Manganese in autism spectrum disorder and attention deficit hyperactivity disorder: The state of the art.

The objective of the present narrative review was to synthesize existing clinical and epidemiological findings linking manganese (Mn) exposure biomarkers to autism spectrum disorder (ASD) and attention deficit hyperactivity disorder (ADHD), and to discuss key pathophysiological mechanisms of neurodevelopmental disorders that may be affected by this metal. Existing epidemiological data demonstrated both direct and inverse association between Mn body burden and ASD, or lack of any relationship. In contrast, the majority of studies revealed significantly higher Mn levels in subjects with ADHD, as well as direct relationship between Mn body burden with hyperactivity and inattention scores in children, although several studies reported contradictory results. Existing laboratory studies demonstrated that impaired attention and hyperactivity in animals following Mn exposure was associated with dopaminergic dysfunction and neuroinflammation. Despite lack of direct evidence on Mn-induced neurobiological alterations in patients with ASD and ADHD, a plethora of studies demonstrated that neurotoxic effects of Mn overexposure may interfere with key mechanisms of pathogenesis inherent to these neurodevelopmental disorders. Specifically, Mn overload was shown to impair not only dopaminergic neurotransmission, but also affect metabolism of glutamine/glutamate, GABA, serotonin, noradrenaline, thus affecting neuronal signaling. In turn, neurotoxic effects of Mn may be associated with its ability to induce oxidative stress, apoptosis, and neuroinflammation, and/or impair neurogenesis. Nonetheless, additional detailed studies are required to evaluate the association between environmental Mn exposure and/or Mn body burden and neurodevelopmental disorders at a wide range of concentrations to estimate the potential dose-dependent effects, as well as environmental and genetic factors affecting this association.

Application of Single Cell Gene Expression Technologies to Neurotoxicology.

Neurotoxicological research faces the challenge of linking biological changes resulting from exposures to neuronal function. An additional challenge is understanding cell-type specific differences and selective vulnerabilities of distinct neuronal populations to toxic insults. Single cell RNA-sequencing (scRNA-seq) allows for measurement of the transcriptome of individual cells. This makes it a valuable tool for validating and characterizing cell types present in multicell type samples in complex tissue or cell culture models, but also for understanding how different cell types respond to toxic insults. Pathway analysis of differentially expressed genes can provide in depth insights into underlying cell type-specific mechanisms of neurotoxicity. Toxicological data often has to be translated to outcomes for human health which requires an understanding of inter-species differences. Transcriptomic data aids in understanding these differences, including understanding developmental timelines of different species. We believe that scRNA-seq holds exciting promises for future neurotoxicological research.

Intestinal microbiota protects against methylmercury-induced neurotoxicity.

Methylmercury (MeHg) remains a global public health issue because of its frequent presence in human food sources obtained from the water. The excretion of MeHg in humans occurs slowly with a biological half-time of 32-47 days. Short-term MeHg exposure may cause long-lasting neurotoxicity. The excretion through feces is a major route in the demethylation of MeHg. Accumulating evidence suggests that the intestinal microbiota plays an important role in the demethylation of MeHg, thereby protecting the host from neurotoxic effects. Here, we discuss recent developments on the role of intestinal microbiota in MeHg metabolism, based on in vitro cell culture experiments, experimental animal studies and human investigations. Demethylation by intestinal bacteria is the rate-limiting step in MeHg metabolism and elimination. The identity of bacteria strains responsible for this biotransformation is currently unknown; however, the non-homogenous distribution of intestinal microbiota may lead to different demethylation rates in the intestinal tract. The maintenance of intestinal barrier function by intestinal microbiota may afford protection against MeHg-induced neurotoxicity, which warrant future investigations. We also discuss studies investigating the effects of MeHg exposure on the population structural stability of intestinal microbiota in several host species. Although this is an emerging area in metal toxicity, current research suggests that a change in certain phyla in the intestinal microbiota may indicate MeHg overexposure.

Mitochondria in the Spotlight: C. elegans as a Model Organism to Evaluate Xenobiotic-Induced Dysfunction.

