Association between lead and circulating markers of inflammation among traffic enforcers in Metro Manila, Philippines: the MMDA traffic enforcer's health study.

PURPOSE: Several epidemiological studies have linked lead (Pb) exposure to induced oxidative stress and the promotion of inflammatory response. We performed a within-subjects study (repeated measures study) to evaluate the relationship between the concentration of blood lead (B-Pb) and toenail lead (T-Pb) and circulating markers of inflammation. METHODS: We evaluated the associations between B-Pb concentrations and T-Pb concentrations and circulating markers of inflammation, soluble intracellular adhesion molecule-1 (s-ICAM-1), soluble vascular adhesion molecule-1 (s-VCAM-1), and high-sensitivity C-reactive protein (hs-CRP) on 158 traffic enforcers from the Metropolitan Manila Development Authority (MMDA) traffic enforcer's health study. Linear mixed-effects models with random subject-specific intercepts were fitted to estimate the association between B-Pb and T-Pb exposure and circulating markers of inflammation, adjusting for confounding factors. RESULTS: Traffic enforcers were middle-aged men (89.4%) with a mean age (± SD) of 37.1 years ± 8.9 years and had a total of 293 valid markers of inflammation measurements. B-Pb concentration was related to increased hs-CRP levels. A 10% increase in B-Pb was associated with a 5.7% increase in hs-CRP level [95% confidence interval (95% CI): 1.3-10.1]. However, B-Pb was not associated with s-ICAM-1 and s-VCAM-1. Furthermore, no associations were observed between T-Pb and all the circulating markers of inflammation. CONCLUSIONS: Low-level B-Pb may increase hs-CRP among traffic enforcers. Moreover, the study suggests that Pb via the oxidative and inflammation pathways may have an essential role in the development of cardiovascular disease. Furthermore, MMDA and the Department of Labor and Employment can use our study's findings as evidence to conduct routine screening of blood heavy metals, especially Pb, among MMDA and other traffic enforcers as part of their yearly medical examination.

Genomic Basis for Individual Differences in Susceptibility to the Neurotoxic Effects of Diesel Exhaust.

Air pollution is a known environmental health hazard. A major source of air pollution includes diesel exhaust (DE). Initially, research on DE focused on respiratory morbidities; however, more recently, exposures to DE have been associated with neurological developmental disorders and neurodegeneration. In this study, we investigated the effects of sub-chronic inhalation exposure to DE on neuroinflammatory markers in two inbred mouse strains and both sexes, including whole transcriptome examination of the medial prefrontal cortex. We exposed aged male and female C57BL/6J (B6) and DBA/2J (D2) mice to DE, which was cooled and diluted with HEPA-filtered compressed air for 2 h per day, 5 days a week, for 4 weeks. Control animals were exposed to HEPA-filtered air on the same schedule as DE-exposed animals. The prefrontal cortex was harvested and analyzed for proinflammatory cytokine gene expression (Il1ß, Il6, Tnfα) and transcriptome-wide response by RNA-seq. We observed differential cytokine gene expression between strains and sexes in the DE-exposed vs. control-exposed groups for Il1ß, Tnfα, and Il6. For RNA-seq, we identified 150 differentially expressed genes between air and DE treatment related to natural killer cell-mediated cytotoxicity per Kyoto Encyclopedia of Genes and Genomes pathways. Overall, our data show differential strain-related effects of DE on neuroinflammation and neurotoxicity and demonstrate that B6 are more susceptible than D2 to gene expression changes due to DE exposures than D2. These results are important because B6 mice are often used as the default mouse model for DE studies and strain-related effects of DE neurotoxicity warrant expanded studies.

Genetic differences in ethanol consumption: effects on iron, copper, and zinc regulation in mouse hippocampus.

