We would like to acknowledge Linus D. Sun and Jeenah Jung, who originated the use of computer-designed, 3-D printed drives in our lab for recording from mice. In addition, we would like to acknowledge all past and present Wilson Lab members for their contributions to micro-drive and tetrode technology development in the lab.

<ol>
<li>Micro-drive array overview A complete micro-drive array consists of several main components (Fig. 1):<ol>
<li>A computer designed plastic base that holds 21 independently movable micro-drives. </li>
<li>A cannula at the bottom of the base in which a set of stationary tetrode guide tubes is collected.</li>
<li>21 micro-drives that each carry one tetrode and which can be individually driven using a screw.</li>
<li>An electrode interface board for interconnection between tetrodes and pre-amplifiers.</li>
<li>A...

<ol>
	<li>Yamamoto, J., Wilson, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large-scale+chronically+implantable+precision+motorized+microdrive+array+for+freely+behaving+animals.">Large-scale chronically implantable precision motorized microdrive array for freely behaving animals.</a> J. Neurophysiol. 100 (4), 2430-2440 (2008).</li></ol>

This protocol describes the general features of the micro-drive building process. It has been successfully modified to produce drive arrays with multiple recording targets or smaller arrays for recordings in mice. In addition to recording from the hippocampus, many researchers use these drives to target cortical and sub-cortical structures. The use of longer screws may be required in order to record from deeper brain structures. With extensive modification to the drive base, motorized drives my be used in lieu of manuall...

<table>
<tbody>
	<tr>
<td>3D-printed plastic parts</td>
<td>Material</td>
<td>American Precision Prototyping (www.approto.com)</td>
<td>&nbsp;</td>
<td>Parts are printed by stereolithography (SLA) using DSM 14120 'ABS-like', DSM 11120 'Watershed' or Accura 55 resins. Request high resolution, no sandblasting.</td>
</tr>
<tr>
<td>stainless steel tubing</td>
<td>Material</td>
<td>Small Parts (www.smallparts.com)</td>
<td>&nbsp;</td>
<td>Cut to custom length and deburred. Sizes: 13 Ga (1pc @ 10 mm), 15 Ga (1pc @ 50 mm), 20 Ga thin wall (21pcs @ 5 mm), 23 Ga (21pcs @ 14 mm) </td>
</tr>
<tr>
<td>Dental cement powder</td>
<td>Material</td>
<td>A-M Systems (www.a-msystems.com)</td>
<td>525000</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>Dental cement solvent</td>
<td>Material</td>
<td>A-M Systems</td>
<td>526000</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>Electrode interface board</td>
<td>Material</td>
<td>Neuralynx (www.neuralynx.com)</td>
<td>EIB-81</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>micro-drive screws</td>
<td>Material</td>
<td>Advanced Machining and Tooling (www.amtmfg.com)</td>