Mitochondria play a crucial role in cellular respiration, ATP production, and the regulation of various cellular processes. Mitochondrial dysfunctions have been directly linked to pathophysiological conditions, making them a significant target of interest in toxicological research. In recent years, there has been a growing need to understand the intricate effects of xenobiotics on human health, necessitating the use of effective scientific research tools. Caenorhabditis elegans (C. elegans), a nonpathogenic nematode, has emerged as a powerful tool for investigating toxic mechanisms and mitochondrial dysfunction. With remarkable genetic homology to mammals, C. elegans has been used in studies to elucidate the impact of contaminants and drugs on mitochondrial function. This review focuses on the effects of several toxic metals and metalloids, drugs of abuse and pesticides on mitochondria, highlighting the utility of C. elegans as a model organism to investigate mitochondrial dysfunction induced by xenobiotics. Mitochondrial structure, function, and dynamics are discussed, emphasizing their essential role in cellular viability and the regulation of processes such as autophagy, apoptosis, and calcium homeostasis. Additionally, specific toxins and toxicants, such as arsenic, cadmium, and manganese are examined in the context of their impact on mitochondrial function and the utility of C. elegans in elucidating the underlying mechanisms. Furthermore, we demonstrate the utilization of C. elegans as an experimental model providing a promising platform for investigating the intricate relationships between xenobiotics and mitochondrial dysfunction. This knowledge could contribute to the development of strategies to mitigate the adverse effects of contaminants and drugs of abuse, ultimately enhancing our understanding of these complex processes and promoting human health.

IGF/mTORC1/S6 Signaling Is Potentiated and Prolonged by Acute Loading of Subtoxicological Manganese Ion.

The insulin-like growth factor (IGF)/insulin signaling (IIS) pathway is involved in cellular responses against intracellular divalent manganese ion (Mn2+) accumulation. As a pathway where multiple nodes utilize Mn2+ as a metallic co-factor, how the IIS signaling patterns are affected by Mn2+ overload is unresolved. In our prior studies, acute Mn2+ exposure potentiated IIS kinase activity upon physiological-level stimulation, indicated by elevated phosphorylation of protein kinase B (PKB, also known as AKT). AKT phosphorylation is associated with IIS activity; and provides direct signaling transduction input for the mammalian target of rapamycin complex 1 (mTORC1) and its downstream target ribosomal protein S6 (S6). Here, to better define the impact of Mn2+ exposure on IIS function, Mn2+-induced IIS activation was evaluated with serial concentrations and temporal endpoints. In the wild-type murine striatal neuronal line STHdh, the acute treatment of Mn2+ with IGF induced a Mn2+ concentration-sensitive phosphorylation of S6 at Ser235/236 to as low as 5 µM extracellular Mn2+. This effect required both the essential amino acids and insulin receptor (IR)/IGF receptor (IGFR) signaling input. Similar to simultaneous stimulation of Mn2+ and IGF, when a steady-state elevation of Mn2+ was established via a 24-h pre-exposure, phosphorylation of S6 also displayed higher sensitivity to sub-cytotoxic Mn2+ when compared to AKT phosphorylation at Ser473. This indicates a synergistic effect of sub-cytotoxic Mn2+ on IIS and mTORC1 signaling. Furthermore, elevated intracellular Mn2+, with both durations, led to a prolonged activation in AKT and S6 upon stimulation. Our data demonstrate that the downstream regulator S6 is a highly sensitive target of elevated Mn2+ and is well below the established acute cytotoxicity thresholds (<50 µM). These findings indicate that the IIS/mTORC1 pathways, in which Mn2+ normally serves as an essential co-factor, are dually responsible for the cellular changes in exposures to real-world Mn2+ concentrations.

Association between Heavy Metals, Metalloids and Metabolic Syndrome: New Insights and Approaches.

Metabolic syndrome (MetS) is an important public health issue that affects millions of people around the world and is growing to pandemic-like proportions. This syndrome is defined by the World Health Organization (WHO) as a pathologic condition characterized by abdominal obesity, insulin resistance, hypertension, and hyperlipidemia. Moreover, the etiology of MetS is multifactorial, involving many environmental factors, including toxicant exposures. Several studies have associated MetS with heavy metals exposure, which is the focus of this review. Environmental and/or occupational exposure to heavy metals are a major risk, contributing to the development of chronic diseases. Of particular note, toxic metals such as mercury, lead, and cadmium may contribute to the development of MetS by altering oxidative stress, IL-6 signaling, apoptosis, altered lipoprotein metabolism, fluid shear stress and atherosclerosis, and other mechanisms. In this review, we discuss the known and potential roles of heavy metals in MetS etiology as well as potential targeted pathways that are associated with MetS. Furthermore, we describe how new approaches involving proteomic and transcriptome analysis, as well as bioinformatic tools, may help bring about an understanding of the involvement of heavy metals and metalloids in MetS.

