Fiber Photometry: An Introductory Guide to This Revolutionary Technique, Part 3

Fiber Photometry: An Introductory Guide to This Revolutionary Technique, Part 3

In part two of this series, we introduced all of the technical components needed to successfully perform Fiber Photometry and collect reliable, accurate data. In part three, we will briefly discuss innovations in Fiber Photometry hardware and the wireless TeleFipho system.

Part 3: Embracing Innovation: Wireless Fiber Photometry

Advancing research techniques is a cornerstone of all scientific pursuits, leading to groundbreaking discoveries, as new tools allow researchers to clear hurdles that were previously thought as insurmountable. In the field of neuroscience, a more complete understanding of the relationship between neural activity and behavior remains a key goal. The challenging part of this endeavor is creating an experimental environment that closely replicates the natural environment of research subjects. Despite Fiber Photometry representing a significant leap forward in understanding how neural activity and behavior are linked, most conventional systems come with the limitation of being tethered, leading to a few drawbacks. When animals are connected to a tethered system, it limits their freedom of movement which can impede completion of a behavioral task and make interpretation of results difficult. Additionally, tethered Fiber Photometry systems are subject to noise due to the movement of the cable itself. Another issue facing tethered systems is the need for chaining multiple optical fibers together using connectors, with each joint, the light intensity weakens which therefore requires a complex and expensive light source. Therefore, these features can limit experimental design and reduce the accuracy of recorded data. Advancements in wireless technology have allowed for the design of wireless Fiber Photometry systems which eliminate some of the issues inherent to using tethered systems.

Now, we will briefly introduce the TeleFipho Wireless Fiber Photometry System—an innovation to Fiber Photometry that allows real-time identification of neuronal activity in free-moving animals.

Introducing the TeleFipho System

In TeleFipho, all the essential fiber photometry components – from the light source to photodetectors and optical filters – are integrated into a lightweight compact headstage. This wireless headstage communicates seamlessly with a base station via radio waves. The base station is easily connected to a PC for quick and accessible data acquisition. This wireless setup permits research animals to move unrestrictedly, navigate tunnels, and doorways, and even engage in social interactions.

 

 

Benefits of Wireless Fiber Photometry

By mounting the headstage directly onto fiber optic cannulas and eliminating the need for long optical cables that can introduce noise and attenuate signal strength, the TeleFipho system simplifies the light path and can achieve unprecedented precision in detecting changes in fluorescence. This means that the data collected is not only more accurate but also provides a clearer window into the intricacies of how neural activity encodes behavior.

Working with data from a system like TeleFipho Wireless Fiber Photometry is made easy by removing the cable, which eliminates motion artifacts caused by rotary joints and long flexible waveguides. This means that you won’t have to process your data to correct for these artifacts.
TeleFipho software provides a real-time view of the data, allowing you to optimize light levels and detection sensitivity quickly. Timestamps and notes can be added manually to the data, or you can connect your behavioral equipment’s outputs to TeleFipho and automatically align behavioral events with the fluorescence data.

In essence, the wireless capabilities of TeleFipho not only represent a leap forward in technological innovation but also mark a significant advance in the way behavioral neuroscience experiments are conducted. By enabling subjects to move freely, researchers can now explore neural activity in a context that more closely mirrors real-life scenarios. This advancement is poised to unlock new insights and pave the way for a deeper understanding of the intricate interplay between neural function and behavior.

If you missed part one or part two of this series make sure to check them out. Additionally, if you would like a more in-depth review of the TeleFipho Wireless Photometry System take a look at our comprehensive Wireless Photometry eBook.

Fiber Photometry: An Introductory Guide to This Revolutionary Technique, Part 2

Fiber Photometry: An Introductory Guide to This Revolutionary Technique, Part 2

In part one of this series, we introduced the fundamental concepts of Fiber Photometry and discussed how this revolutionary technique is advancing the field of neuroscience. In part two, we will transition into discussing the inner workings of Fiber Photometry and how the process works on a technical level.

