The Limit of Photoreceptor Sensitivity: Molecular Mechanisms of Dark Noise in Retinal Cones

Detection threshold in cone photoreceptors requires the simultaneous absorption of several photons because single photon photocurrent is small in amplitude and does not exceed intrinsic fluctuations in the outer segment dark current (dark noise). To understand the mechanisms that limit light sensitivity, we characterized the molecular origin of dark noise in intact, isolated bass single cones. Dark noise is caused by continuous fluctuations in the cytoplasmic concentrations of both cGMP and Ca 2 that arise from the activity in darkness of both guanylate cyclase (GC), the enzyme that synthesizes cGMP, and phosphodiesterase (PDE), the enzyme that hydrolyzes it. In cones loaded with high concentration Ca 2 buffering agents, we demonstrate that variation in cGMP levels arise from fluctuations in the mean PDE enzymatic activity. The rates of PDE activation and inactivation determine the quantitative characteristics of the dark noise power density spectrum. We developed a mathematical model based on the dynamics of PDE activity that accurately predicts this power spectrum. Analysis of the experimental data with the theoretical model allows us to determine the rates of PDE activation and deactivation in the intact photoreceptor. In fish cones, the mean lifetime of active PDE at room temperature is 55 ms. In nonmammalian rods, in contrast, active PDE lifetime is 555 ms. This remarkable difference helps explain why cones are noisier than rods and why cone photocurrents are smaller in peak amplitude and faster in time course than those in rods. Both these features make cones less light sensitive than rods. key words: phototransduction • phosphodiesterase • guanylate cyclase • cGMP • calcium I N T R O D U C T I O N Rod photoreceptors reliably signal single absorbed photons (Baylor et al., 1979), but detection threshold is higher in cones. The limit of light detection varies among cones in different species, and even between cone subtypes in the same species (Miller and Korenbrot, 1993). Still, in complete darkness, even the most sensitive cones require the coincident absorption of several photons (some four to seven) to generate a detectable signal (Schnapf et al., 1990; Miller and Korenbrot, 1993; Donner et al., 1998). To be detected, light-dependent currents must exceed in amplitude intrinsic fluctuations of the photoreceptor dark membrane current referred to as “dark noise. ” For example, in single cones of striped bass, the extrapolated single photon peak current amplitude is 0.14 pA (this report) and it is 0.03 pA in macaque cones (Schnapf et al., 1990). These signals are undetectable because they do not exceed the dark current noise variance, 0.28 pA 2 in bass and 0.12 pA 2 in macaque (0–20 Hz range) (Schnapf et al., 1990). In rods, in contrast, single photon peak photocurrent amplitude is 1 pA in toads (Baylor et al., 1979) and 0.7 pA in macaque (Baylor et al., 1984). These signals are readily detected above dark current noise of variance 0.035 pA 2 in toads (Baylor et al., 1980) and 0.030 pA 2 in macaque (0–20 Hz range) (Baylor et al., 1984). To elucidate the molecular mechanisms underlying dark noise in cones and, hence, the limit of their sensitivity, we investigated the properties and molecular mechanisms of membrane current noise in dark-adapted, green-sensitive single cones of the striped bass. Cone dark noise has been previously characterized both as fluctuation in membrane voltage (Lamb and Simon, 1977; Schneeweis and Schnapf, 1999) and in membrane current (Schnapf et al., 1990; Rieke and Baylor, 2000; Sampath and Baylor, 2002). These studies have shown that dark noise consists of light-sensitive and -insensitive components. The light-sensitive component originates from two principal molecular sources: (1) occasional, spontaneous thermal activation of visual pigment (VP) molecules and (2) continuous fluctuations in the cytoplasmic concentration of cGMP and Ca 2 , as well as in the activity of cGMP-gated (CNG) ion channels. The two components are readily distinguished from each other because of their distinct kinetics: noise that originates from VP excitation has a time course identical Correspondence to Juan I. Korenbrot: juan@itsa.ucsf. edu D. Holcman’s permanent address is Department of Mathematics and Computer Science, Weizmann Institute of Science, Rehovot 76100, Israel. Abbreviations used in this paper: GC, guanylate cyclase; CNG, cGMPgated; PDE, phosphodiesterase; VP, visual pigment. on April 30, 2019 jgp.rupress. org Downloaded from http://doi.org/10.1085/jgp.200509277 Published Online: 31 May, 2005 | Supp Info: