Periodic Reporting for period 1 - HyFiPhotometry (Hyperspectral Fiber Photometry for Flexible, Multiplexed Optical Measurements of Brain Function)
Periodo di rendicontazione: 2024-06-01 al 2025-11-30
Understanding how the brain works requires tools that can both monitor and influence brain activity at the same time, but doing this reliably is still difficult. To address this challenge, we develop new technologies that overcome challenges that limit our ability to study brain function. One such method, fiber photometry, allows us to observe and control the activity of specific groups of brain cells in freely moving animals. Fiber photometry works by using a thin optical fiber implanted in the brain to detect light signals produced by special fluorescent markers that report cellular activity. This technique is already widely used to study normal brain function as well as disorders such as addiction, neurodegenerative diseases, and to support drug development.
However, currently available fiber photometry systems have important limitations. Most commercial systems can only measure one or two optical channels at a time, which restricts the complexity of experiments we can perform. While open-source alternatives exist, they often require advanced technical expertise and are therefore not accessible to many laboratories.
Our goal was to develop an improved, affordable system that can measure a broader range of light signals to expand research possibilities. This system could also have applications beyond neuroscience, including industrial quality control and medical diagnostics, making it valuable not only for science but also for society more broadly.
We previously developed a new approach called Fused Fiber Photometry (FFP). Instead of using bulky optical filters, FFP employs a specially designed fused optical fiber that both delivers light to the brain and collects fluorescent signals in return. This simpler and more elegant design makes the system highly adaptable, allowing researchers to easily change how and what they measure without rebuilding the entire setup.
A major advantage of this new approach is that it captures the full range of light signals rather than just a few predefined ones. This enables a more advanced method called hyperspectral fiber photometry, which provides much richer information about brain activity and underlying biological processes. For example, it becomes possible to monitor changes in blood oxygenation in the brain, something that was not feasible with conventional systems.
Beyond its scientific advantages, the FFP design is also practical and cost-effective. A basic commercial version could be produced at less than half the cost of existing systems, while offering far greater flexibility. Because of its modular design, it can be easily adapted to different research needs at low cost.
A key goal was to establish a method to monitor brain activity across a broad and continuous range of wavelengths. This would allow more detailed and accurate tracking of different biological signals in the brain at the same time. To achieve this, we were collaborating with academic and industry partners to further develop and optimize the components needed for hyperspectral fiber photometry.
The first priority was to refine and scale up production of the fused optical fiber at the core of the system. In parallel, we aimed to complete a basic, user-friendly version of the technology that can be commercialized.
However, currently available fiber photometry systems have important limitations. Most commercial systems can only measure one or two optical channels at a time, which restricts the complexity of experiments we can perform. While open-source alternatives exist, they often require advanced technical expertise and are therefore not accessible to many laboratories.
Our goal was to develop an improved, affordable system that can measure a broader range of light signals to expand research possibilities. This system could also have applications beyond neuroscience, including industrial quality control and medical diagnostics, making it valuable not only for science but also for society more broadly.
We previously developed a new approach called Fused Fiber Photometry (FFP). Instead of using bulky optical filters, FFP employs a specially designed fused optical fiber that both delivers light to the brain and collects fluorescent signals in return. This simpler and more elegant design makes the system highly adaptable, allowing researchers to easily change how and what they measure without rebuilding the entire setup.
A major advantage of this new approach is that it captures the full range of light signals rather than just a few predefined ones. This enables a more advanced method called hyperspectral fiber photometry, which provides much richer information about brain activity and underlying biological processes. For example, it becomes possible to monitor changes in blood oxygenation in the brain, something that was not feasible with conventional systems.
Beyond its scientific advantages, the FFP design is also practical and cost-effective. A basic commercial version could be produced at less than half the cost of existing systems, while offering far greater flexibility. Because of its modular design, it can be easily adapted to different research needs at low cost.
A key goal was to establish a method to monitor brain activity across a broad and continuous range of wavelengths. This would allow more detailed and accurate tracking of different biological signals in the brain at the same time. To achieve this, we were collaborating with academic and industry partners to further develop and optimize the components needed for hyperspectral fiber photometry.
