optical systems.
• Current systems often lack high light collection efficiency, spatial/temporal resolution, and wide field of view.
• Need for advanced microscopy for cellular state recording and control.

Purpose of the Study:

• To describe the 'Firefly' microscope, an optical system designed for enhanced cellular imaging and stimulation.
• To achieve high light collection efficiency, resolution, and patterned optical stimulation in a wide field of view.
• To enable high-throughput all-optical electrophysiology for neuronal phenotyping and drug screening.

Main Methods:

• Designed and built the 'Firefly' microscope using mostly commercially available optical components.
• Optimized the optical system for simultaneous photostimulation and fluorescence imaging in cultured cells.
• Evaluated light collection efficiency compared to commercial microscopes.

Main Results:

• The 'Firefly' microscope achieves a Ø6 mm field of view with 10-fold higher light collection efficiency than comparable commercial systems.
• Demonstrated all-optical electrophysiology ('Optopatch') in cultured neurons with unprecedented throughput and information content.
• Successfully applied the system for voltage and calcium recordings in human induced pluripotent stem cell-derived cardiomyocytes.

Conclusions:

• The 'Firefly' microscope offers a powerful, efficient, and versatile platform for advanced cellular research.
• Enables high-throughput neuronal phenotyping, disease modeling, and drug screening.
• Provides new capabilities for studying cellular function in various cell types, including cardiomyocytes.