Publications by authors named "Richard Nass"

25 Publications

  • Page 1 of 1

The role of charge in the toxicity of polymer-coated cerium oxide nanomaterials to Caenorhabditis elegans.

Comp Biochem Physiol C Toxicol Pharmacol 2017 10 6;201:1-10. Epub 2017 Sep 6.

Department of Plant and Soil Sciences, University of Kentucky, Lexington, KY, United States. Electronic address:

This study examined the impact of surface functionalization and charge on ceria nanomaterial toxicity to Caenorhabditis elegans. The examined endpoints included mortality, reproduction, protein expression, and protein oxidation profiles. Caenorhabditis elegans were exposed to identical 2-5nm ceria nanomaterial cores which were coated with cationic (diethylaminoethyl dextran; DEAE), anionic (carboxymethyl dextran; CM), and non-ionic (dextran; DEX) polymers. Mortality and reproductive toxicity of DEAE-CeO was approximately two orders of magnitude higher than for CM-CeO or DEX-CeO. Two-dimensional gel electrophoresis with orbitrap mass spectrometry identification revealed changes in the expression profiles of several mitochondrial-related proteins and proteins that are expressed in the C. elegans intestine. However, each type of CeO material exhibited a distinct protein expression profile. Increases in protein carbonyls and protein-bound 3-nitrotyrosine were also observed for some proteins, indicating oxidative and nitrosative damage. Taken together the results indicate that the magnitude of toxicity and toxicity pathways vary greatly due to surface functionalization of CeO nanomaterials.
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http://dx.doi.org/10.1016/j.cbpc.2017.08.009DOI Listing
October 2017

G protein-coupled receptor kinase-2 (GRK-2) regulates serotonin metabolism through the monoamine oxidase AMX-2 in .

J Biol Chem 2017 04 17;292(14):5943-5956. Epub 2017 Feb 17.

From the Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107,

G protein-coupled receptors (GPCRs) regulate many animal behaviors. GPCR signaling is mediated by agonist-promoted interactions of GPCRs with heterotrimeric G proteins, GPCR kinases (GRKs), and arrestins. To further elucidate the role of GRKs in regulating GPCR-mediated behaviors, we utilized the genetic model system Our studies demonstrate that loss-of-function strains are egg laying-defective and contain low levels of serotonin (5-HT) and high levels of the 5-HT metabolite 5-hydroxyindole acetic acid (5-HIAA). The egg laying defect could be rescued by the expression of wild type but not by catalytically inactive or by the selective expression of in hermaphrodite-specific neurons. The addition of 5-HT or inhibition of 5-HT metabolism also rescued the egg laying defect. Furthermore, we demonstrate that AMX-2 is the primary monoamine oxidase that metabolizes 5-HT in , and we also found that loss-of-function strains have abnormally high levels of AMX-2 compared with wild-type nematodes. Interestingly, GRK-2 was also found to interact with and promote the phosphorylation of AMX-2. Additional studies reveal that 5-HIAA functions to inhibit egg laying in a manner dependent on the 5-HT receptor SER-1 and the G protein GOA-1. These results demonstrate that GRK-2 modulates 5-HT metabolism by regulating AMX-2 function and that 5-HIAA may function in the SER-1 signaling pathway.
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http://dx.doi.org/10.1074/jbc.M116.760850DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5392585PMC
April 2017

RNA-Seq Reveals Acute Manganese Exposure Increases Endoplasmic Reticulum Related and Lipocalin mRNAs in Caenorhabditis elegans.

J Biochem Mol Toxicol 2016 Feb 29;30(2):97-105. Epub 2015 Sep 29.

A. I. Virtanen Institute for Molecular Sciences, Department of Neurobiology, University of Eastern Finland, Kuopio 70211, Finland.

Manganese (Mn) is an essential nutrient; nonetheless, excessive amounts can accumulate in brain tissues causing manganism, a severe neurological condition. Previous studies have suggested oxidative stress, mitochondria dysfunction, and impaired metabolism pathways as routes for Mn toxicity. Here, we used the nematode Caenorhabditis elegans to analyze gene expression changes after acute Mn exposure using RNA-Seq. L1 stage animals were exposed to 50 mM MnCl2 for 30 min and analyzed at L4. We identified 746 up- and 1828 downregulated genes (FDR corrected p < 0.05; two-fold change) that included endoplasmic reticulum related abu and fkb family genes, as well as six of seven lipocalin-related (lpr) family members. These were also verified by qRT-PCR. RNA interference of lpr-5 showed a dramatic increase in whole body vulnerability to Mn exposure. Our studies demonstrate that Mn exposure alters gene transcriptional levels in different cell stress pathways that may ultimately contribute to its toxic effects.
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http://dx.doi.org/10.1002/jbt.21768DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5054866PMC
February 2016

Systemic and cerebral iron homeostasis in ferritin knock-out mice.

