Name: multi-fiber photometry to record neural activity in freely-moving animals
Uploaded: 2019-10-20T14:30:00
Description: Watch this Scientific Journal Video about Multi-Fiber Photometry to Record Neural Activity in Freely-Moving Animals at JoVE.com

This work was supported by a grant from the Natural Sciences and Engineering Research Council of Canada (NSERC: RGPIN-2017-06131) to C.P. C. P. is a FRSQ Chercheur-Boursier. We also thank the Plateforme d&rsquo;Outils Mol&eacute;culaires (https://www.neurophotonics.ca/fr/pom) for the production of the viral vectors used in this study.

All experiments were done in accordance with the Institutional Animal Care and Use Committees of the University of California, San Diego, and the Canadian Guide for the Care and Use of Laboratory Animals and were approved by the Universit&#233; Laval Animal Protection Committee.&#160;
1. Alignment of the optical path between the CMOS (complementary metal oxide semiconductor) camera and the individual or branching patch cord
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	<li>Loosen all screws on the 5-axis translator (11, <strong cl...

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Fiber photometry is an accessible approach that allows researchers to record bulk-calcium dynamics from defined neuronal populations in freely-moving animals. This method can be combined with a wide range of behavioral tests, including &#8220;movement heavy&#8221; tasks such as forced swim tests2, fear-conditioning18, social interactions1,4, and others7,8<su...

The ability to correlate neural responses with specific aspects of an animal&rsquo;s behavior is critical to understand the role a particular group of neurons plays in directing or responding to an action or stimulus. Given the complexity of animal behavior, with the myriad of internal states and external stimuli that can affect even the simplest of actions, recording a signal with single-trial resolution equips researchers with the necessary tools to overcome these limitations.
Fiber photometry has become the technique of choice for many researchers in the field of systems neuroscience because of its relative simplicity compared to other in vivo recording techniques, its high signal-to-noise ratio, and the ability to record in a variety of behavioral paradigms1,2,3,4,5,6,7,8. Unlike traditional electrophysiological methods, photometry is the optical approach most commonly used in conjunction with genetically-encoded calcium indicators (GECIs, the GCaMP series)9. GECIs change their ability to fluoresce based on whether or not they are bound to calcium. Because the internal concentration of calcium in neurons is very tightly regulated and voltage-gated calcium channels open when a neuron fires an action potential, transient increases in internal calcium concentration, which result in transient increases in the ability of a GECI to fluorescence, can be a good proxy for neuronal firing9.
With fiber photometry, excitation light is directed down a thin, multimode optic fiber into the brain, and an emission signal is collected back up through the same fiber. Because these optic fibers are lightweight and bendable, an animal can move largely unhindered, making this technique compatible with a wide array of behavioral tests and conditions. Some conditions, such as rapid movements or bending of the fiber-optic patch cord beyond the radius at which it can maintain total internal reflection, can introduce signal artifacts. To disambiguate signal from noise, we can exploit a property of GCaMP known as the &ldquo;isosbestic point.&rdquo; Briefly, with GCaMP, as the wavelength of the excitation light is shifted to the left, its emission in the calcium-bound state decreases and the emission in the calcium-unbound state marginally increases. The point at which the relative intensity of these two emissions are equal is termed the isosbestic point. When GCaMP is excited at this point, its emission is unaffected by changes in internal calcium concentrations, and variance in the signal is most often due to attenuation of the signal from overbending of the fiber-optic patch cord or movement of the neural tissue relative to the implanted fiber.
Single unit electrophysiology is still the gold standard for freely-moving in vivo recordings due to its single-cell and single-spike level resolution. However, it can be difficult to pinpoint the molecular identity of the cells being recorded, and the post-hoc analysis can be quite laborious. While fiber photometry does not have single-cell resolution, it does allow researchers to ask questions impossible to address with traditional techniques. Combining viral strategies with transgenic animals, the expression of GECIs can be directed to genetically-defined neuronal types to record population- or projection-defined neural activity, which can be performed by monitoring calcium signal directly at axon terminals10,11. Moreover, by implanting multiple fiber-optic cannulas, it is possible to simultaneously monitor neural activity from several brain regions and pathways in the same animal12,13.
In this manuscript, we describe a technique for single and multi-fiber photometry, how to correct for calcium-independent artifacts, and detail how to perform mono- and dual-color recordings. We also provide examples of the types of questions it enables one to ask and their increasing levels of complexity (see Figure 1). The fiber photometry setup for multi-fiber recordings detailed in this protocol can be built using a list of materials found at https://sites.google.com/view/multifp/hardware (Figure 2).
It is essential that the system be equipped for both 410 nm and 470 nm excitation wavelengths for calcium-independent and calcium-dependent fluorescence emission from GCaMP6 or its variants. For custom-built setups or if there is no available software to run the system, the free, open source program Bonsai (http://www.open-ephys.org/bonsai/) can be used. Alternatively, fiber photometry can be run through MATLAB (e.g., https://github.com/deisseroth-lab/multifiber)12 or other programming language14. The software and hardware of the system should allow manipulation of both the 410 nm and 470 nm LEDs and the camera, extraction of images (Figure 2), and calculation of the mean fluorescent intensity in the regions of interest (ROIs) drawn around the fibers on the images. The output should be a table of mean intensity values recorded with the 470 nm and 410 nm LEDs from each fiber in the patch cord. When performing multi-fiber experiments, 400 &micro;m bundled fibers may limit the movement of mice. In such cases, we recommend using 200 &micro;m patch cords, which provide more flexibility. It may also be possible to use smaller dummy cables during training of mice.
It is crucial to be able to extract time points for events of interest during fiber photometry acquisition. If the system does not readily provide a built-in system to integrate TTLs for specific events, an alternative strategy is to assign a time stamp to individual time points recorded to align with specific times and events during the experiment. Time stamping can be done using the computer clock.

