Long-term behavioral and electrophysiological changes associated with lead exposure at different stages of brain development in the rat

Developmental Brain Research, 29 (1986) 101-110 Elsevier

101

BRD 50443

Long-Term Behavioral and Electrophysiological Changes Associated with Lead Exposure at Different Stages of Brain Development in the Rat LINDA J. BURDE'Iq'E 1and ROBERT GOLDSTEIN 2 t Department of Neurology, Graduate Hospital, Philadelphia, PA and 2Department of Psychology, : Washington University, St. Louis, MO (U.S.A.) (Accepted March 25th, 1986) Key words: lead - - hippocampus - - FEP - - activity - - EEG - - theta

The present investigation was conducted to assess the behavioral and electrophysiologicalimpairments exhibited by adult male rats as a function of the developmental stage during which lead exposure occurred. Dams were given either a lead acetate (0.3%) or a control drinking solution during days 16-23 of gestation, days 1-8 or days 9-16 of nursing. The temporal and spatial activity patterns exhibited by gestationally exposed offspring in the open field between 42-45 days of age was distinguished from all other groups by the absence of a decrement in peripheral field activity across days and by increased exploration of the center field. Although open field activity proved sensitive to the timing of lead exposure, power spectral analyses of hippocampal and cortical EEG activity at 70-72 days of age revealed that lead selectively depressed 6-7 Hz energy in the hippocampus, independent of the developmental stage of exposure; cortical EEG and other hippocampal 0 frequencies were unaffected. The differential sensitivity of open field activity and select hippocampal 0 frequencies to the timing of lead administration suggests that the identification of toxic consequences depends on the function assessed and the developmental stage during which lead exposure occurred. INTRODUCTION Lead poisoning exerts its most severe consequences during d e v e l o p m e n t . Cell mitosis is compromised, restricting both the onset and the extent of cell proliferation 2°'38'44. A l t e r e d cell metabolism 9"35 may contribute to a reduction in dendritic branching 3°'42, which, in turn, may limit the n u m b e r of available synaptic sites 35. Finally, there is in vitro evidence that the dynamics of synaptic transmission t2,2s,33 also are affected. Because lead administered to pregnant 36,37 or nursing dams 22'25 is transmitted to the offspring, the fetus and neonate are particularly vulnerable to these lead poisoning effects. There is limited evidence that vulnerability to lead toxicity may vary even during early development. Lead administration to pregnant dams during the first two trimesters results in spontaneous abortion and increased incidence of fetal death 19'24'37. Crofton et a1.14 identified d e v e l o p m e n t a l delays in exploratory activity resulting from prenatal, relative to perina-

tal, lead exposure. In a comparison of differential sensitivity during nursing, Brown 8 found that a learning deficit exhibited by offspring exposed throughout the nursing period could be attributed to lead effects during the first 10 days of nursing. These d a t a suggest the possibility that brain structures that develop late in gestation and during early postnatal life may be especially susceptible to lead toxicity, The late m a t u r a t i o n of the hippocampus may contribute to the substantial lead-induced p a t h o l o g y that is observed in this area relative to other brain structures; the reduction in the width of cell layers 32.41 and in the complexity of dendrites and synaptic profiles3'11'26 are especially p r o m i n e n t in the hippocampus. Despite this and o t h e r evidence of pathology 1'2, there has been little investigation of altered hippocampai function. M c C a r r e n 34 discovered that lead administered to rat pups during nursing results in a shift of the m o d a l frequency of h i p p o c a m p a l 0 ( 4 - 1 2 Hz) from 7 H z to 5 Hz in the adult, leaving 8 - 1 2 Hz activity unaffected. The effects of lead exposure on

Correspondence: L.J. Burdette, Graduate Hospital, Department of Research/Neurology, 415 South 19th Street, Philadelphia, PA 19146, U.S.A. 0165-3806/86/$03.50 © 1986 Elsevier Science Publishers B.V. (Biomedical Division)

