The differential effects of early-life phlebotomy-induced anemia on regional brain metabolism and behavior

University of Minnesota Ph.D. dissertation. November 2017. Major: Neuroscience. Advisor: Michael Georgieff. 1 computer file (PDF); 137 pages.Oxygen and iron are critical metabolic substrates for rapidly developing brain structures in infancy and childhood. Anemia, which causes a deficit of both, is a common pediatric problem associated with long-term neurodevelopmental impairment. Estimates indicate that 30-50% of the world’s children are anemic, including over 300,000 preterm neonates per year in the US who become profoundly anemic due to phlebotomy-induced anemia (PIA) in the neonatal intensive care unit and are at high risk for neurodevelopmental impairment. PIA risks shortchanging the young brain of oxygen and iron, and the brain structures that are most rapidly developing during the period of anemia are likely to be affected. Few if any clinical studies have directly assessed the short and long-term neurodevelopmental effects of PIA in preterm infants. However, extensive studies in humans and preclinical rodent models have shown the long-term effects of early-life dietary iron deficiency anemia, supporting the hypothesis that early-life anemia is detrimental to the brain. One intervention which may help to alleviate the anemia and lessen these changes is administration of recombinant human erythropoietin which stimulates red blood cell production, thereby increasing tissue oxygen delivery, though potentially drawing iron away from the brain as a result. Using a developmentally and physiologically-relevant mouse model of PIA, my thesis sought to determine the effect of anemia during early-life on the metabolism of four brain regions critical to normal cognitive development: cerebellum, striatum, hippocampus and prefrontal cortex, as well as the long-term outcomes following recovery from anemia. This was achieved using: 1) in vivo ultra-high field 1H NMR spectroscopy to measure regional steady-state concentrations of critical metabolites, 2) western blot of proteins in the metabolic-sensing mTOR pathway, 3) PCR to determine differential gene expression, 4) ex vivo extracellular flux analysis to assess neuronal bioenergetics, and 5) targeted behavioral assays to determine functional deficits in adulthood. The objective was to determine whether there were differential region-specific alterations to metabolism in PIA mice as compared to non-bled controls within regions that were rapidly growing during the period of anemia (hippocampus, striatum) compared with those whose growth spurt occurred before or after the period of anemia