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			<td>1/4&#34;-20 Stainless Steel Cap Screw, 1&#34; Long</td>
			<td>Thorlabs</td>
			<td>SH25S100</td>
			<td></td>
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			<td>1/4&#34;-20 Stainless Steel Cap Screw, 1/2&#34; Long</td>
			<td>Thorlabs</td>
			<td>SH25S050</td>
			<td></td>
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		<tr>
			<td>1/4&#34;-20 Stainless Steel Cap Screw, 3/8&#34; Long</td>
			<td>Thorlabs</td>
			<td>SH25S038</td>
			<td></td>
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		<tr>
			<td>1000 &#181;m, 0.50 NA, SMA-SMA Fiber Patch Cable</td>
			<td>Thorlabs</td>
			<td>M59L01</td>
			<td></td>
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		<tr>
			<td>12.7 mm Optical Post</td>
			<td>Thorlabs</td>
			<td>TR30/M</td>
			<td></td>
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			<td>12.7 mm Pedestal Post Holder</td>
			<td>Thorlabs</td>
			<td>PH20EM</td>
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			<td>15 V, 2.4 A Power Supply Unit with 3.5 mm Jack Connector for T-Cube</td>
			<td>Thorlabs</td>
			<td>KPS101</td>
			<td></td>
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		<tr>
			<td>20x objective</td>
			<td>Thorlabs</td>
			<td>RMS20X</td>
			<td>#10 in Figure 2, #11 in Figure 5</td>
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		<tr>
			<td>30 mm Cage Cube with Dichroic Filter Mount</td>
			<td>Thorlabs</td>
			<td>CM1-DCH/M</td>
			<td>#8-9 in Figure 2, #8-10 in Figure 5</td>
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			<td>405 nm LED</td>
			<td>Doric Lenses</td>
			<td>CLED_405</td>
			<td>#2 in Figure 2</td>
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		<tr>
			<td>410 nm bandpass filter</td>
			<td>Thorlabs</td>
			<td>FB410-10</td>
			<td>#5 in Figure 2; #7 in Figure 5</td>
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		<tr>
			<td>465 nm. LED</td>
			<td>Doric Lenses</td>
			<td>CLED_465</td>
			<td>#1 in Figure 2</td>
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		<tr>
			<td>470 nm bandpass filter</td>
			<td>Thorlabs</td>
			<td>FB470-10</td>
			<td>#4 in Figure 2; #6 in Figure 5</td>
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		<tr>
			<td>560 nm bandpass filter</td>
			<td>Semrock</td>
			<td>FF01-560/14-25</td>
			<td>#5 in Figure 5</td>
		</tr>
		<tr>
			<td>560 nm LED</td>
			<td>Doric Lenses</td>
			<td>CLED_560</td>
			<td>#1 in Figure 3</td>
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			<td>5-axis kinematic Mount</td>
			<td>Thorlabs</td>
			<td>K5X1</td>
			<td>#11 in Figure 2, #12 in Figure 5</td>
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			<td>Achromatic Doublet</td>
			<td>Thorlabs</td>
			<td>AC254-035-A-ML</td>
			<td>#7 in Figure 2</td>
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			<td>Adaptor for 405 collimator</td>
			<td>Thorlabs</td>
			<td>AD11F</td>
			<td>#3 in Figure 2; #4 in Figure 5</td>
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			<td>Adaptor for ajustable collimator</td>
			<td>Thorlabs</td>
			<td>AD127-F</td>
			<td>#3 in Figure 2; #4 in Figure 5</td>
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		<tr>