<td>&nbsp;</td>
<td>Custom made screws with M1.2x0.25 threads (1.2mm major diameter; 0.25 mm thread pitch) </td>
</tr>
<tr>
<td>'Tap-Ease' tapping lubricant </td>
<td>Tool</td>
<td>AGS Company</td>
<td>TA-2</td>
<td>Available from MSC Industrial Supply (www.mscdirect.com)</td>
</tr>
<tr>
<td>Polyimide tubing </td>
<td>Material</td>
<td>IWG High Performance Conductors (www.iwghpc.com)</td>
<td>&nbsp;</td>
<td>Sizes:
30 Ga (outer diam: 0.0071", inner diam: 0.0116") 
5 mil (nominal outer diam: 0.0055", wall: 0.0010") </td>
</tr>
<tr>
<td>Cordless drill/driver </td>
<td>Tool</td>
<td>DeWalt (www.dewalt.com)</td>
<td>DC750KA </td>
<td>Any drill/driver with low-speed setting will do.</td>
</tr>
<tr>
<td>Variable-Speed Rotary Tool Kit with Flex-Shaft </td>
<td>Tool</td>
<td>Dremel (www.dremel.com)</td>
<td>3956-02</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>Razor blades </td>
<td>Tool</td>
<td>VWR (www.vwr.com)</td>
<td>55411-050</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>Watch glass </td>
<td>Tool</td>
<td>VWR</td>
<td>66112-107</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>Tap for custom screws</td>
<td>Tool</td>
<td>Balax</td>
<td>01302</td>
<td>M1.2x0.25 Thredfloer thread-forming tap. Available from MSC Industrial Supply.</td>
</tr>
<tr>
<td>Taps</td>
<td>Tool</td>
<td>Small Parts</td>
<td>HSBT-0172, HSBT-0080 </td>
<td>1-72, 0-80 bottoming taps</td>
</tr>
<tr>
<td>Pin vise</td>
<td>Tool</td>
<td>Vargus</td>
<td>3SHC1--I </td>
<td>To hold taps. Available from MSC Industrial Supply</td>
</tr>
<tr>
<td>Drill bits</td>
<td>Tool</td>
<td>Small Parts</td>
<td>&nbsp;</td>
<td>Sizes: 65, 61, 1.55mm</td>
</tr>
<tr>
<td>Benchtop vise</td>
<td>Tool</td>
<td>Panavise</td>
<td>301</td>
<td>Available from DigiKey (www.digikey.com).</td>
</tr>
<tr>
<td>No.2 Dumont forceps</td>
<td>Tool</td>
<td>Stoelting (www.stoeltingco.com)</td>
<td>52100-27</td>
<td>Used for inserting 20 Ga thin wall sleeves.</td>
</tr>
<tr>
<td>5-minute epoxy</td>
<td>Tool</td>
<td>Allied Electronics (www.alliedelec.com)</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
</tr>
<tr>
<td>Accu-Tek Carbofib Tip Tweezers</td>
<td>Tool</td>
<td>Aven (www.aveninc.com)</td>
<td>18768 (Pattern 304)</td>
<td>Used for handling polyimide tubing.</td>
</tr>
<tr>
<td>Micro Dissecting Scissors</td>
<td>Tool</td>
<td>Biomedical Research Instruments (www.biomedinstr.com)</td>
<td>25-1000</td>
<td>Used for cutting polyimide carrier tubes</td>
</tr>
<tr>
<td>Xcelite 378M Pliers</td>
<td>Tool</td>
<td>Newark</td>
<td>96F8903</td>
<td>Used for handling stainless steel tubing and for construction of custom screwdriver.</td>
</tr>
<tr>
<td>mini pin vise</td>
<td>Tool</td>
<td>MSC Industrial Supply (www.mscdirect.com)</td>
<td>00920314 </td>
<td>Used as a handle for custom made screwdriver.</td>
</tr>		
</tbody>
</table>