One common characteristic of neurodegenerative diseases is dysregulation of iron, usually with observed increases in its concentration in various regions. Heavy alcohol consumption is believed to contribute to such iron dysregulation in the brain with accompanying dementia. To examine this effect and related genetic-based individual differences in an animal model, we subjected female mice from 12 BXD recombinant inbred strains to 16 weeks of alcohol consumption using the drinking in the dark (DID) method. Daily consumption was recorded and at the end of 16 weeks hippocampus tissues harvested. Concentrations of iron, copper and zinc were measured using X-ray fluorescence technology. The results showed that, DID increased iron overall across all strains, ranging from 3 to 68%. Copper and Zinc both decreased, ranging from 0.4-42 and 5-35% respectively. Analysis of variance revealed significant strain by treatment interactions for all three metals. Additionally, in the DID group, we observed strain differences in reduction of hippocampus mass. These findings are particularly interesting to us because high alcohol consumption in humans has been associated with neurodegeneration and dementia related to disruption of iron regulation. The findings of alcohol consumption associated decreases in copper and zinc are novel. The role of copper regulation and neurological function related to alcohol consumption is as yet largely unexplored. The role of zinc is better known as a neuromodulator in the hippocampus and appears to be protective against neurological damage. It would seem then, that the alcohol-related decrease in zinc in the hippocampus would be of concern and warrants further study.

Ventral striatum regulates behavioral response to ethanol and MDMA combination.

Our previous studies consistently showed that MDMA-induced locomotor hyperactivity is dramatically increased by coadministration of ethanol (EtOH) in rats, indicating possible potentiation of MDMA abuse liability. Thus, we aimed to identify the brain region(s) and neuropharmacological substrates involved in the pharmacodynamics of this potentiation. We first showed that potentiation of locomotor activity by the combination of ip administration of EtOH (1.5 g/kg) and MDMA (6.6 mg/kg) is delay sensitive and maximal when both drugs are injected simultaneously. Then, we used the 2-deoxyglucose quantitative autoradiography technique to assess the impact of EtOH, MDMA, or their combination on local cerebral metabolic rates for glucose (CMRglcs). We showed a specific metabolic activation in the ventral striatum (VS) under MDMA + EtOH versus MDMA or EtOH alone. We next tested if reversible (tetrodotoxin, TTX) or permanent (6-hydrodoxyopamine, 6-OHDA) lesion of the VS could affect locomotor response to MDMA and MDMA + EtOH. Finally, we blocked dopamine D1 or glutamate NMDA receptors in the VS and measured the effects of MDMA and MDMA + EtOH on locomotor activity. We showed that bilateral reversible inactivation (TTX) or permanent lesion (6-OHDA) of the VS prevented the potentiation by EtOH of MDMA-induced locomotor hyperactivity. Likewise, blockade of D1 or NMDA receptors in the VS also reduced the potentiation of MDMA locomotor activity by EtOH. These data indicate that dopamine D1 and glutamate NMDA receptor-driven mechanisms in the VS play a key role in the pharmacodynamics of EtOH-induced potentiation of the locomotor effects of MDMA.

Genome-wide transcriptome architecture in a mouse model of Gulf War Illness.

Gulf War Illness (GWI) is thought to be a chronic neuroimmune disorder caused by in-theater exposure during the 1990-1991 Gulf War. There is a consensus that the illness is caused by exposure to insecticides and nerve agent toxicants. However, the heterogeneity in both development of disease and clinical outcomes strongly suggests a genetic contribution. Here, we modeled GWI in 30 BXD recombinant inbred mouse strains with a combined treatment of corticosterone (CORT) and diisopropyl fluorophosphate (DFP). We quantified transcriptomes from 409 prefrontal cortex samples. Compared to the untreated and DFP treated controls, the combined treatment significantly activated pathways such as cytokine-cytokine receptor interaction and TNF signaling pathway. Protein-protein interaction analysis defined 6 subnetworks for CORT + DFP, with the key regulators being Cxcl1, Il6, Ccnb1, Tnf, Agt, and Itgam. We also identified 21 differentially expressed genes having significant QTLs related to CORT + DFP, but without evidence for untreated and DFP treated controls, suggesting regions of the genome specifically involved in the response to CORT + DFP. We identified Adamts9 as a potential contributor to response to CORT + DFP and found links to symptoms of GWI. Furthermore, we observed a significant effect of CORT + DFP treatment on the relative proportion of myelinating oligodendrocytes, with a QTL on Chromosome 5. We highlight three candidates, Magi2, Sema3c, and Gnai1, based on their high expression in the brain and oligodendrocyte. In summary, our results show significant genetic effects of the CORT + DFP treatment, which mirrors gene and protein expression changes seen in GWI sufferers, providing insight into the disease and a testbed for future interventions.

