Physiological Features of the S- and M-cone Photoreceptors of Wild-type Mice from Single-cell Recordings

Abstract

Cone cells constitute only 3% of the photoreceptors of the wild-type (WT) mouse. While mouse rods have been thoroughly investigated with suction pipette recordings of their outer segment membrane currents, to date no recordings from WT cones have been published, likely because of the rarity of cones and the fragility of their outer segments. Recently, we characterized the photoreceptors of Nrl −/− mice, using suction pipette recordings from their “inner segments” (perinuclear region), and found them to be cones. Here we report the use of this same method to record for the first time the responses of single cones of WT mice, and of mice lacking the α-subunit of the G-protein transducin (G tα−/−), a loss that renders them functionally rodless. Most cones were found to functionally co-express both S- (λmax = 360 nm) and M- (λmax = 508 nm) cone opsins and to be maximally sensitive at 360 nm (“S-cones”); nonetheless, all cones from the dorsal retina were found to be maximally sensitive at 508 nm (“M-cones”). The dim-flash response kinetics and absolute sensitivity of S- and M-cones were very similar and not dependent on which of the coexpressed cone opsins drove transduction; the time to peak of the dim-flash response was ∼70 ms, and ∼0.2% of the circulating current was suppressed per photoisomerization. Amplification in WT cones (A ∼4 s−2) was found to be about twofold lower than in rods (A ∼8 s−2). Mouse M-cones maintained their circulating current at very nearly the dark adapted level even when >90% of their M-opsin was bleached. S-cones were less tolerant to bleached S-opsin than M-cones to bleached M-opsin, but still far more tolerant than mouse rods to bleached rhodopsin, which exhibit persistent suppression of nearly 50% of their circulating current following a 20% bleach. Thus, the three types of mouse opsin appear distinctive in the degree to which their bleached, unregenerated opsins generate “dark light.”

Postreceptoral contributions to the light-adapted ERG of mice lacking b-waves

“…3). M-cone photoreceptor currents, recorded from mouse outer segments are observed to peak around 70 ms after a weak flash and to recover to baseline in around 200 ms (Nikonov et al, 2006). Times to peak for stronger flashes such as we used would be earlier.…”

“…This duration is very prolonged compared to results from suction electrode recordings from mouse M-cones. In suction electrode recordings, outer segment currents were reduced only for about 200 ms for weak stimuli, with a peak reduction around 70 ms after a brief flash (Nikonov et al, 2006). In the present study, in order to isolate the cone photoreceptor contribution to the ERG, the paired flash approach (Pepperberg et al, 1997) was used to derive the photocurrent.…”

Regulation of photoreceptor gene expression by Crx-associated transcription factor network

“…Cone function in Nrl knockout mice is preserved as late as 31 weeks ). Single-cell electrophysiology showed responses driven by both S-and M-opsin in all cells tested from Nrl knockout mice , as seen for the wild-type cones that co-express both opsins [ (Applebury et al, 2000;Nikonov et al, 2006); see cone subtypes and gradients section below]. Although the dorsal/ventral M/S-cone gradient is preserved in Nrl −/− mice, more S-opsin sensitivity is seen in Nrl −/− cones than in wild-type cones from comparable dorsal/ventral levels of the retina (Nikonov et al, 2006).…”

“…Single-cell electrophysiology showed responses driven by both S-and M-opsin in all cells tested from Nrl knockout mice , as seen for the wild-type cones that co-express both opsins [ (Applebury et al, 2000;Nikonov et al, 2006); see cone subtypes and gradients section below]. Although the dorsal/ventral M/S-cone gradient is preserved in Nrl −/− mice, more S-opsin sensitivity is seen in Nrl −/− cones than in wild-type cones from comparable dorsal/ventral levels of the retina (Nikonov et al, 2006). Consistent with the above electrophysiological measures, the ultrastructural analysis showed that the Nrl −/− retina has much shorter cone-like outer segments with disrupted morphology and cone-like nuclei with decondensed chromatin Daniele et al, 2005).…”

Chromatic Bipolar Cell Pathways in the Mouse Retina

“…For these two wavelengths, the intensities were equivalent to photo-isomerization rates (in s Ϫ1 per cone) of 2730 -14,800 for blue in S-cones and 1670 -9000 for green in M-cones (assuming exclusive M-opsin expression). This was calculated using the relative sensitivities of the mouse cone opsins at the stimulus wavelengths via an opsin-template (Stockman and Sharpe, 2000) fitted to the peak sensitivities (360 and 511 nm) (Jacobs et al, 1991) and assuming the light collection area of a cone a c ϭ 0.2 m 2 (Nikonov et al, 2006). We lightadapted the slices during the experiment to the low photopic range by presenting a background intensity of Ϸ10 5 photons ⅐ s Ϫ1 ⅐ m…”