Part 2: How Does Fiber Photometry Work?

Understanding the Components

To better understand how Fiber Photometry allows researchers to record neural activity in real time, it is first important to break the typical Fiber Photometry system down into its individual components. From the hardware of the optical components, the surgical implantation of cannulae, and the software based data analysis, these must all come together in order to produce quality data. There are 4 key components for a working Fiber Photometry system, each of which will be discussed briefly below:

1. Light Source

The choice of light source is pivotal in ensuring accurate and reliable fiber photometry measurements. It must emit a precise wavelength of light that matches the excitation spectrum of the fluorescent indicator being used. Additionally, considerations for light intensity and stability are crucial. A stable and well-calibrated light source guarantees consistent excitation, which directly impacts the quality and reliability of the recorded signals. Advanced light sources may also offer modulation capabilities, enabling researchers to perform more sophisticated experiments by synchronizing light pulses with specific events or behaviors.

2. Optical Fiber

The optical fiber acts as the conduit through which the excitation light travels from the source to the target area within the brain. It must be carefully selected based on factors such as numerical aperture, core diameter, and material composition. A higher numerical aperture allows for better light-gathering efficiency, while a larger core diameter permits more flexibility in positioning the fiber. Additionally, specialized coatings can enhance the fiber’s durability and protect against potential damage during implantation or use. Researchers must also consider factors like numerical aperture matching with the collection optics and the compatibility with chosen light sources.

3. Photodetector

The photodetector is responsible for capturing the emitted fluorescent signals from the neural tissue. It must be highly sensitive to the specific wavelengths emitted by the fluorescent indicator. Additionally, factors like quantum efficiency, noise levels, and dynamic range are critical considerations. A high quantum efficiency ensures that a larger proportion of emitted photons are converted into electrical signals, enhancing the detector’s sensitivity. Low noise levels are crucial for detecting weak signals amidst background noise, while a wide dynamic range allows for accurate recording of both strong and weak signals without saturation or loss of resolution.

4. Fluorescent Indicator

Genetically encoded fluorescent indicators are the cornerstone of fiber photometry. These indicators are typically proteins engineered to fluoresce in response to changes in calcium ion or neurotransmitter concentrations, providing a direct readout of neuronal activity. Choosing the right indicator is a crucial decision, as it determines the sensitivity and specificity of the measurements. Factors to consider include the indicator’s excitation and emission spectra, its target affinity and kinetics, as well as potential interactions with other cellular processes (Shen et.al. 2020). Additionally, researchers must ensure that the chosen indicator is compatible with the chosen excitation wavelength and photodetector.

Expression of Genetic Sensors

Advances in genetic engineering have been hand in hand with advances in imaging techniques. As such, researchers are now able to express complex fluorescent protein structures in a highly efficient cell type specific manner, allowing for unprecedented specificity when collecting neural activity data. Achieving success in fiber photometry relies heavily on obtaining optimal expression of genetically encoded fluorescent indicators (GEFIs) within your selected animal model. This critical step determines the quality and reliability of the fluorescent signals that will be measured, ultimately influencing the accuracy of the neural activity data obtained. Now we will briefly discuss the two primary methods for achieving optimal GEFI expression: viral expression and transgenic mouse models.

Day-Cooney et al., (2023) J.Neurochemistry.

