Neurotoxic Effects of Neurotoxic Effects of SolventsSolvents

William BoyesWilliam Boyes

Neurotoxicology DivisionNeurotoxicology DivisionNational Health and Environmental National Health and Environmental

Effects Research LaboratoryEffects Research LaboratoryOffice of Research and Development, Office of Research and Development,

EPAEPAboyes.william@epa.govboyes.william@epa.gov

Chemical StructuresChemical Structures

Benzene Toluene

Trichloroethylene (TCE)

Tetrachloroethylene

(Perchloroethylene) or “Perc”

SolventsSolvents

LipophilicLipophilic Organic chemical compositionOrganic chemical composition Distribute to body lipid-rich tissuesDistribute to body lipid-rich tissues Easily cross lipid membranes & barriersEasily cross lipid membranes & barriers

VolatilityVolatility Inhalation is a significant route of exposureInhalation is a significant route of exposure Delivery to tissues depends upon the Delivery to tissues depends upon the

blood:tissue partition coefficientblood:tissue partition coefficient

Solvent NeurotoxicitySolvent Neurotoxicity

CNS depressionCNS depression EuphoriaEuphoria Sensory, cognitive, Sensory, cognitive,

motor deficitsmotor deficits Reversible after Reversible after

exposureexposure Causes accidents or Causes accidents or

injuriesinjuries Good animal modelsGood animal models Several mechanistic Several mechanistic

targetstargets

Organic brain disorderOrganic brain disorder Originally observed in Originally observed in

Scandinavian paintersScandinavian painters Sensory, cognitive, Sensory, cognitive,

motor deficitsmotor deficits Years of high level Years of high level

exposureexposure No good animal No good animal

modelsmodels Mechanisms unknownMechanisms unknown

Acute Chronic

Volatile Organic Volatile Organic CompoundsCompounds

Widely used in Widely used in industry and industry and commercecommerce

Emission SourcesEmission Sources Factories & chemical Factories & chemical

plants plants Smaller shopsSmaller shops Mobile sourcesMobile sources Indoor air sourcesIndoor air sources Consumer productsConsumer products Pesticide “inerts”Pesticide “inerts” Drinking waterDrinking water Hazardous waste sitesHazardous waste sites

Uses include:Uses include: Organic solventsOrganic solvents Cleaning & degreasingCleaning & degreasing GasolineGasoline Paints & gluesPaints & glues Dry cleaningDry cleaning Printing Printing Paint strippers, nail Paint strippers, nail

polish removerpolish remover Microelectronic Microelectronic

manufacturemanufacture

Upset EmissionsUpset Emissions

UpsetsUpsets Startup, maintance, Startup, maintance,

shutdownshutdown In some cases, upset In some cases, upset

releases exceeded releases exceeded annual releases several annual releases several thousand foldthousand fold

7,533 upset events 7,533 upset events reported to Texas in reported to Texas in 20042004

Some facilities report Some facilities report upset events on average upset events on average every other dayevery other day

Public Citizen, 2005

Small Gasoline EnginesSmall Gasoline Engines

2-cycle engines burn 2-cycle engines burn only about 60% of the only about 60% of the fuelfuel

The remainder is The remainder is emitted as a breathable emitted as a breathable hydrocarbon misthydrocarbon mist

Contains BTEXContains BTEX Benzene, toluene Benzene, toluene

ethylbenzene and xyleneethylbenzene and xylene Hundreds of other Hundreds of other

compounds in small compounds in small amountsamounts

The Washington Post, Aug. 14, 2002

Dry CleanersDry Cleaners

Perchloroethylene Perchloroethylene Detected in co-Detected in co-

located located apartmentsapartments Especially in NYCEspecially in NYC

Residents may Residents may show poor visual show poor visual function or other function or other problemsproblems

Acute Solvent Actions are a Acute Solvent Actions are a Function of Lipid SolubilityFunction of Lipid Solubility

Lipophilic nature Lipophilic nature measured as LogPmeasured as LogP Octanol/water partition Octanol/water partition

coefficientcoefficient Increased partitioning Increased partitioning

into brain and nerve into brain and nerve membranesmembranes

Seen as evidence for Seen as evidence for membrane fluidity membrane fluidity mechanism of actionmechanism of action

Goodman & Gilman, 1990

How are VOCs Causing How are VOCs Causing Acute Effects?Acute Effects?

