Name: implantation and recording of wireless electroretinogram and visual evoked potential in conscious rats
Uploaded: 2016-06-29T15:00:00
Description: Watch this Scientific Journal Video about Implantation and Recording of Wireless Electroretinogram and Visual Evoked Potential in Conscious Rats at JoVE.com

JC would like to acknowledge the David Hay Memorial Fund, The University of Melbourne for financial support in writing this manuscript. Funding for this project was provided by an ARC Linkage grant 100200129 (BVB, AJV, CTON).

Ethics statement: Animal experiments were conducted in accordance with the Australian Code for the Care and Use of Animals for Scientific Purposes (2013). Animal ethics approval was obtained from the Animal Ethics Committee, University of Melbourne.&#160;The materials herein are for laboratory experiments only, and not intended for medical or veterinary use.
1. Preparing Electrodes
Note: A three channel transmitter is used for surgical implantation which enables 2 ERG and 1 VEP recording to b...

<ol>
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K., Van Luijtelaar, E. L., Coenen, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cortical+and+thalamic+visual+evoked+potentials+during+sleep-wake+states+and+spike-wave+discharges+in+the+rat.">Cortical and thalamic visual evoked potentials during sleep-wake states and spike-wave discharges in the rat.</a> Electroencephalogr Clin Neurophysiol. 108, 306-319 (1998).</li><li>Nair, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+common+anesthetics+on+eye+movement+and+electroretinogram.">Effects of common anesthetics on eye movement and electroretinogram.</a> Doc Ophthalmol. 122, 163-176 (2011).</li><li>Amouzadeh, H. R., Sangiah, S., Qualls, C. W., Cowell, R. 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Due to the minimally invasive nature of visual electrophysiology, ERG and VEP recordings in human patients are conducted under conscious conditions and only require the use of topical anesthetics for electrode placement. In contrast, visual electrophysiology in animal models is conventionally conducted under general anesthesia to enable stable electrode placement by eliminating voluntary eye and body movements. However, commonly used general anesthetics alter the ERG and VEP responses as shown by our previous publication...

