Curvilinear Relationship Between Blood Lead Level and Reaction Time: Differential Association with Blood Lead Fractions Derived from Exogenous and Endogenous Sources

Bleecker, Margit L. MD, PhD; Lindgren, Karen N. PhD; Tiburzi, Michael J. MA; Ford, D. Patrick MD, MPH

Journal of Occupational & Environmental Medicine:
Original Article

Prior studies demonstrate an inconsistent relationship between occupational inorganic lead exposure and simple reaction time (SRT) performance. In this study, we administered a computerized SRT test to 78 currently employed lead smelter workers and then investigated the relationship between different measures of blood lead and components of SRT performance. The measures of blood lead included current blood lead (PbB) and mathematically derived blood lead fractions from the environment (PbB-env) and from bone (PbB-bn). Measures of SRT performance, obtained from 44 trials with interstimulus intervals (ISIs) ranging from 1 to 10 seconds, included median SRT (SRT-md), mean SRT for ISIs between 1 and 5 seconds (SRT-1-5), and mean SRT for ISIs between 6 and 10 seconds (SRT-6-10). Multiple regression analysis after adjustment for age and education revealed a curvilinear relationship between PbB and SRT-md. As PbB increased from 0 to 30 µg dl-1, SRT-md decreased, and only with PbB levels above 30 µg dl-1 did SRT-md increase. PbB terms accounted for 13.7% of the variance in this SRT measure (P < 0.01). The longer ISI variable, SRT-6-10, was found to be more strongly related to PbB, to have lesser variability across ISIs, and to be unrelated to age. Additional multiple regression analysis to examine the relationship between components of SRT and the PbB fractions, PbB-env and PbB-bn, showed only PbB-env to account for significant variance in SRT-md, (14.4%, P < 0.01), SRT-1-5 (9.7%, P < 0.03), and SRT-1-6 (15%, P < 0.01). We conclude that the relationship between PbB and SRT is U-shaped, that the SRT measure SRT-6-10 has properties that make it the preferred measure of SRT performance in future studies, and finally that only PbB-env, and not PbB-bn, is related to components of SRT.

Author Information

From the Center for Occupational and Environmental Neurology, Baltimore, Md.

Address correspondence to: Margit L. Bleecker, MD, PhD, Center for Occupational and Environmental Neurology, Children's Hospital Professional Building, 3901 Greenspring Avenue, Suite 101, Baltimore, MD 21211.

Article Outline

Simple reaction time (SRT), a measure of readiness to respond, attention, and vigilance, is part of many neurobehavioral batteries used to detect the presence of nervous-system effects associated with exposure to neurotoxic coumpounds.1-13 The World Health Organization's Neurobehavioral Core Test Battery (WHO-NCTB) included SRT because it fulfilled the criteria of (1) ability to detect early evidence of central nervous system effects, (2) resistance to cultural differences because language requirements are minimal, and (3) ease of administration. 14 SRT from the WHO-NCTB was programmed for computer use as part of the Milan Automated Neurobehavioral System (MANS), eliminating the need to manually record and enter data.15 Unfortunately, few neurotoxicity investigations have used this version of SRT to allow for interstudy comparisons.

The literature on SRT performance in the setting of occupational inorganic lead exposure reports variable results, ranging from no change to both slowed and faster reaction times.1-13 Even when the SRT is significantly different between lead-exposed groups and control groups, a dose-effect relationship between SRT performance and lead exposure is rarely found.2,5,7,8,11,13 One explanation for these variable findings could be the possibility of a nonlinear relationship between SRT and PbB.

PbB, the biomarker used routinely by industry to monitor occupational inorganic lead exposure, is a composite of lead contribution from the environment and from internal stores, primarily bone.16 One recent report established that for the lead workers in the study presented here, the contribution of bone lead to blood lead is 0.17 µg Pb dl-1 (µg Pb (g bone mineral)-1)-1, eg, for a bone lead level of 100 µg Pb (g bone mineral)-1, 17 µg Pb dl-1 of the PbB present at that time derives from internal bone stores, with the remainder derived from environmental sources.17 It has been proposed that lead released from bone remains primarily in the serum compartment, rather than binding to the red blood cell. As such, it is expected to diffuse more readily into target tissues and pose a greater toxicological threat than environmentally absorbed lead, which is largely bound in the red blood cell.18 If lead released from bone is more biologically available to produce health effects, then a differential association between SRT and blood lead derived from bone lead stores versus blood lead absorbed from the ambient environment would be expected.