Impact of Genetic Variation on Stress-Related Ethanol Consumption.

BACKGROUND: The effect of stress on alcohol consumption in humans is highly variable, and the underlying processes are not yet understood. Attempts to model a positive relationship between stress and increased ethanol (EtOH) consumption in animals have been only modestly successful. Our hypothesis is that individual differences in stress effects on EtOH consumption are mediated by genetics. METHODS: We measured alcohol consumption, using the drinking-in-the-dark (DID) paradigm in females from 2 inbred mouse strains, C57BL/6J (B6) and DBA/2J (D2), and 35 of their inbred progeny (the BXD family). A control group was maintained in normal housing and a stress group was exposed to chronic mild stress (CMS), consisting of unpredictable stressors over 7 weeks. These included predator, social, and environmental perturbations. Alcohol intake was measured over 16 weeks in both groups during baseline (preceding 5-week period), CMS (intervening 7-week period), and post-CMS (final 4-week period). RESULTS: We detected a strong effect of CMS on alcohol intake. A few strains demonstrated CMS-related increased alcohol consumption; however, most showed decreased intake. We identified 1 nearly significant quantitative trait locus on chromosome 5 that contains the neuronal nitric oxide synthase gene (Nos1). The expression of Nos1 is frequently changed following alcohol exposure, and variants in this gene segregating among the BXD population may modulate alcohol intake in response to stress. CONCLUSIONS: The results we present here represent the first study to combine chronic stress and alcohol consumption in a genetic reference population of mice. Differences in susceptibility to the effects of stressful environments vis-à-vis alcohol use disorders would suggest that the differences have at least some basis in genetic constitution. We have also nominated a likely candidate gene underlying the large individual differences in effects of stress on alcohol consumption.

Orbitofrontal Neuroadaptations and Cross-Species Synaptic Biomarkers in Heavy-Drinking Macaques.

Cognitive impairments, uncontrolled drinking, and neuropathological cortical changes characterize alcohol use disorder. Dysfunction of the orbitofrontal cortex (OFC), a critical cortical subregion that controls learning, decision-making, and prediction of reward outcomes, contributes to executive cognitive function deficits in alcoholic individuals. Electrophysiological and quantitative synaptomics techniques were used to test the hypothesis that heavy drinking produces neuroadaptations in the macaque OFC. Integrative bioinformatics and reverse genetic approaches were used to identify and validate synaptic proteins with novel links to heavy drinking in BXD mice. In drinking monkeys, evoked firing of OFC pyramidal neurons was reduced, whereas the amplitude and frequency of postsynaptic currents were enhanced compared with controls. Bath application of alcohol reduced evoked firing in neurons from control monkeys, but not drinking monkeys. Profiling of the OFC synaptome identified alcohol-sensitive proteins that control glutamate release (e.g., SV2A, synaptogyrin-1) and postsynaptic signaling (e.g., GluA1, PRRT2) with no changes in synaptic GABAergic proteins. Western blot analysis confirmed the increase in GluA1 expression in drinking monkeys. An exploratory analysis of the OFC synaptome found cross-species genetic links to alcohol intake in discrete proteins (e.g., C2CD2L, DIRAS2) that discriminated between low- and heavy-drinking monkeys. Validation studies revealed that BXD mouse strains with the D allele at the C2cd2l interval drank less alcohol than B allele strains. Thus, by profiling of the OFC synaptome, we identified changes in proteins controlling glutamate release and postsynaptic signaling and discovered several proteins related to heavy drinking that have potential as novel targets for treating alcohol use disorder.SIGNIFICANCE STATEMENT Clinical research identified cognitive deficits in alcoholic individuals as a risk factor for relapse, and alcoholic individuals display deficits on cognitive tasks that are dependent upon the orbitofrontal cortex (OFC). To identify neurobiological mechanisms that underpin OFC dysfunction, this study used electrophysiology and integrative synaptomics in a translational nonhuman primate model of heavy alcohol consumption. We found adaptations in synaptic proteins that control glutamatergic signaling in chronically drinking monkeys. Our functional genomic exploratory analyses identified proteins with genetic links to alcohol and cocaine intake across mice, monkeys, and humans. Future work is necessary to determine whether targeting these novel targets reduces excessive and harmful levels of alcohol drinking.