micro-drive array for chronic in vivo recording: drive fabrication

Chronic recording of large populations of neurons is a valuable technique for studying the function of neuronal circuits in awake behaving rats. Lightweight recording devices carrying a high density array of tetrodes allow for the simultaneous monitoring of the activity of tens to hundreds of individual neurons. Here we describe a protocol for the fabrication of a micro-drive array with twenty one independently movable micro-drives. This device has been used successfully to record from hippocampal and cortical neurons in our lab. We show how to prepare a custom designed, 3-D printed plastic base that will hold the micro-drives. We demonstrate how to construct the individual micro-drives and how to assemble the complete micro-drive array. Further preparation of the drive array for surgical implantation, such as the fabrication of tetrodes, loading of tetrodes into the drive array and gold-plating, is covered in a subsequent video article.

Watch this Scientific Journal Video about Micro-drive Array for Chronic in vivo Recording: Drive Fabrication at JoVE.com

Micro-drive Array for Chronic in vivo Recording: Drive Fabrication

In this protocol we demonstrate how to fabricate a micro-drive array for chronic electrophysiological recordings in rats.

Micro-drive Array for Chronic in vivo Recording: Drive Fabrication

Chronic recording of large populations of neurons is a valuable technique for studying the function of neuronal circuits in awake behaving rats. ...

micro-drive-array-for-chronic-in-vivo-recording-drive-fabrication

Research

JoVE Journal

Biology

25.1K ビュー.  MIT - Massachusetts Institute of Technology. このプロトコルでは、ラットの慢性的な電気生理学的記録のためのマイクロドライブのアレイを作製する方法を示します。

ビデオ: Micro-drive Array for Chronic in vivo Recording: Drive Fabrication

In this protocol we demonstrate how to fabricate a micro-drive array for chronic electrophysiological recordings in...

micro-drive array for chronic in vivo recording: tetrode assembly

Micro-drive Array for Chronic in vivo Recording: Tetrode Assembly

In this protocol we demonstrate how to fabricate and condition tetrodes for use with a micro-drive array, which was designed for chronic electrophysiological recordings in rats.  In addition, we illustrate the final stages of micro-drive array construction, which includes installing ground wires and a protective...

Micro-drive Array for Chronic in vivo Recording: Tetrode Assembly

large-scale recording of neurons by movable silicon probes in behaving rodents

Large-scale Recording of Neurons by Movable Silicon Probes in Behaving Rodents

We describe methods for large-scale recording of multiple single units and local field potential in behaving rodents with silicon probes. Drive fabrication, probe attachment to the drive and probe implantation processes are illustrated in sufficient details for easy...

the td drive: a parametric, open-source implant for multi&#45;area electrophysiological recordings in behaving and sleeping rats

Author Spotlight: Deciphering Memory and Learning Through Neural Implants for Multi&amp;#45;Region Brain Studies

Here, we present a unique, 3D-printable implant for rats, named TD Drive, capable of symmetric, bilateral wire electrode recordings, currently in up to ten distributed brain areas simultaneously.

The TD Drive: A Parametric, Open-Source Implant for Multi&#45;Area Electrophysiological Recordings in Behaving and Sleeping Rats

a lightweight drive implant for chronic tetrode recordings in juvenile mice

Author Spotlight: A Lightweight Drive Implant for Chronic Tetrode Recordings in Juvenile Mice  

Here, we describe a micro-drive design, surgical implantation procedure, and post-surgery recovery strategy that allow for chronic field and single-unit recordings from multiple brain regions simultaneously in juvenile and adolescent mice across a critical developmental window from postnatal day 20 (p20) to postnatal day 60 (p60) and...

Micro-Drive Implant for Recording Daily Neural Activity in Juvenile Mice

construction of an improved multi-tetrode hyperdrive for large-scale neural recording in behaving rats

Construction of an Improved Multi-Tetrode Hyperdrive for Large-Scale Neural Recording in Behaving Rats

We present the construction of a 3D-printable hyperdrive with eighteen independently adjustable tetrodes. The hyperdrive is designed to record brain activity in freely behaving rats over a period of several...

Open&amp;#45;Source Implant for Multi&amp;#45;Region Electrophysiological Monitoring in Rats

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chronic implantation of multiple flexible polymer electrode arrays

Chronic Implantation of Multiple Flexible Polymer Electrode Arrays

Described below is a method for implantation of multiple polymer electrode arrays across anatomically distant brain regions for chronic electrophysiological recording in freely moving rats. Preparation and surgical implantation are described in detail, with emphasis on design principles to guide adaptation of these methods for use in other...

A Lightweight Drive Implant for Chronic Tetrode Recordings in Juvenile Mice

simultaneous recordings of cortical local field potentials, electrocardiogram, electromyogram, and breathing rhythm from a freely moving rat

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

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...

the dream implant: a lightweight, modular, and cost-effective implant system for chronic electrophysiology in head-fixed and freely behaving mice

Author Spotlight: Understanding Naturalistic Neuronal Computation Through Virtual Reality and Electrophysiology in Animals

Here, we introduce a lightweight, cost-effective probe implant system for chronic electrophysiology in rodents optimized for ease of use, probe recovery, experimental versatility, and compatibility with...

The DREAM Implant: A Lightweight, Modular, and Cost-Effective Implant System for Chronic Electrophysiology in Head-Fixed and Freely Behaving Mice

a micro-drive array method for electrophysiological recording from multiple brain regions

The video demonstrates simultaneous electrophysiological recording from multiple brain regions in a rat using an integrative micro-drive array. Recording electrodes are placed in the olfactory bulb, reference electrodes in the frontal cortex, and micro-drive array electrodes in the hippocampus. The rat is allowed to forage freely, and simultaneous recording from all the implanted electrodes is recorded using the...