PLoS One 2015 28;10(1):e0117435. Epub 2015 Jan 28.

Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, Indiana, 46202, United States of America.

Ferritin, a 24-mer heteropolymer of heavy (H) and light (L) subunits, is the main cellular iron storage protein and plays a pivotal role in iron homeostasis by modulating free iron levels thus reducing radical-mediated damage. The H subunit has ferroxidase activity (converting Fe(II) to Fe(III)), while the L subunit promotes iron nucleation and increases ferritin stability. Previous studies on the H gene (Fth) in mice have shown that complete inactivation of Fth is lethal during embryonic development, without ability to compensate by the L subunit. In humans, homozygous loss of the L gene (FTL) is associated with generalized seizure and atypical restless leg syndrome, while mutations in FTL cause a form of neurodegeneration with brain iron accumulation. Here we generated mice with genetic ablation of the Fth and Ftl genes. As previously reported, homozygous loss of the Fth allele on a wild-type Ftl background was embryonic lethal, whereas knock-out of the Ftl allele (Ftl-/-) led to a significant decrease in the percentage of Ftl-/- newborn mice. Analysis of Ftl-/- mice revealed systemic and brain iron dyshomeostasis, without any noticeable signs of neurodegeneration. Our findings indicate that expression of the H subunit can rescue the loss of the L subunit and that H ferritin homopolymers have the capacity to sequester iron in vivo. We also observed that a single allele expressing the H subunit is not sufficient for survival when both alleles encoding the L subunit are absent, suggesting the need of some degree of complementation between the subunits as well as a dosage effect.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0117435PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4309591PMC
February 2016

The putative multidrug resistance protein MRP-7 inhibits methylmercury-associated animal toxicity and dopaminergic neurodegeneration in Caenorhabditis elegans.

J Neurochem 2014 Mar 25;128(6):962-74. Epub 2013 Nov 25.

Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, Indiana, USA.

Parkinson's disease (PD) is the most prevalent neurodegenerative motor disorder worldwide, and results in the progressive loss of dopamine (DA) neurons in the substantia nigra pars compacta. Gene-environment interactions are believed to play a significant role in the vast majority of PD cases, yet the toxicants and the associated genes involved in the neuropathology are largely ill-defined. Recent epidemiological and biochemical evidence suggests that methylmercury (MeHg) may be an environmental toxicant that contributes to the development of PD. Here, we report that a gene coding for the putative multidrug resistance protein MRP-7 in Caenorhabditis elegans modulates whole animal and DA neuron sensitivity to MeHg. In this study, we demonstrate that genetic knockdown of MRP-7 results in a twofold increase in Hg levels and a dramatic increase in stress response proteins associated with the endoplasmic reticulum, golgi apparatus, and mitochondria, as well as an increase in MeHg-associated animal death. Chronic exposure to low concentrations of MeHg induces MRP-7 gene expression, while exposures in MRP-7 genetic knockdown animals results in a loss of DA neuron integrity without affecting whole animal viability. Furthermore, transgenic animals expressing a fluorescent reporter behind the endogenous MRP-7 promoter indicate that the transporter is expressed in DA neurons. These studies show for the first time that a multidrug resistance protein is expressed in DA neurons, and its expression inhibits MeHg-associated DA neuron pathology.
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http://dx.doi.org/10.1111/jnc.12515DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3951531PMC
March 2014

Methylmercury exposure increases lipocalin related (lpr) and decreases activated in blocked unfolded protein response (abu) genes and specific miRNAs in Caenorhabditis elegans.

Toxicol Lett 2013 Oct 18;222(2):189-96. Epub 2013 Jul 18.

A. I. Virtanen Institute for Molecular Sciences, Department of Neurobiology, University of Eastern Finland, Kuopio, Finland. Electronic address:

Methylmercury (MeHg) is a persistent environmental and dietary contaminant that causes serious adverse developmental and physiologic effects at multiple cellular levels. In order to understand more fully the consequences of MeHg exposure at the molecular level, we profiled gene and miRNA transcripts from the model organism Caenorhabditis elegans. Animals were exposed to MeHg (10 μM) from embryo to larval 4 (L4) stage and RNAs were isolated. RNA-seq analysis on the Illumina platform revealed 541 genes up- and 261 genes down-regulated at a cutoff of 2-fold change and false discovery rate-corrected significance q < 0.05. Among the up-regulated genes were those previously shown to increase under oxidative stress conditions including hsp-16.11 (2.5-fold), gst-35 (10.1-fold), and fmo-2 (58.5-fold). In addition, we observed up-regulation of 6 out of 7 lipocalin related (lpr) family genes and down regulation of 7 out of 15 activated in blocked unfolded protein response (abu) genes. Gene Ontology enrichment analysis highlighted the effect of genes related to development and organism growth. miRNA-seq analysis revealed 6-8 fold down regulation of mir-37-3p, mir-41-5p, mir-70-3p, and mir-75-3p. Our results demonstrate the effects of MeHg on specific transcripts encoding proteins in oxidative stress responses and in ER stress pathways. Pending confirmation of these transcript changes at protein levels, their association and dissociation characteristics with interaction partners, and integration of these signals, these findings indicate broad and dynamic mechanisms by which MeHg exerts its harmful effects.
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http://dx.doi.org/10.1016/j.toxlet.2013.07.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3816353PMC
October 2013

The Nrf2/SKN-1-dependent glutathione S-transferase π homologue GST-1 inhibits dopamine neuron degeneration in a Caenorhabditis elegans model of manganism.

Neurotoxicology 2013 Sep 27;38:51-60. Epub 2013 May 27.

Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, IN 46202, USA.

Exposure to high levels of manganese (Mn) results in a neurological condition termed manganism, which is characterized by oxidative stress, abnormal dopamine (DA) signaling, and cell death. Epidemiological evidence suggests correlations with occupational exposure to Mn and the development of the movement disorder Parkinson's disease (PD), yet the molecular determinants common between the diseases are ill-defined. Glutathione S-transferases (GSTs) of the class pi (GSTπ) are phase II detoxification enzymes that conjugate both endogenous and exogenous compounds to glutathione to reduce cellular oxidative stress, and their decreased expression has recently been implicated in PD progression. In this study we demonstrate that a Caenorhabditis elegans GSTπ homologue, GST-1, inhibits Mn-induced DA neuron degeneration. We show that GST-1 is expressed in DA neurons, Mn induces GST-1 gene and protein expression, and GST-1-mediated neuroprotection is dependent on the PD-associated transcription factor Nrf2/SKN-1, as a reduction in SKN-1 gene expression results in a decrease in GST-1 protein expression and an increase in DA neuronal death. Furthermore, decreases in gene expression of the SKN-1 inhibitor WDR-23 or the GSTπ-binding cell death activator JNK/JNK-1 result in an increase in resistance to the metal. Finally, we show that the Mn-induced DA neuron degeneration is independent of the dopamine transporter DAT, but is largely dependent on the caspases CED-3 and the novel caspase CSP-1. This study identifies a C. elegans Nrf2/SKN-1-dependent GSTπ homologue, cell death effectors of GSTπ-associated xenobiotic-induced pathology, and provides the first in vivo evidence that a phase II detoxification enzyme may modulate DA neuron vulnerability in manganism.
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http://dx.doi.org/10.1016/j.neuro.2013.05.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3773487PMC
September 2013

miRNAs and their putative roles in the development and progression of Parkinson's disease.

Front Genet 2012 9;3:315. Epub 2013 Jan 9.

Department of Neurobiology, A.I. Virtanen Institute, University of Eastern Finland Kuopio, Finland.

Small regulatory RNAs, such as miRNAs, are increasingly being recognized not only as regulators of developmental processes but contributors to pathological states. The number of miRNAs determined experimentally to be involved in Parkinson's disease (PD) development and progression is small and includes regulators of pathologic proteins, neurotrophic factors, and xenobiotic metabolizing enzymes. PD gene-association studies have also indicated miRNAs in the pathology. In this review, we present known miRNAs and their validated targets that contribute to PD development and progression. We also incorporate data mining methods to link additional miRNAs with non-experimentally validated targets and propose additional roles of miRNAs in neurodegenerative processes. Furthermore, we present the potential contribution of next-generation-sequencing approaches to elucidate mechanisms and etiology of PD through discovery of novel miRNAs and other non-coding RNA classes.
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http://dx.doi.org/10.3389/fgene.2012.00315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3540391PMC
January 2013

C. elegans as a genetic model system to identify Parkinson's disease-associated therapeutic targets.

CNS Neurol Disord Drug Targets 2012 Dec;11(8):957-64

Department of Neurobiology, AI Virtanen Institute, University of Eastern Finland, Yliopistoranta 1, Kuopio 70210, Finland.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by motor and non-motor symptoms and the selective loss of dopaminergic neurons. The etiology of idiopathic PD is likely a combination of genetic and environmental factors. Despite findings from mammalian studies that have provided significant insight into the disorder, the molecular mechanisms underlying its pathophysiology are still poorly understood. The nematode Caenorhabditis elegans (C. elegans) is a powerful system for genetic analysis. Considering C. elegans short lifespan, fully sequenced genome, high genetic and neurobiochemical conservation with humans, as well as the availability of facile genetic tools, the nematode represents a highly efficient and effective model system to explore the molecular basis of PD. In this review we describe the utility of C. elegans for PD research, and the opportunity the model system presents to identify therapeutic targets.
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http://dx.doi.org/10.2174/1871527311211080004DOI Listing
December 2012

The metal transporter SMF-3/DMT-1 mediates aluminum-induced dopamine neuron degeneration.

