Successful fiber photometry in mazes and other complex environments

Successful fiber photometry in mazes and other complex environments

by | Oct 15, 2021 | Fiber Photometry

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

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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

by | May 27, 2021 | Fiber Photometry, Optogenetics, Wireless Neuroscience

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

by | Sep 23, 2020 | Fiber Photometry, Microdialysis

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.

Microdialysis Fiber 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
Sensitivity Picomolar Nanomolar
Multiple analytes Many analytes can be measured in each sample with the little added difficulty Data processing and experimental protocol are both more difficult
Types of analytes Most types of molecules. Limited by available fluorescent sensors
Type of measurement Absolute concentration or % change relative to baseline % Change relative to baseline
Data analysis Relatively simple Often quite complex
Sampling period ~30 seconds to hours Milliseconds
Sampling duration Days Days, but difficult to monitor slow changes
Targeting Brain region/structure The brain region, specific projection, cellular subtype, axon vs cell body
Animal movement Tether required Tether 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?

Ask Amuza

GECIs and GEFIs: A Guide for Choosing Fluorescent biosensors for fiber photometry

GECIs and GEFIs: A Guide for Choosing Fluorescent biosensors for fiber photometry

by | Jul 29, 2020 | Fiber Photometry

Fiber Photometry is a rapidly advancing field, with biosensors for more analytes and with better sensitivity being announced almost every month. We would like to share information about sensors that should be compatible with fiber photometry when using excitation with blue (~480 nm) light and measuring green (~525 nm) fluorescence. This is the most commonly used wavelength pair and is offered with TeleFipho wireless fiber photometry.

We will update this guide as more information becomes available.

Overview of Fluorescent indicators: Structure and considerations for use.

Genetically encoded fluorescent indicators (GEFIs) are used in conjunction with fiber photometry to report on changes in concentrations of molecules in vivo in real-time.

Most fluorescent biosensors comprise a fluorescent protein yoked to an analyte binding protein, constructed so that binding of the analyte causes a dramatic increase in fluorescence.

Akerboom, Rivera, Guilbe, Malavé, Hernandez, Tian, Hires, Marvin, Looger, Schreiter ER / CC BY (https://creativecommons.org/licenses/by/3.0)

When used with fiber photometry in behaving animals, the sensors are usually introduced by injecting viral expression vectors. The virus is used to both express the sensor and to control its location: targeting sequences allow the researcher to choose a specific cellular subtype, and even the cellular localization, such as axon or soma, where the sensor will be expressed. Anterograde and retrograde localization can also target only a specific projection or circuit in a target region. Transgenic animals are also available for expressing some GEFIs.

Binding kinetics helps determine the range of concentrations the sensor will respond to, and its ability to report fast events. A sensor with high affinity (low Kd) and a long dissociation time can measure very low concentrations of a molecule, but this happens at the expense of being able to resolve more frequent events and a narrower useful range of concentrations. Fast dissociation improves time resolution, but sensitivity usually suffers.

The brightness of the sensor, partially expressed as the ratio of the increase in fluorescence when bound to the analyte compared to baseline (∆F/F or ∆F/F0), is the other major factor to consider. Brighter sensors can generate a useful signal when expressed at lower levels or when used with less illumination when compared to less bright sensors. They can also be used with narrower fibers. Some of our users are stepping down to 250 microns from 400-micron core diameter when using the latest generations of GCaMP type sensors for fiber photometry.

Most biosensors are already available from AddGene: some as plasmids, others as aliquots of ready-to-use viral vectors. The newest biosensors listed here can be sourced directly from the laboratories which invented them. We included the best source we could find, and the original publication describing the sensor in the table below.

Calcium

Calcium Sensors Affinity (Kd or EC50) dissociation
Kinetics (Mean life,
1/Koff)		 ∆F/F0 (% increase) Source for vector or plasmid Reference
GCaMP6s 147 nM 1796 ms 1680
Addgene
Chen, 2013
GCaMP6f 375 nM 400 ms 1314
Addgene
Chen, 2013
jGCaMP7s 68 nM 1260 ms
Janelia
Dana, 2019
jGCaMP7f 150 nM 270 ms 3100
Janelia
Dana, 2019
jGCaMP8f 334 nM 27 ms 7880
Addgene
Janelia
jGCaMP8m 108 nM 55 ms 4570
Addgene
Janelia
jGCaMP8s 46 nM 272 ms 4950
Addgene
Janelia
GCaMP-X
Addgene
Yang, 2018

The GCaMP6 series of genetically encoded Ca2+ indicators (GECIs) are the most popular tools for examining action potentials and have been used extensively with TeleFipho photometry. GCaMP6f is optimized for fast decay kinetics, necessary for monitoring quick events, while GCaMP6s have higher sensitivity and slower decay kinetics. If you are starting a new project, consider the latest generation – jGCaMP7 – which offers higher sensitivity and a larger range of kinetics.

