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9710892-6638/98/0012-0971/$02.25 Q FASEB
/ 382d 0012 Mp 971 Friday Aug 07 08:43 AM LP–FASEB 0012
Neurobehavioral effects from exposure to dental
amalgam Hg
o
: new distinctions between recent
exposure and Hg body burden
DIANA ECHEVERRIA,*
,†,1
H. VASKEN APOSHIAN,
‡
JAMES S. WOODS,*
,†
NICHOLAS J. HEYER,
†
MARY M. APOSHIAN,
‡
ALVAH C. BITTNER, JR.,*
,†
RODERICK K. MAHURIN, AND MARGARET CIANCIOLA
§
*Battelle Centers for Public Health Research and Evaluation, Seattle, Washington 98105, USA;
†
Department of Environmental Health, University of Washington, Seattle, Washington 98195, USA;
‡
Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona 85721, USA;
§
Department of Psychiatry, University of Washington, Seattle, Washington 98195, USA
ABSTRACT
Potential toxicity from exposure to
mercury vapor (Hg
o
) from dental amalgam fillings is
the subject of current public health debate in many
countries. We evaluated potential central nervous sys-
tem (CNS) toxicity associated with handling Hg-con-
taining amalgam materials among dental personnel
with very low levels of Hg
o
exposure (i.e., urinary Hg
õ4 mg/l), applying a neurobehavioral test battery to
evaluate CNS functions in relation to both recent ex-
posure and Hg body burden. New distinctions be-
tween subtle preclinical effects on symptoms, mood,
motor function, and cognition were found associated
with Hg body burden as compared with those asso-
ciated with recent exposure. The pattern of results,
comparable to findings previously reported among
subjects with urinary Hg ú50 mg/l, presents convinc-
ing new evidence of adverse behavioral effects asso-
ciated with low Hg
o
exposures within the range of
that received by the general population.—Echever-
ria, D., Aposhian, H. V., Woods, J. S., Heyer, N. J.,
Aposhian, M. M., Bittner, A. C., Jr., Mahurin, R. K.
Neurobehavioral effects from exposure to dental
amalgam Hg
o
: new distinctions between recent ex-
posure and Hg body burden. FASEB J. 12, 971–980
(1998);
Key Words: behavior · elemental mercury · dentists · DMPS
T
HE CRITICAL TARGET
organ of elemental mercury va-
por (Hg
o
)
2
is the central nervous system (CNS)(1).
Although there is little debate regarding the toxicity
of exposure to Hg
o
associated with urinary Hg con-
centrations above 50 mg/l, no consensus exists with
respect to a safe lower Hg
o
exposure level among ei-
ther dental populations that handle Hg amalgam or
the general population with amalgam restorations.
Hg
o
exposures in this study are relevant to both
groups, since they were assessed in a dental popula-
tion but extend over a continuum of urinary Hg lev-
els from 0 to 4 mg/l, comparable to the low exposure
levels observed in the general U.S. population. Gen-
eral population levels provided by Dr. P. Factor (per-
sonal communication) and Dr. A. Kingman (2) range
respectively from 1.3 to 18 mg/l (meanÅ9 mg/l cre-
atinine corrected) and from 0 to 34 (meanÅ3.1 mg/
l). Thus, this study addresses public health concerns
for Hg
o
toxicity of dental amalgams.
Interpretation of health effects observed among
people with Hg
o
exposures resulting in urinary Hg
levels of less than 50 mg/l has previously been ham-
pered by the inability to distinguish behavioral ef-
fects associated with recent exposure vs. those asso-
ciated with chronic body burden. This study adopted
a novel approach to distinguish between these effects
by examining differences in behavior in relation to
urinary Hg concentrations measured both before
(prechelation) and after (postchelation) treatment
of subjects with the Hg mobilizing agent, sodium
2,3-dimercapto-propane-1-sulfonate (DMPS). Urinary
mercury levels (HgU) subsequent to DMPS chal-
lenge have been reviewed extensively (3, 4) and
shown to constitute a better approximation of Hg
body burden (5).
A central question is the validity of using preche-
lation HgU as a proxy for CNS dose. This indirect
measure has been commonly accepted, because the
lipophilic property and low vapor pressure of Hg
o
(0.005 mm Hg
o
at 377C) permit 76–80% of the vapor
to be absorbed through the lungs. The dissolved va-
por is oxidized primarily in erythrocytes into mercu-
ric ions by the hydrogen peroxide-catalase pathway
(i.e., Hg
o
r Hg
1/
r Hg
2/
) (6). The oxidation process
1
Correspondence: Battelle Centers for Public Health Re-
search and Evaluation, 4000 NE 41st St., Seattle, WA 98105,
USA
2
Abbreviations: NES, Neurobehavioral Evaluation System;
Hg
o
, mercury vapor; CNS, central nervous system; DMPS, so-
dium 2,3-dimercapto-propane-1-sulfonate; HgU, urinary mer-
cury levels; POMS, Profile on Mood State.