			<td>Aluminum Breadboard</td>
			<td>Thorlabs</td>
			<td>MB1824</td>
			<td></td>
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			<td>Clamping Fork</td>
			<td>Thorlabs</td>
			<td>CF125</td>
			<td></td>
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			<td>Cube connector</td>
			<td>Thorlabs</td>
			<td>CM1-CC</td>
			<td></td>
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		<tr>
			<td>Dual 493/574 dichroic</td>
			<td>Semrock</td>
			<td>FF493/574-Di01-25x36</td>
			<td>#10 in Figure 5</td>
		</tr>
		<tr>
			<td>Emission filter for GCaMP6</td>
			<td>Semrock</td>
			<td>FF01-535/22-25</td>
			<td>#6 in Figure 2</td>
		</tr>
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			<td>Enclosure with Black Hardboard Panels</td>
			<td>Thorlabs</td>
			<td>XE25C9</td>
			<td></td>
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			<td>Externally SM1-Threaded End Cap for Machining</td>
			<td>Thorlabs</td>
			<td>SM1CP2M</td>
			<td></td>
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		<tr>
			<td>Fast-change SM1 Lens Tube Filter Holder</td>
			<td>Thorlabs</td>
			<td>SM1QP</td>
			<td>#4-6 in Figure 2, #5-7 in Figure 5</td>
		</tr>
		<tr>
			<td>Fixed Collimator for 405 nm light</td>
			<td>Thorlabs</td>
			<td>F671SMA-405</td>
			<td>#3 in Figure 2; #4 in Figure 5</td>
		</tr>
		<tr>
			<td>Fixed collimator for 470 and 560 nm light</td>
			<td>Thorlabs</td>
			<td>F240SMA-532</td>
			<td>#3 in Figure 2; #4 in Figure 5</td>
		</tr>
		<tr>
			<td>Green emission filter</td>
			<td>Semrock</td>
			<td>FF01-520/35-25</td>
			<td>In light beam splitter</td>
		</tr>
		<tr>
			<td>High-Resolution USB 3.0 CMOS Camera</td>
			<td>Thorlabs</td>
			<td>DCC3260M</td>
			<td>#13 in Figure 2, #15 in Figure 5</td>
		</tr>
		<tr>
			<td>Light beam splitter</td>
			<td>Neurophotometrics</td>
			<td>SPLIT</td>
			<td>#14 in Figure 5</td>
		</tr>
		<tr>
			<td>Longpass Dichroic Mirror, 425 nm Cutoff</td>
			<td>Thorlabs</td>
			<td>DMLP425R</td>
			<td>#8 in Figure 2, #9 in Figure 5</td>
		</tr>
		<tr>
			<td>Longpass Dichroic Mirror, 495 nm Cutoff</td>
			<td>Semrock</td>
			<td>FF495-Di03</td>
			<td>#9 in Figure 2, #8 in Figure 5</td>
		</tr>
		<tr>
			<td>Metabond dental cement</td>
			<td>C&#38;B</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>M8 - M8 cable</td>
			<td>Doric Lenses</td>
			<td>Cable_M8-M8</td>
			<td></td>
		</tr>
		<tr>
			<td>Optic fiber cannulas</td>
			<td>Doric Lenses</td>
			<td></td>
			<td>Need to specify that these will be used to photometry experiments requiring low autofluorescence</td>
		</tr>
		<tr>
			<td>Optic fiber Patchcords</td>
			<td>Doric Lenses</td>
			<td></td>
			<td>Need to specify that these will be used to photometry experiments requiring low autofluorescence</td>
		</tr>
		<tr>
			<td>Red emission filter</td>
			<td>Semrock</td>
			<td>FF01-600/37-25</td>
			<td>In light beam splitter</td>
		</tr>
		<tr>
			<td>T7 LabJack</td>
			<td>LabJack</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>T-cube LED Driver</td>
			<td>Thorlabs</td>
			<td>LEDD1B</td>
			<td></td>
		</tr>
		<tr>
			<td>USB 3.0 I/O Cable, Hirose 25, for DCC3240</td>
			<td>Thorlabs</td>
			<td>CAB-DCU-T3</td>
			<td></td>
		</tr>
	</tbody>
</table>