  • Viral Expression: One approach to achieve targeted GEFI expression involves viral expression, a technique that involves injecting a specially engineered virus carrying the genetic code for the desired GEFI directly into the brain. This method offers the advantage of precision, allowing researchers to target specific cell types or regions within the brain for GEFI expression. However, the effectiveness of viral expression hinges on a critical consideration: virus selection. Different types of viruses are commonly used in neuroscience research, each with specific characteristics that make them valuable tools for manipulating and studying neural circuits. Adeno-Associated Viruses (AAVs) are widely employed due to their efficiency in transducing neurons without causing significant damage. While lentiviruses are useful for studies requiring prolonged expression of genetic constructs as they can integrate their genetic material into the host genome. Additionally, Retrograde Rabies Viruses have gained significant attention in recent years (Osakada et al. 2011). These modified viruses are particularly valuable for tracing neuronal pathways. By introducing a genetically engineered rabies virus into a specific target area, researchers can trace its path backward to identify the upstream neurons that project to that region. This technique provides crucial insights into the connectivity and communication between different brain regions, offering a powerful tool for mapping neural circuits. Finally, achieving the desired level of expression requires careful consideration of virus dilution. Dilution significantly influences the balance between optimal GEFI expression and potential drawbacks such as background noise or over-expression, which can result in inaccurate data interpretation. Researchers must carefully fine-tune the concentration of the injected virus to strike the right balance between signal intensity and stability in order to collect reliable neural activity data.
  • Transgenic Mouse Models: Another effective strategy involves utilizing transgenic mouse models. These models are genetically engineered to express GEFIs uniformly across the entire brain or in specific brain regions. This approach offers versatility and convenience, as it eliminates the need for viral injections and allows researchers to explore neural activity across multiple brain areas simultaneously. However, it’s important to note that transgenic models may have variations in GEFI expression levels between different brain regions. Researchers must account for these potential differences when analyzing the data and drawing conclusions about neural circuitry and behavior. Furthermore, the choice of transgenic mouse model is a critical aspect of the experimental design. Different lines may have distinct patterns of GEFI expression, allowing researchers to select models that align with the specific brain regions or cell populations under investigation. Additionally, advancements in genetic engineering techniques have enabled the development of more sophisticated transgenic models, including those that allow for cell-type-specific expression of GEFIs. These models provide an even higher level of specificity, allowing researchers to target specific neuronal populations with precision.

Implanting the Optical Cannula

In fiber photometry, successful expression of GEFIs represents a critical step, laying the foundation for the subsequent implantation of an optical cannula. This step is of paramount importance as it provides access to the fluorescent signal emitting from specific regions of interest within the brain. Optical cannulas are instrumental in this process, capturing the emitted light and conveying it to a photodetector for subsequent analysis. Their compact design serves to minimize tissue disruption and damage during the implantation, preserving the neural tissue’s integrity.

The significance of mitigating tissue damage and inflammation during the implantation process is crucial in neuroscience research, particularly in techniques like fiber photometry. Inflammation and tissue trauma can trigger a cascade of unintended physiological responses, including immune activation and the release of signaling molecules that have the potential to impact neighboring cells and brain regions. This phenomenon introduces unwanted signals and noise into the recorded data, complicating the task of accurately interpreting the neural activity of interest. Therefore, attention to detail and precision in the implantation of optical cannulas are imperative to uphold the integrity of the experimental system. Things to consider are locating precise stereotaxic coordinates, using fine tip micropipettes for injection, slow and controlled placement of the injection needle and injection rate, controlled viral delivery using a pneumatic pump, minimizing total volume injected, and implementing a post-injection wait period. This diligence ensures minimal tissue damage ensuring that subsequent fiber photometry measurements yield precise, reliable insights into neural dynamics. Join us for part 3, where we will discuss novel approaches and recent innovations in the realm of Fiber Photometry.

If you missed Part 1 of this introduction to Fiber Photometry take a look here. Learn more about Amuza’s cutting-edge Wireless Fiber Photometry system, TeleFipho.

Learn more about Amuza’s cutting-edge Wireless Fiber Photometry system, TeleFipho.

References

Shen et al., (2020) ‘Engineering genetically encoded fluorescent indicators for imaging of neuronal activity: progress and prospects’, Neurosci. Res., 152, pp. 3-14. doi: 10.1016/j.neures.2020.01.01

Osakada et al., (2011) ‘New rabies virus variants for monitoring and manipulating activity and gene expression in defined neural circuits’, Neuron, Aug 25;71(4):617-31. doi: 10.1016/j.neuron.2011.07.005.