The jGCaMP7 series was introduced in 2019 as a collaborative effort between Loren Looger at Janelia and other research institutes. The jGCaMP7 GECIs have several-fold higher ∆F/F0 and a wider range of kinetics when compared to the earlier GCaMP6 sensors. Some GCaMP7 variants that will interest fiber photometry users include jGCaMP7s (highest sensitivity, but slower kinetics), and jGCaMP7f which has the fastest kinetics. We hope to have calcium data generated using jGCaMP7s and TeleFipho wireless fiber photometry soon.

The  jGCAMP8 series from The Looger Lab and the GENIE Project Team at HHMI Janelia was introduced in late 2020 and is the most recent set of GECIs available with improved sensitivity and speed. Compared to jGCAMP7f, the new series all have a faster rise time. 8f (fast) has a 4x faster rise time and a 2.5x faster decay time. 8m (medium) again has about a 4x faster rise time but is also 3.5x more sensitive. 8s (sensitive) is 2x more sensitive and 2x faster. A bonus of using these newer, brighter sensors for fiber photometry is that narrower fibers can be used. Some of our users are stepping down from 400 microns to 250-micron core diameter fibers, allowing for less traumatic surgeries while targeting smaller areas.

GCaMP-X The calmodulin GCaMP based calcium sensors have been shown to cause side effects during some in-vivo uses, such as interference with the function of L-type calcium channels, nuclear accumulation, and cytotoxicity. Changes largely addressed these issues to the design of GCaMP-X.

Dopamine

Dopamine is rapidly becoming the second most common target for imaging and photometry in neuroscience thanks to two sensors introduced in 2018, dLight and GRABDA. The intensity of illumination used with dLight and GRABDA is typically 20 – 30 μW, the same range as is used with GCaMP6.

Dopamine sensors Affinity
(Kd or EC50)		 dissociation
Kinetics
(residence time,
τ = 1/Koff)		 ∆F/F0
(% increase)		 Source for vector or plasmid Reference
dLight1.1 330 nM NA 230
Addgene
Patriarchi, 2018
dLight1.2 770 nM 90 ms 340
Addgene
Patriarchi, 2018
dLight1.3b 1680 nM 930
Addgene
Patriarchi, 2018
GRABDA1m 130 nM 700 ms 90
Addgene
Sun, 2018
GRABDA2m 90 nM NA 340
Yulong Li Lab
Sun, 2020
GRABDA1h 10 nm 2500 ms 90
Addgene
Sun, 2018
GRABDA2h 7 nM NA 280
Yulong Li Lab
Sun, 2020

dLight1.1 and dLight1.2, developed by the Tian lab, have both been used extensively with fiber photometry, with settings similar to those used for GCaMP6.

GRABDA DA2M, DA2H

GRABDA (GPCR-Activation Based DA) was first introduced by Yulong Li’s lab in 2018 and has just been updated to increase Δf/f and increase the range of kinetics. The recent versions are DA2H (high affinity) and DA2M (medium affinity). Both GRABDA2m and GRABDA2H have already been used with fiber photometry, but so far results have only been communicated via preprints.

Norepinephrine and Serotonin

More from the Yulong Li lab, though as of yet their characterization is only available through preprints. GRABNE1m and GRAB5-HT1.0 have both already been used to measure norepinephrine and serotonin via fiber photometry in mice.

Sensors Analyte Affinity
(Kd or EC50)		 dissociation Kinetics
( τ = 1/Koff))		 ∆F/F0
(% increase)

Source for
vector or plasmid

 Reference
GRABNE1h Norepinephrine 83 nM 2000 ms 130
Yulong Li Lab
Feng, 2019
GRABNE1m Norepinephrine 930 nM 750 ms 250
Yulong Li Lab
Feng, 2019
GRAB-5HT1.0 Serotonin 22 nM 3.1 s 280
Yulong Li Lab
Wan, 2020
iSeroSnFr Serotonin EC50 1.5 µm
Tian Lab
Unger, 2020

Biosensors for endo cannabinoids (GRABeCB), ATP, cholecystokinin (CCK), vasoactive intestinal peptide (VIP), somatostatin (SST), vasopressin/oxytocin, ghrelin, and orexin were also announced by the Li lab at Neuroscience 2019, and are still being validated. The best way to keep up with the Li lab may check #GRABSensors on Twitter!
More information on iSeroSnFr from the Tian lab should be available soon.

