A short period of early Pb exposure produced impairments in learning, attention, inhibitory control, and arousal regulation, paralleling the areas of dysfunction seen in children with low to moderate elevations in blood Pb levels. As in children, these impairments were lasting; that is, they were apparent long past the period of Pb exposure, indicative of lasting neurologic dysfunction as a result of the early exposure. The pervasiveness of the impairment was related to the intensity of Pb exposure; for the Mod-Pb group, dysfunction was limited to learning ability, whereas the High-Pb group exhibited impairments in learning, inhibitory control, arousal regulation, and attention. Importantly, succimer treatment significantly alleviated some Pb-induced neurobehavioral deficits, although the magnitude of the benefit varied as a function of both the level of the Pb exposure and the specific area of dysfunction. In contrast, succimer treatment of rats not previously exposed to Pb produced lasting cognitive and affective dysfunction, similar in magnitude and pervasiveness to that produced by the High-Pb exposure regimen. The basis for each of these conclusions is provided below and summarized in .
Each of the areas of impairment seen in the High-Pb, Mod-Pb, and succimer-only groups, followed by the specific finding upon which the functional inference is based.
Pb-induced learning deficits and efficacy of succimer treatment
Both the High-Pb and Mod-Pb groups learned the basic rules of the visual discrimination task and attention task 1 more slowly than the controls, indicating lasting impairment in associative ability as a result of a short period of early Pb exposure, as previously reported (e.g., Garavan et al. 2000
). Succimer treatment of the Pb-exposed animals improved learning rate, although the degree of benefit was greater for the Mod-Pb group than for the High-Pb group. Succimer treatment significantly improved the learning rate of the rats in the Mod-Pb group for both the initial visual discrimination task and attention task 1; in both tasks, the Mod-Pb–succimer group learned significantly faster than the Mod-Pb group and did not differ significantly from the controls. For the High-Pb exposure regimen, succimer treatment did not improve learning rate in the visual discrimination task, but a small benefit was seen in attention task 1. It is likely that more prolonged succimer treatment would have produced an even greater cognitive benefit for these more heavily exposed animals. This suggestion is based on the facts that a
) reductions in brain Pb levels lag significantly behind reductions in blood Pb following chelation or cessation of Pb exposure (Cremin et al. 1999
; Smith et al. 1998
; Stangle et al. 2004
) blood and brain Pb levels were still quite elevated in the High-Pb group at the end of chelation (approximately equal to the nonchelated Mod-Pb rats); and c
) in a previous study from our laboratory, a second 3-week succimer regimen offered significant benefit over one regimen in terms of both blood and brain Pb reductions (Stangle et al. 2004
). Therefore, these results likely underestimate the potential benefit of succimer treatment for the more heavily Pb-exposed animals.
Heightened reaction to errors in the Pb-exposed rats is abolished by succimer treatment
For the four tasks presented here, the rate of all types of errors was significantly greater on trials that followed an error than on trials following a correct response. Similarly, the latency to enter the testing alcove at trial onset was significantly longer on trials following an error than on trials that followed a correct response, a pattern also seen for the latency to make a response after cue onset. The disruption produced by committing an error was significantly greater for the High-Pb animals than for controls for several dependent measures: In the sustained attention task, the percentage of omission errors was significantly higher for the High-Pb group than for controls during mid- to late-session trials following an error—but not following a correct response—a pattern that indicates the combined influences of the disruptive effects of committing an error and the changing motivational and attentional state of the animals across each session. Similarly, in attention task 1, the High-Pb rats took significantly longer to enter the testing alcove and to make a response than controls, but only early in the task when error rate was very high and the rats had not yet learned the task rules. Finally, the drop in performance on trials following an error was also more pronounced for the High-Pb rats than for controls in a conditional olfactory discrimination task with periodic reward omission, an additional task administered subsequently to these rats (Beaudin et al. 2006
The interpretation of the heightened sensitivity to errors of the High-Pb rats is informed by previous studies that examined performance changes as a function of an error on the previous trial, all involving human subjects. In some of these studies, the error rate on post-error trials was exceptionally low (e.g., Laming 1979