A Micro-drive Array Method for Electrophysiological Recording from Multiple Brain Regions

micro-drive system assembly and electrode array implantation for multichannel recording in mice

Micro-Drive System Assembly and Electrode Array Implantation for Multichannel Recording in Mice  

Then, implant the two BR electrodes just deep enough for the tips of the screws to make contact with the brain, which is about 2 millimeters deep. Use between six and eight full turns of the...

hybrid microdrive system with recoverable opto-silicon probe and tetrode for dual-site high density recording in freely moving mice

Hybrid Microdrive System with Recoverable Opto-Silicon Probe and Tetrode for Dual-Site High Density Recording in Freely Moving Mice

This protocol describes the construction of a hybrid microdrive array that allows implantation of nine independently adjustable tetrodes and one adjustable opto-silicon probe in two brain regions in freely moving mice. Also demonstrated is a method for safely recovering and reusing the opto-silicon probe for multiple...

recovery after micro-drive implantation

Recovery after Micro-Drive Implantation  

micro-drive design and construction

Micro-Drive Design and Construction  

electrophysiology of laminar cortical activity in the common marmoset

Author Spotlight: Optimizing Micro-Drive Systems for Precise Electrode Positioning in Marmoset

Custom-built micro-drives enable the sub-millimeter targeting of cortical recording sites with linear silicon...

Development of an X-Y Electrode Stage for Precise Neural Recording in Marmosets

https://cloudfront.jove.com/CDNSource/teasers/65397_b.jpg

A Cost-Effective, Versatile Probe Implant System for Electrophysiology in Mice

https://cloudfront.jove.com/CDNSource/teasers/66867_b.jpg

multichannel extracellular recording in freely moving 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.

Extracellular Electrophysiological Recording in the Motor Cortex of Mice

Population Replacement Strategies for Controlling Vector Populations and the Use of Wolbachia pipientis for Genetic Drive

In this video, Jason Rasgon discusses population replacement strategies to control vector-borne diseases such as malaria and dengue. "Population replacement" is the replacement of wild vector populations (that are competent to transmit pathogens) with those that are not competent to transmit pathogens. There are several theoretical strategies to accomplish this. One is to exploit the maternally-inherited symbiotic bacteria Wolbachia pipientis. Wolbachia is a widespread reproductive parasite that spreads in a selfish manner at the extent of its host's fitness. Jason Rasgon discusses, in detail, the basic biology of this bacterial symbiont and various ways to use it for control of vector-borne diseases.

In this interview, Jason Rasgon explains the concept of genetic drive and the characteristics of an effective gene drive system. The use of the endosymbiotic bacterium, Wolbachia pipientis, as a means to spread genes through mosquito populations, is hypothesized.

In this video, Jason Rasgon discusses population replacement strategies to control vector-borne diseases such as malaria and dengue. "Population ...

Application of Light-cured Dental Adhesive Resin for Mounting Electrodes or Microdialysis Probes in Chronic Experiments

In chronic recording experiments, self-curing dental acrylic resins have been used as a mounting base of electrodes or microdialysis-probes. Since these acrylics do not bond to the bone, screws have been used as anchors. However, in small experimental animals like finches or mouse, their craniums are very fragile and can not successfully hold the anchors. In this report, we propose a new application of light-curing dental resins for mounting base of electrodes or microdialysis probes in chronic experiments. This material allows direct bonding to the cranium. Therefore, anchor screws are not required and surgical field can be reduced considerably. Past experiences show that the bonding effect maintains more than 2 months. Conventional resin's window of time when the materials are pliable and workable is a few minutes. However, the window of working time for these dental adhesives is significantly wider and adjustable.

In this report, we propose a new application of light-curing dental resins for mounting base of electrodes or microdialysis probes in chronic experiments. This material allows direct bonding to the cranium.

In chronic recording experiments, self-curing dental acrylic resins have been used as a mounting base of electrodes or microdialysis-probes. Since these ...

