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Iron deficiency anemia in early life alters the development and functioning of the dopamine neurotransmitter system, but data regarding the specific effects of brain iron loss on dopamine D(2) receptor regulation are lacking. Cell culture and animal models were employed in this study to determine whether D(2) receptor expression is altered when cellular iron levels are depleted. Endogenous D(2) receptor-expressing PC12 cells exposed to increasing concentrations of the iron chelator desferrioxamine (25-100 micromol/L) exhibited dose-dependent decreases in total D(2) receptor protein concentrations (20-65%), but there were minimal effects on D(2) receptor mRNA levels. When iron-deficient cells were repleted with ferric ammonium citrate for 24 h, D(2) receptor protein densities were similar to control. Dietary iron deficiency for 6 wk in weanling rats also reduced regional iron concentrations by nearly 50% in the ventral midbrain and caudate but did not affect D(2) receptor mRNA levels in the ventral midbrain. Iron deficiency significantly reduced membrane D(2) receptor protein levels by >70% in caudate, whereas cytosolic concentrations showed only 25% losses. D(2) receptor protein densities and regional iron concentrations were restored within 2 wk of dietary iron repletion. These results support the concept that D(2) receptor gene expression is not significantly changed by iron deficiency, whereas dopamine receptor trafficking is affected and is likely related to known dopamine system alterations in iron deficiency. [ABSTRACT FROM AUTHOR]

Iron deficiency in early life is associated with delayed development as assessed by a number of clinical trials using similar global scales of development; this poor development during infancy persists in most cases after iron therapy has corrected iron status. If iron deficiency occurs in preschool and older children, the consequences appear reversible with treatment. The biologic understanding of this relationship between development, brain iron status, and functioning is sparse though animal studies repeatedly demonstrate alterations in dopamine metabolism and in the myelination process. Dietary iron deficiency can rapidly deplete brain iron concentrations and repletion is able to normalize them. Residual alterations in striatal dopamine metabolism and myelin production persist if neonatal animals are used. Future studies with more specific measures of neurodevelopment in iron-deficient human infants, and animal models, will allow investigators to more clearly define causal roles of brain iron in neural development and functioning. [ABSTRACT FROM AUTHOR]

Background: Very limited data suggest that meat consumption by older people may promote skeletal muscle hypertrophy in response to resistance training (RT). Objective: The objective of this study was to assess whether the consumption of an omnivorous (meat-containing) diet would influence RT-induced changes in whole-body composition and skeletal muscle size in older men compared with a lactoovovegetarian (LOV) (meat-free) diet. Design: Nineteen men aged 51-69 y participated in the study. During a 12-wk period of RT, 9 men consumed their habitual omnivorous diets, which provided approximately 50% of total dietary protein from meat sources (beef, poultry, pork, and fish) (mixed-diet group). Another 10 men were counseled to self-select an LOV diet (LOV-diet group). Results: Maximal strength of the upper- and lower-body muscle groups that were exercised during RT increased by 10-38% (P < 0.001), independent of diet. The RT-induced changes in whole-body composition and skeletal muscle size differed significantly between the mixed- and LOV-diet groups (time-by-group interactions, P < 0.05). With RT, whole-body density, fat-free mass, and whole-body muscle mass increased in the mixed diet group but decreased in the LOV- diet group. Type II muscle fiber area of the vastus lateralis muscle increased with RT for all men combined (P < 0.01), and the increase tended to be greater in the mixed-diet group (16.2 +/= 4.4 %) than in the LOV diet group (7.3 +/= 5.1 %). Type I fiber area was unchanged with RT in both diet groups. Conclusion: Consumption of a meat-containing diet contributed to greater gains in fat-free mass and skeletal muscle mass with RT in older men than did an LOV diet. Copyright (c) 1999 American Society for Clinical Nutrition [ABSTRACT FROM AUTHOR]

Nutritional benefits of cultivated oat (Avena sativa L., 2n = 6x = 42, AACCDD) are well recognized; however, seed protein levels are modest and resources for genetic improvement are scarce. The wild tetraploid, A. magna Murphy et Terrell (syn A. maroccana Gdgr., 2n = 4x = 28, CCDD), which contains approximately 31% seed protein, was hybridized with cultivated oat to produce a domesticated A. magna. Wild and cultivated accessions were crossed to generate a recombinant inbred line (RIL) population. Although these materials could be used to develop domesticated, high-protein oat, mapping and quantitative trait loci introgression is hindered by a near absence of genetic markers. Objectives of this study were to develop high-throughput, A. magna-specific markers; generate a genetic linkage map based on the A. magna RIL population; and map genes controlling oat domestication. A Diversity Arrays Technology (DArT) array derived from 10 A. magna genotypes was used to generate 2,688 genome-specific probes. These, with 12,672 additional oat clones, produced 2,349 polymorphic markers, including 498 (21.2%) from A. magna arrays and 1,851 (78.8%) from other Avena libraries. Linkage analysis included 974 DArT markers, 26 microsatellites, 13 SNPs, and 4 phenotypic markers, and resulted in a 14-linkage-group map. Marker-to-marker correlation coefficient analysis allowed classification of shared markers as unique or redundant, and putative linkage-group-to-genome anchoring. Results of this study provide for the first time a collection of high-throughput tetraploid oat markers and a comprehensive map of the genome, providing insights to the genome ancestry of oat and affording a resource for study of oat domestication, gene transfer, and comparative genomics.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease with complicated pathogenesis with variable presentation and disease progression. There is a critical need for a panel of biomarkers to provide clinicians and researchers with additional information. In this study, multiplex immunoassays were used to screen a number of cytokines, growth factors, and iron-related proteins. ALS patients had significantly higher plasma levels of L-ferritin and lower concentrations of transferrin when compared to healthy controls and together classified a test group of subjects with 82% accuracy. Duration of ALS symptoms correlated positively with levels of monocyte chemoattractant protein 1 (MCP-1) and negatively with levels of granulocyte-macrophage colony stimulating factor (GM-CSF). The biomarker profile suggests iron homeostasis is disrupted in ALS patients, and changes in ferritin and transferrin (Tf) appear to be indicators of ongoing inflammatory processes. The data demonstrate a plasma biomarker profile in ALS patients that may differ from published reports of cerebrospinal fluid biomarkers.