GABA

GABA Sensors Affinity (Kd or EC50 dissociation
Kinetics
(τ = 1/Koff)		 ∆F/F0 (% increase) Source Reference
iGABASnFR 9 μM 250
Addgene
Marvin, 2019

Glutamate

There are two main sensor types available for monitoring glutamate: iGluSnFR and iGlu. The original iGluSnFR has slower kinetics, while iGluf (fast) and iGluu (ultrafast) are much faster. The new SF-iGluSnFR variants offer higher brightness and a range of different kinetics compared to the original.

Glutamate Sensors Affinity (Kd or EC50 dissociation
Kinetics
(τ = 1/Koff)		 >∆F/F0 (% increase) Source Reference
iGluSnFR 4.9 μM 92 ms 100
Addgene
Marvin, 2018
iGluf 137 μM 2.1 ms
Addgene
Helassa, 2018
iGluu 600 μM 0.7
Addgene
Helassa, 2018

Acetylcholine

Acetylcholine Sensors Affinity (Kd or EC50 dissociation
Kinetics
(τ = 1/Koff)		 ∆F/F0 (% increase) Source Reference
iACHSnFR 1.3 µM 1200
Addgene
Borden, 2020
ACh3.0 2 μM 3.7s
Yulong Li Lab
Jing, 2019
ACh4.3
Yulong Li Lab

iACHSnFR is one of the most recent GEFIs created by the Loren Looger lab at Janelia, along with collaborators.

GACH and GRABACh3.0
While the initial version of the GRAB type acetylcholine indicator (GACH) was not sensitive enough for measuring physiological levels of ACh using fiber photometry (personal communication), the version described in a preprint from Dec. 2019 (ACh3.0) has been used successfully with fiber photometry. The Li lab is also supplying researchers with an even newer version, ACh4.3.

Other Analytes

Adenosine and ATP (Extracellular)

Sensors Analyte Affinity (Kd or EC50 dissociation
Kinetics
(τ = 1/Koff)		 ∆F/F0 (% increase) Source for vector or plasmid Reference
GRABATP1.0 ATP EC50 ~45 nM 9 ms 500-1000
Yulong Li Lab
Wu, 2021
iATPSnFR1 ATP EC50 of ~50 nM 190
Addgene
Lobas, 2019
GRABAdo Adenosine 60 nM 63 ms 120
Yulong Li Lab
Peng, 2020

Adenosine
Peng et al. monitored adenosine in the mouse basal forebrain using fiber photometry and GRABAdo, also called Ado1.0

Please let us know if you have any corrections or additions to this list!

Telefipho Wireless Fiber Photometry

Telefipho Wireless Fiber Photometry

by | Mar 4, 2020 | Fiber Photometry, Neuroscience

Telefipho Wireless Fiber Photometry

We would like to tell you about Amuza’s new wireless fiber photometry (TeleFipho). It’s our newest way to track calcium and neurotransmitters, and it works in real-time in freely moving animals. It’s easy to use, easy to process your data, and it’s ready to go right out of the box.

Fiber photometry is one of the newest tools available to neuroscientists who wish to correlate behavior with neural activity. It’s a powerful, ultra-fast technique used to measure calcium, neurotransmitters and other molecules in vivo in real-time. But until now the technique has required a connection – a fiber optic cable connecting the research animal to the rest of the optical hardware outside of the cage. This cable limits animals’ freedom of movement and social interactions. It is also fragile and can make your data noisier. This, in turn, can limit the design of your experiments.

Fiber Photometry Schematic

Telefipho Schematic

With Telefipho all of the optical hardware – light source, optical filters, and photodetector – are combined in a small headstage. The rechargeable headstage communicates by radio with a base station. Your animals can move through tunnels and doorways and interact freely during experiments. Telefipho can also improve video tracking: tracking software often confuses a swaying cable with a mouse that is still moving. This can complicate the scoring of freezing behavior during fear conditioning.

The headstage mounts directly on an FC size fiber optic cannula without any intervening cables or interconnects. This maximizes light transmission and minimizes noise and artifacts. This headstage weighs just 3 grams and has been tested with both mice and rats.

To use the system, an optical fiber is implanted with the tip placed at the brain region of interest. Blue light from an LED in the headstage is sent through the fiber to excite the fluorescent sensors expressed in the target region. The sensor molecules fluoresce in proportion to the concentration of the analytes, and some of the fluorescent light travels back through the fiber to be measured by a photodetector. A fluorescence filter cube, combining bandpass filters for excitation, a bandpass filter for emission, and dichroic mirrors are used to remove extraneous wavelengths and separate the light paths. The headstage transmits the data to the base station and then to your computer.

Since only a single narrow fiber is implanted inside the skull, the technique is much less invasive than imaging, especially for targets deep inside the brain.
The fluorescent signal is recorded and allows tracking of changes in analyte levels on a subsecond time scale.