Developmental Pb exposure increases AD risk via altered intracellular Ca2+ homeostasis in hiPSC-derived cortical neurons.

Exposure to environmental chemicals such as lead (Pb) during vulnerable developmental periods can result in adverse health outcomes later in life. Human cohort studies have demonstrated associations between developmental Pb exposure and Alzheimer's disease (AD) onset in later life which were further corroborated by findings from animal studies. The molecular pathway linking developmental Pb exposure and increased AD risk, however, remains elusive. In this work, we used human iPSC-derived cortical neurons as a model system to study the effects of Pb exposure on AD-like pathogenesis in human cortical neurons. We exposed neural progenitor cells derived from human iPSC to 0, 15, and 50 ppb Pb for 48 h, removed Pb-containing medium, and further differentiated them into cortical neurons. Immunofluorescence, Western blotting, RNA-sequencing, ELISA, and FRET reporter cell lines were used to determine changes in AD-like pathogenesis in differentiated cortical neurons. Exposing neural progenitor cells to low-dose Pb, mimicking a developmental exposure, can result in altered neurite morphology. Differentiated neurons exhibit altered calcium homeostasis, synaptic plasticity, and epigenetic landscape along with elevated AD-like pathogenesis markers, including phosphorylated tau, tau aggregates, and Aß42/40. Collectively, our findings provide an evidence base for Ca dysregulation caused by developmental Pb exposure as a plausible molecular mechanism accounting for increased AD risk in populations with developmental Pb exposure.

The role of hypoxia-inducible factor 1 alpha (HIF-1α) modulation in heavy metal toxicity.

Hypoxia-inducible factor 1 (HIF-1) is an oxygen-sensing transcriptional regulator orchestrating a complex of adaptive cellular responses to hypoxia. Several studies have demonstrated that toxic metal exposure may also modulate HIF-1α signal transduction pathway, although the existing data are scarce. Therefore, the present review aims to summarize the existing data on the effects of toxic metals on HIF-1 signaling and the potential underlying mechanisms with a special focus on prooxidant effect of the metals. The particular effect of metals was shown to be dependent on cell type, varying from down- to up-regulation of HIF-1 pathway. Inhibition of HIF-1 signaling may contribute to impaired hypoxic tolerance and adaptation, thus promoting hypoxic damage in the cells. In contrast, its metal-induced activation may result in increased tolerance to hypoxia through increased angiogenesis, thus promoting tumor growth and contributing to carcinogenic effect of heavy metals. Up-regulation of HIF-1 signaling is mainly observed upon Cr, As, and Ni exposure, whereas Cd and Hg may both stimulate and inhibit HIF-1 pathway. The mechanisms underlying the influence of toxic metal exposure on HIF-1 signaling involve modulation of prolyl hydroxylases (PHD2) activity, as well as interference with other tightly related pathways including Nrf2, PI3K/Akt, NF-κB, and MAPK signaling. These effects are at least partially mediated by metal-induced ROS generation. Hypothetically, maintenance of adequate HIF-1 signaling upon toxic metal exposure through direct (PHD2 modulation) or indirect (antioxidant) mechanisms may provide an additional strategy for prevention of adverse effects of metal toxicity.

Impaired Reorganization of Centrosome Structure Underlies Human Infantile Dilated Cardiomyopathy.