Iron imbalances in the brain, including excess accumulation and deficiency, are associated with neurological disease and dysfunction; yet, their origins are poorly understood. Using systems genetics analysis, we have learned that large individual differences exist in brain iron concentrations, even in the absence of neurological disease. Much of the individual differences can be tied to the genetic makeup of the individual. This genetic-based differential regulation can be modeled in genetic reference populations of rodents. The work in our laboratory centers on iron regulation in the brain and our animal model consists of 25 BXD/Ty recombinant inbred mouse strains. By studying naturally occurring variation in iron phenotypes, such as tissue iron concentration, we can tie that variability to one or more genes by way of quantitative trait loci (QTL) analysis. Moreover, we can conduct genetic correlation analyses between our phenotypes and others previously measured in the BXD/Ty strains. We have observed several suggestive QTL related to ventral midbrain iron content, including one on chromosome 17 that contains btbd9, a gene that in humans has been associated with restless legs syndrome and serum ferritin. We have also observed gene expression correlations with ventral midbrain iron, including btbd9 expression and dopamine receptor expression. In addition, we have observed significant correlations between ventral midbrain iron content and dopamine-related phenotypes. The following is a discussion of iron regulation in the brain and the contributions a systems genetics approach can make toward understanding the genetic underpinnings and relation to neurological disease.

Restless leg syndrome (RLS) is a sensorimotor disorder. Clinical studies have implicated the dopaminergic system in RLS, while others have suggested that it is associated with insufficient levels of brain iron. To date, alterations in brain iron status have been demonstrated but, despite suggestions from the clinical literature, there have been no consistent findings documenting a dopaminergic abnormality in RLS brain tissue. In this study, the substantia nigra and putamen were obtained at autopsy from individuals with primary RLS and a neurologically normal control group. A quantitative profile of the dopaminergic system was obtained. Additional assays were performed on a catecholaminergic cell line and animal models of iron deficiency. RLS tissue, compared with controls, showed a significant decrease in D2R in the putamen that correlated with severity of the RLS. RLS also showed significant increases in tyrosine hydroxylase (TH) in the substantia nigra, compared with the controls, but not in the putamen. Both TH and phosphorylated (active) TH were significantly increased in both the substantia nigra and putamen. There were no significant differences in either the putamen or nigra for dopamine receptor 1, dopamine transporters or for VMAT. Significant increases in TH and phosphorylated TH were also seen in both the animal and cell models of iron insufficiency similar to that from the RLS autopsy data. For the first time, a clear indication of dopamine pathology in RLS is revealed in this autopsy study. The results suggest cellular regulation of dopamine production that closely matches the data from cellular and animal iron insufficiency models. The results are consistent with the hypothesis that a primary iron insufficiency produces a dopaminergic abnormality characterized as an overly activated dopaminergic system as part of the RLS pathology.

Background: A new index to determine body iron promises a simpler approach to monitoring iron deficiency (ID) prevalence., Objective: Our objective was to compare ID defined as body iron <0 mg/kg and calculated from the log ratio of transferrin receptor to ferritin (the body iron model) to ID defined as >/=2 of 3 abnormal concentrations in ferritin, transferrin saturation, or erythrocyte protoporphyrin (the ferritin model)., Design: We used measures of iron status and inflammation from 486 children aged 1-2 y, 848 children aged 3-5 y, and 3742 nonpregnant females aged 12-49 y from the National Health and Nutrition Examination Survey 2003-2006., Results: ID prevalences (+/-SE) based on the body iron model in children (1-2 and 3-5 y) and in females (12-19 and 20-49 y) were 14.4 +/- 1.9%, 3.7 +/- 0.8%, 9.3 +/- 1.0%, and 9.2 +/- 1.6%, respectively. ID prevalences based on the ferritin model in children (3-5 y) and females (12-19 and 20-49 y) were 4.5 +/- 0.9%, 15.6 +/- 1.2%, and 15.7 +/- 0.8%, respectively. The kappa statistics for agreement between the 2 models were 0.5-0.7. Among females (12-49 y) the positive predictive values of ID based on the body iron model and the ferritin model for identifying anemia were 43 +/- 3% and 30 +/- 2%, respectively, whereas negative predictive values did not differ. C-reactive protein was elevated in 28.8 +/- 3.1% of females with ID by the ferritin model but not by the body iron model and in 0% of persons with ID by the body iron model but not by the ferritin model., Conclusions: The agreement between the 2 indexes was fair to good. Among females, the body iron model produced lower estimates of ID prevalence, better predicted anemia, and appeared to be less affected by inflammation than the ferritin model.

Monoamine metabolism in the central nervous system is altered by dietary iron deficiency, with a stronger effect seen during the active than rest span of the circadian cycle. In this report, we examined changes in intracellular and extracellular monoamine levels, synthetic enzymes, transporter and receptor densities, and responses to amphetamine-induced dopamine (DA) efflux in iron-deficient and iron-sufficient mice. Extracellular striatal DA levels were 15-20% higher in all groups during the active dark phase compared to the inactive light phase, with correspondingly lower dopamine transporter (DAT) and higher tyrosine hydroxylase levels. Iron deficiency decreased DAT density by 20% and 28% in the light and dark phases, respectively, and elevated the DOPAC/DA ratio only in the dark, indicating that iron deficiency does interact with the normal diurnal cues for cyclicity. Enhanced DA efflux after amphetamine stimulation indicates no limitation on monoamine synthesis and release and is consistent with altered synaptic efficacy and perhaps recycling of DA in iron deficiency. These experimental findings provide new evidence that brain iron insufficiency does have a differential effect on the DA system at different biological times of the day and night and may be causally related to the phasic motor symptoms observed in Restless Legs Syndrome.

