Integrating Microscopy Methods to Study Gene and Protein Expression alongside Metal Ion Distribution and Speciation: A Case Study of Iron within Pyramidal Neurons from Distinct Hippocampal CA1 Subregions

Understanding the role of metal ions in normal and abnormal
cell
function continues to emerge as a critical research area in the biological
and biochemical sciences. This is especially true in the context of
brain health and neurodegenerative diseases, as the brain is especially
enriched in metal ions. A range of microscopy and bioanalytical techniques
are available to assist in characterizing and observing changes to
the brain metallome. As is the case in many other scientific fields,
the integration of multiple analytical methods often yields a more
complete chemical picture and deeper biological understanding. Herein,
we present a case study applying 4 different analytical methods to
provide spatially resolved characterization of chemical and biochemical
parameters relating to the iron (Fe) metallome within a specific brain
region, cornu ammonis sector 1 (CA1) of the hippocampus. The CA1 hippocampal
sector was chosen for investigation due to its known endogenous enrichment
in Fe and its selective vulnerability to neurodegeneration. The 4
analytical techniques applied were X-ray fluorescence microscopy (to
quantify Fe distribution); X-ray absorption near-edge structure (XANES)
spectroscopy to reveal information on Fe oxidation state and coordination
environment; immuno-fluorescence to reveal relative abundance of Fe
storage proteins (heavy chain ferritin and mitochondrial ferritin);
and spatial transcriptomics to reveal gene expression pathways relevant
to Fe homeostasis. Collectively, the results highlight that although
pyramidal neurons in lateral and medial regions of the hippocampal
CA1 sector are morphologically similar, key differences in the Fe
metallome are evident. The observed differences within the hippocampal
CA1 sector potentially indicate a higher oxidative environment and
higher metabolic turnover in medial CA1 neurons relative to lateral
CA1 neurons, which may account for the heightened vulnerability to
neurodegeneration that is observed in the medial CA1 sector.