Fundamental Development of In-vivo NMR Technology to Understand Environmental Stress

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Present environmental policies are based largely on apical endpoints such as death, movement and reproduction, but provide little information on sub-lethal impacts, toxic mechanisms or synergistic effects. Exposure to low-level contaminant mixtures is the norm rather than the exception, and subtle sub-lethal effects, are arguably more hazardous, to animal, plant and human populations, as they are often only detected, after physical symptoms become widespread. The medical field have recently identified environmental exposure as the predominant cause of many neurodegenerative diseases including links to heavy metals and pesticides, but the exact environmental causes are not yet known. A traditional top down approach, which involves identifying all contaminants, their degradation products, and then assessing toxicities on an individual compound basis is extremely challenging given the complexity of our environment. Conversely, a bottom-up approach focuses on the organisms themselves and asks questions such as; is a population stressed?; what are the stressor/stressors causing the stress?; and which biochemical pathways are impacted? Nuclear Magnetic Resonance (NMR) is one of the most powerful tools in modern research and can be applied in-vivo. As such the living organism becomes the "ultimate biosensor" responding in real-time to its environment, while the NMR spectrometer interprets the biochemical changes, providing information explaining sub-lethal toxicity at the molecular-level. The ultimate goal is to understand and categorize multifaceted stress responses in complex media (e.g. industrial effluent, waste water) and the natural aquatic environment, such that, one day, these molecular fingerprints can be used to identify the exact cause of environmental stress in native organisms. In turn, the knowledge as to the most problematic stressors impacting environmental and human health is essential to improving monitoring, targeted remediation, prevention and policies. Aquatic neonates and eggs are especially susceptible to toxins, and if compromised, entire populations and ecosystems can be impacted. However, due to its low sensitivity, current NMR approaches require 100's of neonates/eggs, making routine screening impractical. In this proposal, micro-coil NMR, micro-coil arrays, novel targeted experiments and applications will be developed to extend in-vivo NMR to single aquatic neonates and eggs. Due to the increased signal and ability to multiplex many samples, the experiment time can be reduced from months down to hrs/mins. To our knowledge no other techniques come close to providing such a wealth of molecular information in-vivo. This, along with the fact that scientific questions are becoming increasingly complex and biological relevance is always at the forefront, it is clear the high resolution in-vivo NMR methods developed here, will be key to the next generation of scientific research in general.