Day-Cooney et al., (2023) Genetically encoded fluorescent sensors for imaging neuronal dynamics in vivo. J. Neurochemistry, 164, 284–308. Available from: https://doi.org/10.1111/jnc.15608

Fiber Photometry: An Introductory Guide to This Revolutionary Technique

Fiber Photometry: An Introductory Guide to This Revolutionary Technique

Welcome to a comprehensive introduction to Fiber Photometry, an innovative technique revolutionizing the landscape of the neuroscience field. Emerging in the mid-2010s, fiber photometry has rapidly ascended as a preeminent tool for real-time neural circuit measurements during behavioral experiments. Its major impact lies in its ability to offer unprecedented insights into the dynamic activity of neural populations in vivo. This article serves as a guide, providing a look into the fundamentals of Fiber Photometry, elucidating its intricate workings and capabilities. By the conclusion of this analysis, you will not only grasp the fundamentals of this methodology but also appreciate its distinctive advantages over traditional imaging techniques. The pivotal role that Fiber Photometry plays in shaping the future of neuroscience cannot be overstated, as it empowers researchers to unravel the complexities of brain function with an unprecedented level of precision and temporal resolution.

Part 1: Exploring Fiber Photometry

Understanding Neural Dynamics Through Fiber Photometry

In the realm of modern neuroscience, Fiber Photometry emerged as a significant innovation in the mid-2010s, swiftly garnering attention for its capacity to provide real-time measurements of neural circuits during behavioral experiments. Its roots trace back to pioneering work in optogenetics and fiber optic technology. The marriage of these technologies led to the birth of fiber photometry, offering researchers an unprecedented ability to monitor calcium signals within specific populations of neurons. This breakthrough allowed for a deeper understanding of neural circuits and their dynamic responses in live, behaving organisms. Over the years, fiber photometry has continued to evolve, becoming an indispensable tool in neuroscience labs worldwide. This chapter introduces the fundamentals of Fiber Photometry, differentiates it from traditional imaging techniques, and highlights its pivotal role in neuroscience.

Decoding Neural Activity with Calcium Imaging

Calcium imaging has emerged as a pivotal tool for decoding neural activity. Neurons rely on calcium ions as crucial signaling agents, participating in a multitude of cellular processes. Through cutting-edge techniques like Fiber Photometry, scientists have gained the ability to not only visualize but also precisely quantify the functionality of intracellular calcium dynamics. This groundbreaking method affords us the opportunity to closely monitor patterns of calcium ion fluctuations within specific neuronal populations. This level of granularity provides unprecedented insights into the dynamic responses of individual neurons and offers a window into the elaborate web of signals that define neural communication in the brain. By employing this powerful approach, researchers have uncovered essential information about neuronal behavior, from excitatory and inhibitory responses to intricate circuit dynamics (Grienberger & Konnerth, 2012).

In addition to calcium imaging, the field of neuroscience has witnessed the advent of fluorescent neurotransmitter sensors. These sensors offer a unique perspective by directly visualizing the release of specific neurotransmitters, such as dopamine, serotonin, and glutamate, providing invaluable insights into synaptic communication and neuromodulation. This complementary approach complements calcium imaging, offering a more comprehensive understanding of neural dynamics. As a result, the integration of fluorescent calcium and neurotransmitter sensors into the neuroscientist’s toolkit has further enriched our understanding of the complex workings of the brain.

A New Era of Imaging Technologies

Traditional Fiber Photometry methods often entail complex setups and confined animal conditions, hindering behavioral exploration and experimental conditions. While the introduction of optical fiberscopes and miniscopes has transformed this landscape and enabled calcium and neurotransmitter imaging in animals with increased freedom and precision, these systems are not without their drawbacks.

Measuring Neural Activity with Fiber Photometry

Unlike more traditional methods that focus on recording activity from individual neurons, such as whole cell electrophysiology. Fiber Photometry captures activity from a collective population of neurons expressing fluorescent indicators. While electric field potential recordings of neural activity is no novel concept, one key advantage of modern techniques like fiber photometry is the ability to target very specific cell populations using genetically expressed fluorescent sensors.