The genetic dissection of Myo7a gene expression in the retinas of BXD mice.

Purpose: Usher syndrome (US) is characterized by a loss of vision due to retinitis pigmentosa (RP) and deafness. US has three clinical subtypes, but even within each subtype, the severity varies. Myosin VIIA, coded by Myo7a, has been identified as one of the causal genes of US. This study aims to identify pathways and other genes through which Myo7a interacts to affect the presentation of US symptoms. Methods: In this study, we used the retinal tissue of BXD recombinant inbred (RI) mice to examine the expression of Myo7a and perform genetic mapping. Expression quantitative trait locus (eQTL), single nucleotide polymorphism (SNP), and gene correlation analysis were performed using GeneNetwork. Gene set enrichment analysis was performed using WebGestalt, and gene network construction was performed using the Gene Cohesion Analysis Tool. Results: We found Myo7a to be cis-regulated, with varied levels of expression across BXD strains. Here, we propose a genetic network with 40 genes whose expression is highly correlated with Myo7a. Among these genes, six have been linked to retinal diseases, three to deafness, and five share a transcription factor with Myo7a. Gene ontology and pathway analysis revealed a strong connection among ion channel activity, Myo7a, and US. Conclusions: Although Myo7a is a causal gene of US type I, this gene works with many other genes and pathways to affect the severity of US. Many of the genes found in the genetic network, pathways, and gene ontology categories of Myo7a are related to either deafness or blindness. Further investigation is needed to examine the specific relationships between these genes, which may assist in the treatment of US.

A novel heat shock protein alpha 8 (Hspa8) molecular network mediating responses to stress- and ethanol-related behaviors.

Genetic differences mediate individual differences in susceptibility and responses to stress and ethanol, although, the specific molecular pathways that control these responses are not fully understood. Heat shock protein alpha 8 (Hspa8) is a molecular chaperone and member of the heat shock protein family that plays an integral role in the stress response and that has been implicated as an ethanol-responsive gene. Therefore, we assessed its role in mediating responses to stress and ethanol across varying genetic backgrounds. The hippocampus is an important mediator of these responses, and thus, was examined in the BXD family of mice in this study. We conducted bioinformatic analyses to dissect genetic factors modulating Hspa8 expression, identify downstream targets of Hspa8, and examined its role. Hspa8 is trans-regulated by a gene or genes on chromosome 14 and is part of a molecular network that regulates stress- and ethanol-related behaviors. To determine additional components of this network, we identified direct or indirect targets of Hspa8 and show that these genes, as predicted, participate in processes such as protein folding and organic substance metabolic processes. Two phenotypes that map to the Hspa8 locus are anxiety-related and numerous other anxiety- and/or ethanol-related behaviors significantly correlate with Hspa8 expression. To more directly assay this relationship, we examined differences in gene expression following exposure to stress or alcohol and showed treatment-related differential expression of Hspa8 and a subset of the members of its network. Our findings suggest that Hspa8 plays a vital role in genetic differences in responses to stress and ethanol and their interactions.

The Defecation Index as a Measure of Emotionality: Questions Raised by HPA Axis and Prolactin Response to Stress in the Maudsley Model.

The Maudsley Reactive and Maudsley Non-Reactive strains have been selectively bred for differences in open-field defecation (OFD), a putative index of stress. We investigated whether variations in the hypothalamic-pituitary-adrenal (HPA) axis are correlated with strain differences in OFD in the Maudsley model. Exposure to the open-field test did not result in increases in ACTH in male rats of either strain and there were no strain differences in the large increases in ACTH and corticosteroid that occurred in response to intermittent footshock. Parallel studies of prolactin showed that Maudsley Reactive rats had greater response to the open-field and to footshock than Maudsley Non-Reactive rats. The lack of correlation between strain differences in OFD and reactivity of the HPA axis is consistent with the idea that HPA response to stress and OFD reflect the output of different neural systems and that individual differences in emotionality, as indexed by OFD do not influence other measures of stress-reactivity in a simple manner, if at all. The reactivity of the prolactin system to the open-field test and lack of response of ACTH to the same situation is consistent with the idea that the prolactin system is sensitive to lower levels of stress than the HPA axis, a finding at variance with the presumed extreme sensitivity of the latter system. Earlier comparisons of the HPA axis in these strains implicate local factors such as neuropeptide-Y peptide in the adrenal in attenuating the response of the adrenal cortex to ACTH and hints at the complexity of regulation of the HPA axis.