The ERG and VEP are minimally invasive in vivo tools to assess the integrity of retinal and visual pathways respectively in both the laboratory and clinic. The full-field ERG yields a characteristic waveform which can be broken down into different components, with each element representing different cell classes of the retinal pathway1,2. The classic full-field ERG waveform consists of an initial negative slope (a-wave), which has been shown to represent photoreceptor activity post light exposure2-4. The a-wave is followed by a substantial positive waveform (b-wave) which reflects electrical activity of middle retina, predominantly the ON-bipolar cells5-7. Furthermore, one can vary luminous energy and inter-stimulus-interval to isolate cone from rod responses8.
The flash VEP represents electrical potentials of the visual cortex and brain stem in response to retinal light stimulation9,10. This waveform can be broken down into early and late components, with the early component reflecting activity of neurons of the retino-geniculo-striate pathway11-13 and the late component representing cortical processing performed in various V1 laminae in rats11,13. Therefore simultaneous measurement of the ERG and VEP returns comprehensive assessment of the structures involved in the visual pathway.
Currently, in order to record electrophysiology in animals, anesthesia is employed to enable stable placement of electrodes. There have been attempts to measure ERG and VEP in conscious rats14-16 but these studies employed a wired setup, which can be cumbersome and may lead to animal stress by restricting animal movement and natural behavior17. With recent advances in wireless technology including improved miniaturization and battery life, it is now possible to implement a telemetry approach for ERG and VEP recording, decreasing the stress associated with wired recordings and improving long term viability. Fully internalized stable implantations of telemetry probes have proven to be successful for chronic monitoring of temperature, blood pressure18, activity19 as well as electroencephalography20. Such advances in technology will also assist with repeatability and stability of conscious recordings, increasing the platform&#39;s utility for chronic studies.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Bioamplifier</td>
			<td>ADInstruments</td>
			<td>ML 135</td>
			<td>Amplifies ERG and VEP signals</td>
		</tr>
		<tr>
			<td>Carboxymethylcellulose sodium 1.0%</td>
			<td>Allergan</td>
			<td>CAS 0009000-11-7</td>
			<td>Maintain corneal hydration during surgery</td>
		</tr>
		<tr>
			<td>Carprofen 0.5%</td>
			<td>Pfizer Animal Health Group</td>
			<td>CAS 53716-49-7</td>
			<td>Post-surgery analgesia, given with injectable saline for fluid replenishment</td>
		</tr>
		<tr>
			<td>Chlorhexadine 0.5%</td>
			<td>Orion Laboratories</td>
			<td>27411, 80085</td>
			<td>Disinfection of surgical instrument</td>
		</tr>
		<tr>
			<td>Cyanoacrylate gel activator</td>
			<td>RS components</td>
			<td>473-439</td>
			<td>Quickly dries cyanoacrylate gel</td>
		</tr>
		<tr>
			<td>Cyanocrylate gel&#160;</td>
			<td>RS components</td>
			<td>473-423</td>
			<td>Fix stainless screws to skull</td>
		</tr>
		<tr>
			<td>Dental burr</td>
			<td>Storz Instruments, Bausch and Lomb</td>
			<td>E0824A</td>
			<td>Miniature drill head of ~0.7mm diameter for making a small hole in the skull over each hemisphere to implant VEP screws</td>
		</tr>
		<tr>
			<td>Drill</td>
			<td>Bosch</td>
			<td>Dremel 300 series</td>
			<td>Automatic drill for trepanning</td>
		</tr>
		<tr>
			<td>Enrofloxin</td>
			<td>Troy Laboratories</td>
			<td></td>
			<td>Prophylactic antibiotic post surgey</td>
		</tr>
		<tr>
			<td>Ganzfeld integrating sphere</td>
			<td>Photometric Solutions International</td>
			<td></td>
			<td>Custom designed light stimulator: 36 mm diameter, 13 cm aperture size</td>
		</tr>
		<tr>
			<td>Gauze swabs</td>
			<td>Multigate Medical Products Pty Ltd</td>
			<td>57-100B</td>
			<td>Dries surgical incision and exposed skull surface during surgery</td>
		</tr>
		<tr>
			<td>Isoflurane 99.9%</td>
			<td>Abbott Australasia Pty Ltd</td>
			<td>CAS 26675-46-7</td>
			<td>Proprietory Name: Isoflo(TM) Inhalation anaaesthetic. Pharmaceutical-grade inhalation anesthetic mixed with oxygen gas for VEP electrode implant surgery</td>
		</tr>
		<tr>
			<td>Kenacomb ointment</td>
			<td>Aspen Pharma Pty Ltd</td>
			<td></td>
			<td>To reduce skin irritation and itching after surgery</td>
		</tr>
		<tr>
			<td>Luxeon LEDs</td>
			<td>Phillips Lighting Co.</td>
			<td></td>
			<td>For light stimulation, twenty 5 watt and one 1 watt LEDs, controlled by Scope software</td>
		</tr>
		<tr>
			<td>Needle (macrosurgery)</td>
			<td>World Precision Instruments</td>
			<td>501959</td>
			<td>for suturing abdominal and head surgery, used with 3-0 suture, eye needle, cutting edge 5/16 circle Size 1, 15mm</td>
		</tr>
		<tr>
			<td>Needle holder (macrosurgery)</td>
			<td>World Precision Instruments</td>
			<td>500224</td>
			<td>To hold needle during abdominal and head surgery</td>
		</tr>
		<tr>
			<td>Needle holder (microsurgery)</td>
			<td>World Precision Instruments</td>
			<td>555419NT</td>
			<td>To hold needle during ocular surgery</td>
		</tr>
		<tr>
			<td>Optiva catheter</td>
			<td>Smiths Medical International LTD</td>
			<td>16 or 21 G</td>
			<td>Guide corneal active electrodes from skull to conjunctiva</td>
		</tr>
		<tr>
			<td>Povidone iodine 10%</td>
			<td>Sanofi-Aventis</td>
			<td>CAS 25655-41-8</td>
			<td>Proprietory name: Betadine, Antiseptic to prepare the shaved skin for surgery 10%, 500 mL</td>
		</tr>
		<tr>
			<td>Powerlab data acquisition system</td>
			<td>ADInstruments</td>
			<td>ML 785</td>
			<td>Acquire signal from telemetry transmitter, paired to telemetry data converter</td>
		</tr>
		<tr>
			<td>Proxymetacaine 0.5%</td>
			<td>Alcon Laboratories&#160;</td>
			<td>CAS 5875-06-9</td>
			<td>Topical ocular analgesia</td>
		</tr>
		<tr>
			<td>Restrainer</td>
			<td>cutom made</td>
			<td></td>
			<td>Front of the restrainer is tapered to minimize head movement, length can be adjusted to accommodate different rat length, overall diameter is 60 mm.&#160;</td>
		</tr>
		<tr>
			<td>Scapel blade</td>
			<td>R.G. Medical Supplies</td>
			<td>SNSM0206</td>
			<td>For surgical incision</td>
		</tr>
		<tr>
			<td>Scissors (macrosurgery)</td>
			<td>World Precision Instruments</td>
			<td>501225</td>
			<td>for cutting tissue on the abodmen and forhead</td>
		</tr>
		<tr>
			<td>Scissors (microsurgery)</td>
			<td>World Precision Instruments</td>
			<td>501232</td>
			<td>To dissect the conjunctiva for electrode attachment</td>
		</tr>
		<tr>
			<td>Scope Software</td>
			<td>ADInstruments</td>
			<td>version 3.7.6</td>
			<td>Simultaneously triggers the stimulus via the ADI Powerlab system and collects data</td>
		</tr>
		<tr>
			<td>Shaver</td>
			<td>Oster</td>
			<td>Golden A5</td>
			<td>Shave fur from surgical areas</td>
		</tr>
		<tr>
			<td>Stainless streel screws&#160;</td>
			<td>MicroFasteners</td>
			<td>L001.003CS304</td>
			<td>0.7 mm shaft diameter, 3 mm in length&#160;</td>
		</tr>
		<tr>
			<td>Stereotaxic frame</td>
			<td>David Kopf</td>
			<td>Model 900</td>
			<td>A small animal stereotaxic instrument for locating the implantation landmarks on the skull</td>
		</tr>
		<tr>
			<td>Surgical drape</td>
			<td>Vital Medical Supplies</td>
			<td>GM29-612EE</td>
			<td>Ensure sterile enviornment during surgery</td>
		</tr>
		<tr>
			<td>Suture (macrosurgery)</td>
			<td>Ninbo medical needles</td>
			<td>3-0</td>
			<td>for suturing abdominal and head surgery, sterile silk braided, 60cm</td>
		</tr>
		<tr>
			<td>Suture needle (microsurgery)</td>
			<td>Ninbo medical needles</td>
			<td>8-0 or 9-0</td>
			<td>for ocular surgery including, suturing electrode to sclera and closing conjunctival wound, nylon suture, 3/8 circle 1&#215;5, 30cm</td>
		</tr>
		<tr>
			<td>Telemetry data converter&#160;</td>
			<td>DataSciences International</td>
			<td>R08</td>
			<td>allows telemetry signal to interface with data collection software</td>
		</tr>
		<tr>
			<td>Telemetry Data Exchange Matrix</td>
			<td>DataSciences International</td>
			<td></td>
			<td>Gathers data from transmitters, pair with receiver</td>
		</tr>
		<tr>
			<td>Telemetry data receiver</td>
			<td>DataSciences International</td>
			<td>RPC-1</td>
			<td>Receives telemetry data from transmitter</td>
		</tr>
		<tr>
			<td>Telemetry transmitter</td>
			<td>DataSciences International</td>
			<td>F50-EEE</td>
			<td>3 channel telemetry transmitter</td>
		</tr>
		<tr>
			<td>Tropicamide 0.5%</td>
			<td>Alcon Laboratories&#160;</td>
			<td></td>
			<td>Iris dilation</td>
		</tr>
		<tr>
			<td>Tweezers (macrosurgery)</td>
			<td>World Precision Instruments</td>
			<td>500092</td>
			<td>Manipulate tissues during abdominal and head surgery</td>
		</tr>
		<tr>
			<td>Tweezers (microsurgery)</td>
			<td>World Precision Instruments</td>
			<td>500342</td>
			<td>Manipulate tissues during ocular surgery</td>
		</tr>
	</tbody>
</table>