Telefipho Fiber Photometry gCaMP signal graph
This approach offers several advantages:

  1. Implanted Optical Cannula: Fiber Photometry uses an implanted optical cannula, capturing signals from multiple neurons without the need for detailed visualization of individual cells.
  2. Ease of Implementation: Fiber Photometry is accessible for many labs aiming to integrate calcium and neurotransmitter imaging into their research, offering streamlined data output, reduced complexity, and cost-effectiveness.
  3. Diverse Applications: Fiber Photometry is suited for extended behavioral experiments, exploratory studies, and multi-region imaging.
  4. Specificity: As mentioned above, specific subpopulations of neurons, glia, etc. can all be studied independently using genetically expressed fluorescent sensors.

As we delve deeper into the intricacies of Fiber Photometry, we transition into the next chapter, where we will explore the underlying mechanisms and workings of this revolutionary technique. Understanding how Fiber Photometry operates at a fundamental level is key to harnessing its full potential in unraveling the complexities of neural circuits and behavior. From the targeted expression of fluorescent indicators to the implementation of implanted optical cannulas, each facet contributes to the technique’s efficacy and versatility. Join us for the next chapter as we explore the inner workings of Fiber Photometry, shedding light on its transformative impact in the field of neuroscience.

If you’re eager to explore further, we invite you to learn about Amuza’s cutting-edge Wireless Fiber Photometry system, TeleFipho.

References

Grienberger, C. and Konnerth, A. (2012) ‘Imaging calcium in neurons’, Neuron, 73(5), pp. 862–885. doi:10.1016/j.neuron.2012.02.011.

Successful fiber photometry in mazes and other complex environments

Successful fiber photometry in mazes and other complex environments

Fiber photometry is most commonly used to monitor activity and chemical signaling at a specific location in the brains of freely moving animals. The technique uses fluorescent sensor proteins to report on the changing local concentration of calcium and neurotransmitters in real-time. To capture the fluorescent signal, an optical fiber is implanted in the brain with the tip just above the region of interest.

Fiber photometry: Wireless vs patch cord

Traditionally, a fiber optic patch cord (patch cable) is used to both deliver the excitation light to the animal and return the fluorescent signal to a fluorescence cube and a detector outside the cage. However, patch cords can create severe limitations for researchers. Sometimes the cord can directly affect the behavior of the animals: mice become confused, tilt their head, gnaw on the cord, or otherwise react to its presence. The patch cord can also run into objects in the environment, limiting the animal’s ability to move or carry out tasks. Patch cords can also introduce noise and artifacts into the signal when used with fiber photometry. Patch cords also complicate scoring of freezing behavior in fear conditioning experiments. Even after an animal freezes (stops moving), the patch cord can keep swaying, and some video tracking software will confuse this with the continued movement of the animal.

Many mazes, for example, elevated zero mazes and elevated plus mazes, have high walls which can trap the cable as a mouse or rat moves from an open area to an enclosed area.

Zero Maze

Elevated Plus Maze

Combining fiber photometry with behavioral measures for anxiety

Rodents typically prefer enclosed locations, and become anxious in unenclosed, wide-open spaces will limit their time spent exploring wide-open spaces. Anxiety levels play a strong role in how willing mice are to explore the unenclosed areas of a maze, so the zero maze is frequently used to study if a drug or other intervention is anxiolytic or anxiogenic. But a patch cord catching on the end of a wall could prevent a mouse from returning to the enclosed alley, so the researcher may have to be present to manipulate the cord.

Patch cords hanging from above a maze can get caught in doorways and at the ends of alleys.

In contrast, users have found mice fitted with wireless headstages such as those used with Amuza fiber photometry, optogenetics, and EEG have no trouble navigating mazes, and users report that the animals behave naturally.