Both during and after a period of iron deficiency (ID), iron-dependent neural processes are affected, which raises the potential concern that the anemia commonly experienced by many growing infants could have a protracted effect on the developing brain. To further investigate the effects of ID on the immature brain, 49 infant rhesus monkeys were evaluated across the first year of life. The mothers, and subsequently the infants after weaning, were maintained on a standardized diet containing 180 mg/kg of iron and were not provided other iron-rich foods as treats or supplements. As the infants grew, they were all screened with hematological tests, which documented that 16 (33.3%) became markedly ID between 4 and 8 months of age. During this anemic period and subsequently at 1 year of age, cerebrospinal fluid (CSF) specimens were collected to compare monoamine activity in the ID and iron-sufficient infants. Monoamine neurotransmitters and metabolite levels were normal at 4 and 8 months of age, but by 1 year the formerly anemic monkeys had significantly lower dopamine and significantly higher norepinephrine levels. These findings indicate that ID can affect the developmental trajectory of these two important neurotransmitter systems, which are associated with emotionality and behavioral performance, and further that the impact in the young monkey was most evident during the period of recovery., ((c) 2009 Wiley Periodicals, Inc.)

Background: Iron deficiency is most commonly found in women of reproductive age and infants worldwide, but the influence of maternal iron deficiency on infant development is underexplored., Objective: The objective was to examine the relation between maternal iron status and mother-child interactions in a randomized, double-blind, intervention trial conducted in South Africa., Design: Women were recruited into the study from a health clinic at 6-8 wk postpartum and were classified as either iron-deficient anemic (IDA) or iron-sufficient after blood analysis. IDA mothers received iron supplements of 125 mg FeSO(4) (IDA-Fe; n = 34) or placebo (IDA-PL; n = 30) daily from 10 wk to 9 mo postpartum. The control group (n = 31) consisted of iron-sufficient mothers. Free-play mother-child interaction sessions were videotaped in the clinic at 10 wk (n = 80) and 9 mo (n = 66) postpartum and coded per the Emotional Availability Scales (4 maternal scales: sensitivity, structuring, nonintrusiveness, and nonhostility; 2 infant scales: responsiveness and involvement)., Results: At 10 wk, scores for maternal sensitivity and child responsiveness were significantly greater in the control group than in the IDA groups (P = 0.028 and 0.009, respectively). At 9 mo, the control and IDA-Fe groups no longer differed. These 2 groups scored significantly better on the maternal sensitivity, structuring, and nonhostility scales and on the child responsiveness scale than did the IDA-PL group (P = 0.007-0.032), whose iron status remained low., Conclusion: These data indicate that maternal iron deficiency negatively affects mother-child interactions and that iron supplementation protects against these negative effects.

Objective: The aim of this study was to ascertain whether high-dose intravenous (IV) iron sucrose could improve symptoms and change brain iron concentrations in idiopathic RLS., Methods: The study was a randomized, parallel-group double-blind study of 1000mg iron sucrose given IV versus placebo. Primary measures of the clinical status were global rating scale (GRS) and periodic leg movements of sleep (PLMS). Primary measures of brain iron status were CSF ferritin and MRI-determined iron in the substantia nigra., Results: At the time of the interim analysis there were 7 placebo and 11 iron-treated subjects. At 2-weeks post-treatment, iron treatment resulted in a small but significant increase in CSF ferritin and a decrease in RLS severity (GRS) but did not change PLMS or MRI iron index. None of the secondary outcomes changed with treatment. There was no single case of clear treatment benefit in any of the patients. This interim analysis revealed an effect size that was too small to allow for adequate power to find significant differences with the planed 36-subject enrollment for either the primary objective outcome of PLMS or any of the secondary outcomes. The study was stopped at this planned break-point given the lack of both adequate power and any indication for clinically significant benefit., Conclusions: High-dose IV iron failed to demonstrate the robust changes reported in three prior open-label studies. Differences in iron formulation, dosing regiment, and peripheral iron status may explain some of the discrepancies between this and previous IV iron treatment studies.

Background: Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease with complicated pathogenesis that poses challenges with respect to diagnosis and monitoring of disease progression., Objectives: To identify a biomarker panel that elucidates ALS disease pathogenesis, distinguishes patients with ALS from neurologic disease controls, and correlates with ALS disease characteristics, and to determine the effect of HFE gene variants, a potential risk factor for sporadic ALS, on the biomarker profile., Methods: We obtained CSF samples by lumbar puncture from 41 patients with ALS and 33 neurologic disease controls. All patients were genotyped for HFE polymorphisms. We performed a multiplex cytokine and growth factor analysis and immunoassays for iron-related analytes. Classification statistics were generated using a support vector machine algorithm., Results: The groups of patients with ALS and neurologic disease controls were each associated with distinct profiles of biomarkers. Fourteen biomarkers differed between patients with ALS and the control group. The five proteins with the lowest p values differentiated patients with ALS from controls with 89.2% accuracy, 87.5% sensitivity, and 91.2% specificity. Expression of IL-8 was higher in those patients with lower levels of physical function. Expression of beta2-microglobulin was higher in subjects carrying an H63D HFE allele, while expression of several markers was higher in subjects carrying a C282Y HFE allele., Conclusions: A CSF inflammatory profile associated with amyotrophic lateral sclerosis (ALS) pathogenesis may distinguish patients with ALS from neurologic disease controls, and may serve as a biomarker panel to aid in the diagnosis of ALS pending further validation. Some of these biomarkers differ by HFE genotype.

