Probing Role of Tetrahydrobiopterin in Cerebral Palsy by Using Transgenic Rabbits

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PROJECT SUMMARY/ABSTRACT Children with movement disorders are a big burden to society. The burden of disease is very high because of the life-long consequences to the patient, caretakers, and social institutions. Currently there are no cures or preventative treatments for cerebral palsy (CP), as the mechanisms of disease remain poorly defined. Human mutations in key enzymatic pathways constitute genetic causes of childhood movement disorders. With the advent of transgenic rabbit models, a golden opportunity has arisen to study the pathogenetic mechanisms in brain leading to movement disorders, as rabbits are more likely to present with movement disorders mimicking that of humans. Rabbits are perinatal brain developers like humans. Mutations in enzymes of tetrahydrobiopterin pathway result in movement disorders. Tetrahydrobiopterin is an enzyme co-factor and its supplementation in congenital deficiency disorders ameliorates the movement disorder. Thus, there may be a critical role of tetrahydrobiopterin in the development of movement disorders, such as CP. We developed a knockout rabbit that introducing a specific mutation in one of the tetrahydrobiopterin synthesis enzymes, sepiapterin reductase. Following fetal hypoxia-ischemia, newborn rabbits present with hypertonia and difficulty with balance. Fetal rabbits showing low developmental tetrahydrobiopterin in discrete brain regions have a greater disposition to develop hypertonia. Magnetic resonance imaging (MRI) allows us to predict which fetuses will develop postnatal hypertonia. This advance allows the identification of early critical pathways causing hypertonia. Our objective is to elucidate molecular mechanisms of perinatal brain injury in human mutations causing childhood movement disorders, by decreasing tetrahydrobiopterin levels using a hetero- and homozygous knockout transgenic approach in the rabbit. The main question asked in this proposal is whether tetrahydrobiopterin in selective brain regions contributes to the development of motor disorders with a severity determined by an added prenatal insult such as hypoxia-ischemia or inflammation. Using genetic knockout of sepiapterin reductase, we can further lower the tetrahydrobiopterin levels in brain and investigate whether the resulting motor deficits are increased or that we need less degree of insult to achieve the same motor deficits. The first Aim determines whether an added fetal insult, hypoxia-ischemia or inflammation from lipopolysaccharide, enhances movement disorders in the sepiapterin het/homozygous reductase knockout rabbit. The second Aim will determine if neuronal or oligodendroglial injury explains the development of movement disorders in the knockout rabbit. We use innovative pre- and postnatal MRI biomarkers of hypertonia with tissue flow cytometry and high-performance liquid chromatography with electrochemical detection. By conducting a time-dependent, organ-specific and cell-specific pathogenetic study, we will obtain a comprehensive picture of the role of this cofactor in perinatal pathogenesis of movement disorders.