For example, the Dimitrov lab at Rosalind Franklin University studies how stress and pain pathways interact in the brain. They used fiber photometry to monitor calcium fluorescence in the mPFC (medial prefrontal cortex) of mice traversing an elevated Zero maze while subject to inflammatory pain. They simultaneously monitored anxiety-like behavior by using video tracking to determine how much time the mice spent in exposed vs enclosed areas of the maze. They found that calcium fluorescence increased in male mice placed in the maze while subject to inflammatory pain, but there was no change in female mice under those conditions. Combined with other results, this finding suggests that sex-linked differences in the neural circuit between the locus coeruleus and mPFC are related to the differences in behavior and cognition displayed by male and female mice when subjected to inflammatory pain.

fiber photometry

FREE Fiber Photometry eBook

Amuza offers a FREE Wireless Fiber Photometry eBook. This ebook introduces topics and references critical for using fiber photometry during behavioral experiments

Cardenas A, Papadogiannis A, Dimitrov E. The role of medial prefrontal cortex projections to locus ceruleus in mediating the sex differences in behavior in mice with inflammatory pain. FASEB J. 2021 Jul;35(7):e21747.
https://pubmed.ncbi.nlm.nih.gov/34151467/

Animals: male and female mice
Sensor: GCaMP6f (calcium)
Vector: pAAV5.Syn.GCaMP6f, pAAV5.Syn.GCaMP6f.WPRE.SV40
Target Region: right mPFC (medial prefrontal cortex)
Coordinates: 1.8, ±0.4, and −2.2 mm in respect to bregma
Fiber: fiber core 400 μm NA 0.39, length 3 mm
Behavior test: Elevated O-maze
Fiber photometry data analysis: Amuza TeleFipho software
Model: Injection of complete Freund’s adjuvant (CFA) as a model for inflammatory pain.
Results: Inflammatory pain altered the calcium fluorescence signal from the mPFC of male mice placed on an elevated 0-maze, but did not alter the signal in female mice.

My lab has been using TeleFipho wireless photometric system for the past two years. The system is simple to use, durable and reliable. The practicality of TeleFipho allowed us to collect in vivo data about the neuronal activity of various limbic regions of the CNS during behavioral tests in mice.

Eugene Dimitrov MD, PhD

Assistant Professor, Department of Physiology and Biophysics Center for the Neurobiology of Stress Resilience and Psychiatric Disorders Rosalind Franklin University of Medicine and Science

Other mazes which require a mouse to pass through or a doorway or tunnel, such as the puzzle box maze, Light/Dark box, and many social interaction tests, can also present difficulties when using patch cords. Yunlei Yang’s lab used Amuza fiber photometry while mice explored a light/dark box to help characterize an anxiogenic circuit between septal OXTr neurons and the HDB, and identified a possible cause of OXT therapy side effects.

Huang, Tuanjie, Fangxia Guan, Julio Licinio, Ma-Li Wong, and Yunlei Yang. “Activation of septal OXTr neurons induces anxiety-but not depressive-like behaviors.” Molecular Psychiatry (2021): 1-10.
https://pubmed.ncbi.nlm.nih.gov/34489531/

Animals: Male and female C57BL/6 J and Oxtr-Cre mice.
Sensor: GCaMP6s (calcium)
Vectors: AAV1-hSyn1-GCaMP6s-P2A-nls-dTomato, AAV1-hSyn1-axon-GCaMP6s
Target Region: Fiber: vHPC (ventral hippocampus) Vector: lateral septum.

Fiber: fiber core 400 μm diameter, NA 0.39
Behavior test: light-dark box, Elevated Plus Maze
Fiber photometry data analysis: Amuza TeleFipho software
Results: Anxiogenic conditions activate the vHPC and vHPC projections to the lateral septum, as shown by increased calcium levels.

Problems with tethers also apply when using optogenetics and EEG, which is why we recommend using wireless optogenetics and EEG with freely moving animals.

