A Study of Toxicity Evaluation Using the iMScope TRIO - Analysis of Localization of Amiodarone in Rat Lungs

Importance of Topic

Mass spectrometry imaging (MSI) offers a label-free approach to map the spatial distribution of drug compounds and their metabolites directly in tissue sections. This capability is critical in drug discovery and toxicity evaluation, as it provides insight into pharmacokinetics, drug accumulation, and pathological correlations without the need for radioactive or fluorescent labels.

Study Objectives and Overview

This study aimed to evaluate the localization of amiodarone, an antiarrhythmic agent known to induce phospholipidosis at high doses, in rat lung tissue. By combining MSI with histological reference, the work sought to determine whether unchanged amiodarone and its N-deethylated metabolite accumulate in regions exhibiting foamy macrophage infiltration.

Methodology

Animals received three consecutive daily oral doses of amiodarone hydrochloride at 1000 mg/kg. Fresh-frozen lung sections (10 µm thickness) were prepared and coated with α-cyano-4-hydroxycinnamic acid (CHCA) via sublimation (0.7 µm thickness). MSI was conducted in positive-ion mode over m/z 500–700, using a laser diameter of 5 µm and spatial resolution of 5 µm.

Used Instrumentation

• Imaging Mass Microscope: iMScope TRIO
• Matrix Coating Device: iMLayer

Results and Discussion

High-resolution MSI detected a prominent signal at m/z 646.0, matching the molecular weight of amiodarone. Imaging revealed co-localization of this ion with areas of foamy macrophage infiltration identified by hematoxylin-eosin (HE) staining on adjacent sections. Additionally, a peak at m/z 618.0, corresponding to N-deethylated amiodarone, was observed and displayed a similar spatial distribution. These findings confirm that both the parent drug and its metabolite accumulate specifically in pathological lung regions.

Challenges in aligning MSI data with histology arise because HE staining cannot be performed on the same section. The current workflow uses serial sections and landmark-based image registration. A proposed improvement involves removing the matrix from an imaged section, then performing HE staining on that same slide to achieve perfect registration.

Benefits and Practical Applications

• Label-free detection reduces cost and avoids alteration of pharmacokinetics by labels.
• Simultaneous mapping of parent drug and metabolite enhances understanding of in situ biotransformation.
• High spatial resolution (5 µm) enables correlation with microscopic pathological features.
• Applicable for preclinical toxicity assessment and mechanistic studies in drug development.

Future Trends and Potential Applications

Advancements are expected in integrated workflows that combine MSI and histology on the same section, improving spatial co-registration. Automated image alignment tools and novel matrix removal techniques will streamline analysis. Expansion of MSI to a wider range of drugs, endogenous metabolites, and multi-modal imaging approaches will further enhance its role in pharmacology, toxicology, and precision medicine.

Conclusion

This study demonstrates the power of iMScope TRIO MSI to localize amiodarone and its metabolite within rat lung lesions. The label-free method provides valuable insights into drug distribution and toxicity mechanisms, supporting its adoption in drug discovery and safety evaluation.