J Neurochem 2013 Jan 21;124(1):147-57. Epub 2012 Nov 21.

Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, IN 46202, USA.

Aluminum (Al(3+)) is the most prevalent metal in the earth's crust and is a known human neurotoxicant. Al(3+) has been shown to accumulate in the substantia nigra of patients with Parkinson's disease (PD), and epidemiological studies suggest correlations between Al(3+) exposure and the propensity to develop both PD and the amyloid plaque-associated disorder Alzheimer's disease (AD). Although Al(3+) exposures have been associated with the development of the most common neurodegenerative disorders, the molecular mechanism involved in Al(3+) transport in neurons and subsequent cellular death has remained elusive. In this study, we show that a brief exposure to Al(3+) decreases mitochondrial membrane potential and cellular ATP levels, and confers dopamine (DA) neuron degeneration in the genetically tractable nematode Caenorhabditis elegans (C. elegans). Al(3+) exposure also exacerbates DA neuronal death conferred by the human PD-associated protein α-synuclein. DA neurodegeneration is dependent on SMF-3, a homologue to the human divalent metal transporter (DMT-1), as a functional null mutation partially inhibits the cell death. We also show that SMF-3 is expressed in DA neurons, Al(3+) exposure results in a significant decrease in protein levels, and the neurodegeneration is partially dependent on the PD-associated transcription factor Nrf2/SKN-1 and caspase Apaf1/CED-4. Furthermore, we provide evidence that the deletion of SMF-3 confers Al(3+) resistance due to sequestration of Al(3+) into an intracellular compartment. This study describes a novel model for Al(3+)-induced DA neurodegeneration and provides the first molecular evidence of an animal Al(3+) transporter.
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http://dx.doi.org/10.1111/jnc.12072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3725953PMC
January 2013

Chemosensory cue conditioning with stimulants in a Caenorhabditis elegans animal model of addiction.

Behav Neurosci 2012 Jun;126(3):445-56

Department of Psychology, Indiana University Purdue University, USA.

The underlying molecular mechanisms of drug abuse and addiction behaviors are poorly understood. Caenorhabditis elegans (C. elegans) provide a simple, whole animal model with conserved molecular pathways well suited for studying the foundations of complex diseases. Historically, chemotaxis has been a measure used to examine sensory approach and avoidance behavior in worms. Chemotaxis can be modulated by previous experience, and cue-dependent conditioned learning has been demonstrated in C. elegans, but such conditioning with drugs of abuse has not been reported. Here we show that pairing a distinctive salt cue with a drug (cocaine or methamphetamine) results in a concentration-dependent change in preference for the cue that was paired with the drug during conditioning. Further, we demonstrate that pairing of either drug with a distinctive food type can also increase preference for the drug-paired food in the absence of the drug. Dopamine-deficient mutants did not develop drug-paired, cue-conditioned responses. The findings suggest that, like vertebrates, C. elegans display a conditioned preference for environments containing cues previously associated with drugs of abuse, and this response is dependent on dopamine neurotransmission. This model provides a new and powerful method to study the genetic and molecular mechanisms that mediate drug preference.
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http://dx.doi.org/10.1037/a0028303DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3367381PMC
June 2012

SKN-1/Nrf2 inhibits dopamine neuron degeneration in a Caenorhabditis elegans model of methylmercury toxicity.

Toxicol Sci 2010 Dec 20;118(2):613-24. Epub 2010 Sep 20.

Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.

