Makino Research Content
We are investigating the earliest events of vision wherein light is converted into an electrical signal by the retinal photoreceptors.
What sets the speed of the photoresponse?
Rods contain an enormous number of rhodopsin molecules densely packed in membranous disks, in order to achieve high sensitivity. But dense packing could impair phototransduction by restricting the lateral mobility of rhodopsin as well as that of other proteins in the membrane. To test the hypothesis that the speed of the photoresponse is limited by the rates with which molecules collide with each other on the membrane, we recorded photoresponses of single rods containing fewer rhodopsins in their membranes. This condition was achieved by hemizygous knockout of the rod opsin gene in transgenic mice (R+/-). Photoresponses were accelerated by the reduction in rhodopsin packing density, supporting the hypothesis (Fig. 1). The key step in the activation of the photoresponse was identified as the collision between rhodopsin and transducin. Response recovery appears to be limited by the collision rate between rhodopsin and rhodopsin kinase and/or that between between transducin and RGS9. These results predict that under scotopic conditions, humans heterozygous for a null mutation in rhodopsin will have higher than normal temporal resolution.
Figure 1. Top panels: membrane surfaces of wild type and transgenic mouse rod membranes. Rhodopsins and transducins are depicted by red and blue circles, respectively. The green area shows the surface area “sampled” by a photoexcited rhodopsin (green star) every few milliseconds as it diffuses. The area expands and the number of transducins contacted increases as the packing density of rhodopsins decreases. Bottom panels: flash responses of wild type (left) and R+/- (right) rods recorded with suction electrode. Responses of R+/- rods rise and recover more rapidly than those of the wild type rods.
To find out how much the speed of the photoresponse slows with more rhodopsin in the membranes, recordings were made from mutant mice induced to overexpress rhodopsin. Surprisingly, the rods expanded their diameter to accommodate the extra rhodopsins without increasing membrane crowding. The rising phase of the photoresponse was still slowed, however, due to the increased intracellular volume and the dispersal of transducins over a greater membrane surface area.
What determines the spectral sensitivity of a photoreceptor?
The spectral sensitivity of a photoreceptor is largely determined by the visual pigment that it contains. Although photoreceptors were generally thought to express only one type of pigment, there are a number of examples where a cone has been shown to contain two different types of pigments. However, we discovered that the UV-sensitive cone in tiger salamander contains three different types of pigments, all of which are functional (Fig. 2). In addition to a UV-absorbing pigment, the pigments of blue- and red-sensitive cones are also present. Selectivity of pigment expression is not completely lacking in the UV-sensitive cone because the green-sensitive rod pigment is absent. Interestingly, the blue-sensitive cone pigment is also expressed in a blue-sensitive rod (Fig. 2), the first example of a rod and a cone utilizing the same visual pigment.
Figure 2. Averaged spectral sensitivities of salamander UV-sensitive cones (violet circles), blue-sensitive rods (blue diamonds) and cones (blue circles) and red-sensitive cones (red circles). Continuous lines show fits of the spectra with a template. The fit to the UV-sensitive cone spectrum was made assuming that the long wavelength limb included components that were the same as those giving rise to the spectra of the other two cone types.
