Comparison of Functional Properties of Single M– and S–Cones from Wt And Gt–/– Mice

Abstract

Purpose: : To compare functional properties of single S(UV)– and M– cones in WT and Gα_t^–/–mice.

Methods: : Cones of WT and Gα_t^–/– (transducin alpha knockout) mice were studied with suction pipette recordings. Cells were maintained at 35–37 C and stimulated with steps and flashes of light of varied intensity and wavelength. Rod activity was suppressed with 501 nm light.

Results: : Most mouse cones co–express S– and M–opsins, with M–opsin expression following a dorsal–ventral gradient. Responses of cones from the most ventral region (n=8, WT) were S–dominant, from the dorsal region M–dominant (n=5, WT; n=4, Gα_t^–/–). WT M–dominant cones (n=8) were from 4 to 9 times more sensitive to 501 nm than to 361 nm light, indicating low levels (if any) of S–opsin co–expression. The kinetics of S– and M–opsin driven responses of individual cones were nearly identical for all cones. WT M–dominant cones appear to be slightly faster and less sensitive than S–dominant cones, with response time to peak (t_peak), dominant recovery time constant (τ _D), and half–saturating flash intensity (Q_1/2) as follows: t_peak = 63 ± 6 ms, τ _D = 69 ± 22 ms, Q_1/2=7,900 ± 1,700 photons µm^–2 (M–dominant); t_peak = 73 ± 5 ms, τ _D = 73 ± 10 ms, Q_1/2=5,400 ± 1,500 photons µm^–2 (S–dominant). The responses of M– (n=6) and S–dominant (n=8) WT cones to steps of light at lower light levels followed a hyperbolic response vs. intensity relation with I_1/2 ∼ 100,000 – 200,000 photons s^–1 µm^–2. However, increasing the background intensity up to 3,500,000 photons s^–1 µm^–2 did not cause complete suppression of the circulating current in 5 out of 6 M–dominant cones. Calculations suggest this escape from saturation is caused by reduction in light capture due to bleaching of the M–pigment. Even after bleaching up to 90% of the M–pigment, in absence of light the amplitude of the circulating current remained essentially unchanged from its pre–bleach level. S– and M–cones of Gα_t^–/– mice were slower and more sensitive than WT cones, with t_peak and τ _D ∼ 100 ms, Q_1/2 ∼ 4,000 photons µm^–2 and I_1/2 ∼ 30,000 photons s^–1 µm^–2.

Conclusions: : Pigment bleaching appears to be one of the mechanisms allowing cones to escape saturation by a background light. Very large amounts of bleached pigment per se produce little (if any) suppression of the circulating current of M–dominant mouse cones. The difference in kinetics and sensitivity between WT and Gα_t^–/– cones may be explained by adaptation to rod suppressing background; however, the difference in step sensitivity of WT and Gα_t^–/– cones suggests that functional rods may be required for fully normal cone light adaptation behavior.

Keywords: photoreceptors • electrophysiology: non-clinical • opsins