Methylmercury (MeHg) exposure from occupational, environmental, and food sources is a significant threat to public health. MeHg poisonings in adults may result in severe psychological and neurological deficits, and in utero exposures can confer embryonic defects and developmental delays. Recent epidemiological and vertebrate studies suggest that MeHg exposure may also contribute to dopamine (DA) neuron vulnerability and the propensity to develop Parkinson's disease (PD). In this study, we describe a Caenorhabditis elegans model of MeHg toxicity that shows that low, chronic exposure confers embryonic defects, developmental delays, decreases in brood size and animal viability, and DA neuron degeneration. Toxicant exposure results in the robust induction of the glutathione-S-transferases (GSTs) gst-4 and gst-38 that are largely dependent on the PD-associated phase II antioxidant transcription factor SKN-1/Nrf2. We also demonstrate that the expression of SKN-1, a protein previously localized to a small subset of chemosensory neurons and intestinal cells in the nematode, is also expressed in the DA neurons, and a reduction in SKN-1 gene expression increases MeHg-induced animal vulnerability and DA neuron degeneration. These studies recapitulate fundamental hallmarks of MeHg-induced mammalian toxicity, identify a key molecular regulator of toxicant-associated whole-animal and DA neuron vulnerability, and suggest that the nematode will be a useful in vivo tool to identify and characterize mediators of MeHg-induced developmental and DA neuron pathologies.
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http://dx.doi.org/10.1093/toxsci/kfq285DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3003544PMC
December 2010

Global microRNA expression profiling of Caenorhabditis elegans Parkinson's disease models.

J Mol Neurosci 2010 May 21;41(1):210-8. Epub 2010 Jan 21.

Department of Biosciences, Kuopio University, Kuopio, Finland.

MicroRNAs (miRNAs) play an important role in human brain development and maintenance. To search for miRNAs that may be involved in the pathogenesis of Parkinsons disease (PD), we utilized miRNA microarrays to identify potential gene expression changes in 115 annotated miRNAs in PD-associated Caenorhabditis elegans models that either overexpress human A53T alpha-synuclein or have mutations within the vesicular catecholamine transporter (cat-1) or parkin (pdr-1) ortholog. Here, we show that 12 specific miRNAs are differentially regulated in the animals overexpressing alpha-synuclein, five in cat-1, and three in the pdr-1 mutants. The family of miR-64 and miR-65 are co-underexpressed in the alpha-synuclein transgenic and cat-1 strains, and members of let-7 family co-underexpressed in the alpha-synuclein and pdr-1 strains; mdl-1 and ptc-1 genes are target candidates for miR-64 and miR-65 and are overexpressed in alpha-synuclein transgenic as well as miR-64/65 (tm3711) knockout animals. These results indicate that miRNAs are differentially expressed in C. elegans PD models and suggest a role for these molecules in disease pathogenesis.
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http://dx.doi.org/10.1007/s12031-009-9325-1DOI Listing
May 2010

The divalent metal transporter homologues SMF-1/2 mediate dopamine neuron sensitivity in caenorhabditis elegans models of manganism and parkinson disease.

J Biol Chem 2009 Dec;284(51):35758-68

Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.

Parkinson disease (PD) and manganism are characterized by motor deficits and a loss of dopamine (DA) neurons in the substantia nigra pars compacta. Epidemiological studies indicate significant correlations between manganese exposure and the propensity to develop PD. The vertebrate divalent metal transporter-1 (DMT-1) contributes to maintaining cellular Mn(2+) homeostasis and has recently been implicated in Fe(2+)-mediated neurodegeneration in PD. In this study we describe a novel model for manganism that incorporates the genetically tractable nematode Caenorhabditis elegans. We show that a brief exposure to Mn(2+) increases reactive oxygen species and glutathione production, decreases oxygen consumption and head mitochondria membrane potential, and confers DA neuronal death. DA neurodegeneration is partially dependent on a putative homologue to DMT-1, SMF-1, as genetic knockdown or deletion partially inhibits the neuronal death. Mn(2+) also amplifies the DA neurotoxicity of the PD-associated protein alpha-synuclein. Furthermore, both SMF-1 and SMF-2 are expressed in DA neurons and contribute to PD-associated neurotoxicant-induced DA neuron death. These studies describe a C. elegans model for manganism and show that DMT-1 homologues contribute to Mn(2+)- and PD-associated DA neuron vulnerability.
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http://dx.doi.org/10.1074/jbc.M109.051409DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2791006PMC
December 2009

Caenohabditis elegans in Parkinson's disease drug discovery: addressing an unmet medical need.

Mol Interv 2008 Dec;8(6):284-93

Department of Pharmacology and Toxicology, Center for Environmental Health, and Stark Neuroscience Research Institute, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.