102 the strong relationship between specific 0 frequencies and the degree of locomotor activity has not been studied, although hyperactivity has been observed in both hippocampally damaged 1°'27"45 and lead-poisoned w'18 animals. The objectives of the present study were twofold: to determine if behavioral and electrophysiological measures are altered irreversibly by brief exposure to low lead dosages during development: and to identify the pattern or severity of CNS impairment associated with lead exposure during specific developmental stages. The developmental sequence of the hippocampus was used to delineate 3 periods during which offsprings were exposed to lead via the dams: days 16-23 of gestation (peak pyramidal cell proliferation45"49); days 1-8 of nursing (peak granule cell proliferationt3'48'49), or days 9-16 of nursing (peak synaptogenesis of cholinergic afferents6'7131. As the majority of lead clears the bloodstream within 8 days following exposure termination 5~. an 8-day poisoning duration, together with a 23-day nursing period, allowed equivalent exposure to milk lead residue for each of the 3 groups. Temporal and spatial activity patterns in the open field were chosen as behavioral measures affected by lead treatment and hippocampal damage. Finally, direct assessment of hippocampal integrity was provided by a power spectral analysis of hippocampal EEG activity. Alterations in the hippocampal power spectrum then were compared with that obtained from concurrent visual cortex recordings in order to contrast toxic effects in two areas that differ markedly in their maturation sequence.

MATERIALS AND METHODS

Dosing regimen Dams were assigned randomly to receive lead acetate or control solutions during one of 3 periods: days 16-23 of gestation (G); days 1-8 (1N) of nursing; or days 9-16 (2N) of nursing. The following protocol was used during the 8-day experimental session. Three grams of lead acetate were mixed vigorously with distilled water to yield a 1000-ml solution. To prevent the formation of a lead precipitate, 0.3 ml of glacial acetic acid were added while stirring. Control solutions were prepared in an identical manner, with the omission of the lead acetate. The volume re-

quired to deliver the prescribed do~age ot 270 n~g lead acetate/kg body wt/day was calculated on the basis of the dam's weight. This was the only fluid a~adable to the dam from 18.00 to (J8.(10 h. If the pre~ scribed volume was consumed during this period, the dam was permitted ad lib access to tap water m volumetric Richter tubes for the remaining 1(I h. If the dam had not finished the prescribed volume h~ the end of the 14 h. the remaining solution was the ont~ fluid available to the animal for the subsequent ~(I h, Any solution remaining at the end ,,~f24 h was added to the volume administered during the next 10-h perv od. A daily index of fluid intake was computed by summing the volume of experimental solution and water consumed. When the dams were not scheduled for the experimental protocol, ad hh access to tap water was permitted, and daily water consumption and body weight were recorded.

Sublects Forty-eight prim~parous Wistar rats. bred in the laboratory facilities at Malcolm Bli~s Mental Health Center. were impregnated; gestation Day 1 was designated by the presence of a vaginal plug. Only male pups were used for subsequent behavioral and elcctrophysiological testing to avoid potential confounding introduced by fluctuations in the estrous cycle. Litters were culled to 10 pups at birth with care taken to maintain an equal sex ratio whenever possible. Dams and their litters were housed in cages constructed to prevent pup access to the dam's drinking solution. With the above noted exceptions, all animals were permitted ad lib access to Purina rat chow and water, and were maintained on a 12-h light-dark cycle. Offsprings were weaned at z,~ days of age. Pups were weighed individually on days 1.9 and 17 of nursing, marking the termination of the G. IN and 2N experimental sessions, respectively. Since no attempt was made to identify subjects until weaning, an average weight/litter was computed on these occasions to provide a general index of litter growth. Following weaning, the weights of individual male pups were recorded at 3-day intervals: this procedure also familiarized the animals with handling prior to behavioral testing. At 37 days ot age, male subjects were separated from their tittermates and were housed individually for the duration of the experiment.

