NSF Award
0234388
0234388<br/>Levin<br/><br/>Understanding developmental pattern formation is fundamental to cell and evolutionary biology; it will be a vital part of biomedicine addressing birth defects, cancer, & organ/tissue regeneration by achieving fine-scale control over the structure/function of living tissues. Research in the post-genomic age requires a synthesis of genetics and molecular biology with functional physiology in vivo. Through interdisciplinary molecular and computer modeling techniques, the Levin lab seeks a detailed understanding of cellular signals that control large-scale pattern formation by modulation of gene activity and the biophysics of membrane flux. The vertebrate body-plan possesses bilateral symmetry; however, embryos also exhibit a striking left-right asymmetry (LRA) of the viscera and brain which is highly conserved across species. Why does asymmetry exist at all? What are the implications of asymmetry for the normal physiology of the heart, gut, and brain? Why are all normal individuals not only asymmetric, but asymmetric to the same direction? When, in evolution, did handed asymmetry appear? Is it connected to chirality in plants and mollusc shells? At what developmental stages is asymmetry initiated in vertebrate embryos? What are the implications of brain asymmetry for human cognition? And, how can the left-right axis be consistently oriented with respect to the other two axes in the absence of any macroscopic feature of the world which distinguishes left from right? Are quantum parity violations relevant to mesoscopic biological shape? These issues await a detailed understanding, at the molecular, genetic, and biochemical levels, of biased asymmetry in embryos since early LR steps are poorly understood.<br/><br/>In pursuing the mechanisms of LRA, the involvement of a novel and important aspect was discovered: serotonin (5HT). 5HT is a key neurotransmitter in the central nervous system that regulates psychodynamic function and is directly implicated in both normal cognition and syndromes such as memory impairment, sleep disorders, schizophrenia, dementia, aggression, depression, etc. Thus serotonin is, potentially, a biochemical correlate of conscious states which can be used to probe the physical basis of cognition. Preliminary studies in our lab utilized the frog embryo (Xenopus) to show that (1) some members of the 5HT pathway are present in the cleavage-stage embryo (i.e., long prior to the formation of neurons) in dynamic patterns some of which display a consistent LRA, 2) 5HT signaling is functionally implicated in early steps of LRA by pharmacological experiments, and 3) the L and R sides of the embryo show differential responses to intracellular serotonin. Thus, the 5HT pathway is a promising and completely novel entrypoint into early steps of LR patterning. The proposed research will seek to understand extremely basic properties of serotonergic signaling in early embryos by capitalizing on the many advantages of Xenopus embryos over rodent and human systems for these studies, and collaborations in the neurobiology community. The aims are: (1) characterize at the embryonic and subcellular level, the localization of all LR-relevant 5HT pathway members at each stage of early development, (2) determine precisely which embryonic stages in LR patterning utilize 5HT signaling, (3) characterize a novel cytoplasmic site of functional 5HT receptors, and (4) test two specific molecular models of the role of 5HT signaling with known early steps in LR patterning. <br/><br/>Intellectual merit and broader impacts: A direct benefit will be the characterization of a novel mechanism in embryonic left-right asymmetry, in the context of known steps in LR patterning. Importantly, since this work identifies a marker of LR asymmetry within 2 hours of fertilization, it shows that embryos know left from right far earlier than heretofore suspected in the field, as well as providing reagents which set up a direct attack on the elusive "step 1". Perhaps the most interesting are the numerous long-term, wider impacts of this interdisciplinary work. Techniques will be developed and introduced for functional developmental studies of small molecule flux in vivo. These novel embryological techniques and approaches will be taught directly to graduate students and post-docs in our lab, as well as be published and presented at scientific meetings to help disseminate valuable ideas to the scientific community which can form the basis of new future work in many labs. Particularly exciting is the fact that an early role for this neurotransmitter may suggest that serotonergic synapses are an evolutionary co-opting of far more primitive cell:cell signaling events, which will have interesting implications for evolutionary developmental biology as well as neurobiology. This provides a ready context within which to train future scientists in interdisciplinary approaches to biological problems. These studies will thus initiate an understanding of a new role for a signaling molecule well-known for its physiological relevance in adult tissue. This information will benefit society by shedding light on very basic problems of neuroscience, embryology, evolution, and the safety of pharmaceutically popular 5HT- modulating drugs during pregnancy.