; Robertson et al. 1997
), indicating the operation of an executive error-correction system localized to the anterior cingulate cortex (e.g., Bush et al. 2000
; Fernandez-Duque et al. 2000
). However, the finding consistently seen in our rodent studies—increased error rate on trials following an error (reviewed by Strupp and Beaudin 2006
)—has also been reported in some human studies (e.g., Elliott et al. 1996
; Rabbitt and Rodgers 1977
) and likely reflects an emotional response to the error. This interpretation is supported by the finding that an electrophysiologic marker of error detection, termed error-related negativity, varies as a function of individual differences in negative affect and emotionality (e.g., Luu et al. 2000
; also see Beaudin et al. 2006
Importantly, this heightened disruption following an error was very responsive to succimer treatment: In all cases, the High-Pb–succimer group was indistinguishable from controls (; Beaudin et al. 2006
). These results provide encouragement that Pb-induced dysfunction in the area of arousal or emotion regulation can be significantly alleviated by succimer treatment. This type of dysfunction may be an important contributor of the behavioral problems and increased delinquency rates seen in Pb-exposed children and adolescents (e.g., Bellinger et al. 1994
; Dietrich et al. 2001
; Needleman et al. 1996
), based on evidence that individuals vulnerable to faulty regulation of negative emotion are at risk for violence and aggression (Davidson et al. 2000
Succimer was effective in alleviating some, but not all, types of Pb-induced attentional dysfunction
The sustained attention task revealed two types of attentional dysfunction in the High-Pb group. First, the early-session increase in omission errors (relative to mid-session), seen for all groups across all sessions on this task, was more pronounced for the High-Pb group than for the controls, with group differences being largest for trials with the longest cue duration (700 msec). This pattern of results may indicate that lapses in attention were more common for the High-Pb rats than for controls during this early part of the session. More frequent attentional lapses in the High-Pb group would have the consequence of flattening the slope across cue duration and making group differences largest on trials for which performance of the controls was best (i.e., those with the longest cues). One interpretation of this pattern is that early Pb exposure may impair the ability to rapidly engage in a new task when transitioning between activities, manifested here as an increased tendency to miss the cue (an attentional deficit).
A second type of attentional dysfunction observed in the High-Pb rats was evident later in each testing session. The incidence of omission errors increased for all groups across each daily testing session, the classic pattern seen when sustained attention is taxed (Parasuraman and Warm 1998
). However, as discussed above, the High-Pb rats committed a significantly higher percentage of omission errors than controls during mid- to late-session trials, specifically on trials that followed an error. This pattern of effects, coupled with the similar pattern seen in the omission of reward task administered subsequently to these same animals (Beaudin et al. 2006
), indicates that as attention and motivation waned across the session, the disruptive effect of an error on attention and persistence increased, and that this disruption was more pronounced for the Pb-exposed rats than for controls.
The effectiveness of succimer chelation varied for these different types of attentional deficits. As discussed above, the heightened attentional disruption seen in the High-Pb rats following an error was completely normalized by succimer treatment, as the chelated High-Pb rats differed significantly from the High-Pb group and did not differ from controls. In contrast, succimer treatment only partially alleviated the attentional lapses seen early in each test session; the High-Pb–succimer group was intermediate to the other two, not differing from either.
Deficient inhibitory control in the High-Pb rats
In the sustained attention task, the percentage of premature responses was significantly higher for the High-Pb rats than for controls, indicating deficient inhibitory control. Converging evidence for this area of dysfunction was provided by the higher percentage of premature responses committed by the High-Pb rats in the final block of trials of attention task 1, a point at which the basic rules of the task had already been learned. Succimer treatment was ineffective in alleviating this area of dysfunction: In both instances, the percentage of premature responses for the High-Pb–succimer rats was significantly greater than that of the control rats, and not different from the High-Pb group.