BACKGROUND: During cardiomyocyte maturation, the centrosome, which functions as a microtubule organizing center in cardiomyocytes, undergoes dramatic structural reorganization where its components reorganize from being localized at the centriole to the nuclear envelope. This developmentally programmed process, referred to as centrosome reduction, has been previously associated with cell cycle exit. However, understanding of how this process influences cardiomyocyte cell biology, and whether its disruption results in human cardiac disease, remains unknown. We studied this phenomenon in an infant with a rare case of infantile dilated cardiomyopathy (iDCM) who presented with left ventricular ejection fraction of 18% and disrupted sarcomere and mitochondria structure. METHODS: We performed an analysis beginning with an infant who presented with a rare case of iDCM. We derived induced pluripotent stem cells from the patient to model iDCM in vitro. We performed whole exome sequencing on the patient and his parents for causal gene analysis. CRISPR/Cas9-mediated gene knockout and correction in vitro were used to confirm whole exome sequencing results. Zebrafish and Drosophila models were used for in vivo validation of the causal gene. Matrigel mattress technology and single-cell RNA sequencing were used to characterize iDCM cardiomyocytes further. RESULTS: Whole exome sequencing and CRISPR/Cas9 gene knockout/correction identified RTTN, the gene encoding the centrosomal protein RTTN (rotatin), as the causal gene underlying the patient's condition, representing the first time a centrosome defect has been implicated in a nonsyndromic dilated cardiomyopathy. Genetic knockdowns in zebrafish and Drosophila confirmed an evolutionarily conserved requirement of RTTN for cardiac structure and function. Single-cell RNA sequencing of iDCM cardiomyocytes showed impaired maturation of iDCM cardiomyocytes, which underlie the observed cardiomyocyte structural and functional deficits. We also observed persistent localization of the centrosome at the centriole, contrasting with expected programmed perinuclear reorganization, which led to subsequent global microtubule network defects. In addition, we identified a small molecule that restored centrosome reorganization and improved the structure and contractility of iDCM cardiomyocytes. CONCLUSIONS: This study is the first to demonstrate a case of human disease caused by a defect in centrosome reduction. We also uncovered a novel role for RTTN in perinatal cardiac development and identified a potential therapeutic strategy for centrosome-related iDCM. Future study aimed at identifying variants in centrosome components may uncover additional contributors to human cardiac disease.

Acute manganese exposure impairs glutamatergic function in a young mouse model of Alzheimer's disease.

Manganese (Mn) is an essential metal that serves as a cofactor for metalloenzymes important in moderating oxidative stress and the glutamate/glutamine cycle. Mn is typically obtained through the diet, but toxic overexposure can occur through other environmental or occupational exposure routes such as inhalation. Mn is known to accumulate in the brain following exposure and may contribute to the etiology of neurodegenerative disorders such as Alzheimer's disease (AD) even in the absence of acute neurotoxicity. In the present study, we used in vitro primary cell culture, ex vivo slice electrophysiology and in vivo behavioral approaches to determine if Mn-induced changes in glutamatergic signaling may be altered by genetic risk factors for AD neuropathology. Primary cortical astrocytes incubated with Mn exhibited early rapid clearance of glutamate compared to saline treated astrocytes but decreased clearance over longer time periods, with no effect of the AD genotype. Further, we found that in vivo exposure to a subcutaneous subacute, high dose of Mn as manganese chloride tetrahydrate (3 ×50 mg/kg MnCl2·4(H2O) over 7 days) resulted in increased expression of cortical GLAST protein regardless of genotype, with no changes in GLT-1. Hippocampal long-term potentiation was not altered in APP/PSEN1 mice at this age and neither was it disrupted following Mn exposure. Mn exposure did increase sensitivity to seizure onset following treatment with the excitatory agonist kainic acid, with differing responses between APP/PSEN1 and control mice. These results highlight the sensitivity of the glutamatergic system to Mn exposure. Experiments were performed in young adult APP/PSEN1 mice, prior to cognitive decline or accumulation of hallmark amyloid plaque pathology and following subacute exposure to Mn. The data support a role of Mn in pathophysiology of AD in early stages of the disease and support the need to better understand neurological consequences of Mn exposure in vulnerable populations.

Epigenetics and Methylmercury-Induced Neurotoxicity, Evidence from Experimental Studies.

MeHg is an environmental neurotoxin that can adversely affect the development of the nervous system. The molecular integrity of chromatin in the nucleus is an important target of MeHg. Low levels of MeHg trigger epigenetic mechanisms that may be involved in long-lasting and transgenerational neurotoxicity after exposure. Emerging evidence has shown that these mechanisms include histone modification, siRNA, and DNA methylation. The MeHg-induced inhibition of neurodifferentiation and neurogenesis are mechanistically associated with epigenetic alterations in critical genes, such as neurotrophin brain-derived neurotrophic factor (BDNF). Further, MeHg exposure has been shown to alter the activity and/or expression of the upstream regulators of chromatin structure, including histone deacetylases (HDACs) and DNA methyltransferase (DNMTs), which may trigger permanent alterations in histone modifications and DNA methylation. MeHg-exposure also alters several species of miRNA that are associated with neurodevelopment. Genetic studies in the C. elegans model of MeHg-induced toxicity proposes a potential interplay between exogenous RNAi and antioxidant defense. In this review, we discuss the molecular basis for MeHg exposure-induced alterations in chromatin structure and the roles of histone modifications, siRNA, and DNA methylation in MeHg-induced neurotoxic effects.