Iron movement between organ pools involves a dynamic equilibrium of iron efflux and uptake, and homeostatic mechanisms are likely involved in providing iron to cells and organs when required. Daily iron levels in the plasma pool fluctuate with the diurnal cycle, but clear explanations regarding the objectives and regulation of the flux are lacking. The association between diurnal cycle and iron flux is relevant in the disease of restless legs syndrome (RLS), where individuals display diurnal deficits in motor control, have impaired brain iron metabolism, and perhaps altered iron uptake from the plasma pool. The goal of the present study was to examine diurnal variations in peripheral and regional brain iron to evaluate iron flux between organs in iron-sufficient and iron-deficient mice. In mice fed control diet, liver iron was elevated 30-40%, and plasma iron was reduced 20-30% in the active dark period compared with the inactive light phase. Dietary iron deficiency eliminated this variation in liver iron in male and female mice and in plasma iron in male mice. Reductions in ventral midbrain and nucleus accumbens iron and ferritin were apparent in iron-deficient mice during both diurnal phases, but only during the light phase was an approximately 25% reduction in whole brain iron observed, suggesting different brain iron requirements between phases. These data demonstrate that iron flux between organs is sensitive to diurnal regulatory biology. Importantly, variations in brain iron may have temporal implications regarding neural functioning and may contribute to the diurnal cycle-dependent symptoms of RLS.

Infants who experience iron deficiency during the first 6-12 mo of life are likely to experience persistent effects of the deficiency that alter functioning in adulthood. A lack of sufficient iron intake may significantly delay the development of the central nervous system as a result of alterations in morphology, neurochemistry, and bioenergetics. Depending on the stage of development at the time of iron deficiency, there may be an opportunity to reverse adverse effects, but the success of repletion efforts appear to be time dependent. Publications in the past several years describe the emerging picture of the consequences of iron deficiency in both human and animal studies. The mechanisms for iron accumulation in the brain and perhaps redistribution are being understood. The data in human infants are consistent with altered myelination of white matter, changes in monoamine metabolism in striatum, and functioning of the hippocampus. Rodent studies also show effects of iron deficiency during gestation and lactation that persist into adulthood despite restoration of iron status at weaning. These studies indicate that gestation and early lactation are likely critical periods when iron deficiency will result in long-lasting damage.

Iron deficiency (ID) disrupts brain dopamine (DA) and norepinephrine (NE) metabolism including functioning of monoamine transporters and receptors. We employed caudate microdialysis and no net flux (NNF) in post-weaning rats to determine if ID decreased the extraction fraction (E(d)). Five micromolar quinpirole, a dopamine D(2) receptor agonist, resulted in 80% decrease in extracellular DA and 45% higher E(d) in control animals. The D(2) agonist had no effect on E(d) in ID animals despite a reduction in basal DA. DAT mRNA levels were reduced by 58% with ID, while DAT protein in ventral midbrain and caudate and membrane associated DAT were also reduced by ID. Carbidopa/l-DOPA was administered to determine if elevated extracellular DA in ID was due to increased release. The DA response to l-DOPA in ID rats was 50% smaller and delayed, whereas the NE response was threefold higher. The caudate concentration of NE was also elevated in ID. Elevated dopamine-beta-hydroxylase activity in ID provides a tentative explanation for the increased NE response to l-DOPA. These experiments provide new evidence that ID results in altered synthesis and functioning of DAT and perhaps suggests some compensatory changes in NE metabolism.

Our purpose in the present study was to examine how two different sets of stressors, one representing the physical environment and the other representing the social environment, related to perceived stress among new mothers served by a health clinic in Khayelitsha, South Africa. We found that among the chronic urban poverty-environmental stressors related to water, housing, transportation, toileting, and lack of food, that lack of drinkable water in the home had the strongest correlation with perceived stress. In terms of social stressors we found that 60% of new mothers had no partner, and 43% of those with a partner reported that they currently were not coresiding. In terms of the social stressors, the inability to depend on a partner in times of trouble had the strongest relationship to perceived stress. Other findings relating to partner support are discussed as well as sample and community characteristics. Given the importance of partner support, it is argued that the conditions of poverty itself serve to destabilize relationships, which in turn contributes to the cycle of poverty experienced by many residents of periurban settlements like Khayelitsha.

Objective: Studies using cerebrospinal fluid, magnetic resonance imaging, and autopsy tissue have implicated a primary role for brain iron insufficiency in restless legs syndrome (RLS). If the abnormalities of brain iron regulation reflect a basic disturbance of iron metabolism, then this might be expressed at least partially in some peripheral systems. Thus the study aim was to determine whether patients with RLS and control subjects show differences in lymphocyte iron regulator proteins., Methods: Fasting morning blood samples were used to obtain common serum measures of iron status and to determine lymphocyte iron management proteins. Twenty-four women with early-onset RLS and 25 control women without RLS symptoms were studied., Results: RLS and control subjects were matched for age, hemoglobin, and serum iron profile. However, transferrin receptor (TfR) and DMT1 (divalent metal transporter 1 protein) levels in lymphocytes were significantly higher for RLS patients than for controls. No significant differences in ferritin subtypes or transferrin levels were found. No significant correlations were found between lymphocyte and serum indices of iron status., Interpretation: RLS lymphocytes showed an increase in ferroportin, implying increased cellular iron excretion, in the face of increased iron need (increased TfR and DMT1). In the absence of changes in H-ferritin, the findings indicate a balance between input and output with no net iron change but probable overall increase in iron turnover. The lack of any significant correlation between serum and lymphocyte iron indices indicates that iron management proteins from lymphocytes are at a minimum an alternative and independent marker of cellular iron metabolism.

