Transretinal ERG Recordings from Mouse Retina: Rod and Cone Photoresponses

Summary

We describe a relatively simple method of transretinal electroretinogram (ERG) recordings for obtaining rod and cone photoresponses from intact mouse retina. This approach takes advantage of the block of synaptic transmission from photoreceptors to isolate their light responses and record them using field electrodes placed across the isolated flat-mounted retina.

Abstract

There are two distinct classes of image-forming photoreceptors in the vertebrate retina: rods and cones. Rods are able to detect single photons of light whereas cones operate continuously under rapidly changing bright light conditions. Absorption of light by rod- and cone-specific visual pigments in the outer segments of photoreceptors triggers a phototransduction cascade that eventually leads to closure of cyclic nucleotide-gated channels on the plasma membrane and cell hyperpolarization. This light-induced change in membrane current and potential can be registered as a photoresponse, by either classical suction electrode recording technique^1,2 or by transretinal electroretinogram recordings (ERG) from isolated retinas with pharmacologically blocked postsynaptic response components^3-5. The latter method allows drug-accessible long-lasting recordings from mouse photoreceptors and is particularly useful for obtaining stable photoresponses from the scarce and fragile mouse cones. In the case of cones, such experiments can be performed both in dark-adapted conditions and following intense illumination that bleaches essentially all visual pigment, to monitor the process of cone photosensitivity recovery during dark adaptation^6,7. In this video, we will show how to perform rod- and M/L-cone-driven transretinal recordings from dark-adapted mouse retina. Rod recordings will be carried out using retina of wild type (C57Bl/6) mice. For simplicity, cone recordings will be obtained from genetically modified rod transducin α-subunit knockout (Tα^-/-) mice which lack rod signaling⁸.

Protocol

1. Making Electrodes

1. Prepare glass electrodes. Weigh 120 mg agar and mix it in 10 mL distilled water (final agar concentration 1.2%). Melt the agar solution in hot water bath. Fill glass capillaries (we use Word Presision Instruments TW100-4 capillaries with the following dimensions: length = 100 mm, OD/ID = 1/0.75 mm, and internal volume = 44 μL) with the agar solution using plastic syringe. Solidify the agar at room temperature for 10 min. Thus, up to ~200 capillaries can be filled (the rest of the agar mixture can be stored at -4 °C and re-used several times).
2. Cut the capillaries into halves using a diamond knife.
3. Soak the resulting ....

Discussion

The method of rod- and cone-driven transretinal ERG recordings described above is becoming a powerful tool for investigating the function of mouse photoreceptors in both wild type and genetically modified animals. In addition to the easy characterization of basic photoresponse properties, this simple technique provides great response stability during long-lasting experiments performed on close-to-intact retina preparations. Both dark-adapted rod maximal response amplitude and photosensitivity in wild type mice are stable.......