Pattern of succimer effects in the Pb-exposed animals
The efficacy of the succimer treatment varied as a function of both the level of Pb exposure and the specific functional domain tested. Interestingly, the pattern seen for succimer efficacy parallels the apparent sensitivity of specific functional domains to the Pb exposure. Learning was impaired by both Pb exposure regimens, whereas impaired regulation of arousal or emotion was seen only in the High-Pb group, suggesting that disruption in learning occurs at lower exposures than dysregulation of arousal or emotion. The efficacy of succimer across these domains corresponded to the degree to which chelation reduced brain Pb levels in the two Pb exposure groups. For the Mod-Pb group, succimer treatment almost completely removed Pb from the brain and effectively alleviated the learning dysfunction. In contrast, brain Pb levels were still moderately elevated in the High-Pb group following chelation; these chelated High-Pb rats exhibited the same pattern of dysfunction as the unchelated Mod-Pb group, which also had moderately elevated brain Pb levels on PND52. Both groups also exhibited impaired learning but not affective dysfunction. This constellation of findings suggests that succimer treatment of the High-Pb rats reduced tissue (i.e., brain) Pb levels below that which produces lasting impairment in emotion regulation, but not enough to alleviate the learning dysfunction.
The mechanistic basis for this pattern of effects is unknown, but likely reflects complex interactions between the timing of the Pb exposure and succimer treatment relative to the ontogenetic stage of specific neural systems, the extent to which succimer treatment reduced Pb levels in specific neural systems (Cremin et al. 1999
), and the inherent sensitivity of those developmental processes and/or associated functional domains to Pb exposure.
Effects of succimer in the absence of Pb exposure
The present study also revealed the unexpected finding that a single 3-week course of succimer treatment during early development produced lasting dysfunction in cognition and arousal regulation in rats not previously exposed to Pb. Note that these four behavioral tests were administered across a 7-month period following cessation of succimer treatment, suggesting lasting brain changes. These results corroborate preliminary results from a similar study in nonhuman primates (Laughlin 2001
). The succimer-only group learned the initial visual discrimination task more slowly than the controls. They also committed a higher rate of inaccurate responses than controls in attention task 1, indicative of attentional dysfunction. In both the sustained attention task and the selective attention task, their performance benefited less by the lengthening of the visual cue than did the controls, indicative of lapses in attention, as discussed above. Additionally, in the selective attention task, the succimer-only group committed a higher rate of inaccurate responses than controls, an effect that was most pronounced for trials on which a distractor was presented and that followed an error, indicating impairments in both selective attention and arousal regulation. Finally, a comparison of performance on trials without distractors (during the selective attention task) to performance on the baseline task revealed that the succimer-only rats experienced a generalized disruption of performance in the selective attention task that extended beyond the trials on which a distractor was presented. This finding, too, implicates impaired arousal regulation in the succimer-only group. These various impairments were similar in magnitude to those produced by the High-Pb exposure.
The mechanism(s) responsible for these adverse succimer effects is not known. One possibility is that succimer, a metal-chelating agent, altered essential metal homeostasis and increased metal diuresis, as has been suggested by a number of clinical and nonhuman primate studies (Aposhian et al. 1989
; Fournier et al. 1988
; Smith et al. 2000a
Extrapolation of treatment regimens to humans
Both Pb-exposed groups exhibited low blood Pb levels (≈25–35 μg/dL) during the first 3 weeks of the 4-week exposure period. During the fourth and final week of exposure, both groups experienced an increase in tissue Pb levels because of the direct ingestion of Pb-adulterated water. Although the blood Pb levels produced during this final week of exposure were higher than commonly reported for children (see Supplemental materials
available online at http://www.ehponline.org/docs/2006/9263/suppl.pdf
), direct comparison of blood Pb levels across species should be cautioned. The available evidence indicates that the blood Pb levels required to produce overt toxicity (i.e., malaise, coma, convulsions, death) or lasting neurobehavioral effects (i.e., effects that last beyond the period of exposure) are higher in both rats and nonhuman primates than in humans (Garavan et al. 2000
; Levin and Bowman 1986
; Morgan et al. 2001
; Rice 1988
). Consistent with these earlier studies, none of the animals in the present study exhibited any signs of overt toxicity throughout the study and could not be distinguished from controls throughout the 8 months of daily handling. The Mod-Pb group exhibited very circumscribed and subtle behavioral/cognitive effects, and the High-Pb group exhibited subtle, although more widespread, neurobehavioral effects. As such, the regimens used in the present study may be viewed as modeling subclinical or asymptomatic Pb exposure. Similarly, although the succimer regimen used in the present study closely modeled the duration and dose (weight-normalized) of the regimens used clinically in children, it is possible that it was more “aggressive” than the clinical regimen, when allometrically scaled across species.