It has been over forty years since dopamine neuron degeneration in the substantia nigra and Lewy body formation within surviving cells were described as the pathological hallmarks of Parkinson's disease (PD). Although research in the intervening decades particularly in the last twenty-five years has yielded a variety of robust animal models and invaluable mechanistic insights into PD-associated neuronal dysfunction and cell death, therapeutic agents have not been forthcoming to alter the course of PD. Recently, the screening of experimental therapeutics for PD has been pursued through the use of genetically tractable models, such as the nematode Caenorhabditis elegans. This simple worm remarkably recapitulates the basic cellular and molecular pathways associated with PD, is amenable to facile genetic methods, and through the use of high-throughput screening technologies, provides powerful new opportunities for the in vivo identification of therapeutic targets. In this review we briefly describe the utility that the C. elegans model system may have for PD drug discovery.
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http://dx.doi.org/10.1124/mi.8.6.6DOI Listing
December 2008

Use of non-mammalian alternative models for neurotoxicological study.

Neurotoxicology 2008 May 25;29(3):546-55. Epub 2008 Apr 25.

Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.

The field of neurotoxicology needs to satisfy two opposing demands: the testing of a growing list of chemicals, and resource limitations and ethical concerns associated with testing using traditional mammalian species. National and international government agencies have defined a need to reduce, refine or replace mammalian species in toxicological testing with alternative testing methods and non-mammalian models. Toxicological assays using alternative animal models may relieve some of this pressure by allowing testing of more compounds while reducing expense and using fewer mammals. Recent advances in genetic technologies and the strong conservation between human and non-mammalian genomes allow for the dissection of the molecular pathways involved in neurotoxicological responses and neurological diseases using genetically tractable organisms. In this review, applications of four non-mammalian species, zebrafish, cockroach, Drosophila, and Caenorhabditis elegans, in the investigation of neurotoxicology and neurological diseases are presented.
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http://dx.doi.org/10.1016/j.neuro.2008.04.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2702842PMC
May 2008

Neurotoxic potential of depleted uranium effects in primary cortical neuron cultures and in Caenorhabditis elegans.

Toxicol Sci 2007 Oct 16;99(2):553-65. Epub 2007 Jul 16.

Department of Physiology and Pharmacology, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157-1083, USA.

Depleted uranium (DU) is an extremely dense metal that is used in radiation shielding, counterbalances, armor, and ammunition. In light of the public concerns about exposure to DU and its potential role in Gulf War Syndrome (GWS), this study evaluated the neurotoxic potential of DU using focused studies on primary rat cortical neurons and the nematode Caenorhabditis elegans. We examined cell viability, cellular energy metabolism, thiol metabolite oxidation, and lipid peroxidation following exposure of cultured neurons to DU, in the form of uranyl acetate. We concurrently evaluated the neurotoxicity of uranyl acetate in C. elegans using various neuronal-green fluourescent protein reporter strains to visualize neurodegeneration. Our studies indicate that uranyl acetate has low cytotoxic potential, and uranium exposure does not result in significant changes in cellular energy metabolism, thiol metabolite oxidation, or lipid peroxidation. Furthermore, our C. elegans studies do not show any significant neurodegeneration following uranyl acetate exposure. Together, these studies suggest that DU, in the form of uranyl acetate, has low neurotoxic potential. These findings should alleviate the some of public concerns regarding DU as an etiologic agent of neurodegenerative conditions associated with GWS.
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http://dx.doi.org/10.1093/toxsci/kfm171DOI Listing
October 2007

The nematode C. elegans as an animal model to explore toxicology in vivo: solid and axenic growth culture conditions and compound exposure parameters.

Curr Protoc Toxicol 2007 Feb;Chapter 1:Unit1.9

Vanderbilt University Medical Center, Nashville, Tennessee, USA.

Significant limitations in vertebrate animal model systems include the time involved, the expense, the fact that in vitro results may not reflect live animal pathology, difficulties in transporting the toxin past the blood brain barrier, and the inability to identify the mechanism of action without some a priori knowledge of the toxin's target. The availability of the complete genome sequence of the nematode C. elegans, coupled with the worm's size, growth rate, ease of culturing, and the realization that basic biological mechanisms and disease processes between worms and humans are highly conserved, makes this genetically tractable model a remarkable opportunity to dissect and identify in vivo the cellular processes involved in toxin-induced cell dysregulation and death. This unit includes protocols for culturing worms on solid and axenic media and acute and chronic exposure parameters for Parkinson's disease-associated toxins and hemin chloride. These methods provide the groundwork for using this powerful model system to further elucidate and understand the molecular mechanisms involved in nutrition as well as toxicological responses relevant to human diseases.
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http://dx.doi.org/10.1002/0471140856.tx0109s31DOI Listing
February 2007

Identification of gene expression changes in transgenic C. elegans overexpressing human alpha-synuclein.

Neurobiol Dis 2006 Jun 19;22(3):477-86. Epub 2006 Apr 19.

Department of Neurobiology, A.I. Virtanen Institute, Kuopio, 70211, Finland.

