Using the Electroretinogram to Assess Function in the Rodent Retina and the Protective Effects of Remote Limb Ischemic Preconditioning

Summary

The electroretinogram (ERG) is an electrical potential generated by the retina in response to light. This paper describes how to use the ERG to assess retinal function, in dark-adapted rats, and how it can be can be used to assess a neuroprotective intervention, in the present case remote ischemic preconditioning.

Abstract

The ERG is the sum of all retinal activity. The ERG is usually recorded from the cornea, which acts as an antenna that collects and sums signals from the retina. The ERG is a sensitive measure of changes in retinal function that are pan-retinal, but is less effective for detecting damage confined to a small area of retina. In the present work we describe how to record the ‘flash’ ERG, which is the potential generated when the retina is exposed to a brief light flash. We describe methods of anaesthesia, mydriasis and corneal management during recording; how to keep the retina dark adapted; electrode materials and placement; the range and calibration of stimulus energy; recording parameters and the extraction of data. We also describe a method of inducing ischemia in one limb, and how to use the ERG to assess the effects of this remote-from-the-retina ischemia on retinal function after light damage. A two-flash protocol is described which allows isolation of the cone-driven component of the dark-adapted ERG, and thereby the separation of the rod and cone components. Because it can be recorded with techniques that are minimally invasive, the ERG has been widely used in studies of the physiology, pharmacology and toxicology of the retina. We describe one example of this usefulness, in which the ERG is used to assess the function of the light-damaged retina, with and without a neuroprotective intervention; preconditioning by remote ischemia.

Introduction

The ERG is an electrical potential generated by the retina in response to light, and recorded from the corneal surface of the eye. When conditions of recording are managed carefully, the ERG can be used in a variety of ways to assess retinal function. Here we described how to record the ‘flash ERG’, the potential generated when the retina is exposed to a brief, bright flash presented in a Ganzfeld background. The Ganzfeld disperses the light homogenously and the flash of light reaches the whole retina approximately uniformly. If the retina is dark adapted before recording, and the dark-adaptation is maintained as the animal is prepared for recording, the E....

Protocol

This protocol follows the animal care guidelines of University of Sydney.

1. Making Electrodes

1. Construct the positive electrode (the one which will contact the cornea) from a short (5 cm) length of platinum wire 1-2 mm in diameter. Fashion it into a loop a few mm in diameter. Connect this loop to a conventional lead, long enough to reach the input stage of your amplifier (see Figure 4).
2. Construct the negative electrode (which will go in the animal’s mouth) using an Ag/AgCl pellet 1-2 mm in diameter, also connected to a convention lead (see Figure 4).
3. As a reference elec....

Results

The protocol can be used to measure visual function of rodent retina in vivo. The a-wave, a measure of photoreceptor function, and the b-wave, a measure of inner retina function, are annotated in Figure 1.

The rod-dominated ERG signal increases with the increasing light stimulus, as shown in Figure 2A. The a-wave becomes apparent at ~0.4 log scot cd.s.m^-2 and the amplitude of the a-wave increases until saturation at 2.5 log scot cd.s.m^.......

Discussion

The dark-adapted flash ERG method described above is a reliable method for assessing retinal function in rats. Both the a-wave and b-wave were reduced by light damage. Remote ischemic preconditioning mitigated light damage-induced reductions in the a-wave and b-wave. This preservation of retinal function suggests that remote ischemic preconditioning has induced neuroprotection, resembling other forms of protective preconditioning such as hypoxia, ischemia and exercise^8-10. The ERG signal recorded is determined.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
PC computer
Powerlab, 4 channel acquistion hardware	AD Instruments	PL 35044	Acquistion of ERG
Animal Bio Amp	AD Instruments	FE 136	Amplifier for ERG
Lab chart	AD Instruments		Signal collection software
Ganzfield	Photometric solutions	FS-250A	Light stimulus
Ganzfield operating system	Photometric solutions
Research Radiometer	International light technologies	ILT-1700	calibrate light series
Lux meter		LX-1010B	check red light illumanation
Excel	Microsoft
Lead wires	AD Instruments		Connect postive, negative ground electrodes to amplifier
Lead wires - alligator	AD Instruments		ground ganzfield and acquistion hardware to computer
Platinum wire 95%	A&E metals		postive electrode
Mouth electrode Ag/AgCl Pellet	SDR	E205	negative electode
26 G needle	BD		ground electode
Water pump
Water bath
Tubing
Homeothermic blanket system with flexible probe	Harvard Appartus	507222F
Atropine 1% w/v	Bausch & Lomb		topical mydriasis
Proxmethycaine 0.5% w/v	Bausch & Lomb		topical anaesthetic
Visco tears eye drops	Novartis		carbomer polymer
Thread			retract eye lid
Tweezers
Reusable adhesive	Blu tac		Dim red headlamp. Affix electrodes
Absorbent bedding
Ketamil - ketamine 100 mg/ml - 50 ml	Troy Laboratories Pty Ltd		dissociative
Xylium - Xylazine 100 mg/ml - 50 ml	Troy Laboratories Pty Ltd		muscle relaxant
Scale