103

Open field activity Activity was monitored in a sound-shielded room that was illuminated with a 15-W bulb suspended over the center of a 116 x 116 cm field, divided into 25 equal squares. Two measures of open field behavior, peripheral field (PFA) and center field (CFA) activity, were recorded without knowledge of the animal's experimental history. The center and perimeter of the field were defined, respectively, as the 9 interior and 16 border squares. Entry into a square was tallied when both front paws were placed on the square. Totals of center and peripheral square crossings were recorded at 1-min intervals for 5 min, Testing was conducted between 13.00 and 16.00 h for 3 consecutive days when animals were between 40 and 42 days of age.

Surgery At 60 days of age, etherized animals were implanted stereotaxically with bipolar leads in the CA3 hippocampal field. This recording site was selected to minimize variability in signal energy resulting from the sensitivity of the bipolar recording configuration to the changing phase relationship between the two 0 generators (CA1 and the dentate gyrus) with electrode depth 46. Twisted 0.25 mm insulated stainless-steel wires with exposed tips spaced vertically about 0.75 mm apart were positioned 3 mm posterior to bregma, 3 mm left of midline and 3.3 mm below the dural surface (with a 5-mm bite bar elevation above the interaural line). Three screw electrodes were placed over the right visual cortex (2 mm lateral to lambda), right frontal cortex (4 mm anterior to bregma, 3 mm lateral to midline) and left frontal cortex (4 mm anterior to bregma, 2 mm lateral to midline) to serve as the active, reference and ground recording sites, respectively. Hippocampal wires were crimped to Amphenol connecting pins inserted into a miniature strip plug; screw heads were wrapped with stainless-steel wires that were wrapped at the other end around the connecting pins. The entire assembly then was embedded in dental acrylic and the wound was closed. A 10-day postoperative recovery elapsed before testing.

Electroencephalographic activity Electrophysiological data collection began at 70 days of age. Following cable attachment, the animal

was placed in a cylinder (26.5 cm diameter) for i min before recording 3 min of hippocampal and cortical E E G activity. The duration of the recording period was limited by the daily administration of several protocols and by the decision to complete the study within a 4-month period to minimize seasonal influences on lead absorption 23. Hippocampal and cortical E E G signals were passed to two Tektronix differential amplifiers, with band pass settings at 0.1 and 300 Hz; hippocampal and cortical signals were amplified by 5 K and 10 K, respectively. These data were recorded with a Honeywell instrumentation tape recorder, calibrated with linear output for frequencies below 100 Hz, for off-line computer analysis. Animals were tested at the same time each evening, between 19.00 and 24.00 h for 3 consecutive days. At the end of the experiment, low-pass filtered (0-50 Hz) signals were submitted to power spectrum analysis using a PDP 11/40 computer. To prevent aliasing, a common artifact in power spectral analyses, an A/D sampling rate of 128 Hz met the criterion of being at least twice that of the highest component frequency. A 4-s period defined 0.25 Hz as the lowest frequency analyzed. Alternate periods of E E G data were visually inspected and a total of 21 (7 periods/min X 3 min) artifact-free periods were submitted to the analysis. Stability of the power spectrum measure was achieved by averaging both within and across sampling periods. Within each period, frequencies were sorted into 11 bands, the width of each band chosen according to the frequencies of interest; since the primary objective of the power spectrum analysis was to identify shifts in power within the dominant hippocampal 0 range, 4-12 Hz activity was averaged in 2-Hz bands. For 12-20 Hz activity, band width was 3 Hz and for higher frequencies, band width was 10 Hz. The power in each frequency band was expressed as a proportion of total energy, and then was averaged across 7 periods to provide a stable power spectrum measure for each minute of each daily recording session.