Developmental Methylmercury Exposure Induced and Age-Dependent Glutamatergic Neurotoxicity in Caenorhabditis elegans.

Developmental methylmercury (MeHg) exposures cause latent neurotoxic effects in adults; however, the mechanisms underlying the latent neurotoxicity are not fully understood. In the current study, we used C. elegans as an animal model to investigate the latent neurotoxic effects of developmental MeHg exposures on glutamatergic neurons. The young larvae stage 1 worms were exposed to MeHg (0.05 ~ 5 µM) for 48 h. The morphological and behavioral endpoints of glutamatergic neurons were compared when worms reached to adult stages including the young adult stage (day 1 adult) and the old adult stage (day 10 adult). Here, we showed that C. elegans glutamatergic neurons were morphologically intact following low or medium MeHg exposures (0.05 ~ 0.5 µM). The morphological damage of glutamatergic neurons appeared to be pronounced in day 10 adults developmentally exposed to 5 µM MeHg. Behavioral assays also showed an age-dependent latent effect of MeHg. In the nose touch response assay, only day 10 adult worms exhibited a functional decline following prior 5 µM MeHg exposure. Moreover, the disruption of NaCl memory appeared only in day 1 adults following MeHg exposures but not in day 10 adults. The expression of C. elegans homologs of mammalian vesicular glutamate transporter (eat-4) was repressed in day 1 adults, while the glutamate receptor homolog (glr-1) was upregulated in day 10 adults with 5 µM MeHg. In the comparison of age-dependent changes in the insulin-like pathway (daf-2/age-1/daf-16) following MeHg exposures, we showed that the daf-2/age-1/daf-16 pathway was mobilized in day 1 adults but repressed in day 10 adults. Collectively, our data supports a conclusion that MeHg-induced glutamatergic neurotoxicity exhibits an age-dependent pattern, possibly related to the prominent changes in age-dependent modulation in the glutamatergic neurotransmission and metabolic pathways.

Treatment with THI, an inhibitor of sphingosine-1-phosphate lyase, modulates glycosphingolipid metabolism and results therapeutically effective in experimental models of Huntington's disease.

Huntington's disease (HD) is a fatal neurodegenerative disorder with no effective cure currently available. Over the past few years our research has shown that alterations in sphingolipid metabolism represent a critical determinant in HD pathogenesis. In particular, aberrant metabolism of sphingosine-1-phosphate (S1P) has been reported in multiple disease settings, including human postmortem brains from HD patients. In this study, we investigate the potential therapeutic effect of the inhibition of S1P degradative enzyme SGPL1, by the chronic administration of the 2-acetyl-5-tetrahydroxybutyl imidazole (THI) inhibitor. We show that THI mitigated motor dysfunctions in both mouse and fly models of HD. The compound evoked the activation of pro-survival pathways, normalized levels of brain-derived neurotrophic factor, preserved white matter integrity, and stimulated synaptic functions in HD mice. Metabolically, THI restored normal levels of hexosylceramides and stimulated the autophagic and lysosomal machinery, facilitating the reduction of nuclear inclusions of both wild-type and mutant huntingtin proteins.

Caenorhabditis elegans as a Model to Study Manganese-Induced Neurotoxicity.

Caenorhabditis elegans (C. elegans) is a nematode present worldwide. The worm shows homology to mammalian systems and expresses approximately 40% of human disease-related genes. Since Dr. Sydney Brenner first proposed C. elegans as an advantageous experimental worm-model system for genetic approaches, increasing numbers of studies using C. elegans as a tool to investigate topics in several fields of biochemistry, neuroscience, pharmacology, and toxicology have been performed. In this regard, C. elegans has been used to characterize the molecular mechanisms and affected pathways caused by metals that lead to neurotoxicity, as well as the pathophysiological interrelationship between metal exposure and ongoing neurodegenerative disorders. Several toxic metals, such as lead, cadmium, and mercury, are recognized as important environmental contaminants, and their exposure is associated with toxic effects on the human body. Essential elements that are required to maintain cellular homeostasis and normal physiological functions may also be toxic when accumulated at higher concentrations. For instance, manganese (Mn) is a trace essential element that participates in numerous biological processes, such as enzymatic activities, energy metabolism, and maintenance of cell functions. However, Mn overexposure is associated with behavioral changes in C. elegans, which are consistent with the dopaminergic system being the primary target of Mn neurotoxicity. Caenorhabditis elegans has been shown to be an important tool that allows for studies on neuron morphology using fluorescent transgenic worms. Moreover, behavioral tests may be conducted using worms, and neurotransmitter determination and related gene expression are likely to change after Mn exposure. Likewise, mutant worms may be used to study molecular mechanisms in Mn toxicity, as well as the expression of proteins responsible for the biosynthesis, transport, storage, and uptake of dopamine. Furthermore, this review highlights some advantages and limitations of using the experimental model of C. elegans and provides guidance for potential future applications of this model in studies directed toward assessing for Mn neurotoxicity and related mechanisms.