This article examines SRT in smelter workers to determine the nature of the relationship between PbB and measures of SRT and further queries as to whether measures of SRT are differentially associated with PbB derived from an endogenous source versus PbB derived from an exogenous source.

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The group of subjects consisted of 80 currently employed lead smelter workers who were participants in a neurotoxicologic evaluation of a lead smelter. Other aspects of this study have been described elsewhere.17,19 Demographic data for these subjects are presented in Table 1.

All workers volunteered for the study and signed an informed consent form; the bone lead protocol was approved by the Human Subject Committee at the University of Maryland, Baltimore, MD.

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The lead smelter, in operation since 1966, had a sinter plant, treated gases in a contact sulfuric acid plant, and used standard kettle methods for lead refining. PbB, obtained on the same day that SRT testing was performed, was analyzed by a Centers for Disease Control and Prevention-approved laboratory using atomic absorption spectroscopy with a graphite furnace. Bone lead analysis, quantitated at the mid-tibia with K-XRF measurements, was performed at the University of Maryland Toxicology Program laboratories, using methods described by Chettle et al.20 Daily calibrations, both before and after subject measurements, were made against standard-addition plaster of Paris phantoms. Negative values for bone lead were possible because of measurement error. Bone lead levels, PbB, and SRT were all measured within 1 day.

Bone lead measurements were used to fractionate PbB into components derived from endogenous (bone) stores (PbB-bn) vs PbB derived from the environment (PbB-env). The fractionation was performed using the aforementioned relationship, Pb-bn = 0.178 ([µg dl-1] [µg Pb {g bone mineral-1}]-1), which predicts the contribution of a given bone lead level to a given blood lead level.17 This relationship, established with members of the present cohort, used PbB obtained after a 10-month strike in 1991, during which time there was no occupational exposure. The slope of this PbB vs bone lead established the relationship of bone lead in predicting PbB.17 Therefore, for each subject, in the present analysis, PbB-bn was determined using this relationship, and PbB-env was calculated by subtracting PbB-bn from PbB (Pb-env = PbB - Pb-bn) (Table 1).

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Simple Reaction Time

SRT was measured using a modified MANS15 loaded on a standard desktop computer. The stimulus, a red square, was presented on the screen of the computer monitor, with interstimulus intervals (ISIs) ranging from 1 to 10 seconds. The subject responded to the presentation of the stimulus by depressing the spacebar of the computer keyboard as quickly as possible after the appearance of the stimulus. The subject maintained contact between the space bar and dominant index finger to minimize movement time. Reaction times were recorded on the screen to motivate the subject. Eleven practice trials were followed by 44 responses. The major modification of the testing system was that the ISIs were not allowed to be pseudorandom but were predetermined to ensure that each subject was exposed to the same pattern of ISIs. Also all ISIs from 1 to 10 seconds were analyzed, not just ISIs between 1 and 5 seconds as in the MANS.

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Hypothesized confounders included age, education, current depressive symptomatology, current alcohol abuse, and peripheral nerve function. These confounders were measured as follows: age in years, education in years of schooling, depressive symptomatology in Profile of Mood States Depression subscale score, alcohol abuse in ounce-equivalents of alcohol consumed weekly, and peripheral nerve function in Current Perception Threshold (CPT) score in milliamperes (mA).

CPT is a means of quantifying peripheral nerve function. A stimulation frequency of 2000 Hz for the large myelinated fibers provided the CPT (in mA) for the ring finger and second toe on the nondominant side. Nondominant side was allowed because the peripheral neurotoxic effects of lead are expected to be bilateral. Forced-choice paradigm used to determine the threshold occurred with a consistent report of feeling a stimulus at one level of intensity and not at a slightly lower intensity.

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Data Analysis

The Statistical Package for the Social Sciences (SPSS-PC; SPSS Inc., Chicago, IL) was used for the data analyses.

