The mechanism of GPCR signalling in zebrafish semicircular canal morphogenesis

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The semicircular canals of the inner ear detect turning movements of our head, triggering muscular reflexes that enable us to maintain our balance. There are three canals in each inner ear, and these are each arranged at right angles to one another, so that they can detect movement in any direction. Understanding how these canals develop in the embryo, together with the rest of the intricate structure of the inner ear-rightly also known as the labyrinth-is an interesting problem. We study this process in the zebrafish embryo, but there are many similarities to the sequence of events during human development. The zebrafish ear initially starts out in the early embryo as a simple fluid-filled ball of cells. Projections of tissue then form that grow out into the fluid-filled space inside this ball, where they touch and fuse to make pillars, which become the hubs of the semicircular canal ducts. We have recently identified a receptor protein that appears to be involved in triggering a whole series of events to enable to the projections to recognise one another, fuse and rearrange accordingly. When the gene coding for this receptor is disrupted, the projections fail to touch and fuse: they appear to slide past one another without recognition. As a result, the semicircular canals do not form properly, the ear becomes misshapen and swollen, and the fish grow up with mild balance defects. The receptor is a member of an interesting class of proteins called GPCRs that are located in the cell membrane, projecting from its surface. A GPCR receptor does not act in isolation, however, but usually responds to a signalling molecule at the cell surface, and then triggers events inside the cell that activate new genes. One of the aims of this project is to identify the other players in this process. We have a number of good clues from fish strains in which the semicircular canals fail to form correctly. We will aim to identify any new genes involved in the same process. We will also gather information about when and where the receptor protein is present, how it is regulated, and how it functions during the development process. We have already identified some of the genes that are regulated by the receptor. One class of these, the versicans, remain highly active when receptor function is missing, when they should normally be turned off. We will test whether the versicans are the players that actually control how the projections move and fuse in the developing zebrafish ear. A final aim is to search for small drug-like molecules that can affect the process of semicircular canal formation in the ear. There are many GPCR proteins, and these comprise targets for many of the drugs that are on the market today. There is, therefore, considerable commercial interest in understanding more about GPCRs and in identifying new drugs that affect their activity. We will aim to carry out a small-scale study to identify new activators and inhibitors of the receptor, which we will then be able to develop further in future projects. Although this project aims to understand processes that happen in the zebrafish ear, the GPCR is active in other areas of the embryo, and so the conclusions are likely to aid in our understanding of the development of other organ systems, such as the heart and nervous system.