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is dose dependent in that low doses result in a higher
proportion of Hg
2/
in blood than do higher doses
approaching saturation (7). Blood Hg levels in dental
personnel populations range from 1.2 to 14 mg/l (8),
well below saturation levels, assuring that the oxida-
tive pathways in the kidney and brain are also below
saturation. Thus, HgU varies with occupational Hg
o
exposure and provides an effective measure of cur-
rent dose. With respect to the CNS, the rate of oxi-
dation is slower than circulation time from the lung
to the brain, allowing unoxidized Hg
o
to pass
through the blood–brain barrier, where it is then ox-
idized to the divalent form (Hg
2/
), complexed, and
retained. Controlled radioactive Hg
o
inhalation stud-
ies in humans indicate that the brain retains Hg for
approximately 21 days (9), providing for CNS accu-
mulation and stabilization over approximately 1
month. This compares well to the 2 month half-life
of Hg in urine. These factors have collectively sup-
ported the validity of using prechelation HgU as an
indicator of CNS dose associated with recent sub-
chronic Hg
o
exposure.
Behavioral studies that rely on prechelation HgU
are necessarily limited to evaluations of recent ex-
posure because of the relatively short residence or
half-life of Hg in the urine compared with that ac-
cumulated in soft body tissues. Aposhian et al. (5)
have demonstrated that administering DMPS at a
dose of 300 mg p.o after an 11 h fast effectively mo-
bilizes Hg from soft tissues, which is then excreted
in urine over the subsequent 0–6 and 6–24 h periods.
The postchelation HgU level reflects the decrease of
Hg in both the kidney and cellular fraction of blood,
suggesting that DMPS reduces the renal whole-body
burden of mercury in humans (10). Examining CNS
effects in relation to both pre- and postchelation
measures in this study permitted differentiating CNS
effects associated with recent subchronic Hg
o
expo-
sure from those associated with body burden, which
are attributable to more persistent long-term expo-
sures.
Studies assessing CNS preclinical effects among
subjects with urinary Hg concentrations in the range
of 50–200 mg/l support four aspects of frank mer-
curialism (11): 1) psychosomatic symptoms (saliva-
tion, insomnia, and loss of appetite); 2) alterations in
affect or emotional liability [mood swings, irritability,
fatigue, loss of interest, withdrawal, and sweating and
blushing (erethism)]; 3) motor effects (in the arms,
progressing to uncoordination, imbalance, and cer-
ebella ataxia and tremor in muscles that are highly
enervated and perform fine motor control of extrem-
ities such as fingers, eyelids, and lips); and 4) insidi-
ous loss of mental capacity (progressively affecting
memory, logical reasoning, or intelligence).
Occupational studies assessing urinary Hg
o
levels
between 2 and 200 mg/l have demonstrated impres-
sive consistency with the four aspects of mercurialism
summarized above. Alterations of emotional state,
mood, and symptoms have been the most frequently
reported effects at HgU levels ranging between 30
and 100 mg/l (12–18). Six dental studies have previ-
ously examined mood (19–24) where scores on the
Profile on Mood States (POMS) (25) and aggression
were higher than controls, supporting our choice of
using POMS in this study.
Deficits in motor function were first reported as
finger tremor among felters and later as hand tremor
among chloralkali workers (26). However, losses in
hand steadiness, finger tapping (27, 28), and manual
dexterity (29, 30) have also been reported at lower
levels of exposure. Among dentists with mean urinary
levels of 26 mg/l, statistically significant losses in per-
formance in hand steadiness (known to be correlated
with tremor) were also found (30). These studies sup-
port a comprehensive evaluation of motor function
at even lower levels of exposure as a threshold level
of effect remains to be determined.