Median SRT (SRT-md) was constructed from the 44 trials. Two outliers, defined as individuals with SRT-md greater than 2.5 standard deviations from the mean of the group, were excluded from the analysis. This resulted in a final n of 78.

For each individual, a truncated mean reaction time (SRT-tr) was created for each ISI to minimize the influence of extreme scores. The mean SRT-tr for ISIs 1 through 5 seconds (SRT-1-5) was calculated by averaging the SRT-tr's for each ISI from 1 through 5 seconds; SRT-6-10 was similarly calculated by averaging the SRT-tr's for each ISI from 6 through 10 seconds.

The relationship between each potential confounder and the outcome variables was examined. Only age and education were found to be significantly related and were retained as covariates.

The relationship between the three measures of blood lead (PbB, PbB-env, and PbB-bn) and the three measures of SRT (SRT-md, SRT-1-5, and SRT-6-10) was examined via multiple linear regressions, with the covariates age and education entered as the first block. Blood lead measures were entered as the independent variables in the second and/or third blocks.

The identical procedure was repeated with bone lead as the independent variable entered in the second block of the regression analysis.

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The mean (SD) SRT-md for all ISIs was 262 milliseconds (ms) (38.9 ms), with a range of 179 to 387 ms. The mean (SD) SRT-1-5 was 295 ms (41.2 ms), with a range of 186 to 442 ms, and the mean (SD) SRT-6-10 was 252 ms (37.5 ms), with a range of 187 to 371 ms.

Multiple linear regression of PbB and bone lead on SRT-md showed PbB to account for 10.1% of the variance in SRT-md, after adjustment for age and education (P < 0.01) whereas bone lead level was nonsignificant. However, the relationship between PbB and SRT-md was inverse, with SRT decreasing with increasing PbB. Examination of residuals suggested a curvilinear relationship. To investigate this possibility, a quadratic term, PbB2 was included in the regression equation. PbB2 accounted for an additional 3.6% of the variance in SRT-md (P < 0.08). Addition of the quadratic term resulted in a U-shaped relationship between PbB and SRT-md, with SRT-md decreasing over the range of 0 to 30 µg dl-1 and then increasing as PbB exceeded 30 µg dl-1 (Figure 1).

Trends in performance across the 10 ISIs were examined by plotting the truncated mean RT (SRT-tr) for each interval against the corresponding interval. SRT-tr variably decreased for ISIs 1 through 5 seconds but plateaued for ISIs 6 through 10 seconds (Figure 2).

Two multiple linear regressions examined the relationship of age and education with SRT-1-5 and SRT-6-10. Age accounted for 3.4% of the variance with SRT-1-5 (P = 0.10), but none with SRT-6-10 (Figure 3). Education was not significantly related to either variable but was included as a covariate based on the SRT-md analysis.

The relationship between exposure, PbB and PbB2, and SRT-1-5 and SRT-6-10 was examined by two multiple linear regressions, with age and education forced in as a covariates, as described above. The combined exposure terms contributed 11.5% of the variance to SRT-1-5 (P < 0.01) and 13.9% of the variance to SRT-6-10 (P < 0.01).

The influence of the different fractions of PbB, PbB-bn and PbB-env, on SRT-md was explored with multiple regression. Age and education were entered first, followed by PbB-bn and PbB-bn2 in the next block and PbB-en and PbB-env2 in the final block. The change in r2 for the PbB-bn terms was 1.3% (P = not statistically significant [ns]), whereas that for the PbB-env terms was 14.4% (P < 0.01). The same analysis was repeated for ISI-1-5 and ISI-6-10. For ISI-1-5 the change in r2 for the PbB-bn terms was 1.5% (P = ns) and for the PbB-env terms was 9.7% (P < 0.03). For ISI-6-10 the change in r2 for the PbB-bn terms was 1.3% (P = ns) and for the PbB-env terms was 15% (P < 0.01). The addition of the PbB-env terms after the PbB-bn terms demonstrated the significant contribution of PbB-env to the variance in SRT-md, SRT-1-5, and SRT-6-10, even after accounting for any explanation in the variance due to PbB-bn.