Histology At the termination of electrophysiological recording, etherized animals were killed by transcardial perfusion with isotonic saline, followed by a 10% formalin solution. The brains were sectioned every 100 /~m and sections tracing electrode penetration were

104 stained with Cresyl violet. Statistical analyses

D a t a were analyzed for t r e a t m e n t [lead. control) and d e v e l o p m e n t a l stage IG. 1N. 2N) group effects using analysis of variance ( A N O V A ) : the Geiss e r - G r e e n h o u s e a d j u s t m e n t for heterogeneity of covariance was e m p l o y e d for all r e p e a t e d measures factors. Prior to analysis, possible violations of parametric assumptions of normality were assessed using a transformation comparison test 27. and. d e p e n d i n g on the type of distortion, a p p r o p r i a t e transformations were p e r f o r m e d . To minimize the effects of r e p e a t e d sampling, both a priori and a posterior± levels were adjusted in accordance with the n u m b e r of analyses conducted. The probability assoctated with a particular o u t c o m e is r e p o r t e d with reference to the adjusted a. RESULTS Error term A l t h o u g h the r a n d o m assignment of pups across litters minimizes any bias attributed to between-litter variance, the residual exposure of pups nursing from

gestationally e x p o s e d dams p r e c l u d e d this experimental control. To d e t e r m i n e the most conservative estimate of error variance, within- and between-litter variance were c o m p a r e d for two measures, day-40 body weight and total n u m b e r of square crossings in the open field. F o r the body weight m e a s u r e , between-litter variance was twice that o b s e r v e d within litter. The litter was chosen as the conservative error estimate for this analysis. For total activity scores. the equivalence of the two variances. 72.9. suggested that e x p e r i m e n t a l error was due to individual, rather than litter, variability. Because of the increased power advantage p r o v i d e d by the former error term. behavioral and electrophysiological analyses were conducted using the subject as the e x p e r i m e n t a l unit of variance. B o d y weight and fluid intake measures Except for 3 dams (two in the 1N group and one in the 2N group), all females c o m p l i e d with the dosage regimen for the entire 8-day session. These 3 were conforming to the dosage schedule by the fourth day of the e x p e r i m e n t a l session. The general health of

the dams was assessed by analysis of the percentage of weight change (arc-sine transformed) on the last day of each experimental period ~relative to average body weight on gestation days I1 --15) and bv analysis of average daily fluid intake. A l t h o u g h a significant t r e a t m e n t - b y - t i m e interaction tl:, .~ = 4.2~ " < (1.(125) was identified in the body weight analvs,s. simple effects test indicated thal lead and control dams did not differ at any of the 3 nmc points. Average daily fluid intake was affected, however, b~ the timing at which treatment was administered. Po~;I hoc testing of the c o m p o n e n t s of the stage by time interaction (Fa ~,, - 8.09. P < {).025~ revealed that during the 1N and 2N e x p e r i m e n t a l sessions both lead and control dams significantly decreased average daily fluid intake by 11.5 ml tlt.L(<).} and I1.11 ml { 13.6% I. respectively, relative to dams permitted ad lib access to tap water during nursing. This reduction in fluid intake, however, was nol reflected either in pup growth rate (as m e a s u r e d by average weight/litTer on postnatal days 1 . 9 and 17~ or in b o d y wetghts of male offspring on day 41t when behavioral testing was initiated. Fluid intake and b o d y weight measures for the dams and their offspring are presented m Table 1. Open field exploration To d e t e r m i n e whether brief lead exposure disrupts the temporal or spatial p a t t e r n of o p e n field activitv. peripheral field activity (square root transformed) and center field acuvity (reciprocal transformed) scores each were analyzed for habituation effects within and across test sessions. The most interesting

outcome of the P F A analysis was the significant treatment-by-stage-by-days

interacnon

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TABLE I Dam's body weight as a percentage change at the end o] each period ]rom the mean of gestation days 11 -15 Test days

Controls 16-23 gestation 1-8 nursing u_ 16 nursing Lead 16-23 gestation 1-8 nursing 9-16 nursing