Iron deficiency (ID) anemia during infancy results in long-term neurological consequences, yet the mediating mechanisms remain unclear. Infant monkeys often become naturally anemic during the first 6 months of life, presenting an opportunity to determine the effect of developmental iron deficiency. After weaning, animals were chosen randomly for supplementation with oral iron or, fed a standard commercial chow diet. The control group was never iron deficient. ID anemia was corrected by 12 months in both groups, as indicated by hematological parameters. CSF was collected for proteomic analysis at 12 months of age to assess the impact of developmental ID on the brain. The CSF proteome for both formerly iron deficient groups was similar and revealed 12 proteins with expression levels altered at least twofold. These proteins were identified by matrix assisted laser desorption ionization time-of-flight spectrometry and included prostaglandin D synthase, olfactory receptors and glial fibrillary acidic protein. Thus the proteomic analysis reveals a persistent effect of ID and provides insights into reports of disturbed sleep, hypomyelination and other behavioral alterations associated with ID. Furthermore, alterations in the CSF proteome despite normal hematologic parameters indicate that there is a hierarchical system that prioritizes repletion of red cell mass at the expense of the brain.

Fe, Cu, and Zn are of widespread neurobiological importance, but must be regulated closely as too much or too little of these metals can have adverse effects on brain function. Recent evidence from nutritional models notes that the hippocampus is particularly vulnerable to Fe and Zn deficiencies. We recently performed a quantitative trait loci (QTL) analysis as a preliminary step in identifying genes that contribute to natural variation in hippocampal Fe, Cu, and Zn content. We used ICP-MS to measure the concentrations of these metals in 120-day-old mice from 30 strains of the BXD/TY panel. The BXD/Ty recombinant inbred strain panel is well-suited for complex trait analysis, as all strains are genotyped with a dense marker set and have been phenotyped extensively for neurobehavioral traits and hippocampal gene expression. We observed a wide-range of hippocampal Fe, Cu, and Zn concentrations across the BXD strains. These concentrations were related to systemic Fe status, but not to Fe, Cu, and Zn elsewhere in the brain. The three metals also showed strong covariance, suggestive of overlap in their regulatory pathways. We identified two QTL, on chromosomes 14 and 9, most strongly associated with Cu but also suggestively associated with Fe (chr. 14) and Zn (chr. 9). We also performed genetic correlational analyses with existing data on these strains and revealed associations with cognitive, anxiety-related, and alcohol-related phenotypes. Covariance of these metals with gene expression is also discussed. This work shows that hippocampal Fe, Cu, and Zn are under polygenic influence and that trace metal regulation is associated with hippocampus-related behaviors. Future work will elucidate the genes underlying the two QTL identified, to aid in identifying homologous genetic variants in human populations, which may underlie altered trace metal homeostasis and related neurological disease., ((c) 2008 Wiley-Liss, Inc.)

Rice is commonly consumed in the Philippines; however the contribution of other foods to the diet is not well defined. Our aim was to determine the nutrient intake and food group intake of Philippine nuns and compare their intakes to the current estimated average requirements (EAR), and food-based recommendations, respectively, and assess any differences in nutrient adequacy and energy intakes between body mass index (BMI) categories. Body weight was assessed at baseline and at nine months; three-day weighed food intakes were recorded once every fortnight (n=187). At baseline, the mean (SD) age and BMI of the women was: 25.0 (4.6) years and 21.8 (17.3) kg/m2, respectively. Over the nine months, women with an underweight (n=46;<18.5 kg/m2) and acceptable BMI (n=132; 18.5-25 kg/m2) lost 5.0 kg (p=0.005) and 1.5 kg (p=0.047), respectively, whereas overweight women maintained their weight. Irrespective of BMI, 98% of women consumed less than the adequate intake for calcium, and no one met the folate EAR. The intake of all food groups (e.g., rice, vegetables, fruit, meat, dairy) was lower than food-based recommendations. It is evident that the nutrient density of the Philippine diet is poor. In order to meet nutrient requirements, it is recommended that all women increase intake of fruits, vegetables, fish, meat and dairy products, to reduce risk of micronutrient deficiencies. For the overweight women, these nutrient dense foods also are recommended, however it is important that they be substituted for energy dense foods to promote weight loss and prevent weight gain.

Iron homeostasis is one of the most critical functions in living systems. Too little iron can lead to anemia and tissue-specific disorders, such as splenomegaly. Excessive systemic iron is characteristic of hemochromatosis and is implicated in the brain in Parkinson's disease. With the exception of some single gene diseases like hemochromatosis, we know little about genetic-based, individual differences in iron-related parameters and their impact on biology. To model genetic control of iron homeostasis, we measured liver, spleen, and plasma iron concentrations, hematocrit and hemoglobin, transferrin saturation, and total iron-binding capacity in several BXD/Ty recombinant inbred mouse strains derived from C57BL/6 and DBA/2 progenitors. At 120 days of age, the animals were killed for iron analysis. All measures showed genetic-based variability consistent with polygenic influence. Analysis of principal components of the seven measures revealed three factors that we named availability, transport, and storage. Quantitative trait loci (QTL) analysis revealed one suggestive QTL on chromosome 5 for availability, two suggestive QTL (one on chromosome 1 and the other on chromosome 7) for transport, and one weak QTL on chromosome 2 for storage. The results show that iron homeostasis is a complex trait and is influenced by multiple genes.