2021 Optimizations to Wireless Fiber Photometry and Wireless Optogenetics Products, Based on Customer Feedback

2021 Optimizations to Wireless Fiber Photometry and Wireless Optogenetics Products, Based on Customer Feedback

Amuza has spent the last year listening to researchers who use optogenetics and imaging techniques with mice and rats and learning from their experiences. As customers use our products their suggestions for how to optimize our products have been invaluable, which sent our engineers back to work. The results are six new upgrades, options, and improvements for our wireless products, making them easier to use, allowing experiments to run longer, and providing a better customer experience.

Updates to Wireless Neuroscience Products

1. Removable, Rechargeable Batteries to Keep Experiments Running

One of the first questions researchers ask us about our wireless neuroscience products is how long the batteries will last, since this determines how long the experiment can run before the headstage needs to be swapped for recharging. To date, Amuza wireless neuroscience systems have been powered by integrated rechargeable batteries. We have offered versions with larger batteries, but this does increase the weight.

Our wireless systems are now offered with removable rechargeable batteries in several sizes. The batteries connect using a simple plug: To continue your experiment, just swap the battery for a recharged one. Choose a standard battery for mice (total weight for TeleFipho, 3.3 g), or a larger battery (total weight 5 g) with 60% longer battery life for rats.

TeleFipho headstage (left) and removable batteries (center, right).

2. Wired, but better.

Not entirely an update to our wireless products, but still highly relevant for long experiments and experiments where switching the battery would disturb the animal. Amuza now offers a wired version of our fiber photometry system. Unlike fiber photometry using optical patch cords, the headstage still contains the light source and fluorescence detection system. For the tether: instead of an optical signal, power and the amplified data signal are sent through a slim electrical cable. Because of this, compared to other wired systems, the wired version of TeleFipho does not degrade the signal by passing it through multiple optical connections or a rotary joint on the way to the detector. The cable is also very flexible, and bending it will not introduce artifacts into the signal. Just let us know what type of plug we should use on the cable, and we can make certain it will be compatible with the commutator you choose, preventing the cable from twisting or tangling.

Wired TeleFipho

3. Superior protection from interference for cleaner data on fiber photometry

To optimize the data transmission of our wireless fiber photometry system, we improved the shielding and the antenna on the TeleFipho headstage. This cuts down on both electrical noise in the data, and also makes the radio communication more robust in environments where radio interference has been an issue. In addition to the shielding and antenna improvements, a new directional antenna for TeleFipho provides a great solution for radio interference in more complex environments. If you experienced signal dropouts with TeleFipho’s standard antenna, this antenna should completely eliminate them.

4. New fiber optic cannula sizes for fiber photometry and optogenetics

TeleFipho has now been tested with 200 μm (core) diameter fibers as well as 400 μm. Narrower fibers offer the benefit of causing less physical trauma, as well as being able to target smaller structures. But they collect much less light from the target region, so bright, well expressed fluorescent biosensors are required to get high-quality data. Many of our customers in Japan have now used 200-micron fibers with TeleFipho to detect calcium using newer versions of GCaMP and reported excellent results. We now offer fiber optic cannula with either 200 or 400-micron fibers, cut to the length you specify. The cannula can also be used with our optogenetics system. Please see our blog post on fluorescent biosensors for updated information on sensors used for fiber photometry.

5. Scaling up fiber photometry for more animals

At launch, TeleFipho had four separate radio channels, allowing up to four animals to be monitored in the same room. We added 4 more radio channels to TeleFipho, so now up to 8 systems can be used together simultaneously.

6. Headstage protection chamber prevents equipment damage

Group housing gives animals the opportunity to nibble on or dislodge headstages, plus some cages have wire lids and other surfaces which can catch head-mounted equipment. Our new protection chamber isolates the headstage from the environment but is easy to remove between experiments. We recommend using the protection chamber with rats, non-human primates (NHPs), and other larger animals.

For more information on customizing wireless neuroscience products for your experiments to obtain better data from your wireless system, visit our wireless neuroscience products page.

Which upgrade is the most helpful for your research? What other upgrades would be valuable to you? Leave a comment below.

