Molecular Mechanisms of Photoreceptor Function.
Images focused on the retina of the vertebrate eye are sensed by rod and cone photoreceptors. Our lab studies phototransduction, the mechanism by which light is detected and converted into a chemical signal at the synaptic terminus. Increases in illumination hyperpolarize rods and cones, darkness depolarizes them, and constant illumination leads to adaptation. Photoreceptors are loaded with mitochondria; they have exceptionally high metabolic rates and they stay poised to respond within milliseconds to small changes in illumination. The downside of this metabolic balance seems to be that it makes photoreceptors vulnerable to mutations that affect their metabolism. Mutations in signal transduction enzymes disrupt the metabolic balance of photoreceptors and often cause blindness.
In rods and cones light is absorbed by visual pigments. The photoactivated pigment activates a G protein, transducin, which then stimulates a phosphodiesterase to hydrolyze intracellular cyclic GMP. This closes cation channels, hyperpolarizes the photoreceptor, and slows glutamate release at the synaptic terminal. In this way, the cell detects light and passes on that information to the inner retina and ultimately to the brain.
Each step in this phototransduction pathway is associated with biochemistry that inactivates it. We’re studying several of these biochemical reactions both in vitro and in vivo. For example, we’re investigating how rhodopsin kinase inactivates photoactivated rhodopsin. Light stimulates rapid loss of Ca2+ from the cytoplasm of rod and cone outer segments and this change in intracellular Ca2+ is critical for adaptation. We’re studying how Ca2+ plays a critical role in recovery and adaptation by regulating rhodopsin phospohorylation and synthesis of cGMP.
The tools we use for these investigations are diverse. We use mass spectrometry to analyze phosphorylation that takes place in vivo; we use electroretinography to monitor electrical responses of photoreceptors within the eye and we are setting up to use vision-dependent behaviors to analyze how well the retina conveys sensory information to the brain. Biochemical assays and molecular biological manipulations are widely used in the laboratory. We also use random mutagenesis of zebrafish to screen for novel genes essential for vision and we are using “reverse-genetics” with mice and zebrafish to investigate the functions of known genes.
One of our major interests is to investigate cone phototransduction. Rod photoreceptors have been studied extensively but less is known about how cones function. Cones respond with faster kinetics and they adapt to a much broader range of background illumination than rods. Cones express counterparts of many of the well-studied phototransduction enzymes found in rods. To understand the basis for enhanced versatility of cones, Dr. Susan Brockerhoff has initiated the use of zebrafish as a model organism for investigating cone function. Zebrafish are ideally suited for this because they are amenable to a combination of genetic, biochemical, and physiological strategies.
In general, we subscribe to the philosophy that a productive analysis of an important biological process, such as vision, requires integration of knowledge from multiple scientific disciplines. An important goal of our research is to learn how to effectively integrate ideas and findings from multiple biological and chemical perspectives.