A number of algorithms have been developed to predict the bioavailability of iron from mixed meals and diets, but their direct validity in predicting change in iron status remains questionable. Throughout the course of conducting a large feeding trial in 10 convents in Manila, we collected weighed food intake data (n = 317) and directly compared the performance of these prediction equations to each other and to the change in serum ferritin (SF). Dietary weighed food intakes were measured on d 3 every 2 wk for each woman and iron status determined at baseline, 4.5 mo, and 9 mo. The Monsen and Balintfy equation predicted higher median absorption efficiency (7.3%) than did the equations of Hallberg and Hulthen (6.1%) and Reddy et al. (5.8%). In contrast, the predictions that used the equations of Bhargava et al. (3.8%), Tseng et al. (2.9%), and Du et al. (2.6%) were significantly lower. The iron absorption efficiencies calculated using the Monsen and Balintfy equation correlated with those using the Hallberg and Hulthen equation (r = 0.91, P < 0.001). This slope did not differ from unity, whereas all other equations underestimated iron absorption efficiency relative to Monsen and Balintfy's equation. The median efficiency of absorption, based on change in SF in 114 subjects, was 17.2%, suggesting that these equations underestimate iron absorption. The inhibitory and enhancing factors in the published prediction equations were quantitatively either too large or perhaps too small to correctly predict apparent iron bioavailability over a 9-mo period. The causes of the lack of agreement between change in iron status estimated by SF change and absorption predicted by algorithms are open to discussion and will need to be resolved.

Background: The variability in habitual intakes of most components in the Philippine diet is unknown., Objective: To perform a quantitative evaluation of the traditional Philippine diet using data collected over an extended period of time. We sought to identify seasonal variations and within-subject components of variation in nutrient intake., Methods: A quantitative evaluation of the Philippine diet was conducted in convents in metropolitan Manila as part of an efficacy trial to examine biofortified rice as an approach to improve iron nutritional status. Weighed food intakes were conducted on 54 days in each of more than 300 religious sisters over 9 months in 10 convents. The sisters consumed their habitual diets except for the substitution of one variety of rice for another., Results: More than 40% of calories were derived from rice, with protein from meat and fish comprising 18% of calories. There were significant variations in macronutrient and micronutrient intakes across seasons of the year, with more rice consumed in the wet season and more fruits, eggs, milk, and beverages consumed in the dry season. The day-to-day within-subject variation (CV) in median intake was 23% for energy, 31% for protein, 42% for iron, and 138% for vitamin A., Conclusions: These novel data show that traditional Filipino dietary patterns have substantial individual variation and are inadequate in certain micronutrients. This quantitative evaluation of diet can provide a reference point for dietary adequacy.

Iron deficiency anemia in early childhood causes developmental delays and, very likely, irreversible alterations in neurological functioning. One primary goal for the present study was to determine whether the effects of late gestational iron deficiency on brain monoamine metabolism, iron content, and behavioral phenotypes could be repaired with iron intervention in early lactation. Young pregnant rats were provided iron-deficient or control diets from mid-gestation (G15). At postnatal d 4 (P4), pups from iron-deficient dams were out-fostered either to other ID dams or control dams while pups of control dams were similarly fostered to other control dams. Dietary treatments continued to adulthood (P65) when brain iron and regional monoamines were evaluated. P4 iron repletion normalized body iron status, brain iron concentrations, monoamine concentrations, and monoamine transporter and receptor densities in most brain regions. Dopamine transporter densities in caudate and substantia nigra were lower in ID rats but were normalized with iron repletion. Serotonin transporter levels in most brain regions and open-field exploration were also normalized with iron repletion. The success of this approach of early postnatal iron intervention following iron deficiency in utero contrasts to a relative lack of success when the intervention is performed at weaning. These data suggest that a window of opportunity exists for reversing the detrimental effects of iron deficiency in utero in rats and provides strong support of intervention approaches in humans with iron deficiency during pregnancy.

Background: A number of algorithms have been developed that seek to predict the bioavailability of iron from mixed meals and diets, but their direct validity in predicting change in iron status remains questionable. Throughout the course of conducting a large feeding trial in convents in Manila, we collected weighed food-intake data and have the opportunity to directly compare the performance of these prediction equations., Aims: The specific aims of this particular analysis are as follows: (a) to determine habitual intakes of macro- and micronutrients in religious sisters in Manila, (b) to determine the predicted efficiency of iron absorption from each of the published bioavailability algorithms (Monsen, Hallberg, Reddy, Tseng, Barghava, and Du), and (c) to determine which of these equations best predicts the actual "gain" in iron in religious sisters over the duration of the trial., Design: The efficacy of consuming high-iron rice was tested during a nine-month feeding trial with a double-blinded dietary intervention; these results have been published [7]. Religious sisters living in 10 convents around metropolitan Manila, the Philippines were randomly assigned to consume either high-iron rice (3.21 mg/kg Fe) or a local variety of control rice (0.57 mg/kg Fe) within each convent., Results: Religious sisters in convents consumed a diet that appeared to be typical of habitual intakes of macro-and micronutrients in this part of the Philippines. The analysis of the six equations revealed highly significant differences in predicted efficiency of iron absorption. The Hallberg, Monsen, and Reddy equations all predicted similar median efficiency (6.88, 7.92, and 6.42%). In contrast, Bhargava (4.68%), Tseng (3.23%), and Du (2.92%) were significantly lower. The correlation (r = 0.98) of Monsen to Hallberg was highly significant and the slope was not different than unity. The median efficiency of absorption based on the gain in body iron in 114 subjects over nine months, combined with an estimate of daily iron requirements, was 17.2%. Thus, none of these equations approximated the computed iron absorption based on improvement in serum ferritin and suggests alternative approaches to predicting iron accumulation from diet need to be formulated., Conclusions: Inhibitory factors in the prediction equations either had little effect or had too large of an effect on apparent bioavailability as compared to median absorption over a nine-month period. The causes of the lack of agreement between computed iron gain and predicted absorption are open to discussion and will need to be resolved.