G23

NS

NI 6

29.2(+_1.6t 31.1 {±2.0) 30.0 (+_2.8)

3.01+_I.1) 3,I~ (±2.6) I 21+l.31

1.t 1-+2.21 1.9(±3.2) 3.0 (+-3.0)

25.2(+_1.21 30.3 ( :r-1.9) 29.5 (+-3.1}

2.6(+_1.8) 3.~,(+-2.9) 5 v ()~1.01

2.4(~2.0) 4.3 t-+2.4) 9.1 ~±3.5,'J

105 TABLE II Dam's average daily fluid intake Test days

Controls 16-23 gestation 1-8 nursing 9-16 nursing Lead 16-23 gestation 1-8 nursing 9-16 nursing

G16-G23

N1-N8

N9-N16

39.2 (+2.6) 43.2 (+2.6) 47.6 (+2.6)

57.1 (+3.2) 42.1 (+4.0) 55.6 (+3.1)

79.7 (+4.9) 81.9 (+6.3) 64.2 (+7.4)

44.2 (+2.2) 48.2 (+3.3) 47.8 (+5.2)

55.3 (+2.7) 47.1 (+2.4) 56.8 (+4.2)

81.0 (+4.0) 82.2 (+3.1) 77.3 (+3.1)

Table III A verage litter weights Test days

Controls 16-23 gestation 1-8 nursing 9-16 nursing Lead 16-23 gestation 1-8 nursing 9-16 nursing

Day 1

Day 9

Day 17

Day 40

6.2 (+0.1) 6.0 (+0.3) 6.2 (+0.4)

17.6 (+0.9) 17.5 (+0.2) 17.3 (+1.6)

36.4 (+3.1) 36.2 (+1.9) 32.5 (+2.2)

186.4 (+5.8) 184.6 (+6.3) 189.2 (+5.7)

6.2 (+0.2) 6.3 (+0.4) 6.4 (+0.3)

17.6"(+1.2) 16.2 (+1.0) 19.1 (+0.8)

35.0 (+2.3) 35.2 (+3.0) 36.7 (+1.2)

185.2 (+6.9) 189.9 (+3.1) 169.4 (+6.0)

3.00, P < 0.025) depicted in Fig. 1. Simple effects tests revealed the source of the interaction as the elevated peripheral activity levels on D a y 2 (F1,294 = 16.01, P < 0.008) and D a y 3 (FI,294 = 18.31, P < 0.008) of subjects exposed in utero compared to all other groups. Although no group interactions across days were identified for C F A scores, a significant treatment-by-stage interaction (F2,98 = 3.16, P <

0.025) indicated that subjects exposed in utero were more active overall in the center field relative to all other experimental conditions (Fig. 2). To ascertain that this effect was not due to a general increase in activity, the analysis was repeated using a ratio of center field to total activity (arc-sine transformed). Sim-

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Fig. 2. Center field activity as a function of the developmental stage of lead exposure (means and S.E.M.).

t06 pie effects tests and accompanying mean contrasts of the significant treatment-by-stage interaction. F2,~ = 7.74. confirmed that subjects exposed in utero were proportionately more active in the center field relative to other groups.

Electroencephalographic activiw A typical electrode placement in hippocampat field CA3 is presented in Fig. 3 Although relative signal energy in each of 11 frequency groupings was available (Fig. 4a), interest in the dominant hippocampal theta activity restricted analyses to the 5 bins representing 4-14 Hz. A significant treatment main effect (F1..sl = 7.33. P < 0.01) documented the proportional decrease in relative power (arc-sine transformedl in the 6 - 7 Hz band for lead-treated animals. To ensure that this result did not reflect an artefact of the relative power measure, a post-hoc analysis of absolute 6-7 Hz signal energy confirmed a significant depression consequent to lead exposure (Fig. 4b). All other frequency bands comprising 0 activity were unaffected by experimental manipulation. The characteristics of the power spectra also did not vary

