Background: Circadian systems influence every aspect of an organis's biology, fine-tuning biochemistry, physiology and behaviour to the varying demands of the daily cycle. In our own species, circadian timing has a profound impact in medicine and in agriculture, with shift work, jet-lag and the timed application of drugs being obvious examples of the interaction of biological and environmental time. While recognition of the importance of circadian timing in biology has recently increased, our comprehension of the basic mechanisms involved has lagged. Approaches to understanding circadian organization have concentrated on the mechanisms that generate the circadian rhythm, the environmental inputs regulating circadian phase (entrainment), and the outputs from the biological clock.
In this proposal we will restrict our studies to the input pathway, and investigate the organization and mechanisms of the photic system mediating circadian entrainment in mammals. Several studies have shown that the neural and molecular substrates regulating circadian responses to light are distinct from the visual channels involved in image formation and visual perception, and in this respect many assumptions about the nature of the entrainment pathway have proved to be incorrect. For example, rod photoreceptors were assumed to mediate photic entrainment, however retinally degenerate (rd) mice and transgenic mice which lack rod photoreceptors show unattenuated circadian responses to light. While it is well known that light regulates melatonin secretion by the pineal in humans, recent evidence has shown that secretion of this hormone can also be modulated by photic stimulation in certain subpopulations of blind humans.
We aim to build upon these observations and define the genetic, developmental and functional properties of the entrainment pathway of mammals. Aims and Methods. In order to dissect out the circadian light pathway, our primary strategy will rely on mammals with selective genetic "lesions" within the retina.
To achieve this goal, we will employ natural populations of blind mammals and different strains of mice with hereditary retinal disorders. In addition, we will construct transgenic mice, which lack specific photoreceptors cells or opsins (null mutations). Another key aim of the study will focus on the organization and responses of the light entrainment pathway in primate species. Identifying the circadian photoreceptors and characterizing their connections with the central pacemaker in these models will involve the use of a broad range of experimental approaches which can only be achieved through a large scale coordinated effort involving the complementary expertise of several specialized laboratories. Our research strategies will involve recombinant DNA techniques to isolate, sequence and characterize putative "circadian" opsins. The elements of the entrainment pathway, from the photorecptors to the circadian oscillator (suprachiasmatic nuclei, SCN) will be identified by employing in situ hybridization techniques, immunohistochemistry, ultrastructural analysis, gene expression, and viral and neuroanatomical tract tracing techniques. The functional characteristics of candidate photoreceptors and of entrainment pathway will be assessed using electrophysiological and microspectrophotometric techniques, and by measuring light entrainment of circadian activity, temperature, and sleep-wake cycles. A quantitative and qualitative analysis of the circadian responses to light in blind and normally sighted human subjects will also be undertaken in order to specify the responsecharacteristics of the human photic system.
Potential: Enhancing our knowledge of the mechanisms underlying the transduction of photic information by biological systems for the regulation of temporal physiology is a necessary prerequisite for elaborating future clinical strategies aimed at the improvement and treatment of chronobiological disorders.
We believe this approach will have a major impact in two areas of biology:
1) Light provides the primary environmental entraining agent (Zeitgeber) for most circadian systems, however this area of signal transduction remains poorly understood and the photoreceptors and the detailed nature of their connections with the central oscillator remain unknown. An understanding of these entrainment mechanisms is central to the understanding of circadian organization;
2) Lightprovides the most reliable means of manipulating circadian phase and hence re-aligning biological time with a new environmental phase. The ability to successfully manipulate biological time will have a profound impact on medicine timed drug administration, light treatments of chronobiological disorders, and ocular treatments. Specific benefits are expected to emerge from an understanding of the human circadian system and interaction between the fundamental, clinical, and industrial participants and consultants involved in the program. Keywords: chronobiology, circadian rhythms, visual physiology, retinal degeneration, blindness, molecular biology, recombinant DNA techniques, photoreceptor opsins, primates, transgenic mice.
Funding SchemeCSC - Cost-sharing contracts