The urea cycle is the major pathway for detoxification of ammonia in mammals. In humans, arginase 1 deficiency is characterized clinically by progressive mental impairment, spasticity, and growth retardation, with only periodic episodes of hyperammonemia unlike the other urea cycle disorders where this is much more common. In recent experiments, we have found substantial anatomical, ultrastructural and electrophysiological differences between knockout animals and wild type controls. This includes decreased intrinsic excitability, altered functional synaptic transmission, decreased dendritic arborization and decreased synapse density. These measurable differences at the neuron, synapse, and circuit level have begun to elucidate the functional abnormalities in arginase deficiency. However, a human model is essential as there are critical differences between mice and humans in disease presentation for this disorder. Such a model will allow establishment of high-throughput screens to determine the compounds responsible for the neuronal abnormalities detected and to discover compounds that may improve cognitive and motor functions. In this proposal the Lipshutz Lab will examine the role arginase plays in the normal development of neurons and circuits as they have hypothesized that anatomic and functional abnormalities develop in neurons of arginase deficient patients and that such changes are responsible for the phenotype detected in humans with homozygous deficiency of arginase 1. In addition, the lab will study metabolites hypothesized to be involved in the pathogenesis of the disorder, thus opening doors to potential pharmacological interventions. Preliminary data: This research group has (amongst other findings): 1) constructed and characterized the arginase 1 knockout mouse; 2) demonstrated long-term survival and rescue with liver-specific arginase 1 expression by recombinant adeno-associated viral vectors; 3) demonstrated that only low-level ureagenesis is necessary for long-term survival; 4) shown that single copy (heterozygotes) or double copy (homozygotes) of loss of arginase gene expression results in abnormalities of intrinsic excitability and the dendritic arbor of murine neurons; 5) shown, using an array of tests, that AAV-treated liver-specific arginase knockout animals have persistent abnormalities of neurons at the synapse and dendritic arbor level and in their electrophysiologic response; and 6) shown that peripheral metabolism can result in control of circulating plasma arginine.
In Aim 1, the hypothesis that human pluripotent stem cell (hPSC)-derived neurons with deficient arginase expression will show deficits in synaptogenesis, excitability, and neurite development will be tested.
In Aim 2, we will determine the effect of arginine and other presumptive causative agents on the intrinsic excitability and synaptic physiology of arginase-deficient hPSC-derived neurons. This proposal from a team of investigators with complementary expertise in arginase deficiency, stem cell biology with in vitro neuron development, and electrophysiology will be critical for building and validating this human neuronal culture model.
This project will elucidate the role arginase activity plays in the normal development of neurons utilizing arginase- deficient stem cells and disease-in-a-dish methodology. We will also examine the role that certain biochemical compounds may play in the neuropathology of arginase deficiency.