A Craniotomy Surgery Procedure for Chronic Brain Imaging

Imaging techniques are becoming increasingly important in the study brain function. Among them, two-photon laser scanning microscopy has emerged as an extremely useful method, because it allows the study of the live intact brain. With appropriate preparations, this technique allows the observation of the same cortical area chronically, from minutes to months. In this video, we show a preparation for chronic in vivo imaging of the brain using two-photon microscopy. This technique was initially pioneered by Dr. Karel Svoboda, who is now a Howard Hughes Medical Institute Investigator at Janelia Farm. Preparations like the one shown here can be used for imaging of neocortical structure (e.g., dendritic and axonal dynamics),  to record neuronal activity using calcium-sensitive dyes, to image cortical blood flow dynamics, or for intrinsic optical imaging studies. Deep imaging of the neocortex is possible with optimal cranial window surgeries. Operating under the most sterile conditions possible to avoid infections, together with using extreme care to do not damage the dura mater during the surgery, will result in successful and long-lasting glass-covered cranial windows.


This video and protocol demonstrate how to implant a glass-covered cranial window in rodents. These preparations can be used for chronic in vivo two-photon imaging of the neocortex over time scales of months. It may also be used for other types of imaging, including optical intrinsic signal imaging.

Imaging techniques are becoming increasingly important in the study brain function. Among them, two-photon laser scanning microscopy has emerged as an ...

The tetrode, a bundle of four electrodes, has proven to be a valuable tool for the simultaneous recording of multiple neurons in-vivo.  The differential amplitude of action potential signatures over the channels of a tetrode allows for the isolation of single-unit activity from multi-unit signals.  The ability to precisely control the stereotaxic location and depth of the tetrode is critical for studying coordinated neural activity across brain regions.  In combination with a micro-drive array, it is possible to achieve precise placement and stable control of many tetrodes over the course of days to weeks.  In this protocol, we demonstrate how to fabricate and condition tetrodes using basic tools and materials, install the tetrodes into a multi-drive tetrode array for chronic in-vivo recording in the rat, make ground wire connections to the micro-drive array, and attach a protective cone onto the micro-drive array in order to protect the tetrodes from physical contact with the environment.



In this protocol we demonstrate how to fabricate and condition tetrodes for use with a micro-drive array, which was designed for chronic electrophysiological recordings in rats. In addition, we illustrate the final stages of micro-drive array construction, which includes installing ground wires and a protective cone.

The tetrode, a bundle of four electrodes, has proven to be a valuable tool for the simultaneous recording of multiple neurons in-vivo.  The differential ...

Electrophysiological Recording in the Drosophila Embryo

Drosophila is a premier genetic model for the study of both embryonic development and functional neuroscience. Traditionally, these fields are quite isolated from each other, with largely independent histories and scientific communities. However, the interface between these usually disparate fields is the developmental programs underlying acquisition of functional electrical signaling properties and differentiation of functional chemical synapses during the final phases of neural circuit formation. This interface is a critically important area for investigation. In Drosophila, these phases of functional development occur during a period of &lt;8 hours (at 25&deg;C) during the last third of embryogenesis. This late developmental period was long considered intractable to investigation owing to the deposition of a tough, impermeable epidermal cuticle. A breakthrough advance was the application of water-polymerizing surgical glue that can be locally applied to the cuticle to enable controlled dissection of late-stage embryos. With a dorsal longitudinal incision, the embryo can be laid flat, exposing the ventral nerve cord and body wall musculature to experimental investigation. Whole-cell patch-clamp techniques can then be employed to record from individually-identifiable neurons and somatic muscles. These recording configurations have been used to track the appearance and maturation of ionic currents and action potential propagation in both neurons and muscles. Genetic mutants affecting these electrical properties have been characterized to reveal the molecular composition of ion channels and associated signaling complexes, and to begin exploration of the molecular mechanisms of functional differentiation. A particular focus has been the assembly of synaptic connections, both in the central nervous system and periphery. The glutamatergic neuromuscular junction (NMJ) is most accessible to a combination of optical imaging and electrophysiological recording. A glass suction electrode is used to stimulate the peripheral nerve, with excitatory junction current (EJC) recordings made in the voltage-clamped muscle. This recording configuration has been used to chart the functional differentiation of the synapse, and track the appearance and maturation of presynaptic glutamate release properties. In addition, postsynaptic properties can be assayed independently via iontophoretic or pressure application of glutamate directly to the muscle surface, to measure the appearance and maturation of the glutamate receptor fields. Thus, both pre- and postsynaptic elements can be monitored separately or in combination during embryonic synaptogenesis. This system has been heavily used to isolate and characterize genetic mutants that impair embryonic synapse formation, and thus reveal the molecular mechanisms governing the specification and differentiation of synapse connections and functional synaptic signaling properties.