significantly within or across the short test session, or as a function of the developmental stage of poisoning. Regional specificity of brain dystunction was deduced by a comparison of spectral alterations observed in hippocampus and visual cortex. No significant lead influence was evident at any of the cortical frequency groupings, indicating a selective hippocampal deficit. DISCUSSION The findings of the present investigation confirm that brief exposure to a low lead dosage during development results in behavioral and electrophysiological abnormalities in the young adult rat, and further. that the observed impairment pattern is dependent on both the timing of lead exposure and the function assessed. The differential sensitivity of locomotor activity and hippocampal 0 to the developmental stage during which lead exposure occurred may imply, among other possibilities, that E E G activity is more sensitive than behavioral measures to lead toxicity.

Fig. 3. Representative electrode placement in field CA3 of the hippocampus.

107

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Fig. 4. A: power spectrum of relative signal energy recorded from the hippocampus as a function of lead exposure. B: effects of lead exposure on 6-7-Hz absolute signal energy (means and S.E.M.).

On the other hand, this contrast tends to discount the possibility that altered activity patterns displayed by lead G subjects were a consequence of higher effective dosages experienced in utero. This interpretation is supported by evidence in the literature that lead uptake in dams 51 or lead transfer to offspring25 is greater during nursing than during gestation, an effect attributed to potentiation of lead absorption by lactose 1°. An attempt was made in this study to determine internal dosage by measuring free erythrocyte protoporphyrin, an index of toxic interference with heme synthesis. Although the data (not presented) suggested that lead may suppress or delay the transition from fetal to adult heme synthesis, additional study is needed to determine the magnitude and time

course of this effect. The results of the open field test indicate that the pattern of activity is a more sensitive index of lead exposure than the level of activity. Only those offspring exposed in utero failed to exhibit the normal decrement in peripheral field activity over days and demonstrated increased exploration of the center field. Although the failure to exhibit normal activity decrements over time 18 and increased center field exploration 29 previously have characterized the behavior of poisoned animals, the unique contribution of the present findings lies in the identification of the exposure period of enhanced susceptibility. The failure of poisoned G subjects to exhibit habituation in peripheral field activity over days is consistent with the observation that hippocampal damage results in prolonged habituation to novel environments 16,3t,45. Caution must be exercised, however, in attributing this habituation deficit solely to hippocampal dysfunction. Other evidence indicates that either no difference39'5° or even accelerated 39 habituation may be observed as a function of hippocampal damage. At present, then, the role of the hippocampus in the habituation of exploratory behavior remains unclear. Though the extent of hippocampal involvement in regulating temporal characteristics of exploration still is in dispute, a growing body of evidence suggests that the hippocampus is involved prominently in the formation of spatial memories or maps. Following the observation that single unit firing patterns demonstrate place specificity4°, several authors have reported that hippocampal damage alters spatial response patterns. Harley21 found that animals with hippocampal lesions exhibited random arm selection in a sunburst maze, in contrast to unoperated controls or animals with neocortical lesions who selected maze arms oriented towards the goal. These findings are similar to those indicating that hippocampal lesions interfere with place, relative to response, learning43. More relevant to the present findings, Ruppert et al. 47 noted that animals sustaining CA3 damage as a result of trimethyltin exposure selectively increased activity in the circular, relative to the straight, alleys of a figure-8 maze. The altered temporal and spatial characteristics of open field activity observed in the present study, at the very least, are consistent with behavioral characteristics associated with hippocampal damage.