The first priority was to refine and scale up production of the fused optical fiber at the core of the system. In parallel, we aimed to complete a basic, user-friendly version of the technology that can be commercialized.
During the project, we developed a wideband multimode circulator (WMC), a three-port fused-fiber optical device with an asymmetric design optimized for efficient light delivery and fluorescence collection. The WMC exhibits a flat spectral response, making it particularly well suited for simultaneous excitation and detection of fluorescence in biological tissue. The device was further optimized to minimize intrinsic autofluorescence. Using this approach, we demonstrated enhanced fluorescence detection compared to existing fiber-based methods and successfully applied the WMC to in vivo fiber photometry recordings to infer neuronal activity in a small neuromodulatory brainstem nucleus in awake mice.
Building on this technological development, we worked with an industry partner (Rapp Optoelectronic) to develop a commercial prototype of the fused-fiber photometry device, marking an important step toward broader dissemination and practical deployment of the technology.
In parallel, we made substantial progress toward establishing a dual-color fiber photometry system combined with optogenetic stimulation using a 633 nm laser. During this process, spectral crosstalk emerged as a major limitation, motivating a strategic shift to longer-wavelength stimulation at 650 nm. Although the full dual-recording setup could not be completed within the project timeline, in vivo validation of ChrimsonR activation with the 650 nm laser was successfully achieved, demonstrating the feasibility of this approach with appropriate power adjustments.
Finally, we developed a custom-built spectrometer that achieved higher light collection efficiency and greater flexibility than a commercial Ocean Optics system. This instrument enabled spectrally resolved fiber photometry with performance comparable to standard photodetectors for both slow and fast signals. At the same time, this work revealed important limitations, including constraints in spectral resolution, variability in biosensor emission profiles across animals, and challenges in spectral decomposition. These findings indicate that further improvements in optical design and the use of individualized reference spectra will be necessary to fully exploit the potential of hyperspectral fiber photometry.
Building on this technological development, we worked with an industry partner (Rapp Optoelectronic) to develop a commercial prototype of the fused-fiber photometry device, marking an important step toward broader dissemination and practical deployment of the technology.
In parallel, we made substantial progress toward establishing a dual-color fiber photometry system combined with optogenetic stimulation using a 633 nm laser. During this process, spectral crosstalk emerged as a major limitation, motivating a strategic shift to longer-wavelength stimulation at 650 nm. Although the full dual-recording setup could not be completed within the project timeline, in vivo validation of ChrimsonR activation with the 650 nm laser was successfully achieved, demonstrating the feasibility of this approach with appropriate power adjustments.
Finally, we developed a custom-built spectrometer that achieved higher light collection efficiency and greater flexibility than a commercial Ocean Optics system. This instrument enabled spectrally resolved fiber photometry with performance comparable to standard photodetectors for both slow and fast signals. At the same time, this work revealed important limitations, including constraints in spectral resolution, variability in biosensor emission profiles across animals, and challenges in spectral decomposition. These findings indicate that further improvements in optical design and the use of individualized reference spectra will be necessary to fully exploit the potential of hyperspectral fiber photometry.
As described in the previous section, we expanded the spectral range of a multicolor fiber photometry and optogenetic stimulation system to 650 nm for optogenetic stimulation. This extension preserves a wide spectral window at shorter wavelengths for photometry recordings, while enabling optogenetic stimulation in the long-wavelength range with substantially reduced optical crosstalk. This spectral separation provides an improved framework for combining multicolor recordings with causal manipulations in vivo.
In parallel, a patent application covering the fused-fiber photometry system was filed. While the original goal was to obtain patent approval within the duration of the grant, the examination process raised several outstanding objections. These issues have since been addressed in detail, and a formal rebuttal has been submitted to the patent office. The application is therefore progressing, with approval pending completion of the examination process.
In parallel, a patent application covering the fused-fiber photometry system was filed. While the original goal was to obtain patent approval within the duration of the grant, the examination process raised several outstanding objections. These issues have since been addressed in detail, and a formal rebuttal has been submitted to the patent office. The application is therefore progressing, with approval pending completion of the examination process.