They were once thought to simply They were once thought to simply dissolve lipid cell membranes dissolve lipid cell membranes

New evidence suggests more New evidence suggests more selection disruption of nerve selection disruption of nerve membrane ion channel proteinsmembrane ion channel proteins

But which ones?But which ones?

Patch-Clamp ElectrophysiologyPatch-Clamp Electrophysiology

V = IRV = IRTo equipment with lots To equipment with lots of wires, knobs, of wires, knobs, switches, buttons and switches, buttons and lights !lights !

VVII

• Glutamate•The most common CNS excitatory neurotransmitter

•NMDA receptor•One type of glutamate receptor •located in the visual system

• NMDA receptor activity is inhibited by toluene

•(in vitro studies, Cruz et al., 1998)

NMDA/gly NMDA/gly NMDA/gly

Taken from Cruz et al., 1998

Compound Ion Channel

Solvents NMDA Receptor nAChR GABAA Receptor Glycine Receptor VSCC*

Toluene Inhibits Inhibits Potentiates Potentiates Inhibits/Potentiates

Trichloroethylene ND ND Potentiates Potentiates Inhibits/Potentiates

Perchloroethylene ND Inhibits ND ND Inhibits/Potentiates

1,1,1-Trichloroethane Inhibits ND Potentiates Potentiates Inhibits

Ethanol Inhibits Inhibits/Potentiates Potentiates Potentiates Inhibits

Halothane Inhibits Inhibits Potentiates Potentiates Inhibits

Solvents & Ion Channel Solvents & Ion Channel FunctionFunction

Bushnell et al., 2005

Chronic Solvent Chronic Solvent Encephalopathy?Encephalopathy?

Scandinavian painters Scandinavian painters and other workersand other workers

Chronic exposure Chronic exposure a variety of a variety of

impairments of mood impairments of mood and intellectual function and intellectual function leading eventually to leading eventually to dementiadementia

Early studies Early studies confounded confounded Poorly matched controlsPoorly matched controls Poor documentation of Poor documentation of

exposure historyexposure history

More recent studies More recent studies showshow Increased reaction Increased reaction

timestimes Poor visual functionPoor visual function Impaired auditory Impaired auditory

thresholdsthresholds Impaired motor skillsImpaired motor skills Impaired performance Impaired performance

of cognitive and of cognitive and memory tasksmemory tasks

TolueneToluene Present in paints, glues, gasoline and many other Present in paints, glues, gasoline and many other

productsproducts Subject of over 40 EPA Maximum Achievable Subject of over 40 EPA Maximum Achievable

Control Technology (MACT) / residual risk Control Technology (MACT) / residual risk assessmentsassessments

The substance of choice for glue sniffersThe substance of choice for glue sniffers

CH3

Toluene

COOH

Benzoic acid

COOGly

Hippuric acid

glycine

0 1 2 3 4 5 6-0.1

0.0

0.1

0.2

0.3REPORTED DATA

PREDICTED EFFECT

95% CONFIDENCE LIMITS

DRAFT. DO NOT CITE.