Background: Evidence suggests that brain iron deficiency at any time in life may disrupt metabolic processes and subsequently change cognitive and behavioral functioning. Women of reproductive age are among those most vulnerable to iron deficiency and may be at high risk for cognitive alterations due to iron deficiency., Objective: We aimed to examine the relation between iron status and cognitive abilities in young women., Design: A blinded, placebo-controlled, stratified intervention study was conducted in women aged 18-35 y of varied iron status who were randomly assigned to receive iron supplements or a placebo. Cognition was assessed by using 8 cognitive performance tasks (from Detterman's Cognitive Abilities Test) at baseline (n = 149) and after 16 wk of treatment (n = 113)., Results: At baseline, the iron-sufficient women (n = 42) performed better on cognitive tasks (P = 0.011) and completed them faster (P = 0.038) than did the women with iron deficiency anemia (n = 34). Factors representing performance accuracy and the time needed to complete the tasks by the iron-deficient but nonanemic women (n = 73) were intermediate between the 2 extremes of iron status. After treatment, a significant improvement in serum ferritin was associated with a 5-7-fold improvement in cognitive performance, whereas a significant improvement in hemoglobin was related to improved speed in completing the cognitive tasks., Conclusions: Iron status is a significant factor in cognitive performance in women of reproductive age. Severity of anemia primarily affects processing speed, and severity of iron deficiency affects accuracy of cognitive function over a broad range of tasks. Thus, the effects of iron deficiency on cognition are not limited to the developing brain.

Many genes are likely involved in the control of iron metabolism in brain and in peripheral tissues, and genetically-defined murine strains present the opportunity to investigate genetic variations in iron metabolism. Weanling C57BL/6 (B6) and DBA/2 (D2) mice were divided into two treatment groups receiving distilled water with or without 5000 ppm ferric chloride ad libitum as their sole fluid source for 100 days. Iron overload increased liver, spleen and plasma iron levels in male and female B6 and female D2 mice. In D2 males, liver iron was increased relative to control, but spleen and plasma iron remained unaffected. Brain iron content was not different between control and iron-treated mice in ventral midbrain, caudate, pons or hippocampus, but D2 iron overloaded mice displayed lower iron levels in nucleus accumbens and prefrontal cortex. We conclude that genetic background influences the accumulation of excess iron in the periphery and iron regulation in the central nervous system.

Neurological development and functioning of dopamine (DA) neurotransmission is adversely affected by iron deficiency in early life. Iron-deficient rats demonstrate significant elevations in extracellular DA and a reduction in dopamine transporter (DAT) densities in the caudate putamen and nucleus accumbens. To explore possible mechanisms by which cellular iron concentrations control DAT functioning, endogenous DAT-expressing PC12 cells were used to determine the effect of iron chelation on DAT protein and mRNA expression patterns. In addition, we used human DAT (hDAT)-transfected Neuro2a (N2A) cells to examine DAT degradation and trafficking patterns. A 50 microM treatment for 24 h with the iron chelator, desferrioxamine (DFO), significantly decreased dopamine uptake in a dose-dependent manner, with no apparent change in K(m), in both PC12 and N2A cells. Reduced DA uptake was accompanied by concentration- and time-dependent reductions in total DAT protein levels in both cell lines. Exposure to increasing concentrations of DFO did not significantly alter DAT mRNA in either PC12 or N2A cells. However, DAT degradation rates increased three-fivefold in both cell types exposed to 50 microM DFO for 24 h. Biotinylation studies in N2A cells indicate a more dramatic loss of DAT in the membrane fraction, while OptiPrep fractionation experiments revealed an increase in lysosomal DAT with iron chelation. Inhibition of protein kinase C activation with staurosporin prevented the effect of iron chelation on DAT function, suggesting that in vitro iron chelation affects DAT primarily through the effects on trafficking rather than on synthesis.

Iron deficiency in human infancy reportedly leads to developmental delays and changes in neurobiology that may be irreversible. Using a rodent model, the present study examined whether dietary iron deficiency late in pregnancy and during lactation alters sensorimotor development and brain monoaminergic systems. Rats were assigned to 1 of 4 dietary treatments during gestation and lactation: 1) iron sufficient control; 2) prenatal iron deficiency beginning on gestational d 15 (G15); 3) postnatal iron deficiency beginning on postnatal d 4 (P4); 4) iron deficiency beginning on G15 followed by an iron sufficient diet on P4. Developmental milestones, open field behavior, brain iron and proteins, monoamines, and their transporters were evaluated between P6 and P21. Only G15 iron deficient rats had greater dopaminergic activity than controls as indicated by increased tyrosine hydroxylase levels, phosphorylated tyrosine hydroxylase levels, and cellular dopamine in prefrontal cortex and striatum at P15. These rats also showed delayed eye opening, ear development, and reduced locomotor activity. Iron repletion at P4 returned most measures to control levels by the time of weaning. Postnatal iron deficiency reduced striatal and ventral midbrain iron as well as cellular dopamine levels in prefrontal cortex and striatum at P21. Developmental delays in ear development and achievement in bar holding and surface righting also resulted from postnatal iron deficiency. These results indicate that iron deficiency begun at G15 affects early dopamine neurobiology, the development of specific developmental milestones, and behavior in preweaned rats.

Background: Nutritional surveys use acute phase protein (APP) biomarkers such as C-reactive protein (CRP) and alpha1-acid glycoprotein (AGP) to identify the influence of inflammation on the distribution of iron status biomarkers. Few, however, have examined which biomarker better identifies persons with spurious elevations in iron status markers., Objective: We explored the relations of APP biomarkers to iron-status biomarkers in infants and school-age children., Design: In screening surveys, we identified a sample of African American infants (n = 351) and Guatemalan school-age children (n = 375). We used a common set of APP and iron-status biomarkers to examine the association between the 2 sets of markers (laboratory variables)., Results: The overall prevalence of either inflammation or iron deficiency was <10% in both samples. The log AGP and CRP values were significantly correlated (r = 0.70), but the unexplained variance still was >50%. Serum ferritin-but not transferrin receptor, transferrin receptor index, or serum iron-was related to APP concentrations, but poor positive predictive value (<72%) and low kappa scores were found. Ferritin concentrations >1 geometric SD above the geometric mean were poorly predicted by either elevated AGP or CRP. Qualitative CRP analysis was not effective in identifying persons who had other indications of mild inflammation., Conclusions: These analyses show that a low prevalence of inflammation has little influence on the distribution of ferritin, and 2 common indicators of inflammation do not perform equally well in identifying persons who may have elevations in ferritin due to inflammation.

