Project Summary Vision forms a prominent part of our daily lives, and this sensory process begins with the capture of light by a particular class of photoreceptor called cones, a class well adapted to daylight conditions, to high acuity vision, and to color perception. The basic response properties of cone photoreceptors have been largely described in studies conducted in isolated cells or retinal tissue. Surprisingly, many of these response properties, such as temporal dynamics and intensity sensitivity range, remain controversial in the literature. Moreover, some of them conflict with data acquired in vivo and with human visual performance when measured psychophysically. The primary aims of this grant are to generate a more comprehensive characterization of cone photoreceptors in cone-dominated animals, including primates. This will be achieved by conjoint experiments conducted in intact retinal explants in parallel with in vivo recordings where it is now possible to target single cones with adaptive optics microstimulation. Similar stimulation paradigms will be employed in each experimental approach to the measure intensity response functions, the rapidity of time-to-peak, the decay kinetics following saturating stimuli, and the adaptation rate in response to changing background light levels. These data will be used to develop a complete theoretical model of response properties based on cone phototransduction which will be of great use to vision scientists and may delineate the limits and boundaries of in vivo vs. in vitro experiments on the retina. An additional aim will be to detail how a novel form of cone activation—via pulsed infrared 2-photon excitation—may be used for improved spatial stimulation of cones. The 2-photon experiments are best done interleaved with traditional 1- photon stimulation in order to examine how the 2-photon excitation mechanism differs from 1- photon absorption, and also to determine how much endogenous fluorescence may be generated by such stimuli. As 2-photon absorption may be used to measure the health of the cone visual cycle, understanding the basic response properties of infrared stimulation will be foundational for future studies of cone function and disease.