Vision Assessment

While there are many genetically engineered mice produced specifically to study human ophthalmology, an important piece of knowledge to keep in mind is that with any form of systemic disease, pathology of the eye can follow along as a part of the spectrum. In addition to this, there are many background strains that have retinal degeneration and cataracts, among other ocular diseases, as part of the strain-specific lesions. Researchers should be aware of these complications and have measures in place to counteract or account for the effects of the statistical impediments.

To perform a general visual ability assessment, the visual cliff response (VCR) test is a simple and absolute test that can be conducted on many mice in a short period of time. The VCR is performed in a box with a horizontal surface and a vertical wall with a drop-off, extended with a piece of clear Plexiglass to give the appearance of a vertical cliff face. To emphasize the visual appearance of the box, black-and-white checkered paper can be layered on the horizontal and vertical drop-off surfaces. The mouse in question is placed on a platform at the border between the horizontal surface and the apparent drop-off, giving them a choice of stepping down on either surface. Mice with normal visual acuity will step down mostly on the horizontal surface, avoiding what they see as a vertical drop into space. Blind mice will step down equally on both sides since they cannot see the fake cliff.22

Another simple test of visual acuity is based on the tendency of mice to enter dark places, when placed into an area of high light. Given the choice, mice with normal light/dark perception will enter a darkened chamber placed in front of them, while blind mice will not. Fine tuning this assessment by timing the duration between placement in the light area and entering the dark chamber can help to generalize the amount of light/dark perception available in individual mice.22

A deeper examination of the ophthalmology of transgenic mice involves an examination of the eye. These exams are noninvasive, and the animals can be restrained by hand and without anesthetic, as no pain or major stress is involved. Response to light is evaluated by passing a small beam of light over the eye, causing pupil constriction when exposed to the light source, and pupil dilation as the light moves away. Following a general exam of the external eye structures and surrounding area, the left and right eye pupils are dilated using 1% tropicamide that produces full dilation in the mouse within 5 to 10 min. Direct examination of the left and right eyes is through the use of a Kowa SL-14 portable slit lamp. Using ophthalmoscopy, most of the major structures in the eye can be visualized, including the cornea, lens, retina, and optic nerve. Any deviation from normal structure may be a phenotype, though care must be taken not to confuse background strain affects on eye pathology with genetically engineered changes.

For more specific testing of ophthalmic function, greater effort and sophisticated equipment is required. Two of the more commonly tested areas of visual function are intraocular pressure to detect glaucoma and retinal function to test for visual acuity. Intraocular hydrostatic pressure can be measured using an electrophysiologic approach — the servo-null micropipette system (SNMS). For this procedure, all mice should be under general anesthesia supplemented with proparacaine or other local anesthetic topically applied to the eye. SNMS consists of a micropipette filled with 3M KCl solution to counteract fluid resistance of the extracellular fluid and carboxyfluorescein to enhance visualization, a ground reference placed on the conjunctiva, and a servo-null device. The micropipette tip is placed in the drop of topical anesthetic, overlaying the pupil at an angle of 60 to 70° relative to a tangent to the corneal surface, and then rapidly inserted into the anterior chamber through the cornea. This changes the hydrostatic pressure and forces aqueous humor into the pipette to displace the KCl solution. The resultant increase in electrical resistance is measured via a signal through a vacuum-pressure pump and is the value of the intraocular pressure (IOP). The IOP can be monitored at a rate of 3 to 5 measurements per second.32 Testing for retinal function can be performed through a procedure called electroretinography. The instrument consists of a flashlamp with focusing and filtering optics and amplifiers so that light can be delivered to the eye in single pulses of 20 |isec duration. The electroretinogram (ERG) waveforms are recorded in triplicate and averaged, reflecting the amount of activity within the retina in response to the pulsing light beams. Mice undergoing ERG should be under general anesthesia and have topical anesthesia for the eye as well as a pupil dilator to allow for better light penetration. In preparation for a trial, mice should be dark-adapted overnight and placed under anesthesia in dim red light.34

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