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The curvilinear relationship of PbB and SRT as demonstrated in this study may explain some of the inconsistencies in the PbB-SRT relationships previously reported in the literature.1-13 Review of 13 prior studies revealed seven examples of significantly slowed SRT in lead-exposed groups, compared with control groups 2,5,7,8,10,11,13 in only four of these studies, however, was the slowed SRT found in the highest lead-exposed groups.5,10,11,13 In the remaining six of the 13 studies, four reported no significant difference in SRT between exposed and nonexposed groups3,4,6,9 and two reported significantly faster SRT in the lead-exposed groups.1,12 Examination of the studies showing no significant difference in SRT revealed that mean SRT was frequently faster (although nonsignificantly) in the exposed groups.4,6,9 Repko et al reported significantly slowed SRT in a lead-exposed group compared with a control group, yet within the lead-exposed group, a negative correlation between SRT and lead exposure implied a trend toward faster performance with increasing lead exposure.2 A U-shaped SRT-PbB relationship in the study by Stollery et al showed the lowest mean SRT in the medium lead-exposed group when compared with the low and high lead-exposed groups.11 The average PbB in these studies has been between 40 and 50 µg dl-1, with a range from 32 to 71 µg dl-1.1-13

SRT in the reviewed studies was affected by at least some parameters other than those present in our study. These include movement of the fifth digit,2 response time to a touch stimulus,1 time to extinguish a light,5 and response to an auditory stimulus.6 Sometimes, if the task is long and monotonous, the SRT of even the low-exposed group will increase across repeated trials.11 These design and administration differences in SRT have all contributed to the variable results reported in the literature.

Shorter ISIs have been hypothesized to be more sensitive to neurotoxic exposure.15 This was not supported in this study, with the longer ISI variable (SRT-6-10) being more highly related to lead exposure. This was likely a result of the lesser intrinsic variability of the longer ISIs-making any lead effect easier to discern-and to the lack of relationship of the longer ISI variable (SRT-6-10) with age.

In neurobehavioral testing, age is a significant confounder, sometimes accounting for approximately 20% of the variance in SRT performance21; such an influence of age could potentially mask subtle changes in SRT resulting from exposure. Therefore a test that minimizes the contribution of age, such as the SRT-6-10, has advantages in monitoring for early neurotoxic effects. Also, in the current study, evidence of fatigue-by comparison of SRT scores between the beginning and end of testing-was not found (data not presented). This feature again improves the test's ability to screen for toxic effects by minimizing competing sources of variability.

The fractions of PbB, PbB-env and PbB-bn, were calculated and not directly measured, but they were still expected to provide reasonable approximations of the contribution of environmental and bone lead to the blood compartment. Although it has been hypothesized that bone serves as a source of toxicologically more significant serum lead,18 we found that PbB-bn contributed negligibly to the PbB-SRT relationship whereas PbB-env contributed significantly. Possible explanations for this contradiction are that our method of fractionation of PbB into PbB-bn and PbB-env was invalid, that Pb-bn is only significantly related to SRT at bone lead levels higher than those present in our subjects, that lead from exogenous sources contributes to serum lead to a greater extent than bone lead, or perhaps that RBC-bound lead, rather than serum lead, is the more relevant proximate toxicant affecting this specific neurobehavioral test.

In summary, we have demonstrated in this study a nonlinear U-shaped relationship between PbB and a measure of SRT. Our results suggests that PbB levels below 30 µg dl-1-an often-acknowledged hygienic standard-are activating, decreasing SRT with increasing levels. Yet this finding is not incongruent with prior neurotoxicologic studies and with animal data.22 Additionally, we have provided reasons for choosing the SRT-6-10 as the preferential measure of SRT in future neurotoxicologic studies. Finally, we have shown that, given certain assumptions, exogenously derived lead is more strongly related with measures of SRT than lead acquired from endogenous (bone) sources.

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Support received for this study from New Brunswick Occupational Health and Safety Commission, New Brunswick, Canada. The authors thank Dr Fiona McNeill for performing the bone lead measurements, Dr Bruce Thompson for statistical consultation, and Shirley Thorpe for secretarial help.

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Cited By:

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