Blood Toluene (mg/liter)

Be

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ral

Eff

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t (p

rop

ort

ion

of

ba

seli

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)

Toluene & Acute Behavioral ChangesToluene & Acute Behavioral Changese.g. Choice Reaction Timee.g. Choice Reaction Time

Meta-analysis of 6 studies, Benignus et al., in press

Toluene AbuseToluene Abuse

Neuropathic effects in humansNeuropathic effects in humans Following repeated solvent abuseFollowing repeated solvent abuse Very high dose levelsVery high dose levels Cerebellar damageCerebellar damage Cerebral atrophyCerebral atrophy Multiple symptoms of dementiaMultiple symptoms of dementia Confounded by hypoxia and other exposuresConfounded by hypoxia and other exposures

Toluene at High Doses Causes Toluene at High Doses Causes Outer Haircell Damage in Outer Haircell Damage in

ChochleaChochlea

Johnson (1993)Johnson (1993)

Ethanol (Alcohol)Ethanol (Alcohol)

CNS depressantCNS depressant Legal definition of inebriation based on BACsLegal definition of inebriation based on BACs

Often 0.1% (100 mg/100 ml)Often 0.1% (100 mg/100 ml) Acute exposureAcute exposure

EuphoriaEuphoria Loss of inhibitions / poor judgmentLoss of inhibitions / poor judgment Loss of balance & motor coordinationLoss of balance & motor coordination Impaired vision & visual/motor functionImpaired vision & visual/motor function Ataxia, nausea, vomitingAtaxia, nausea, vomiting Unconsciousness Unconsciousness

CH3—CH2—OH

Ethanol MetabolismEthanol Metabolism

CH3—CH2—OH

EthanolEthanol

CH3—CHO

AcetaldehydeAcetaldehydeNAD

Alcohol Alcohol dehydrogenasedehydrogenase

CatalaseCatalaseHH22OO22

+ NADH

Microsomal ethanol Microsomal ethanol oxidizing system oxidizing system

(MEOS) = CYP2E1(MEOS) = CYP2E1

NADPH

10-30% of 10-30% of metabolized metabolized

ethanolethanol

Rate of metabolism Rate of metabolism is ~ 15-20 mg%/hris ~ 15-20 mg%/hr

< 10%

< 20%

Ethanol MetabolismEthanol Metabolism

CH3—CH2—OH

EthanolEthanol

CH3—CHO

AcetaldehydeAcetaldehydeNAD

Alcohol Alcohol dehydrogenase dehydrogenase

(ADH)(ADH)+ NADH

CH3—COO-

Acetic acidAcetic acid

Acetaldehyde dehydrogenase Acetaldehyde dehydrogenase (ALDH)(ALDH)

NAD

+ NADH

ADH

• males > females

ALDH isozymes

• inactive variant in %50 Asians

•Variant form in Native Americans

• inhibited by Disulfiram

Chronic Ethanol ToxicityChronic Ethanol Toxicity

AlcoholismAlcoholism High % of calories from alcoholHigh % of calories from alcohol Thiamine deficientThiamine deficient Wernicke's encephalopathyWernicke's encephalopathy

Damage to multiple brain areasDamage to multiple brain areas Impaired cognition, motor functionImpaired cognition, motor function

Korsakoff's psychosisKorsakoff's psychosis Acute & ChronicAcute & Chronic

100,000 premature deaths / year in U.S.100,000 premature deaths / year in U.S.

Fetal Alcohol Syndrome Fetal Alcohol Syndrome (FAS)(FAS)

Characterized by:Characterized by: Mental retardationMental retardation MicrocephalyMicrocephaly IrritabilityIrritability Reduced birth weightReduced birth weight Poor muscle coordinationPoor muscle coordination Cranio/Facial anomaliesCranio/Facial anomalies Mechanism poorly understoodMechanism poorly understood

Methanol (Wood Alcohol)Methanol (Wood Alcohol)

Clinical signs (people)Clinical signs (people) Multiphasic syndromeMultiphasic syndrome

Early – like ethanol (Central nervous system Early – like ethanol (Central nervous system depression, weakness, headache, vomiting)depression, weakness, headache, vomiting)

Mid- asymptomatic period (12-24 hr)Mid- asymptomatic period (12-24 hr) Late - Severe metabolic acidosis, optic disc Late - Severe metabolic acidosis, optic disc

edema, and bilateral necrosis of the putamenedema, and bilateral necrosis of the putamen Other adverse effects of methanol in humans Other adverse effects of methanol in humans

include minor skin and eye irritationinclude minor skin and eye irritation Formic acid is the toxic metabolite of methanol. Formic acid is the toxic metabolite of methanol.