Determinations of a lower threshold for cognitive
effects are complicated by mixed results among sev-
eral chloralkali worker studies at low exposure levels
ranging between 0.025 and 0.076 mg/m
3
(10–19.9
mg/l in blood) (31–33). These conflicting results may
be better addressed by studying more uniform sub-
jects, such as dentists, who have similar economic and
educational backgrounds. For example, our own pi-
lot dental studies have detected a reduction in cog-
nitive skills (22–24) similar to that seen in other den-
tal studies (19–21). The largest dental study (19)
conducted in Singapore examined 98 dentists and 54
nondentist controls, where mean exposures of 16.7
mg/m
3
Hg
o
in air were associated with differences in
trailmaking, digit-symbol, digit span, logical memory
delayed recall, and visual reproduction. Two other
dental studies (20, 21) also found associations be-
tween chronic exposure with visuographic memory
deficits by using the Bender-Gestalt (21) (one of four
tests) and Rey’s recurrent figures (20) tests (one of
six tests), which also included the PASAT, Rey’s AVL,
finger tapping, and the grooved peg board. These
findings support placing emphasis on the cognitive
domain.
The evidence for potential impairment among
the four domains provides the basis for test selec-
tion on an anticipated continuum between preclin-
ical effects and clinical deficits. Along this contin-
uum, a preclinical effect is defined as a subtle
adverse change in performance not usually de-
tected by clinical examination because the ob-
served effect falls within the range of normal per-
formance on tasks. However, preclinical effects can
be demonstrated by showing that the variation in
cognitive task performance, though well within the
normal range, is correlated with exposure to Hg.
Preclinical effects range between 3 and 18% when
compared to a zero or low-exposure group. Deficits
EXPOSURE TO DENTAL AMALGAM Hg
o
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exceeding 18% are likely to border on clinical sig-
nificance, and effects of less than 2% are not likely
be occupationally relevant (34).
The broad diversity in clinical effects coupled with
the evidence from epidemiologic studies indicates
more than one mechanism of toxicity is involved, cov-
ering several areas of the brain. Consequently, we
base our behavioral hypothesis on the results dis-
cussed above, which suggest that low-level Hg
o
ex-
posures may increase symptoms, alter mood, de-
crease manual coordination, increase tremor, and
cause deterioration of cognitive skills requiring vi-
sual-spatial memory and attention. We consider ad-
verse effects on these four domains to be selective,
leaving language and retrograde memory largely in-
tact. This justifies the use of each subject’s vocabulary
score as an available measure of premorbid intelli-
gence or a ‘hold test’ not expected to be adversely
affected by exposure. Our test battery was designed
to cover the four domains with adequate redundancy
to detect subtle effects and to discriminate between
areas resistant to Hg insult. Tests were selected for
their sensitivity to Hg
o
; their ability to be adapted for
joint human/animal assessments, which provides a
broader understanding of the results; previous vali-
dation by the World Health Organization (35) and
the Agency for Toxic Substances and Disease Registry
(ATSDR) (36); and use in quantifying neurotoxic ef-
fects attributable to low-level exposures.
MATERIALS AND METHODS
The study population and test procedures
Thirty-four practicing dentists and 15 dental assistants were
selected to participate in this study. The study group was ad-
ministered a pretest questionnaire that medically screened
subjects for preexisting clinical disorders that may interfere
with performance on the test battery such as physical injury,
diabetes, epilepsy, alcoholism, multiple sclerosis, encephalitis,
manic depression, and use of medications that produce drows-
iness or otherwise affect performance. Two subjects were elim-
inated: one was diabetic, the other was an alcoholic. The study
population was predominantly male (69%), Caucasian (92%),
middle-aged (mean ageÅ49), native English speaking, right-
handed (88%), and consumed a moderate number of alco-
holic beverages per week (nÅ3,
SD
Å4.5). This dental popu-
lation is an ideal study population, with characteristics that
improve detection of subtle preclinical effects, as they have
Hg
o
exposures within the range of interest (prechelation HgU
õ 4 mg/l), are highly educated, have excellent test-taking
skills, and have well-developed motor skills.
Prior to the DMPS challenge, participants signed a consent
form in accordance with the Declaration of Geneva of the
World Medical Assembly, and completed a questionnaire cov-
ering occupational and medical work histories and work prac-
tices (37). Subjects also completed an assessment of symptoms
and the POMS. This process was followed by a 1 h test battery
described in greater detail below. After the test session, par-
ticipants were administered DMPS (300 mg, p.o.) after an 11
h fast. Urine was collected from 011 to 0 h prechelation and
from 0 to 6 h postchelation. Test administrators were blind
with respect to subjects’ HgU status.
The behavioral test battery
The test battery described below evaluated symptoms, mood,
motor function, and cognition, using the Neurobehavioral
Evaluation System (NES) vocabulary score (38) as an estimate
of premorbid intelligence.