The perplexing paradox of paraquat: the case for host-based susceptibility and postulated neurodegenerative effects.

Paraquat is an herbicide used extensively in agriculture and has also been proposed to be a risk factor for Parkinson's disease. To date, experimental, clinical, and epidemiological data on paraquat neurotoxicity have been equivocal. In this short review, we discuss some technical and biological mechanisms that contribute to inconsistencies regarding paraquat neurotoxicity. We hypothesize that individual genetic variations in susceptibility generate major differences in neurotoxic risk and functional outcome. Identifying these heritable sources of variation in host susceptibility, and their role in complex gene-environment interactions, is crucial to identify risk biomarkers and to devise better prevention and treatment for those exposed to paraquat and other potential neurotoxicants.

Biomimetic nanoparticles with enhanced rapamycin delivery for autism spectrum disorder treatment via autophagy activation and oxidative stress modulation.

Rationale: Autism spectrum disorder (ASD) represents a complex neurodevelopmental condition lacking specific pharmacological interventions. Given the multifaced etiology of ASD, there exist no effective treatment for ASD. Rapamycin (RAPA) can activate autophagy by inhibiting the mTOR pathway and has exhibited promising effects in treating central nervous system disorders; however, its limited ability to cross the blood-brain barrier (BBB) has hindered its clinical efficacy, leading to substantial side effects. Methods: To address this challenge, we designed a drug delivery system utilizing red blood cell membrane (CM) vesicles modified with SS31 peptides to enhance the brain penetration of RAPA for the treatment of autism. Results: The fabricated SCM@RAPA nanoparticles, with an average diameter of 110 nm, exhibit rapid release of RAPA in a pathological environment characterized by oxidative stress. In vitro results demonstrate that SCM@RAPA effectively activate cellular autophagy, reduce intracellular ROS levels, improve mitochondrial function, thereby ameliorating neuronal damage. SS31 peptide modification significantly enhances the BBB penetration and rapid brain accumulation of SCM@RAPA. Notably, SCM@RAPA nanoparticles demonstrate the potential to ameliorate social deficits, improve cognitive function, and reverse neuronal impairments in valproic acid (VPA)-induced ASD models. Conclusions: The therapeutic potential of SCM@RAPA in managing ASD signifies a paradigm shift in autism drug treatment, holding promise for clinical interventions in diverse neurological conditions.

Epigenetic analysis in a murine genetic model of Gulf War illness.

Of the nearly 1 million military personnel who participated in the 1990-1991 Gulf War, between 25% and 35% became ill with what now is referred to as Gulf War Illness (GWI) by the Department of Defense. Symptoms varied from gastrointestinal distress to lethargy, memory loss, inability to concentrate, depression, respiratory, and reproductive problems. The symptoms have persisted for 30 years in those afflicted but the basis of the illness remains largely unknown. Nerve agents and other chemical exposures in the war zone have been implicated but the long-term effects of these acute exposures have left few if any identifiable signatures. The major aim of this study is to elucidate the possible genomic basis for the persistence of symptoms, especially of the neurological and behavioral effects. To address this, we performed a whole genome epigenetic analysis of the proposed cause of GWI, viz., exposure to organophosphate neurotoxicants combined with high circulating glucocorticoids in two inbred mouse strains, C57BL/6J and DBA/2J. The animals received corticosterone in their drinking water for 7 days followed by injection of diisopropylfluorophosphate, a nerve agent surrogate. Six weeks after DFP injection, the animals were euthanized and medial prefrontal cortex harvested for genome-wide DNA methylation analysis using high-throughput sequencing. We observed 67 differentially methylated genes, notably among them, Ttll7, Akr1c14, Slc44a4, and Rusc2, all related to different symptoms of GWI. Our results support proof of principle of genetic differences in the chronic effects of GWI-related exposures and may reveal why the disease has persisted in many of the now aging Gulf War veterans.