1 (18

A reduction in relative and absolute signal energy of 6 - 7 Hz h i p p o c a m p a l 0 activity was the sole E E G reflection of lead exposure. In addition to highlighting toxic vulnerability of the h i p p o c a m p u s , the data imply that lead exposure selectively interferes with only restricted 0 frequencies. A l t h o u g h differences in the grouping of E E G frequencies prevent a direct comparison between this result and the r e p o r t e d shift in peak 0 energy from 7 Hz to 5 Hz following early lead exposure 34, it is interesting that a depression in energy was noted in both studies at lower 0 frequencies. Functional differentiation of low ( 5 - 7 Hz) and high ( 8 - 1 2 Hz) 0 frequencies has been s u p p o r t e d by behavioral, pharmacological and electrophysiological evidence. The original distinction between these two E E G categories was m a d e by the association with specific classes of behavior: alert immobility is associated with 4 - 7 - H z O, while walking is c o r r e l a t e d with the a p p e a r a n c e of 8 - 1 2 - H z f r e q u e n c t"e s ~"-. Based on results o b t a i n e d with pharmacological probes and electrical stimulation, D e s t r a d e and Ott ~5 conclude that slower 0 frequencies are g e n e r a t e d by cholinergic innervation from the septum. Given the lead-induced disruption of in vitro p e r i p h e r a l cholin• • ~ ~.8. . 33 ergic transmlsston . . it is tempting to interpret the depression in signal energy at 6 - 7 Hz as reflecting interference with s e p t a l - h i p p o c a m p a l afferents. Several arguments caution against such a simple interpretation. First, conclusive evidence of a lead-induced depression in central cholinergic function in viw) is lacking. Second, the absence of any d e v e l o p m e n t a l stage effect would argue against interference by lead with septal innervation of the h i p p o c a m p u s , since cholinergic afferents cannot be identified until the

REFERENCES l Alfano, D.P., LeBoutillier, J.C. and Petit, T.L., Hippocampal mossy fiber pathway development in normal and postnatally lead-exposed rats, Exp. Neurol., 75 (1982") 308-319. 2 Alfano, D.P. and Petit, T.L., Behavioral effects of postnatal lead exposure: possible relationship to hippocampal dysfunction, Behav. Neurol. Biol., 32 (1981) 319-333. 3 Alfano, D.P. and Petit, T.L., Neonatal lead exposure alters dendritic development of hippocampal dentate granule cells, Exp. Neurol., 75 (1982) 275-288. 4 Bayer, S., Development of the hippocampal region in the rat. 1. Neurogenesis examined with 3H-thymidine aurora-

second postnatal d a @ ~ And thirL' ~tlthL)ut!h J weak argument could be m a d e that a del~ression m t ~ c ~ ¢p frequencies may be associated witt~ decreased innno bililv, lhe prolonged explorator} hetmvior o~ uestationally exposed subjects argues ,mr a c o n c o m i t a m increase in higher o frequenclcs. { ~|il turther cnJerla for differentiating Iox~ and high frequency ~ ecnc:rators is available, the selective m~crference oT lead with low frequency o cannot be explained. In summary, chronic behavior~,i ,rod clectrophystological a b e r r a n o n s were detected m earl v adulthood following brief exposure to a Io~v dosage of lead during early development. With reservation, differential sensltlWty among exposure pen<~ds also was a~sociated with specific impairments Iv, ~pen field performance, animals exposed to lcati m utero showed altered temporal and spatial characteristics of acnvity patterns. In contrast to the heha\~oral data the resuits of the power spectrum-analysis suggest thai pr(~cesses involved in the production ¢,l low-frequenc~ o are vulnerable to lead throughout d e v e l o p m e n t , discounting the likelihood of a critical period for toxic insult. W h e t h e r this discrepant} reflects the differential sensitivity of behavioral and elcctrophysiolog~cal measures or differential susceptibility of neurophysiological processes to such in,,utt, the data c~mfirm that the specific consequences of lead exposure d e p e n d on the function assessed and the period ot exposure. ACKNOWLEI)GEMENTS The authors wish to thank Kevin Crofton and Drs. Vernon Benignus. Diane Miller. and Dave Otto for their thoughtful c o m m e n t s about this manuscript.