Symptoms (22)
The symptom questionnaire was adapted from several previ-
ous questionnaires that were designed to evaluate potential
CNS effects of mercury. Responses to persistent symptoms that
last for more than a year were collected on a continuous scale
that permits evaluations of both the extent and severity of
symptoms among subjects.
Profile of mood states (25)
The self-administered mood scales include 65 mood descrip-
tors, which are rated on a 5-point scale from ‘none at all’ to
‘extreme’. The items comprise six mood scales: total mood
(sum of all mood scales except vigor), tension, depression,
anger, fatigue, and confusion.
The hand steadiness battery (39)
This task requires subjects to place pins in a series of holes
with decreasing diameters in a prescribed manner as quickly
as possible, where the number of hits and latencies for eight
holes are recorded.
Simple reaction time NES (38, 40)
This computerized NES task requires subjects to press a button
with the index finger of the right and left hands every time a
stimulus appears on the screen.
Finger tapping NES (38)
This computerized NES task measures motor quickness and
accuracy. It requires subjects to tap a button as many times as
possible in 10 s under three conditions (dominant index fin-
ger, nondominant, and alternate two-button tap with domi-
nant index finger).
Tremor Analysis Test System: acceleration finger tremor (41)
The subject was asked to keep still for three 10 s trials, with a
15 s rest period between trials. Resting tremor (of the domi-
nant and nondominant hands) was recorded by a two-axis mi-
croaccelerometer embedded in the tip of a 12 cm 1 0.8 cm
pencil. The accelerations are normalized by using Fourier
analysis to get the power distribution in the frequency band
of 0.9–15 Hz. The tremor spectra show absolute and relative
power over the range in 0.2 Hz bands, encompassing the 6.5
Hz region previously found to be affected by mercury. Tremor
intensity was determined by the root-mean-square of acceler-
ations over the data collection period. The accelerometers
have a sensitivity of 85 mV per Gauss and a frequency response
rate of 1–830 Hz. A sample rate of 60 Hz reduces noise from
power lines. The fast Fourier transform used a sample size of
512. At 60 Hz, each sample of 512 requires 8.53 s of data col-
974 Vol. 12 August 1998 The FASEB Journal ECHEVERRIA ET AL.
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lection, which is amplified, processed, and transferred to a
portable IBM-compatible computer.
One-hole pins (pins/min) (42)
This computerized task requires subjects to place pins in a
hole in a prescribed manner as fast as possible for five 1 min
trials. The posture of the subject is controlled by having the
dominant hand pick up pins while the other is resting on a
fixed plate. Secondary measures include the time to ‘grasp’,
’move’, ‘position’, and ‘reach’ for each pin.
Vocabulary NES (38)
The NES computerized test is a modification of the Armed
Forces Qualifying Test. Twenty-five words are presented by
computer, and the subject is asked to select, from a set of four
words, the synonym for the word originally presented. This
‘hold’ test was not expected to vary with Hg exposure,
and serves as a measure of premorbid intelligence and user
schooling.
Recognition memory test (for words) (43)
This is a word memory test in which subjects are asked to
correctly recall 50 words that are administered in a fixed se-
quence.
Trailmaking A and B (44)
This paper-and-pencil test is an executive function task assess-
ing cognitive tracking. In A trails, the subject must track a
numeric sequence on a spatial array. In B trails, the subject
must alternate meaningful sequences of numbers and letters
on a spatial array. Both tasks were scored for the number of
errors and response time.
Visual retention test NES (38)
The NES computerized test requires the subject to memorize
a picture and then select the correct one of four possible
choices. Over the course of 12 trials, the pictures become
more complex. This test is a computerized version of the Ben-
ton Memory Task.
The switching task (45)
This computer-presented task requires subjects to press a
‘same’ or ‘different’ button when confronted with a pattern
comparison, semantic letter comparison, or semantic graphi-
cal comparison. Items are presented in apparent random or-
der, but actually follow a complete Latin square procedure
balanced for residual effects. This test modifies the traditional
design by inserting extra trials to achieve greater stability of
the estimates of switching between tasks and to avoid the abil-
ity of subjects to predict the next task. These additions provide
a total of eight repetitions for the six switching combinations
and between 10 to 16 repetitions for the three control con-
ditions.
Symbol-digit NES (38)
This coding task requires the subject to enter the number that
matches a symbol by using a matched set printed at the top of
the screen. The task requires fine manual dexterity, visual
scanning, and motor speed. A visual memory component in-
creases performance speed if used by the subject.
Exposure to urinary mercury
HgU was analyzed by using cold vapor atomic absorption (5).
