Eye Preparation

Dark Adaptation, Dilation and Eye Hydration

To prepare an animal for ERG or VEP testing, consider the required dark/light adaption state of the animal. Most rodent protocols require a dark-adapted setting. Dilation is another critical factor as it maximizes the amount of light that stimulates the retina. The eyes must also remain hydrated over the course of testing to ensure good conductivity throughout. Anesthesia and Setting up a Dark Room are each discussed in separate articles. 

Dark-Adaptation

Most rodent ERG testing is done on dark-adapted animals. For best results, keep the animal enclosed in a completely dark space overnight, or for an absolute minimum of 3-hours. Be sure to review your institution’s policy regarding overnight housing. 

Red lights may be used during dark-adapted testing. It is standard practice to use red lamps, headlights, or red-light caps when setting up a dark-adapted animal for testing. Be sure to also cover the computer monitor with a red filter. Red filters and red-light caps may be purchased from Diagnosys Accessories. For more information about creating a dark-adapted environment, refer to this Testing Environment article.

 

Dilation 

Dilate the animal’s pupils maximally using a combination of Phenylephrine HCl and either Tropicamide or Atropine. Allow the drops to remain on the eyes for approximately 2 minutes before wicking away excess liquid. 

Warning: Too many eye drops could flow into the animal’s nose and lungs which could inadvertently drown the animal! Be particularly careful when applying eye drops to baby mice whose eyes are very close to their noses. 

For testing that requires longer recordings, such as C-wave or DC-ERG responses, corneal anesthetic such as proparacaine is recommended.

Hydration

Hydrating drops must be used to prevent corneal drying and subsequent cataracts. A .3% Hypromellose lubricating gel solution is best. Saline or artificial tears are too thin to stay on the eye and will need frequent reapplication to keep the eye hydrated. While 2.5% (gonioscopic) hypromellose is too thick to sufficiently hydrate the eye and should not be used. Completely avoid using hydration drops with Mineral Oil as a key ingredient. Mineral oil will insulate the eye, where ERG requires a solution that promotes conductivity. 

There are several brands of .3% Hypromellose gel: Systane (US), Genteal (US), Artelac (UK), Evolve (Australia), and several others.

When doing longer-form testing, such as C-wave protocols, the eye may require extra hydration. Adding artificial tear drops onto the eye-electrode connection can help maintain eye hydration for the duration of testing. Drops such as Refresh Tears and other drops that contain active ingredients such as Carboxymethylcellulose Sodium (0.5%) can work well for this purpose. 

Procedural Bibliogrpahy

Kinoshita J, Peachey NS. Noninvasive Electroretinographic Procedures for the Study of the Mouse Retina. Curr Protoc Mouse Biol. 2018 March; 8(1): 1–16. doi:10.1002/cpmo.39

Benchorin G, Calton MA, Beaulieu MO, Vollrath D. Assessment of Murine Retinal Function by Electroretinography. Bio Protoc. 2017 April 5; 7(7): . doi:10.21769/BioProtoc.2218.

Weymouth, A.E., Vingrys, A.J., Rodent electroretinography: Methods for extraction and interpretation of rod and cone responses. Progress Retinal Eye Res (2007), doi:10.1016/j.preteyeres.2007.09.003