Electrophysiological Recording in the Drosophila Embryo

Electrophysiological recordings from Drosophila embryos allow analyses of developing muscle and neuron electrical properties, as well as characterization of functional synaptogenesis at the glutamatergic neuromuscular junction and central cholinergic and GABAergic synapses.

Drosophila is a premier genetic model for the study of both embryonic development and functional neuroscience. Traditionally, these fields are quite ...

In vivo-like Organotypic Murine Retinal Wholemount Culture

Targeted ablations of genes and analysis of animal models is the classical strategy for enrolling specific retinal gene function. However, transgenic, retina-specific or conditional knockout mouse models often display early lethality or suffer from severe malformations, preventing an analysis beyond embryonic or early postnatal stages.
Primary cell culture is an alternative to investigate the effects of exogenously applied recombinant factors, overexpression of genes or siRNA-mediated gene knockdown in a controlled environment. Dissociated cell culture has the advantage that the endogenous signals reaching the target cells are reduced, thereby facilitating the identification of exogenously triggered effects after pharmacological manipulation. However, important cell-cell interactions are initially destroyed by enzymatic digestion or mechanical dissociation, even if re-aggregated retinospheroid cultures1 are used.
By contrast, organotypic retinal wholemount cultures provide a system close to the physiological in vivo situation with neuronal interactions and connections still preserved2-5.
In this video article we provide a step by step demonstration of (1) the establishment of in vivo-like organotypic retinal wholemount cultures including dissection peculiarities of embryonic, postnatal and adult murine eyes and (2) a dissociation and cytospin procedure for analysis of neuronal apoptosis and retinal cell proliferation in organotypic wholemounts, e.g. after culture in the presence of exogenously applied recombinant factors.

In vivo-like Organotypic Murine Retinal Wholemount Culture

This video article demonstrates the establishment of organotypic retinal wholemount cultures and a cytospin procedure for analysis of exogenously induced effects. Organotypic retinal wholemount cultures mimic the in vivo situation and significantly facilitate the accessibility of murine retinas for experimental manipulations while circumventing the disadvantages of classical murine animal models.

Targeted ablations of genes and analysis of animal models is the classical strategy for enrolling specific retinal gene function. However, transgenic, ...

Ambulatory ECG Recording in Mice

Telemetric ECG recording in mice is essential to understanding the mechanisms behind arrhythmias, conduction disorders, and sudden cardiac death. Although the surface ECG is utilized for short-term measurements of waveform intervals, it is not practical for long-term studies of heart rate variability or the capture of rare episodes of arrhythmias. Implantable ECG telemeters offer the advantages of simple surgical implantation, long-term recording of electrograms in ambulatory mice, and scalability with simultaneous recordings of multiple animals. Here, we present a step-by-step guide to the implantation of telemeters for ambulatory ECG recording in mice. Careful adherence to aseptic technique is required for favorable survival results with the possibility of implantation and recording from weeks to months. Thus, implantable ECG telemetry is a valuable tool for detection of critical information on cardiac electrophysiology in ambulatory animal models such as the mouse. 

Telemetric ECG has emerged as an essential tool in evaluating animal models for cardiac arrhythmias and sudden cardiac death. Here, we present a stepwise guide to telemetric ECG recordings for application in long-term ambulatory ECG monitoring in mice.

Telemetric ECG recording in mice is essential to understanding the mechanisms behind arrhythmias, conduction disorders, and sudden cardiac death. Although ...

One-channel Cell-attached Patch-clamp Recording

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate depends on properties intrinsic to each channel protein as well as the mechanism by which it is generated and controlled and represents an important area of current research. Information about the operational dynamics of ion channel proteins can be obtained by observing long stretches of current produced by a single molecule. Described here is a protocol for obtaining one-channel cell-attached patch-clamp current recordings for a ligand gated ion channel, the NMDA receptor, expressed heterologously in HEK293 cells or natively in cortical neurons. Also provided are instructions on how to adapt the method to other ion channels of interest by presenting the example of the mechano-sensitive channel PIEZO1. This method can provide data regarding the channel&#8217;s conductance properties and the temporal sequence of open-closed conformations that make up the channel&#8217;s activation mechanism, thus helping to understand their functions in health and disease.