multi-fiber photometry to record neural activity in freely-moving animals

Recording the activity of a group of neurons in a freely-moving animal is a challenging undertaking. Moreover, as the brain is dissected into smaller and smaller functional subgroups, it becomes paramount to record from projections and/or genetically-defined subpopulations of neurons. Fiber photometry is an accessible and powerful approach that can overcome these challenges. By combining optical and genetic methodologies, neural activity can be measured in deep brain structures by expressing genetically-encoded calcium indicators, which translate neural activity into an optical signal that can be easily measured. The current protocol details the components of a multi-fiber photometry system, how to access deep brain structures to deliver and collect light, a method to account for motion artifacts, and how to process and analyze fluorescent signals. The protocol details experimental considerations when performing single and dual color imaging, from either single or multiple implanted optic fibers.

Neural correlates of behavioral responses can vary depending on a variety of factors. In this example, we used in vivo fiber photometry to measure the activity of axon terminals from the lateral hypothalamic area (LHA) that terminate in the lateral habenula (LHb). Wild type mice were injected with an adeno-associated virus (AAV) encoding GCaMP6s (AAV-hSyn-GCaMP6s) in the LHA and an optic fiber was implanted with the tip immediately above the LHb (Figure 4A). GCaMP6s expressi...

Watch this Scientific Journal Video about Multi-Fiber Photometry to Record Neural Activity in Freely-Moving Animals at JoVE.com

Multi-Fiber Photometry to Record Neural Activity in Freely-Moving Animals

This protocol details how to implement and perform multi-fiber photometry recordings, how to correct for calcium-independent artifacts, and important considerations for dual-color photometry imaging.

Recording the activity of a group of neurons in a freely-moving animal is a challenging undertaking. Moreover, as the brain is dissected into smaller and ...

