Capillary Electrophoresis-High Resolution Mass Spectrometry for Measuring In Vivo Arginine Isotope Incorporation in Alzheimer's Disease Mouse Models

Abstract

Immune-based metabolic reprogramming of arginine utilization in the brain contributes to the neuronal pathology associated with Alzheimer's disease (AD). To enable our long-term goals of differentiation of AD mouse model genotypes, ages, and sexes based on activity of this pathway, we describe here the novel dosing (using uniformly labeled (¹³C₆¹⁵N₄) arginine) and analysis methods using capillary electrophoresis high-resolution accurate-mass mass spectrometry for isotope tracing of metabolic products of arginine. We developed a pseudoprimed infusion-dosing regimen, using repeated injections, to achieve a steady state of uniformly labeled arginine in 135-195 min post bolus dose. Incorporation of stable isotope labeled carbon and nitrogen from uniformly labeled arginine into a host of downstream metabolites was measured in vivo in mice using serially sampled dried blood spots from the tail. In addition to the dried blood spot time course samples, total isotope incorporation into arginine-related metabolites was measured in the whole brain and plasma after 285 min. Preliminary demonstration of the technique identified differences isotope incorporation in arginine metabolites between male and female mice in a mouse-model of sporadic Alzheimer's disease (APOE4/huNOS2). The technique described herein will permit arginine pathway activity differentiation between mouse genotypes, ages, sexes, or drug treatments in order to elucidate the contribution of this pathway to Alzheimer's disease.