Alpha-synuclein containing cellular inclusions are a hallmark of Parkinson Disease, Lewy Body Dementia, and Multiple System Atrophy. A genome wide expression screen was performed in C. elegans overexpressing both wild-type and A53T human alpha-synuclein. 433 genes were up- and 67 genes down-regulated by statistical and fold change (> or <2) criteria. Gene ontology (GO) categories within the regulated gene lists indicated over-representation of development and reproduction, mitochondria, catalytic activity, and histone groups. Seven genes (pdr-1, ubc-7, pas-5, pas-7, pbs-4, RPT2, PSMD9) with function in the ubiquitin-proteasome system and 35 mitochondrial function genes were up-regulated. Nine genes that form histones H1, H2B, and H4 were down-regulated. These results demonstrate the effects of alpha-synuclein on proteasome and mitochondrial complex gene expression and provide further support for the role of these complexes in mediating neurotoxicity. The results also indicate an effect on nuclear protein genes that suggests a potential new avenue for investigation.
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http://dx.doi.org/10.1016/j.nbd.2005.12.021DOI Listing
June 2006

Selective sensitivity of Caenorhabditis elegans neurons to RNA interference.

Neuroreport 2005 Dec;16(18):1995-9

Department of Neurobiology, A.I. Virtanen Institute for Molecular Sciences, University of Kuopio, Kuopio, Finland.

RNA interference is a new approach to knockdown gene expression, but effectiveness varies depending on the organism, cell type or target sequence. Studies with Caenorhabditis elegans have shown that subsets of cells including neurons are often resistant to RNA interference. We measured RNA interference using green fluorescent protein reporter strains and feeding, soaking and injection delivery methods in a number of Caenorhabditis elegans neuron subtypes (dopaminergic, GABAergic, cholinergic, glutamatergic, touch). The sensitivity to RNA interference varied: GABAergic and dopaminergic neurons showed greater resistance while cholinergic, glutamatergic and touch neurons were more sensitive. Dysfunctional RRF-3, a putative RNA-directed RNA polymerase, had a significant effect on increasing neuron sensitivity in most subtypes. These results demonstrate that Caenorhabditis elegans neurons vary in their sensitivity to RNA interference.
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http://dx.doi.org/10.1097/00001756-200512190-00005DOI Listing
December 2005

A genetic screen in Caenorhabditis elegans for dopamine neuron insensitivity to 6-hydroxydopamine identifies dopamine transporter mutants impacting transporter biosynthesis and trafficking.

J Neurochem 2005 Aug 30;94(3):774-85. Epub 2005 Jun 30.

Department of Anesthesiology, Vanderbilt School of Medicine, Nashville, Tennessee 37232-8548, USA.

The presynaptic dopamine (DA) transporter (DAT) is a major determinant of synaptic DA inactivation, an important target for psychostimulants including cocaine and amphetamine, and a mediator of DA neuron vulnerability to the neurotoxins 6-hydroxydopamine (6-OHDA) and 1-methyl-4-phenylpyridinium ion. To exploit genetic approaches for the study of DATs and neural degeneration, we exploited the visibility of green fluorescent protein (GFP)-tagged DA neurons in transgenic nematodes to implement a forward genetic screen for suppressors of 6-OHDA sensitivity. In our initial effort, we identified three novel dat-1 alleles conferring 6-OHDA resistance. Two of the dat-1 alleles derive from point mutations in conserved glycine residues (G55, G90) in contiguous DAT-1 transmembrane domains (TM1 and TM2, respectively), whereas the third allele results in altered translation of the transporter's COOH terminus. Our studies reveal biosynthetic, trafficking and functional defects in the DAT-1 mutants, exhibited both in vitro and in vivo. These studies validate a forward genetic approach to the isolation of DA neuron-specific toxin suppressors and point to critical contributions of the mutated residues, as well as elements of the DAT-1 COOH terminus, to functional expression of catecholamine transporters in neurons.
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http://dx.doi.org/10.1111/j.1471-4159.2005.03205.xDOI Listing
August 2005

Dopaminergic neuronal loss and motor deficits in Caenorhabditis elegans overexpressing human alpha-synuclein.

J Neurochem 2003 Jul;86(1):165-72

Department of Neurobiology, A. I. Virtanen Institute, Kuopio University, Finland.