Fiber photometry CSOM imaging combines genetically encoded custom indicator and optical fibers to monitor neural activity in freely-moving animal. Which is critical to understand how a specific group of neurons play in directing or responding to an action or a stimulus. This technique is an accessible way to record from specific brain regions that are defined by their connection or genetic profiles, with the built in control to separate signal to noise.Demonstrating the processor will be, Ekaterina Martianova, a graduate student in my lab. Begin by loosening all screws on the five axis translator. Screw in the patch cord to the adaptor that is affixed to the five axis translator.Then turn on the 470 nanometer excitation light at low power and position the tip of the fiber pointing to an auto fluorescent plastic slide. Next record from the complimentary metal-oxide semiconductor or CMOS camera in live mode. Increase the gain or adjust the lookup table until the image is visible at the focal point of the objective.Advance the five axis translator towards the objective ensuring that the 470 nanometer light is centered on the fiber at the SMA or FC end of the patch cord, until an image can be resolved on the camera. Finally, adjust the X and Y axes until the image is centered and well resolved. Visualize the light emitted from the variole end of the patch cord, as it should appear as an isotropic circle.Start by turning on all the excitation lights to better visualize the fibers. And adjust the camera gains such that no pixels are saturated, and a clear image of the fibers are present. Then take a preliminary image.Draw regions of interest or ROIS around the fibers. And keep them for the measurement of the mean intensity values during recordings. For multiple fiber recordings, test for independents by live recording from all fibers.Point one fiber towards a light source, and tap with a finger. Observe fluctuations in the channel. Then apply colored tape to the end of the fibers to label which ROI corresponds to which fiber.Finally, set up the recording area by hanging the patch cord above the arena using stands, clamps, and holders. Lastly, make sure that the animal will not be limited in movement by the length of the fiber and can freely move throughout the entire arena. Begin by inspecting the distal end of the fibers of the patch cord by eye, and with a mini fiber microscope.If the surface of the fibers is scratched, re-polish the fibers using fiber polishing film with fine grit. Then clean the distal ends of the patch cord with 70%ethanol and a cotton tip applicator. Clean the fiber optic cannulas using 70%ethanol and a cotton tip applicator.Connect the variole end of the patch cord to the implanted fiber using a ceramic split sleeve covered with a black shrink tube. During the connection, make sure that the sleeve is tight. Allow the animal to recover for a few minutes.Then start recording the optical signal, and run the experiment. While recording keep a careful eye on the live trace to ensure quality recordings. And watch for any jumps in signal, indicating the sleeve is not tight enough.For the dual color recordings, add a 516 nanometer LED to the photometry system to excite the red fluorescent calcium sensor, and appropriate dichroic mirrors and filters. Then add an image splitter between the objective and the CMOS camera to separate the green and red emission wave lengths. Finally, draw ROIS around all fibers in both colors, red and green.Make sure to clearly identify each ROI with the corresponding fiber and channel. Trigger simultaneous excitation with 470 nanometer, and 560 nanometer LEDs, and alternate them with a 410 nanometer LED. Results indicate, the measured fluorescence significantly increased concurrently with the administration of air puffs for mice with activation of the LHA-LHB pathway.However, in mice expressing green fluorescent protein or GFP, no change in the signal was detected during the administration of air puffs. This technique allows researchers to reasonably record from specific type of cells in different brain regions. While an animal is freely behaving.This furthers the ability to dissect the functions of these brain regions as they relate to behavior.

multi-fiber-photometry-to-record-neural-activity-freely-moving

Research

JoVE Journal

Neuroscience

High-density EEG Recordings of the Freely Moving Mice using Polyimide-based Microelectrode

Electroencephalogram (EEG) indicates the averaged electrical activity of the neuronal populations on a large-scale level. It is widely utilized as a noninvasive brain monitoring tool in cognitive neuroscience as well as a diagnostic tool for epilepsy and sleep disorders in neurology. However, the underlying mechanism of EEG rhythm generation is still under the veil. Recently introduced polyimide-based microelectrode (PBM-array) for high resolution mouse EEG1 is one of the trials to answer the neurophysiological questions on EEG signals based on a rich genetic resource that the mouse model contains for the analysis of complex EEG generation process. This application of nanofabricated PBM-array to mouse skull is an efficient tool for collecting large-scale brain activity of transgenic mice and accommodates to identify the neural correlates to certain EEG rhythms in conjunction with behavior. However its ultra-thin thickness and bifurcated structure cause a trouble in handling and implantation of PBM-array. In the presented video, the preparation and surgery steps for the implantation of PBM-array on a mouse skull are described step by step. Handling and surgery tips to help researchers succeed in implantation are also provided.

In this article, we described the surgery procedure and handling tips for implantation of ultra-thin polyimide-based microelectrode array (PBM-array) on the mouse skull for acquisition of high-density encephalography (EEG) in a mouse model.

Electroencephalogram (EEG) indicates the averaged electrical activity of the neuronal populations on a large-scale level. It is widely utilized as a ...

Design and Assembly of an Ultra-light Motorized Microdrive for Chronic Neural Recordings in Small Animals

The ability to chronically record from populations of neurons in freely behaving animals has proven an invaluable tool for dissecting the function of neural circuits underlying a variety of natural behaviors, including navigation1 , decision making 2,3, and the generation of complex motor sequences4,5,6. Advances in precision machining has allowed for the fabrication of light-weight devices suitable for chronic recordings in small animals, such as mice and songbirds. The ability to adjust the electrode position with small remotely controlled motors has further increased the recording yield in various behavioral contexts by reducing animal handling.6,7
 Here we describe a protocol to build an ultra-light motorized microdrive for long-term chronic recordings in small animals. Our design evolved from an earlier published version7, and has been adapted for ease-of use and cost-effectiveness to be more practical and accessible to a wide array of researchers. This proven design 8,9,10,11 allows for fine, remote positioning of electrodes over a range of ~ 5 mm and weighs less than 750 mg when fully assembled. We present the complete protocol for how to build and assemble these drives, including 3D CAD drawings for all custom microdrive components.