Systems genetic analysis of the effects of iron deficiency in mouse brain.

Iron regulation in the brain is both necessary and highly complex. Too little or too much iron can compromise neurological function, yet we still do not know all of the regulatory processes. In our research, we seek to identify genes and gene networks underlying individual differences in brain iron regulation. To this end, we fed mice from 20+ inbred strains a diet low in iron from weaning to 4 months of age. At sacrifice, we measured iron content in the ventral midbrain (VMB). The VMB contains the substantia nigra, a region particularly vulnerable to iron imbalance. The results showed high, inter-strain variability in dietary iron reduction, from almost no loss to more than 40 % vs. control. When we performed quantitative trait loci (QTL) analysis, we observed a significant area on chromosome 2. Within this QTL, we selected glial high-affinity glutamate transporter 1 (Glt1) as the leading candidate. Expression of this gene is both correlated with VMB iron and is also cis-modulated by local sequence variants that segregate in the BXD family. VMB expression differences of Glt1 in six strains covary with differential susceptibility to VMB iron loss.

Systems genetic analysis of multivariate response to iron deficiency in mice.

The aim of this study was to identify genes that influence iron regulation under varying dietary iron availability. Male and female mice from 20+ BXD recombinant inbred strains were fed iron-poor or iron-adequate diets from weaning until 4 mo of age. At death, the spleen, liver, and blood were harvested for the measurement of hemoglobin, hematocrit, total iron binding capacity, transferrin saturation, and liver, spleen and plasma iron concentration. For each measure and diet, we found large, strain-related variability. A principal-components analysis (PCA) was performed on the strain means for the seven parameters under each dietary condition for each sex, followed by quantitative trait loci (QTL) analysis on the factors. Compared with the iron-adequate diet, iron deficiency altered the factor structure of the principal components. QTL analysis, combined with PosMed (a candidate gene searching system) published gene expression data and literature citations, identified seven candidate genes, Ptprd, Mdm1, Picalm, lip1, Tcerg1, Skp2, and Frzb based on PCA factor, diet, and sex. Expression of each of these is cis-regulated, significantly correlated with the corresponding PCA factor, and previously reported to regulate iron, directly or indirectly. We propose that polymorphisms in multiple genes underlie individual differences in iron regulation, especially in response to dietary iron challenge. This research shows that iron management is a highly complex trait, influenced by multiple genes. Systems genetics analysis of iron homeostasis holds promise for developing new methods for prevention and treatment of iron deficiency anemia and related diseases.

Brain-iron deficiency models of restless legs syndrome.

Restless legs syndrome (RLS) is a common sensorimotor disorder for which two main pathological elements are fairly well accepted: Brain iron deficiency (BID) and an altered dopaminergic system. The ability to better understand the causal and consequential factors related to these two pathological elements, would hopefully lead to the development of better therapeutic strategies for treating, if not curing, this disease. The current understanding of the relationship between these two elements is that BID leads to some alterations in neurotransmitters and subsequent changes in the dopaminergic system. Therefore, rodent models based on diet-induced BID, provide a biological substrate to understand the consequences of BID on dopaminergic pathway and on alternative pathways that may be involved. In this review, we present the current research on dopaminergic changes found in RLS subjects and compare that to what is seen in the BID rodent model to provide a validation of the BID rodent model. We also demonstrate the ability of the BID model to predict changes in other neurotransmitter systems and how that has led to new treatment options. Finally, we will present arguments for the utility of recombinant inbred mouse strains that demonstrate natural variation in brain iron, to explore the genetic basis of altered brain iron homeostasis as a model to understand why in idiopathic RLS there can exist a BID despite normal peripheral iron store. This review is the first to draw on 25 years of human and basic research into the pathophysiology of RLS to provide strong supportive data as to the validity of BID model as an important translational model of the disease. As we will demonstrate here, not only does the BID model closely and accurately mimic what we see in the dopaminergic system of RLS, it is the first model to identify alternative systems from which new treatments have recently been developed.