implantation and recording of wireless electroretinogram and visual evoked potential in conscious rats

The full-field electroretinogram (ERG) and visual evoked potential (VEP) are useful tools to assess retinal and visual pathway integrity in both laboratory and clinical settings. Currently, preclinical ERG and VEP measurements are performed with anesthesia to ensure stable electrode placements. However, the very presence of anesthesia has been shown to contaminate normal physiological responses. To overcome these anesthesia confounds, we develop a novel platform to assay ERG and VEP in conscious rats. Electrodes are surgically implanted sub-conjunctivally on the eye to assay the ERG and epidurally over the visual cortex to measure the VEP. A range of amplitude and sensitivity/timing parameters are assayed for both the ERG and VEP at increasing luminous energies. The ERG and VEP signals are shown to be stable and repeatable for at least 4 weeks post surgical implantation. This ability to record ERG and VEP signals without anesthesia confounds in the preclinical setting should provide superior translation to clinical data.

The photoreceptor response is analyzed by fitting a delayed Gaussian to the leading edge of the initial descending limb of the ERG response at the top 2 luminous energies (1.20, 1.52 log c.s.m-2) for each animal, based on the model of Lamb and Pugh22, formulated by Hood and Birch23. This formula returns an amplitude and a sensitivity parameter, (Figure 1C and 1D, respectively). A hyperbolic function was fitted to the lumin...

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Implantation and Recording of Wireless Electroretinogram and Visual Evoked Potential in Conscious Rats

We show surgical implantation and Recording procedures to measure visual electrophysiological signals from the eye (electroretinogram) and brain (visual evoked potential) in conscious rats, which is more analogous to the human condition where recordings are conducted without anesthesia confounds.

The full-field electroretinogram (ERG) and visual evoked potential (VEP) are useful tools to assess retinal and visual pathway integrity in both ...

Research

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Behavior

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