Described here is a procedure for obtaining long stretches of current recording from one ion channel with the cell-attached patch-clamp technique. This method allows for observing, in real time, the pattern of open-close channel conformations that underlie the biological signal. These data inform about channel properties in undisturbed biological membranes.

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate ...

Array Comparative Genomic Hybridization (Array CGH) for Detection of Genomic Copy Number Variants

Array CGH for the detection of genomic copy number variants has replaced G-banded karyotype analysis. This paper describes the technology and its application in a clinical diagnostic service laboratory. DNA extracted from a patient&#8217;s sample (blood, saliva or other tissue types) is labeled with a fluorochrome (either cyanine 5 or cyanine 3). A reference DNA sample is labeled with the opposite fluorochrome. There follows a cleanup step to remove unincorporated nucleotides before the labeled DNAs are mixed and resuspended in a hybridization buffer and applied to an array comprising ~60,000 oligonucleotide probes from loci across the genome, with high probe density in clinically important areas. Following hybridization, the arrays are washed, then scanned and the resulting images are analyzed to measure the red and green fluorescence for each probe. Software is used to assess the quality of each probe measurement, calculate the ratio of red to green fluorescence and detect potential copy number variants.

Array Comparative Genomic Hybridization Array CGH for Detection of Genomic Copy Number Variants

Array CGH for the detection of genomic copy number variants has replaced G-banded karyotype analysis. This paper describes the technology and its application in a diagnostic service laboratory.

Array CGH for the detection of genomic copy number variants has replaced G-banded karyotype analysis. This paper describes the technology and its ...

Measuring DNA Damage and Repair in Mouse Splenocytes After Chronic In Vivo Exposure to Very Low Doses of Beta- and Gamma-Radiation

Low dose radiation exposure may produce a variety of biological effects that are different in quantity and quality from the effects produced by high radiation doses. Addressing questions related to environmental, occupational and public health safety in a proper and scientifically justified manner heavily relies on the ability to accurately measure the biological effects of low dose pollutants, such as ionizing radiation and chemical substances. DNA damage and repair are the most important early indicators of health risks due to their potential long term consequences, such as cancer. Here we describe a protocol to study the effect of chronic in vivo exposure to low doses of &#947;- and &#946;-radiation on DNA damage and repair in mouse spleen cells. Using a commonly accepted marker of DNA double-strand breaks, phosphorylated histone H2AX called &#947;H2AX, we demonstrate how it can be used to evaluate not only the levels of DNA damage, but also changes in the DNA repair capacity potentially produced by low dose in vivo exposures. Flow cytometry allows fast, accurate and reliable measurement of immunofluorescently labeled &#947;H2AX in a large number of samples. DNA double-strand break repair can be evaluated by exposing extracted splenocytes to a challenging dose of 2 Gy to produce a sufficient number of DNA breaks to trigger repair and by measuring the induced (1 hr post-irradiation) and residual DNA damage (24 hrs post-irradiation). Residual DNA damage would be indicative of incomplete repair and the risk of long-term genomic instability and cancer. Combined with other assays and end-points that can easily be measured in such in vivo studies (e.g., chromosomal aberrations, micronuclei frequencies in bone marrow reticulocytes, gene expression, etc.), this approach allows an accurate and contextual evaluation of the biological effects of low level stressors.

Measuring DNA Damage and Repair in Mouse Splenocytes After Chronic In Vivo Exposure to Very Low Doses of Beta- and Gamma-Radiation

A protocol to evaluate changes in DNA damage levels and DNA repair capacity that may be induced by chronic in vivo low dose irradiation in mouse spleen lymphocytes, by measuring phosphorylated histone H2AX, a marker of DNA double-strand breaks, using flow cytometry is presented.

Low dose radiation exposure may produce a variety of biological effects that are different in quantity and quality from the effects produced by high ...