The design, fabrication and assembly of an ultra-light motorized microdrive is described. The device provides a cost-effective and easy-to-use solution for chronic recordings of single units in small behaving animals.

The ability to chronically record from populations of neurons in freely behaving animals has proven an invaluable tool for dissecting the function of ...

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Detection and interpretation of olfactory cues are critical for the survival of many organisms. Remarkably, species across phyla have strikingly similar olfactory systems suggesting that the biological approach to chemical sensing has been optimized over evolutionary time1. In the insect olfactory system, odorants are transduced by olfactory receptor neurons (ORN) in the antenna, which convert chemical stimuli into trains of action potentials. Sensory input from the ORNs is then relayed to the antennal lobe (AL; a structure analogous to the vertebrate olfactory bulb). In the AL, neural representations for odors take the form of spatiotemporal firing patterns distributed across ensembles of principal neurons (PNs; also referred to as projection neurons)2,3. The AL output is subsequently processed by Kenyon cells (KCs) in the downstream mushroom body (MB), a structure associated with olfactory memory and learning4,5. Here, we present electrophysiological recording techniques to monitor odor-evoked neural responses in these olfactory circuits.
First, we present a single sensillum recording method to study odor-evoked responses at the level of populations of ORNs6,7. We discuss the use of saline filled sharpened glass pipettes as electrodes to extracellularly monitor ORN responses. Next, we present a method to extracellularly monitor PN responses using a commercial 16-channel electrode3. A similar approach using a custom-made 8-channel twisted wire tetrode is demonstrated for Kenyon cell recordings8. We provide details of our experimental setup and present representative recording traces for each of these techniques.

Multi-unit Recording Methods to Characterize Neural Activity in the Locust Schistocerca Americana Olfactory Circuits

We demonstrate variations of the extracellular multi-unit recording technique to characterize odor-evoked responses in the first three stages of the invertebrate olfactory pathway. These techniques can easily be adapted to examine ensemble activity in other neural systems as well.

Detection and interpretation of olfactory cues are critical for the survival  of many organisms. Remarkably, species across phyla have strikingly similar ...

A Novel Behavioral Assay to Investigate Gustatory Responses of Individual, Freely-moving Bumble Bees (Bombus terrestris)

Generalist pollinators like the buff-tailed bumble bee, Bombus terrestris, encounter both nutrients and toxins in the floral nectar they collect from flowering plants. Only a few studies have described the gustatory responses of bees toward toxins in food, and these experiments have mainly used the proboscis extension response on restrained honey bees. Here, a new behavioral assay is presented for measuring the feeding responses of freely-moving, individual worker bumble bees to nutrients and toxins. This assay measures the amount of solution ingested by each bumble bee and identifies how tastants in food influence the microstructure of the feeding behavior.
The solutions are presented in a microcapillary tube to individual bumble bees that have been previously starved for 2-4 hr. The behavior is captured on digital video. The fine structure of the feeding behavior is analyzed by continuously scoring the position of the proboscis (mouthparts) from video recordings using event logging software. The position of the proboscis is defined by three different behavioral categories: (1) proboscis is extended and in contact with the solution, (2) proboscis is extended but not in contact with the solution and (3) proboscis is stowed under the head. Furthermore the speed of the proboscis retracting away from the solution is also estimated.
In the present assay the volume of solution consumed, the number of feeding bouts, the duration of the feeding bouts and the speed of the proboscis retraction after the first contact is used to evaluate the phagostimulatory or the deterrent activity of the compounds tested.
This new taste assay will allow researchers to measure how compounds found in nectar influence the feeding behavior of bees and will also be useful to pollination biologists, toxicologists and neuroethologists studying the bumble bee&#39;s taste system.