Both occupational and nonoccupational sources of Hg expo-
sure were covered in questionnaires, including the number of
amalgam fillings in the subject’s mouth and seafood con-
sumption. Hg speciation analysis revealed that organic intake
of Hg was negligible. Personal habits and detailed work prac-
tices were later used in regression models to determine factors
predicting HgU levels.
Statistical methods
Multiple regression was used to evaluate log-linear, dose-effect
behavioral relationships for pre- and postchelation urinary Hg
values, controlling for age, race, gender, vocabulary, alcohol
consumption, and wearing of eyeglasses (46). Regression
models for cognitive outcomes also included the log SRT of
the dominant hand to control for motor subcomponents of
the cognitive tasks. Summary factor scores (30) were created
to reduce multiple outcomes to single scores on select tests
with enhanced reliabilities (hand steadiness, finger tapping,
tremor, and switching attention). Paired t tests between pre-
and postchelation HgU coefficients were used to evaluate
whether the two dose measures differed significantly in pre-
dicting specific test performance. Standardized beta coeffi-
cients are a partial correlation coefficient, indicating the
unique variance associated with each measure of exposure
when all other variables in the model have been accounted
for (47). Standardized beta is a metric that can be appropri-
ately compared across independent variables and different do-
mains.
RESULTS
Work-related and personal factors (including
postchelation HgU levels and number of amalgam
fillings in one’s mouth) were determinants of
prechelation HgU levels
There was an order of magnitude difference between
pre- and postchelation urinary Hg concentrations
(Fig. 1). The correlation between the two samples
(rÅ0.53), although significant, indicated that differ-
ent parameters were being assessed, given very high
reliabilities of urinary Hg assessments across a day in
a previous report (27, 28). HgUs of the dentists and
dental assistants did not differ pre- or postchelation
(prechelation: dentistsÅ0.89 mg/l,
SD
Å0.51; dental
assistantsÅ1.07 mg/l,
SD
Å0.93; postchelation: den-
tistsÅ10.08 mg/l,
SD
Å7.37; dental assistantsÅ8.07 mg/
l,
SD
Å5.99). Several work-related factors that are ame-
nable to intervention were significantly associated
with prechelation HgU as well as one personal source
of exposure (Table 1). These factors include the
number of restorations placed per week, the use of
dispensers vs. capsules, and the irregular use of a
mask while handling Hg, as well as the number of
EXPOSURE TO DENTAL AMALGAM Hg
o
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Figure 1. Histograms of pre- and postchelation urinary mercury levels (HgU).
amalgam fillings in one’s mouth. All personnel wore
gloves.
Pre- and postchelation HgU levels result in distinct
patterns of preclinical effects, providing evidence
of associations with recent exposure and chronic
body burden
The patterns of association between symptoms,
mood, motor function, and cognition with pre- and
postchelation urinary Hg measures were distinct.
Standardized beta coefficients for pre- and postche-
lation urinary Hg levels showed that all four domains
were associated with Hg
o
exposure (Fig. 2). However,
an overview of standardized betas indicates that the
relative sensitivity of the two urinary measures dif-
fered considerably for individual tests.
Persistent symptoms involving memory and head-
aches, lightheadedness, and dizziness were selectively
associated with postchelation HgU (Table 2). In con-
trast, transient mood scores were associated with both
pre-and postchelation HgU values, where the associ-
ations for all five mood scales were robust and uni-
formly associated with prechelation HgU (exceeding
all other test scores).
The motor function results showed that finger tap-
ping is also more strongly associated with prechela-
tion HgU, but has a lesser statistically significant as-
sociation with Hg body burden. Hand steadiness had
mixed associations with pre- and postchelation HgU
levels, but differences in the strength of the associa-
tion between recent exposure and Hg body burden
were less pronounced (betaÅ0.39 vs. betaÅ0.30, re-
spectively). Resting tremor, a clinical measure fre-
quently found affected in cases of severe neurode-
generative disease, was not affected as anticipated.
In contrast, cognitive function as measured by
switching attention, trailmaking A and B, and visual
retention memory was selectively associated with pre-
chelation HgU. One exception to this trend was the
number correct in the Word Recognition Memory
Test, the only test dependent on words and memory,
which was associated with postchelation HgU. Sym-
bol-digit response time was not associated with either
measure of exposure to Hg.
The log-linear relationships throughout the range
of postchelation HgU values are illustrated by finger
tapping (right/left/alternate number of taps) and
the total number of symptoms associated with post-
chelation HgU (Fig. 3). No evidence of a threshold
of effect appeared across the body of our results; re-
lationships were smooth and generally conformed to
a log-linear trend (i.e., there was no evidence of sub-
population clustering).