Accounts for the metabolic acidosis and blindness Accounts for the metabolic acidosis and blindness seen in people following methanol poisoning seen in people following methanol poisoning

CH3—OH

Methanol Metabolism Methanol Metabolism (Primates)(Primates)

CH3—OH

MethanolMethanol

CH2O

FormaldehydeFormaldehydeNAD

Alcohol Alcohol dehydrogenasedehydrogenase

+ NADH

HCOO-

Formic acidFormic acid

Formaldehydealdehyde dehydrogenaseNAD

+ NADHCO2

TetrahydrofolateTetrahydrofolate

LowLow

Methanol Metabolism Methanol Metabolism (Rodents)(Rodents)

CH3—OH

MethanolMethanol

CH2O

FormaldehydeFormaldehyde

CatalaseCatalase

HCOO-

Formic acidFormic acid

Formaldehydealdehyde dehydrogenaseNAD

+ NADHCO2

TetrahydrofolateTetrahydrofolate

HIGHHIGH

Folate deficiency increases the sensitivity of methanol in rodents. Intraretinal metabolism may be important.

Methanol Visual ToxicityMethanol Visual Toxicity

Eells et al., PNAS 2003

n-Hexane and Methyl n-n-Hexane and Methyl n-butyl ketonebutyl ketone

NeurotoxicityNeurotoxicity Sensorimotor polyneuropathySensorimotor polyneuropathy

Sensory numbness and paresthesiaSensory numbness and paresthesia Distal nerves affected firstDistal nerves affected first Clinical signs often delayed for 6-12+ monthsClinical signs often delayed for 6-12+ months Axonal swelling and secondary demyelinationAxonal swelling and secondary demyelination

2,5-hexanedione is common toxic 2,5-hexanedione is common toxic metabolitemetabolite

Spencer & Schaumburg, 2000

Peripheral NeuropathyPeripheral Neuropathy

Sources & ReadingsSources & Readings

Anthony, DC., Montine, T.J., Valentine W.M., and Graham, D.G. Toxic responses of the nervous system. In Casarett and Doull’s Toxicology: the Basic Science of Poisons, Sixth Edition. McGraw-Hill Medical Publishing Division, New York, pp 535-563, 2001

Bushnell, P.J., Shafer, T.J., Bale, A.S., Boyes, W.K., Simmons, J.E., Eklund, C. and Jackson, T.L. Developing an exposure-dose-response model for organic solvents: overview and progress on in vitro models and dosimetry. Environmental Toxicology and Pharmacology, 19: 607–614, 2005.

Benignus, V.A., Bushnell, P.J. and Boyes, W. K. Toward cost-benefit analysis of acute behavioral effects of toluene in humans. Risk Analysis, 25 (2), 447-456, 2005.

Bruckner, J.V. and Warren, D.A., W.K. Toxic effects of solvents and vapors. In Casarett and Doull’s Toxicology: the Basic Science of Poisons, Sixth Edition. McGraw-Hill Medical Publishing Division, New York, pp 869-916, 2001

Schreiber, J S; Hudnell, H K; Geller, A M; House, D E; Aldous, K M; Force, M E; Langguth, K W; Prohonic, E J; Parker, J C (2002) Apartment residents’ and day care workers’ exposures to tetrachloroethylene and deficits in visual contrast sensitivity. Environ Health Perspect 110:655–664.

Arlien-Søborg, P. (1992). Solvent Neurotoxicity. CRC Press, Boca Raton FL.

Spencer PS., and Schaumburg H.H. Experimental and Clinical Neurotoxicology, 2nd Edition, Oxford University Press, 2000.