A Novel Behavioral Assay to Investigate Gustatory Responses of Individual, Freely-moving Bumble Bees Bombus terrestris

A novel behavioral assay is described for investigating the short term gustatory responses of the mouthparts of freely-moving bumble bees (Bombus terrestris) toward nutrients and toxins in solution.

Generalist pollinators like the buff-tailed bumble bee, Bombus terrestris, encounter both nutrients and toxins in the floral nectar they collect from ...

Recording EEG in Freely Moving Neonatal Rats Using a Novel Method

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such as epilepsy and neurodegenerative disorders. However, it is technically difficult to obtain EEG recordings in neonates as it requires specialized handling and great care. Here, we present a novel method to record EEG in neonatal rat pups (P8-P15). We designed a simple and reliable electrode using computer pin loci; it can be easily implanted into the skull of a rat pup to record high-quality EEG signals in the normal and epileptic brain. Pups were given an intraperitoneal (i.p.) injection of the neurotoxin kainic acid (KA) to induce epileptic seizures. The surgical implantation performed in this procedure is less expensive than other EEG procedures for neonates. This method allows one to record high-quality and stable EEG signals for more than 1 week. Furthermore, this procedure can also be applied to adult rats and mice to study epilepsy or other neurological disorders.

Here, we introduce a novel technique designed to record electroencephalography (EEG) in freely moving neonatal epileptic pups and describe its procedures, features, and applications. This method allows one to record EEG for more than 1 week.

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such ...

Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls body functions and internal organ rhythms when animals are exposed to emotional challenges and changes in their living environments. In general experiments, signals from different organs, such as the brain and the heart, are recorded by independent recording systems that require multiple recording devices and different procedures for processing the data files. This study describes a new method that can simultaneously monitor electrical biosignals, including tens of local field potentials in multiple brain regions, electrocardiograms that represent the cardiac rhythm, electromyograms that represent awake/sleep-related muscle contraction, and breathing signals, in a freely moving rat. The recording configuration of this method is based on a conventional micro-drive array for cortical local field potential recordings in which tens of electrodes are accommodated, and the signals obtained from these electrodes are integrated into a single electrical board mounted on the animal's head. Here, this recording system was improved so that signals from the peripheral organs are also transferred to an electrical interface board. In a single surgery, electrodes are first separately implanted into the appropriate body parts and the target brain areas. The open ends of all of these electrodes are then soldered to individual channels of the electrical board above the animal's head so that all of the signals can be integrated into the single electrical board. Connecting this board to a recording device allows for the collection of all of the signals into a single device, which reduces experimental costs and simplifies data processing, because all data can be handled in the same data file. This technique will aid the understanding of the neurophysiological correlates of the associations between central and peripheral organs.

This study introduces a method for the simultaneous recording of local field potentials in the brain, electrocardiograms, electromyograms, and breathing signals of a freely moving rat. This technique, which reduces experimental costs and simplifies data analysis, will contribute to the understanding of the interactions between the brain and peripheral organs.

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls ...

Noninvasive EEG Recordings from Freely Moving Piglets

The method allows the recording of high-quality electroencephalograms (EEGs) from freely moving piglets directly in the pigpen. We use a one-channel telemetric electroencephalography system in combination with standard self-adhesive hydrogel electrodes. The piglets are calmed down without the use of sedatives. After their release into the pigpen, the piglets behave normally&#8212;they drink and sleep in the same cycle as their siblings. Their sleep phases are used for the EEG recordings.

Here, we present a protocol to record telemetric electroencephalograms (EEGs) from freely moving piglets directly in the pigpen without the use of a sedative, making it possible to record typical EEG patterns during non-REM sleep, like spindle bursts.

The method allows the recording of high-quality electroencephalograms (EEGs) from freely moving piglets directly in the pigpen. We use a one-channel ...