Background: Iron deficiency anemia (IDA) has been associated with altered cognitive, motor, and social-emotional outcomes in human infants. We recently reported that rats with chronic perinatal IDA, had altered regional brain iron, monoamines, and sensorimotor skill emergence during early development., Objective: To examine the long-term consequences of chronic perinatal IDA on behavior, brain iron and monoamine systems after dietary iron treatment in rats., Methods: Sixty dams were randomly assigned to iron-sufficient (CN) or low-iron (EID) diets during gestation and lactation. Thereafter, all offspring were fed the iron-sufficient diet, assessed for hematology and behavior after weaning and into adulthood and for brain measures as adults (regional brain iron, monoamines, dopamine and serotonin transporters, and dopamine receptor). Behavioral assessments included sensorimotor function, general activity, response to novelty, spatial alternation, and spatial water maze performance., Results: Hematology and growth were similar for EID and CN rats by postnatal day 35. In adulthood, EID thalamic iron content was lower. Monoamines, dopamine transporter, and dopamine receptor concentrations did not differ from CN. EID serotonin transporter concentration was reduced in striatum and related regions. EID rats had persisting sensorimotor deficits (delayed vibrissae-evoked forelimb placing, longer sticker removal time, and more imperfect grooming chains), were more hesitant in novel settings, and had poorer spatial water maze performance than CN. General activity and spatial alternation were similar for EID and CN., Conclusion: Rats that had chronic perinatal IDA showed behavioral impairments that suggest persistent striatal dopamine and hippocampal dysfunction despite normalization of hematology, growth and most brain measures.

Iron deficiency anemia in early childhood is associated with developmental delays and perhaps, irreversible alterations in neurological functioning. The goals were to determine if dietary induced gestational and lactational iron deficiency alters brain monoamine metabolism and behaviors dependent on that neurotransmitter system. Young pregnant rats were provided iron deficient or control diets from early in gestation through to weaning of pups and brain iron concentration, regional monoamine variables and achievement of specific developmental milestones were determined throughout lactation. Despite anemia during lactation, most brain iron concentrations did not fall significantly until P25, and well after significant changes in monoamine levels, transporter levels, and D2R density changed in terminal fields. The changes in D2R density were far smaller than previously observed models that utilized severe dietary restriction during lactation or after weaning. Iron deficient pups had normal birth weight, but were delayed in the attainment of a number of milestones (bar holding, vibrissae-evoked forelimb placing). This approach of iron deficiency in utero and during lactation sufficient to cause moderate anemia but not stunt growth demonstrates that monaminergic metabolism changes occur prior to profound declines in brain iron concentration and is associated with developmental delays. Similar developmental delays in iron deficient human infants suggest to us that alterations in iron status during this developmental period likely affects developing brain monaminergic systems in these infants.

Diurnal effects on motor control are evident in the human disease of Restless Leg Syndrome (RLS), which is purported to be linked to brain iron deficiency as well as alterations in dopaminergic systems. Thus, we explored the relationship between daily rhythms, the onset of motor dysregulation and brain iron deficiency in an animal model of iron deficiency. Male and female weanling Sprague-Dawley rats consuming control (CN) or iron-deficient (ID) diets were examined weekly for acoustic startle response (ASR) and prepulse inhibition (PPI) for a 5-week period. Iron deficiency reduced the magnitude, but not timing, of the ASR at specific time points. ASR was elevated 60% at the onset of the dark cycle relative to the median of the light cycle in male CN and ID rats. The respective elevation was 400% and 150% in female CN and ID rats during the first 2 weeks of testing. The diurnal cycle of ASR response was attenuated by 3 weeks of testing in both dietary treatment groups. PPI was not affected by iron deficiency, sex, diurnal cycle or the interaction between these factors. These results thus demonstrate that iron deficiency moderately alters ASR signaling although the inhibitory pathways of ASR do not appear to be affected.

Iron deficiency is associated with alterations in dopamine and serotonin transporters as well as changes in dopamine receptor (DR) density, monoamine concentrations, and in vivo extracellular contents of monoamines in terminal fields. Human infants with iron deficiency have both delayed maturation as well as lengthened central conduction times in auditory evoked potential studies. The current study utilizes the magnitude of the acoustic startle response (ASR), prepulse inhibition (PPI), and mean latency to maximum startle response (T(max)), to examine the functional integrity of response to environmental cues. Male and female rats consumed iron deficient (ID) or iron adequate (CN) diets from weaning until adulthood. ID rats of both sexes had 20-60% reductions in ASR when compared to CN rats but there was no effect on PPI. T(max) was significantly longer by 10-20% in females, but not males. Dopamine transporter density was significantly lower in putamen, nucleus accumbens, and olfactory tubercle in males, but not female rats while the serotonin transporter was significantly different from control animal density in five of 14 brain regions. Norepinephrine transporter density was lower in the locus ceruleus of ID male rats but was unaffected in ID female rats. Regression modeling of ASR with brain monoamine transporters and receptors showed hematocrit, norepinephrine transporter (NET) in dentate gyrus, and D1R in the nucleus accumbens account for nearly 49% of the variance in ASR. T(max) was not significantly associated with any of the independent variables. We conclude that iron deficiency affects the startle response, but not the inhibitory circuits involved in prepulse inhibition. Importantly, sex also strongly influenced these behavioral responses. Future studies, perhaps pharmacologic in nature, are necessary to ascertain whether iron deficiency modifies the contribution of monoaminergic systems to responses to environmental stimuli.