A chronic iron-deficient/high-manganese diet in rodents results in increased brain oxidative stress and behavioral deficits in the morris water maze

Abstract

Iron deficiency (ID) is especially common in pregnant women and may even persist following childbirth. This is of concern in light of reports demonstrating that ID may be sufficient to produce homeostatic dysregulation of other metals, including manganese (Mn). These results are particularly important considering the potential introduction of the Mn-containing gas additive, methyl cyclopentadienyl manganese tricarbonyl (MMT), in various countries around the world. In order to model this potentially vulnerable population, we fed female rats fed either control (35 mg Fe/kg chow; 10 mg Mn/kg chow) or low iron/high-manganese (IDMn; 3.5 mg Fe/kg chow; 100 mg Mn/kg chow) diet, and examined whether these changes had any long-term behavioral effects on the animals' spatial abilities, as tested by the Morris water maze (MWM). We also analyzed behavioral performance on auditory sensorimotor gating utilizing prepulse inhibition (PPI), which may be related to overall cognitive performance. Furthermore, brain and blood metal levels were assessed, as well as regional brain isoprostane production. We found that treated animals were slightly ID, with statistically significant increases in both iron (Fe) and Mn in the hippocampus, but statistically significantly less Fe in the cerebellum. Additionally, isoprostane levels, markers of oxidative stress, were increased in the brain stem of IDMn animals. Although treated animals were indistinguishable from controls in the PPI experiments, they performed less well than controls in the MWM. Taken together, our data suggest that vulnerable ID populations exposed to high levels of Mn may indeed be at risk of potentially dangerous alterations in brain metal levels which could also lead to behavioral deficits.

Fig. 1

Average weekly gain. Pregnant dams were weighed weekly as a measure of general health. Loss of apparent body mass at week 2 indicates birth of pups. In general, dams on an iron-deficient/high Mn (IDMn) diet were statistically significantly lighter (P < 0.0001) than controls as determined by linear regression

Fig. 2

Regional brain masses. Brains were removed and dissected into various regions for metal and isoprostane analysis. No statistically significant difference in regional wet mass was determined for each region

Fig. 3

Brain metal accumulation. Metal amount was determined by atomic absorption spectroscopy (AAS). Student’s t-test indicated significant regional accumulation of, a Mn or b iron in treated animals compared to controls. Mn (P = 0.05) and iron (P = 0.0062) were higher in the hippocampus of treated rats. In the cerebellum, iron levels were reduced (P = 0.035), while Mn content approached statistical significance (P = 0.062) compared to cerebella from animals on a control diet

Fig. 4

Regional isoprostane (IsoP) formation. IsoP levels were determined by mass spectrometery as described in the Sect. Material and Methods. Data is presented as percent control IsoP production, mean ± SEM. *P = 0.0487, **P = 0.0064

Fig. 5

Morris water maze. a Acquisition latency is represented as a function of trial block for both control and IDMn groups. There were no significant differences between groups; b Mean search difference (MSD) score is presented as a function of group. The IDMn group demonstrated a significantly lower MSD score on the probe trial as compared to controls, demonstrating these animals searched less frequently in the quadrant formerly containing the platform as compared to controls; c Mean zone difference (MZD) score is presented as a function of group. Similar to the MSD score, the IDMn group demonstrated a significantly lower MZD score on the probe trial as compared to controls, demonstrating they made less visits to the former platform location as compared to controls. *P < 0.05

Fig. 6

Prepulse inhibition. a 73 dB prepulse trials; b 76 dB prepulse trials; c 82 dB prepulse trials. There were no statistically significant differences between groups regardless of the prepulse auditory intensity