Methylmercury exposure-induced reproductive effects are mediated by dopamine in Caenorhabditis elegans.

Methylmercury (MeHg) is a neurotoxicant that exists in the natural environment, which level can be greatly increased because of human activity. MeHg exposures have the risk of being detrimental to the development of the nervous system. Studies on MeHg toxicity have largely focused on the mechanisms of its neurotoxicity following developmental exposures. Additionally, reproductive toxicity of developmental MeHg exposures has been noted in rodent models. The model organism Caenorhabditis elegans (C. elegans) is a self-fertilizing animal which has a short lifespan around 20 days. Most C. elegans are hermaphrodites that can generate both sperm and oocytes. To investigate the effects of developmental MeHg exposures on the reproduction in C. elegans, larvae stage 1 worms were exposed to MeHg (0, 0.01 or 0.05 µM) for 24 h. The laid eggs and oocytes were compared during each day at adult stages for 6 days. We showed that MeHg exposure significantly induced an increased number of eggs in day 1 adults without an effect on the timing of egg laying or the total number of eggs or oocytes over the 6-day period. The expression of dat-1 and cat-2 and dopamine levels were increased in worms exposed to MeHg. Supplementation with 100 µM dopamine recapitulated the effect of MeHg on the number of eggs present in day 1 adults. Furthermore, the effect of MeHg on the number of eggs was abrogated in the cat-2 mutant worms CB1112. The number of oocytes in the 6-day adult stages was decreased by MeHg in the dat-1 mutant RM2702. MeHg exposures did not change the mating rate or the number of offspring from mating. Combined, these novel findings show that developmental exposure to low levels of MeHg has limited effects on the reproduction in C. elegans. Furthermore, our data support a modulatory role of dopamine in MeHg-induced effects on reproduction in this model system.

The Human LRRK2 Modulates the Age-Dependent Effects of Developmental Methylmercury Exposure in Caenorhabditis elegans.

Methylmercury (MeHg) neurotoxicity exhibits age-dependent effects with a latent and/or persistent neurotoxic effect on aged animals. Individual susceptibility to MeHg neurotoxicity is governed by both exposure duration and genetic factors that can magnify or mitigate the pathologic processes associated with this exposure. We previously showed the G2019S mutation of leucine-rich repeat kinase 2 (LRRK2) modulates the response of worms to high levels of MeHg, mitigating its effect on neuronal morphology in pre-vesicles in cephalic (CEP) dopaminergic neurons. Here we sought to better understand the long-term effects of MeHg exposure at low levels (100-fold lower than that in our previous report) and the modulatory role of the LRRK2 mutation. Worms exposed to MeHg (10 or 50 nM) at the larval stage (L1 stage) were compared at adult stages (young age: day 1 adult; middle age: day 5 adult; old age: day 10 adult) for the swimming speeds in M9 buffer, moving speeds during locomotion on an OP50-seeded plate, and the numbers of CEP dopaminergic pre-vesicles, vesicular structures originating from the dendrites of CEP for exportation of cellular content. In addition, the expression levels of Caenorhabditis elegans homologs of dopamine transporter (dat-1) and tyrosine hydroxylase (cat-2) were also analyzed at these adult stages. Our data showed that swimming speeds were reduced in wild-type worms at the day 10 adult stage at 50 nM MeHg level; yet, reduced swimming speeds were noted in the G2019S LRRK2 transgenic line upon MeHg exposures as low as 10 nM. Compared to locomotor speeds, swimming speeds appear to be more sensitive to the behavioral effects of developmental MeHg exposures, as the locomotor speeds were largely intact and indistinguishable from controls following MeHg exposures. Furthermore, we showed an age-dependent modulation of dat-1 and cat-2 expressions, which could also be modified by the LRRK2 mutation. Although MeHg exposures did not change the number of pre-vesicles, the LRRK2 mutation was associated with increased numbers of pre-vesicles in aged worms. Our data suggest that the latent behavioral effects of MeHg are sensitized by the G2019S LRRK2 mutation, and the underlying mechanism likely involves age-dependent changes in dopaminergic signaling.