Genetic analysis of iron-deficiency effects on the mouse spleen.

Iron homeostasis is crucial to many biological functions in nearly all organisms, with roles ranging from oxygen transport to immune function. Disruption of iron homeostasis may result in iron overload or iron deficiency. Iron deficiency may have severe consequences, including anemia or changes in immune or neurotransmitter systems. Here we report on the variability of phenotypic iron tissue loss and splenomegaly and the associated quantitative trait loci (QTLs), polymorphic areas in the mouse genome that may contain one or more genes that play a role in spleen iron concentration or spleen weight under each dietary treatment. Mice from 26 BXD/Ty recombinant inbred strains, including the parent C57BL/6 and DBA/2 strains, were randomly assigned to adequate iron or iron-deficient diets at weaning. After 120 days, splenomegaly was measured by spleen weight, and spleen iron was assessed using a modified spectrophotometry technique. QTL analyses and gene expression comparisons were then conducted using the WebQTL GeneNetwork. We observed wide, genetic-based variability in splenomegaly and spleen iron loss in BXD/Ty recombinant inbred strains fed an iron-deficient diet. Moreover, we identified several suggestive QTLs. Matching our QTLs with gene expression data from the spleen revealed candidate genes. Our work shows that individual differences in splenomegaly response to iron deficiency are influenced at least partly by genetic constitution. We propose mechanistic hypotheses by which splenomegaly may result from iron deficiency.

Genetic Modulation of Initial Sensitivity to Δ9-Tetrahydrocannabinol (THC) Among the BXD Family of Mice.

Cannabinoid receptor 1 activation by the major psychoactive component in cannabis, Δ9-tetrahydrocannabinol (THC), produces motor impairments, hypothermia, and analgesia upon acute exposure. In previous work, we demonstrated significant sex and strain differences in acute responses to THC following administration of a single dose (10 mg/kg, i.p.) in C57BL/6J (B6) and DBA/2J (D2) inbred mice. To determine the extent to which these differences are heritable, we quantified acute responses to a single dose of THC (10 mg/kg, i.p.) in males and females from 20 members of the BXD family of inbred strains derived by crossing and inbreeding B6 and D2 mice. Acute THC responses (initial sensitivity) were quantified as changes from baseline for: 1. spontaneous activity in the open field (mobility), 2. body temperature (hypothermia), and 3. tail withdrawal latency to a thermal stimulus (antinociception). Initial sensitivity to the immobilizing, hypothermic, and antinociceptive effects of THC varied substantially across the BXD family. Heritability was highest for mobility and hypothermia traits, indicating that segregating genetic variants modulate initial sensitivity to THC. We identified genomic loci and candidate genes, including Ndufs2, Scp2, Rps6kb1 or P70S6K, Pde4d, and Pten, that may control variation in THC initial sensitivity. We also detected strong correlations between initial responses to THC and legacy phenotypes related to intake or response to other drugs of abuse (cocaine, ethanol, and morphine). Our study demonstrates the feasibility of mapping genes and variants modulating THC responses in the BXDs to systematically define biological processes and liabilities associated with drug use and abuse.

Paraquat Toxicogenetics: Strain-Related Reduction of Tyrosine Hydroxylase Staining in Substantia Nigra in Mice.

Paraquat (PQ) is a putative risk factor for the development of sporadic Parkinson's disease. To model a possible genetic basis for individual differences in susceptibility to exposure to PQ, we recently examined the effects of paraquat on tyrosine hydroxylase (TH)-containing neurons in the substantia nigra pars compacta (SNc) of six members of the BXD family of mice (n = 2-6 per strain). We injected males with 5 mg/kg paraquat weekly three times. The density of TH+ neurons counted by immunocytochemistry at 200x in eight or more sections through the SNc is reduced in five of the six strains relative to control (N = 4 ± 2 mice per strain). TH+ loss ranged from 0 to 20% with an SEM of 1%. The heritability was estimated using standard ANOVA and jackknife resampling and is 0.37 ± 0.05 in untreated animals and 0.47 ± 0.04 in treated animals. These results demonstrate genetic modulation and GxE variation in susceptibility to PQ exposure and the loss of TH staining in the substantia nigra.