Quantification of Isoflurane Uptake for Immobilization of Ostrich Embryos for Preclinical In Ovo Imaging

Preclinical imaging has recently been expanded through the use of ostrich embryos as an alternative in vivo model. In ovo experiments represent a promising substitute for conventional rodent-based animal testing. For artifact-free dynamic nuclear medicine imaging, reliable immobilization of embryos is essential. Although previous studies have demonstrated the feasibility of isoflurane anesthesia, the kinetics and uptake mechanisms of isoflurane in ostrich embryos remain insufficiently characterized. The aim of this study was to characterize gas exchange dynamics in ostrich eggs and to quantify isoflurane uptake using two complementary approaches: indirect consumption measurements in a closed system and direct quantification by serial blood sampling. Fourteen ostrich eggs, including seven fertilized and seven unfertilized specimens, were analyzed at developmental stages up to day 37 of incubation. Gas exchange was assessed in a sealed container using a clinical anesthesia gas monitoring system to measure oxygen consumption and carbon dioxide excretion. Isoflurane uptake was evaluated during exposure to concentrations of 2%, 4%, or 6%. In a separate experimental series, serial blood samples were collected during and after exposure to the same concentrations to determine systemic uptake. Fertilized embryos showed progressive increases in metabolic activity, with a maximal oxygen consumption and carbon dioxide excretion of 116 mL/h/kg and 93 mL/h/kg on day 37. Indirect measurements demonstrated isoflurane uptake rates of up to 1.1 mL/min at 6%, with proportionally lower values at 4% and 2%. Blood analyses confirmed systemic absorption, peak concentrations of 160 µg/mL, and detectable residual levels for up to 120 min after exposure. These findings refine in ovo imaging.