Microdialysis vs Fiber Photometry for Neurotransmitters

Microdialysis vs Fiber Photometry for Neurotransmitters

Microdialysis and Fiber Photometry are both powerful techniques for monitoring the concentrations
of neurotransmitters in vivo, but each technique has very different advantages. Amuza is the only company that provides both types of equipment to the neuroscience community, putting us in a unique position to help you determine when microdialysis or fiber photometry would be the best choice.

MicrodialysisFiber Photometry
Best for
  • Monitoring long-lasting changes
  • Absolute concentrations
  • Many analytes
  • Sub nanomolar analytes
  • Monitoring fast changes
  • Relative concentrations
  • One or two analytes
  • Targeting cell subtypes
SensitivityPicomolarNanomolar
Multiple analytesMany analytes can be measured in each sample with the little added difficultyData processing and experimental protocol are both more difficult
Types of analytesMost types of molecules.Limited by available fluorescent sensors
Type of measurementAbsolute concentration or % change relative to baseline% Change relative to baseline
Data analysisRelatively simpleOften quite complex
Sampling period~30 seconds to hoursMilliseconds
Sampling durationDaysDays, but difficult to monitor slow changes
TargetingBrain region/structureThe brain region, specific projection, cellular subtype, axon vs cell body
Animal movementTether requiredTether or wireless headstage

Multiple analytes

It is routine to measure many different neurotransmitters or amino acids in each microdialysis sample using HPLC chromatography and electrochemical detection. Samples can also be split and stored before analysis so that additional analytes can be added to the protocol at a later date. Commercially available assays also allow panels of multiple peptides and proteins to be measured in each sample.

Measuring multiple analytes simultaneously using fiber photometry is possible, but it requires expressing multiple fluorescent sensors at the target and the data analysis also becomes much more complex

Types of analytes

Microdialysis is extremely flexible and has been used to measure many types of analytes including amino acids, proteins, peptides, neurotransmitters, sugars, and more. If you can assay for it, you can probably sample for it using microdialysis.

Fiber photometry can now measure most of the principal neurotransmitters but is limited by the fluorescent sensors available. The number of sensors is increasing rapidly, but the options are still smaller than those available to microdialysis users.

Type of measurement

Microdialysis allows the monitoring of absolute concentrations or concentrations relative to basal levels in every sample.

Fiber photometry data yields a change in fluorescence data (∆F/F0). This correlates with changes in the concentration of the analyte relative to baseline but typically isn’t used to determine absolute concentrations.

Data Analysis

HPLC software packages already include all of the functions required to automate the processing of microdialysis data, allowing the monitoring of many analytes simultaneously over time. The analysis can be easily taught to new users. In contrast, fiber photometry data processing is still usually a complex operation, with most labs using multiple software packages and writing their own scripts to deconvolve and analyze multiple streams of data. (TeleFipho photometry data only uses fluorescence data from one wavelength, and is easier to process)

Sampling Period and Duration

While sample times can be as short as 30 seconds, microdialysis excels in measuring long-lasting changes in extracellular concentrations, and for determining absolute concentrations for baseline levels. It is quite routine to continuously sample for many hours or even days during a microdialysis experiment. Furthermore, the microdialysis probe can be removed and replaced with a dummy probe for days or weeks between sampling sessions.

Fiber photometry is at the opposite extreme: fluorescence is measured every 1 to 10 ms, allowing it to record events lasting less than a second. But drift, noise, and photobleaching during fiber photometry experiments complicate observation of slow changes over time.

Targeting

Both microdialysis and fiber photometry are used to target a discrete region or structure within the brain. Fiber photometry also allows retrograde or anterograde targeting, so that only specific projections are monitored. Viral vectors that only express the sensor in a specific cellular subtype or target either the axon or cell body can further narrow the scope of the events recorded.

Animal movement

Microdialysis and fiber photometry both typically require a tether during the experiment. TeleFipho is the exception and uses a wireless rechargeable headstage to gather and transmit fiber photometry data.

Do you have a question about microdialysis or fiber photometry?