TeleFipho data is also easy to work with. Removing the cable means removing the motion artifacts caused by rotary joints and long flexible waveguides, so you won’t have to process your data to correct for them.

TeleFipho software provides a real-time view of the data, allowing you to quickly optimize light levels and detection sensitivity. You can manually add timestamps and notes to the data, or you can connect your behavioral equipments’ outputs to TeleFipho and automatically align behavioral events with the fluorescence data.

You can also send the signal to your own recording equipment and process the data using your own software.

With the ever-increasing number of genetically encoded fluorescent indicators for molecules beyond calcium, such as dopamine, glutamate, acetylcholine, norepinephrine, endocannabinoids, and even cyclic monophosphates, fiber photometry is certain to become a versatile tool in your lab. This is doubly true since the vectors used to express the sensors are becoming increasingly precise at targeting specific cell types and circuits so that results are increasingly specific with less interference from off-target cells and molecules.

If you would like to learn more about TeleFipho fiber photometry, please contact Amuza.

Questions?

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Wireless Fiber Photometry: Measuring Neurochemicals In Vivo in Real Time

Wireless Fiber Photometry: Measuring Neurochemicals In Vivo in Real Time

by | Oct 14, 2019 | Fiber Photometry, Neuroscience, Shows and Events

Amuza and Teleopto launch the first commercial wireless fiber photometry system at Neuroscience 2019

Our wireless optogenetic users have frequently asked us if we could provide wireless photometry – we are happy to announce that now we can!

TeleFipho wireless headstages allow your freely behaving animals to move with true freedom, enabling novel experimental approaches with fiber photometry. The 3-gram headstages are optimized for GCaMP and other GFP-based indicators.

TeleFipho includes all of the components required for fiber photometry – light source, filter cube, photodetector, and wireless transmission hardware – in a 3-gram headstage.

What is Fiber Photometry?

Fiber Photometry is a powerful technique for measuring rapid changes in neuromodulators in vivo via fluorescence. It is most commonly used to measure fast (sub-second) changes in concentrations of calcium in freely behaving animals, but it is now also capable of being used to monitor neurotransmitters and other molecules. To use fiber photometry, genetically encoded fluorescent indicators are first expressed at the location of interest. When excited by light of the right wavelength, these proteins fluoresce – but only while they are bound to their target analyte. As local concentrations of the analyte rise and fall, the fluorescence intensity rises and falls in response. Genetically encoded calcium indicators (GECIs), such as GcAMP, have been the mainstay of fiber photometry and also for calcium imaging, a closely related technique. Recently, dopamine indicators (Dlight1, GRABDA) and norepinephrine indicators (GRABNE) have been introduced, and more neurochemical sensors are in development.

To capture this signal in vivo, an optical fiber is implanted in the target region in the animal. The other end of the fiber is attached to the photometry hardware. First, an LED or laser light source passes light through the fiber to excite the indicator proteins in the target region. The resulting fluorescent light then travels back through the fiber to a photodetector, creating a record of the changing concentrations of the analyte. Careful filtering and splitting of the light traveling through the fiber optic are required to separate the light used for excitation from the fluorescence being sent to the photodetector.

Why use Fiber Photometry?

The most frequent use is to measure changes in calcium levels at synapses as a proxy for changes in neural activity, helping researchers discern the links between behavior states and the firing patterns of neurons. But the same technique is also used to monitor the activity of GPCRs and ion channel drug targets.

When used with freely moving animals, fiber optic tethers can be problematic. The cable can prevent animals from using exercise wheels or shelters or getting tangled in complicated environments, limiting behavioral testing. Cables can also cause artifacts when used with video tracking software. For example, the cable often continues to sway after the animal has stopped moving, making it difficult to recognize freezing behavior during fear conditioning studies. Placing all of the necessary components for fiber photometry in a small lightweight headstage ends these problems.

TeleFipho has been tested with both mice and rats. The data above shows stress-induced (tail pinch) changes in GCaMP signals from hypothalamic orexin neurons in mice. GCaMP is a genetically encoded calcium indicator often used to monitor calcium dynamics. Data is Courtesy of Dr. Daisuke Ono in the Akihiro Yamanaka Lab, Nagoya University.

TeleFipho has been tested with both mice and rats. The data above shows stress-induced (tail pinch) changes in GCaMP signals from hypothalamic orexin neurons in mice. GCaMP is a genetically encoded calcium indicator often used to monitor calcium dynamics. Data is Courtesy of Dr. Daisuke Ono in the Akihiro Yamanaka Lab, Nagoya University.[/caption]

Shrinking the components for fiber photometry has a bonus: it also allows us to shrink the price. Telefipho starts at roughly half of the cost of other commercial fiber photometry systems.

Please stop by our booth during SfN 2019 to ask for a demonstration and visit our product page for more information.

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