Changing iron content of the mouse brain during development

Abstract

Iron is crucial to many processes in the brain yet the percentages of the major iron-containing species contained therein, and how these percentages change during development, have not been reliably determined. To do this, C57BL/6 mice were enriched in 57Fe and their brains were examined by Mössbauer, EPR, and electronic absorption spectroscopy; Fe concentrations were evaluated using ICP-MS. Excluding the contribution of residual blood hemoglobin, the three major categories of brain Fe included ferritin (an iron storage protein), mitochondrial iron (consisting primarily of Fe/S clusters and hemes), and mononuclear nonheme high-spin (NHHS) FeII and FeIII species. Brains from prenatal and one-week old mice were dominated by ferritin and were deficient in mitochondrial Fe. During the next few weeks of life, the brain grew and experienced a burst of mitochondriogenesis. Overall brain Fe concentration and the concentration of ferritin declined during this burst phase, suggesting that the rate of Fe incorporation was insufficient to accommodate these changes. The slow rate of Fe import and export to/from the brain, relative to other organs, was verified by an isotopic labeling study. Iron levels and ferritin stores replenished in young adult mice. NHHS FeII species were observed in substantial levels in brains of several ages. A stable free-radical species that increased with age was observed by EPR spectroscopy. Brains from mice raised on an Fe-deficient diet showed depleted ferritin iron but normal mitochondrial iron levels.

Kinetics of Iron Import into Developing Mouse Organs Determined by a Pup-swapping Method

“…80396) supplemented with 50 mg of 57 Fe III citrate (96.3%, Isoflex USA) per Kg chow, as described (5). Another colony was raised equivalently but on chow supplemented with natural abundance Fe III citrate containing ϳ96% 56 Fe.…”

“…After more than three generations on these diets, the mice were Ͼ90% enriched in either isotope. For simplicity, only these two isotopes were measured and included in the analysis (5 56 Fe litters were also used as controls, but only volumes and iron concentrations in brains were determined. After weaning, pups were fed the diets described above.…”

“…Brain, liver, spleen, heart, and kidneys were dissected and weighed. The concentration of iron in each organ was determined by ICP-MS, as described (5). Starting at 2 weeks and continuing each week thereafter, ϳ50 l of blood was removed from each euthanized animal.…”

“…The supernatant obtained by blood centrifugation was defined as plasma. Blood was extensively flushed from animals using Ringer's buffer (5 Fe. This symmetry allowed us to combine 56357 and 57356 data and relabel isotopes as starting and enriching.…”

“…Young rodent brains experience a growth-associated iron deficiency due to the inability of iron to be imported fast enough to counterbalance the diluting effect of brain growth (4,5).…”