Microdialysis of Excitatory Amino Acids During EEG Recordings in Freely Moving Rats

Microdialysis is a well-established neuroscience technique that correlates the changes of neurologically active substances diffusing into the brain interstitial space with the behavior and/or with the specific outcome of a pathology (e.g., seizures for epilepsy). When studying epilepsy, the microdialysis technique is often combined with short-term or even long-term video-electroencephalography (EEG) monitoring to assess spontaneous seizure frequency, severity, progression and clustering. The combined microdialysis-EEG is based on the use of several methods and instruments. Here, we performed in vivo microdialysis and continuous video-EEG recording to monitor glutamate and aspartate outflow over time, in different phases of the natural history of epilepsy in a rat model. This combined approach allows the pairing of changes in the neurotransmitter release with specific stages of the disease development and progression. The amino acid concentration in the dialysate was determined by liquid chromatography. Here, we describe the methods and outline the principal precautionary measures one should take during in vivo microdialysis-EEG, with particular attention to the stereotaxic surgery, basal and high potassium stimulation during microdialysis, depth electrode EEG recording and high-performance liquid chromatography analysis of aspartate and glutamate in the dialysate. This approach may be adapted to test a variety of drug or disease induced changes of the physiological concentrations of aspartate and glutamate in the brain. Depending on the availability of an appropriate analytical assay, it may be further used to test different soluble molecules when employing EEG recording at the same time.

Here, we describe a method for in vivo microdialysis to analyze aspartate and glutamate release in the ventral hippocampus of epileptic and non-epileptic rats, in combination with EEG recordings. Extracellular concentrations of aspartate and glutamate may be correlated with the different phases of the disease.

Microdialysis is a well-established neuroscience technique that correlates the changes of neurologically active substances diffusing into the brain ...

Intracerebroventricular Delivery of Gut-Derived Microbial Metabolites in Freely Moving Mice

The impact of gut microbiota and their metabolites on host physiology and behavior has been extensively investigated in this decade. Numerous studies have revealed that gut microbiota-derived metabolites modulate brain-mediated physiological functions through intricate gut-brain pathways in the host. Short-chain fatty acids (SCFAs) are the major bacteria-derived metabolites produced during dietary fiber fermentation by the gut microbiome. Secreted SCFAs from the gut can act at multiple sites in the periphery, affecting the immune, endocrine, and neural responses due to the vast distribution of SCFAs receptors. Therefore, it is challenging to differentiate the central and peripheral effects of SCFAs through oral and intraperitoneal administration of SCFAs. This paper presents a video-based method to interrogate the functional role of SCFAs in the brain via a guide cannula in freely moving mice. The amount and type of SCFAs in the brain can be adjusted by controlling the infusion volume and rate. This method can provide scientists with a way to appreciate the role of gut-derived metabolites in the brain.

Gut-derived microbial metabolites have multifaceted effects leading to complex behavior in animals. We aim to provide a step-by-step method to delineate the effects of gut-derived microbial metabolites in the brain via intracerebroventricular delivery via a guide cannula.

The impact of gut microbiota and their metabolites on host physiology and behavior has been extensively investigated in this decade. Numerous studies have ...

Extracellular Electrophysiological Recording in the Motor Cortex of Mice

Multichannel Extracellular Recording in Freely Moving Mice

The protocol aims to uncover the properties of neuronal firing and network local field potentials (LFPs) in behaving mice carrying out specific tasks by correlating the electrophysiological signals with spontaneous and/or specific behavior. This technique represents a valuable tool in studying the neuronal network activity underlying these behaviors. The article provides a detailed and complete procedure for electrode implantation and consequent extracellular recording in free-moving conscious mice. The study includes a detailed method for implanting the microelectrode arrays, capturing the LFP and neuronal spiking signals in the motor cortex (MC) using a multichannel system, and the subsequent offline data analysis. The advantage of multichannel recording in conscious animals is that a greater number of spiking neurons and neuronal subtypes can be obtained and compared, which allows the evaluation of the relationship between a specific behavior and the associated electrophysiological signals. Notably, the multichannel extracellular recording technique and the data analysis procedure described in the present study can be applied to other brain areas when conducting experiments in behaving mice.

Author Spotlight: Advancements in Multichannel Extracellular Recording for Studying Neuronal Activity in Freely Moving Mice 

The protocol describes the methodology of extracellular recording in the motor cortex (MC) to reveal extracellular electrophysiological properties in freely moving conscious mice, as well as the data analysis of local field potentials (LFPs) and spikes, which is useful for evaluating the network neural activity underlying behaviors of interest.

The protocol aims to uncover the properties of neuronal firing and network local field potentials (LFPs) in behaving mice carrying out specific tasks by ...