DISCUSSION
This is the first behavioral study to distinguish recent
Hg exposure from Hg body burden when examining
subtle changes in preclinical behavior associated with
very low levels of Hg
o
exposure. The results are strik-
ing in that statistically significant dose-effect relation-
ships were found with prechelation HgU (ranging
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TABLE 1. Work-related and personal factors associated with prechelation HgU
a
Factors Mean
SD
b
preHgU
SE
b
Postchelation HgU 9.1 6.9 0.16 0.04 0.43***
No. of amalgams placed/wk 16.1 8.2 0.01 0.00 0.34***
Do not wear a mask (7/49) 15% 0.38 0.12 0.32***
No. of amalgams in own mouth 1.6 0.8 0.08 0.03 0.25**
a
The mean and standard deviation for each factor in the model is accompanied by the regression
coefficient for prechelation HgU, the standard error, the standardized Beta coefficient that presents the
unique partial correlation for each factor in the model, and the level of significance. Even controlling
for Hg body burden does not eliminate the importance of associations between recent exposure, current
work-related factors, and the number of amalgams in one’s mouth. *** P õ .005, ** P õ .05; full model
R
21
Å 0.61.
Figure 2. A comparison of standardized beta coefficients across regression models (a partial correlation coefficient) for pre- and
postchelation HgU levels in regression models evaluating symptoms, mood, motor function, and cognition. Statistical differences
between the two urinary measures are noted by (.) where the dark bars represent postchelation HgU levels; the horizonal line
reflects the significance level at a P õ 0.05 for each association for both pre- and postchelation HgU levels (nÅ48).
from 0 to 4 mg/l) and postchelation HgU (ranging
from 1 to 32 mg Hg/l). These prechelation HgU con-
centrations were previously thought to be trivial in
terms of potential health risks. The modest correla-
tion between pre- and postchelation HgU measures
(6) supports the view that the Hg present in body
tissues may contribute to apparent associations with
recent exposure, as measured by prechelation HgU.
Similar increases in symptoms (12–23), alterations
in mood (19–24), reduction in speed and accuracy
in motor function (20, 21, 24, 30), and subtle losses
in memory and visuospatial cognitive skills (19–23)
have been reported in studies of dental professionals
(highly consistent with our present findings). How-
ever, mean urinary Hg levels among subjects in these
studies was higher (ú20 mg/l) than found among
EXPOSURE TO DENTAL AMALGAM Hg
o
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TABLE 2. Associations for pre- and postchelation urinary mercury (HgU) values with symptoms, mood, motor function, and cognition
a
Test variable
Prechelation HgU
Mean
SD
b
inHg
SE
b
Postchelation HgU
b
inHg
SE
b
Symptoms
Mean number of symptoms/subject 0.30 0.13 2.44 4.37 0.09 0.005 0.03 0.29*
Memory 1.39 1.22 2.59 19.52 0.02 14.63 6.91 0.32**
Headaches, lightheaded, dizzy 2.39 1.33 7.12 15.27 0.07 10.88 5.46 0.28**
Mood (POMS)
Total mood 11.65 4.82 011.21 2.80 00.56*** 03.07 1.15 00.41**
Tension 3.04 1.40 03.33 0.83 00.57*** 00.91 0.34 00.42**
Anger 2.08 0.92 01.97 0.52 00.53*** 00.51 0.21 00.37**
Fatigue 2.15 2.15 03.57 1.14 00.40** 01.18 0.44 00.36**
Confusion 0.12 0.33 01.19 0.45 00.38** 00.23 0.20 00.20
Depression 2.02 0.98 01.14 0.63 00.29* 00.24 0.24 00.16
Motor
Finger taps
/
(right/left/alternate) 61.1 9.1 02.66 0.71 00.55*** 00.49 0.27 00.30*
Hand steadiness
/,L
(7, 8, 9 hole/s) 1 5.80 13.70 1.08 0.41 0.39** 0.28 0.15 0.30*
Hand steadiness
/,L
(7, 8, 9 hole/s) 2 5.80 13.70 1.15 0.54 0.34** 0.27 0.19 0.24
Simple reaction time
/,L
(R/L s) 0.355 0.039 1.07 0.70 0.24 0.38 0.23 0.25*
Resting tremor
L
(center frequency) 6.98 0.97 0.31 0.29 0.25 8.19 8.85 0.26
One-hole pins (pins/min) 34 6 00.001 0.17 00.01 00.27 1.75 00.03
Cognition
Visual processing/attention
Switching attention
L
(ms) 0.792 0.198 1.58 0.76 0.38** 0.17 0.30 0.18
Trails A (s) 26.20 7.60 10.13 4.56 0.34** 1.14 1.93 0.10
Trails B (s) 66.10 22.32 25.99 15.22 0.28* 7.99 5.84 0.23
Visual retention test
L
(number correct) 5.30 1.50 0.27 0.13 0.26** 0.03 0.05 0.08
Symbol-digit time
L
(s) 22.60 4.20 0.11 0.09 0.16 0.01 0.03 0.03
Verbal processing/attention
Word memory (number correct) 47.60 2.90 01.37 1.96 00.11 01.38 0.71 00.31*
a
Symptom, mood, and motor function regression models include vocabulary, age, gender, race, alcohol, and wearing of glasses; cognitive
regression models include simple reaction time (dominant hand), vocabulary, age, gender, race, alcohol, and wearing of glasses.