Alterations in metal homeostasis occur prior to canonical markers in Huntington disease.

The importance of metal biology in neurodegenerative diseases such as Huntingtin Disease is well documented with evidence of direct interactions between metals such as copper, zinc, iron and manganese and mutant Huntingtin pathobiology. To date, it is unclear whether these interactions are observed in humans, how this impacts other metals, and how mutant Huntington alters homeostatic mechanisms governing levels of copper, zinc, iron and manganese in cerebrospinal fluid and blood in HD patients. Plasma and cerebrospinal fluid from control, pre-manifest, manifest and late manifest HD participants were collected as part of HD-Clarity. Levels of cerebrospinal fluid and plasma copper, zinc, iron and manganese were measured as well as levels of mutant Huntingtin and neurofilament in a sub-set of cerebrospinal fluid samples. We find that elevations in cerebrospinal fluid copper, manganese and zinc levels are altered early in disease prior to alterations in canonical biomarkers of HD although these changes are not present in plasma. We also evidence that CSF iron is elevated in manifest patients. The relationships between plasma and cerebrospinal fluid metal are altered based on disease stage. These findings demonstrate that there are alterations in metal biology selectively in the CSF which occur prior to changes in known canonical biomarkers of disease. Our work indicates that there are pathological changes related to alterations in metal biology in individuals without elevations in neurofilament and mutant Huntingtin.

Genomic, transcriptomic, and metabolomic profiles of hiPSC-derived dopamine neurons from clinically discordant brothers with identical PRKN deletions.

We previously reported on two brothers who carry identical compound heterozygous PRKN mutations yet present with significantly different Parkinson's Disease (PD) clinical phenotypes. Juvenile cases demonstrate that PD is not necessarily an aging-associated disease. Indeed, evidence for a developmental component to PD pathogenesis is accumulating. Thus, we hypothesized that the presence of additional genetic modifiers, including genetic loci relevant to mesencephalic dopamine neuron development, could potentially contribute to the different clinical manifestations of the two brothers. We differentiated human-induced pluripotent stem cells (hiPSCs) derived from the two brothers into mesencephalic neural precursor cells and early postmitotic dopaminergic neurons and performed wholeexome sequencing and transcriptomic and metabolomic analyses. No significant differences in the expression of canonical dopamine neuron differentiation markers were observed. Yet our transcriptomic analysis revealed a significant downregulation of the expression of three neurodevelopmentally relevant cell adhesion molecules, CNTN6, CNTN4 and CHL1, in the cultures of the more severely affected brother. In addition, several HLA genes, known to play a role in neurodevelopment, were differentially regulated. The expression of EN2, a transcription factor crucial for mesencephalic dopamine neuron development, was also differentially regulated. We further identified differences in cellular processes relevant to dopamine metabolism. Lastly, wholeexome sequencing, transcriptomics and metabolomics data all revealed differences in glutathione (GSH) homeostasis, the dysregulation of which has been previously associated with PD. In summary, we identified genetic differences which could potentially, at least partially, contribute to the discordant clinical PD presentation of the two brothers.

Exposing the role of metals in neurological disorders: a focus on manganese.

Metals are ubiquitous chemical entities involved in a myriad of biological processes. Despite their integral role in sustaining life, overexposure can lead to deleterious neurological outcomes posing a public health concern. Excess exposure to metals has been associated with aberrant neurodevelopmental and neurodegenerative diseases and prominently contributes to environmental risk for neurological disorders. Here, we use manganese (Mn) to exemplify the gap in our understanding of the mechanisms behind acute metal toxicity and their relationship to chronic toxicity and disease. This challenge frustrates understanding of how individual exposure histories translate into preventing and treating brain diseases from childhood through old age. We discuss ways to enhance the predictive value of preclinical models and define mechanisms of chronic, persistent, and latent neurotoxicity.