Overexpression of human alpha-synuclein in model systems, including cultured neurons, drosophila and mice, leads to biochemical and pathological changes that mimic synucleopathies including Parkinson's disease. We have overexpressed both wild-type (WT) and mutant alanine53-->threonine (A53T) human alpha-synuclein by transgenic injection into Caenorhabditis elegans. Motor deficits were observed when either WT or A53T alpha-synuclein was overexpressed with a pan-neuronal or motor neuron promoter. Neuronal and dendritic loss were accelerated in all three sets of C. elegans dopaminergic neurons when human alpha-synuclein was overexpressed under the control of a dopaminergic neuron or pan-neuronal promoter, but not with a motor neuron promoter. There were no significant differences in neuronal loss between overexpressed WT and A53T forms or between worms of different ages (4 days, 10 days or 2 weeks). These results demonstrate neuronal and behavioral perturbations elicited by human alpha-synuclein in C. elegans that are dependent upon expression in specific neuron subtypes. This transgenic model in C. elegans, an invertebrate organism with excellent experimental resources for further genetic manipulation, may help facilitate dissection of pathophysiologic mechanisms of various synucleopathies.
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http://dx.doi.org/10.1046/j.1471-4159.2003.01809.xDOI Listing
July 2003

The Caenorhabditis elegans dopaminergic system: opportunities for insights into dopamine transport and neurodegeneration.

Annu Rev Pharmacol Toxicol 2003 10;43:521-44. Epub 2002 Jan 10.

Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232-6420, USA.

The neurotransmitter dopamine (DA) plays a central role in the coordination of movement, attention, and the recognition of reward. Loss of DA from the basal ganglia, as a consequence of degeneration of neurons in the substantia nigra, triggers postural instability and Parkinson's disease (PD). DA transporters (DATs) regulate synaptic DA availability and provide a conduit for the uptake of DA mimetic neurotoxins, which can be used to evoke neuronal death and Parkinson-like syndrome. Recently, we have explored the sensitivity of DA neurons in the nematode Caenorhabditis elegans to the Parkinsonian-inducing neurotoxin 6-hydroxydopamine (6-OHDA) and found striking similarities, including DAT dependence, to neurodegeneration observed in mammalian models. In this review, we present our findings in the context of molecular and behavioral dimensions of DA signaling in C. elegans with an eye toward opportunities for uncovering DAT mutants, DAT regulators, and components of toxin-mediated cell death.
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http://dx.doi.org/10.1146/annurev.pharmtox.43.100901.135934DOI Listing
September 2003

Neurotoxin-induced degeneration of dopamine neurons in Caenorhabditis elegans.

Proc Natl Acad Sci U S A 2002 Mar 26;99(5):3264-9. Epub 2002 Feb 26.

Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232-6420, USA.

Parkinson's disease is a complex neurodegenerative disorder characterized by the death of brain dopamine neurons. In mammals, dopamine neuronal degeneration can be triggered through exposure to neurotoxins accumulated by the presynaptic dopamine transporter (DAT), including 6-hydroxydopamine (6-OHDA) and 1-methyl-4-phenylpyridinium. We have established a system for the pharmacological and genetic evaluation of neurotoxin-induced dopamine neuronal death in Caenorhabditis elegans. Brief (1 h) exposure of green fluorescent protein-tagged, living worms to 6-OHDA causes selective degeneration of dopamine neurons. We demonstrate that agents that interfere with DAT function protect against 6-OHDA toxicity. 6-OHDA-triggered neural degeneration does not require the CED-3/CED-4 cell death pathway, but is abolished by the genetic disruption of the C. elegans DAT.
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http://dx.doi.org/10.1073/pnas.042497999DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC122507PMC
March 2002

The yeast endosomal Na+/H+ exchanger, Nhx1, confers osmotolerance following acute hypertonic shock.

Microbiology (Reading) 1999 Nov;145 ( Pt 11):3221-3228

Department of Physiology, The Johns Hopkins University School of Medicine, 725 N. Wolfe Street, Baltimore MD 21205, USA1.

Osmotolerance in yeast is regulated by at least two distinct mechanisms. The acquired response occurs following long-term exposure to hypertonic medium and requires the induction of the HOG-MAP (high-osmolarity glycerol mitogen-activated protein) kinase cascade to increase levels of the osmolyte glycerol. The acute response occurs following sudden exposure to high osmotica and appears to be dependent on normal vacuole function. In this study it is reported that the yeast endosomal/prevacuolar Na+/H+ exchanger Nhx1 contributes to osmotolerance following sudden exposure to hyperosmotic media. Vacuolar shrinkage and recovery in response to osmotic shock was altered in the (delta)nhx1 null mutant. Our results also show that the osmotolerance conferred by Nhx1 contributes to the postdiauxic/stationary-phase resistance to osmotic stress and allows for the continued growth of cells until the acquired osmotolerance response can occur.
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http://dx.doi.org/10.1099/00221287-145-11-3221DOI Listing
November 1999
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