†
Factored
scores;
L
log transformed. * P õ 0.10, ** P õ 0.05, *** P õ 0.01.
subjects with comparable preclinical effects reported
here. Likewise, comparable CNS effects have been
consistently reported among subjects with urinary Hg
levels above 50 mg/l (1, 11). These studies collectively
support our prechelation HgU behavioral findings in
each domain and point to a potential log-linear con-
tinuum of health effects from low to high Hg expo-
sure.
However, questions remain unanswered concern-
ing the lower threshold of Hg
o
exposure for behav-
ioral effects, as we found no indication of a lower
boundary in any of our subjective or objective results
(see Fig. 3). This inability to detect a threshold ex-
posure level strengthens the hypothesis that subtle
preclinical effects found at very low levels of Hg
o
ex-
posure appear on a continuum with the far more se-
vere clinical deficits. Of equal importance, we also
found no evidence of special susceptibility within a
subset of this dental population. Behavioral re-
sponses typically increased with exposure in a fairly
uniform manner, indicating a more general re-
sponse, as illustrated in Fig. 3.
The patterns of subjective responses associated
with HgU differed. Persistent symptoms that appear
over a year were selectively associated with Hg body
burden; this finding suggests that symptoms may re-
main undetected in evaluations that rely solely on
prechelation urinary measures of Hg
o
exposure. In
contrast, the more transient nature of the POMS was
found to be more strongly associated with recent ex-
posure, with a smaller but statistically significant con-
tribution from Hg body burden. This pattern sug-
gests that prechelation HgU levels, which are partially
dependent on the 2-month half-life, are more
strongly associated with mood; one may speculate
that the amount of Hg stored as body burden is less
associated, as it may be biologically less available to
the CNS.
This study comprehensively assessed fine manual
speed, accuracy, and coordination, measures of par-
ticular relevance for dental professionals who work
with handheld tools. Among the five motor function
tests, individual and factored performance scores for
finger tapping and hand steadiness were also associ-
ated with recent exposure, as measured by preche-
lation HgU. The standardized beta of 00.55 for fin-
ger tapping was comparable to a beta of 00.56 for
total mood, indicating a relatively strong association.
Similar to the results for mood, performance on both
motor tests also had smaller but detectable associa-
978 Vol. 12 August 1998 The FASEB Journal ECHEVERRIA ET AL.
/ 382d 0012 Mp 978 Friday Aug 07 08:43 AM LP–FASEB 0012
Figure 3. Exposure-effect relationships between postchelation HgU levels and the number of taps/min for
finger tapping (a) and the sum of symptoms reported per subject (b).
tions with Hg body burden. These results suggest that
distinctions between recent and long-term exposure
could prove even more relevant at higher levels of
exposure.
Unlike the other three behavioral domains, the
cognitive regression models included the log SRT of
the nondominant hand to control for potential mis-
interpretation associated with slowed motor re-
sponse. Only one association was observed between
Hg body burden and the number correct for word
recognition memory, a measure of verbal memory.
Associations with recent exposure were found for
switching attention, a measure of selective attention;
trailmaking parts A and B, measures of psychomotor
speed and cognitive flexibility; and the Visual Reten-
tion Memory Test, a measure of visual memory. The
subtle impairments across this set of cognitive tests
suggest diffuse nonspecific alterations in task perfor-
mance that are potentially associated with aspects of
attention. These cognitive effects were selectively as-
sociated with recent exposure, with no detectable
contribution from Hg body burden, whereas verbal
memory was associated only with long-term exposure
or Hg body burden, as measured by postchelation Hg
levels.
The observed pattern of statistically significant re-
sults for prechelation HgU had coefficients that were
relatively strong for POMS mood scores (median
betaÅ0.45, range 0.29–0.56) and moderate for the
more objective domains of motor function (median
betaÅ0.42, rangeÅ0.30–0.55) and cognitive tasks
(median betaÅ0.31, rangeÅ0.26–0.38) (see Table
2). Associations with postchelation HgU showed the
standardized beta coefficients in each domain were
also moderate for symptoms (medianÅ0.31, range
0.28–0.32), relatively strong for mood though less
pronounced than the prechelation coefficients for
mood (medianÅ0.40, range 0.36–0.42); moderate
for motor function (medianÅ0.30, rangeÅ0.25–0.30;
and sparse but moderate for one cognitive task
(betaÅ0.31). One interpretation of these variations
in strength of associations between pre- and post-
chelation HgU is that urinary Hg from recent expo-
sure is more likely to be available to the CNS and
should be more strongly associated with preclinical
effects, with less contribution from stored Hg as body
burden.
Good occupational work practices further reduce
urinary Hg levels given the behavioral effects associ-
ated with low levels of HgU (see Table 1). The vari-
ation of HgU in a national sample of 6925 dentists
participating in the American Dental Association
Health Screening Program was related to the varia-
tion in air Hg levels, and industrial hygiene surveys
among dental offices show that 6–16% of dental prac-
tices exceed exposure levels permissible by OSHA
EXPOSURE TO DENTAL AMALGAM Hg
o
979
/ 382d 0012 Mp 979 Friday Aug 07 08:43 AM LP–FASEB 0012
(37). This reinforces the need to comply with indus-
trial hygiene guidelines in dental offices.
The distribution of HgU in the national HgU sam-
ple had a range of 0 to 104 mg/l and was not distrib-
uted normally: 10% had levels ú10.4 mg/l, 3% had
levels ú18.8 mg/l, 2.5% had levels ú20.4 mg/l, and
1% had levels ú33.4 mg/l. In this study, our mean
prechelation urinary Hg concentration of 0.94 mg/l
(
SD
Å0.50) corresponds to the lowest 10th percentile
of dentists in the United States. Further, as noted ear-
lier, general population HgU levels (P. Factor, per-
sonal communication; ref 2) overlap with these oc-
cupational levels, supporting a public health concern
for very low-level Hg
o
toxicity.
Concern for very low-level Hg
o
toxicity is supported
by our observations of associations at HgU levels well
below the proposed biological standard of 25 mg/l
(16, 17) and below urinary levels that would be ex-
pected at the OSHA permissible exposure limit of 50
mgHg
o
/m
3
in air (48). The low Hg
o
exposures be-
tween 0 and 4 mg/l were partially attributable to the
number of Hg amalgam fillings in the dental group
(as seen in Table 1). The apparent association be-
tween HgU and personal risk factors argues for fu-
ture studies to examine the potential for similar ad-
verse effects in the general population from Hg
amalgam fillings. Some might argue that the present
findings have immediate implications regarding the
continued use of Hg amalgam in dental restorations
(49). We are divided on this issue, inasmuch as there
are currently unanswered toxicological questions re-
garding chronic Hg body burden, dose-rate, and po-
tential differences in modality of exposure derived
from amalgam restorations alone. Two clinical trials
regarding the safety of dental amalgams among chil-
dren, a potentially more susceptible population, will
not be completed for several years (50). Nevertheless,
it is clear from the present study that comparing as-
sociations with pre- and postchelation urinary Hg lev-
els revealed patterns of previously unobserved effects.
These would not have been identified if they had
been evaluated in relation to the traditional preche-
lation urinary Hg levels alone. Thus, the DMPS che-
lation technique enhances interpretation of observed
associations with low-level cumulative Hg
o
exposure.
In conclusion, by using an approach that distin-
guishes recent Hg exposure from Hg body burden,
we have observed subtle associations between Hg and
symptoms, mood, motor function, and nonspecific
cognitive alterations in task performance in an oc-
cupationally exposed group with HgU levels compa-
rable to the general U.S. population. Application of
this approach may be particularly useful in defining
thresholds of Hg
o
toxicity and for establishing safe
limits of exposure to mercury from dental amalgam
material, the restoration itself, diet, and other
sources.
Supported by National Institutes of Health grants DE11712,
ES04696, ES04940, and by the Wallace Research Foundation.
Support was also provided by the University of Washington
Center for Ecogenetics and Environmental Health (P30
ES07033). We thank Heyl Co., Berlin, Germany, for the gift
of DIMAVALt (DMPS).
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Received for publication December 1, 1997.
Revised for publication March 27, 1998.