diography, J. ('ornp. Neurol. t9111t980 ~87-114. 5 Bayer. S.. Development of the hippocampal region in the rat. II Morphogenesis during embryonic and early postnatal life. J. Comp. Neurol., 190119801/15-134. 6 Ben-Barak, J. and Dudai, Y.. Chotinergic binding sitcs m rat hippocampal formation: properties and ontogenesis. Brain Res. 166 119791 245-257. 7 Ben-Barak. J. and Dural. Y.. Early septal lesion: effect on the development of the cholinergic system in the hippocampus. Brain Res.. 185 (198[)) 323-334" x Brown. D.R.. Neonatal lead exposurem the rat: decreased learning as a function of age and blood lead concentration, Toxicol. Appl. Pharmaeol. . 32 (19751 628-637. c) Bull. R.J.. Lutkenhoff. SD.. McCartv. G.E. and Miller.

109 R.G., Delays in the postnatal increase of cerebral cytochrome concentrations in lead-exposed rats, Neuropharmacology, 18 (1979) 83-92. 10 Bushnell, P.J. and DeLuca, H.F., Lactose facilitates the intestinal absorption of lead in weanling rats, Science, 211 (1981) 61-63. 11 Campbell, J.B., Woolley, D.E., Vijayan, V.K. and Overmann, S.R., Morphometric effects of postnatal lead exposure on hippocampal development of the 15-day-old rat, Dev. Brain Res., 3 (1982) 595-612. 12 Carroll, P.T., Silbergeld, E.K. and Goldberg, A.M., Alteration of central cholinergic function by chronic lead acetate exposure, Biochem. Pharmacol., 26 (1977) 397-402. 13 Cowan, W.M., Stanfield, B.B. and Kishi, K., The development of the dentate gyrus, Current Topics Dev. Biol., 15 (1980) 103-157. 14 Crofton, K.H., Taylor, D.H., Bull, R.J., Sivulka, D.J. and Lutkenhoff, S.D., Developmental delays in exploration and locomotor activity in male rats exposed to low level lead, Life Sci., 26 (1980) 823-831. 15 Destrade, C. and Ott, T., Quantitative analyses of hippocampal rhythmical slow activity induced by stimulation of different diencephalic structures, Neuroscience, 6 (1981) 1089-1094. 16 Douglas, R.J., The hippocampus and behavior, Psychol. Rev., 67 (1967) 416-442. 17 Driscoll, J.W. and Stegner, S.E., Behavioral effects of chronic lead ingestion on laboratory rats, Pharmacol. Biochem. Behav., 4 (1976) 411-417. 18 Driscoll, J.W. and Stegner, S.E., Lead-produced changes in the relative rate of open field activity of laboratory rats, Pharmacol. Biochem. Behav., 8 (1978) 743-747. 19 Gerber, G.B., Maes, J. and Deroo, J., Effect of dietary lead on placental blood flow and on fetal uptake of a-amino isobutyrate, Arch. Toxicol., 41 (1978) 125-131. 20 Gerber, G.B., Maes, J., Gilliavod, N. and Casale, G., Brain biochemistry of infant mice and rats exposed to lead, Toxicol. Lett., 2 (1978) 51-63. 21 Harley, C.W., Arm choices in a sunburst maze: effects of hippocampectomy in the rat, Physiol. Behav., 23 (1979) 283-290. 22 Hejtmancik, M.R., Dawson, E.B. and Williams, B.J., Tissue distribution of lead in rat pups nourished by lead-poisoned mothers, J. Toxicol. Environ. Health, 9 (1982) 77-86. 23 Hunter, J.H., The summer disease: some field evidence on seasonality in childhood lead poisoning, Social Sci. Med., 12 (1978) 85-94. 24 Jacquet, P., Early embryonic development in lead-intoxicated mice, Arch. Pathol. Lab. Med., 101 (1977) 641-643. 25 Keller, C.A. and Doherty, R.A., Bone lead mobilization in lactating mice and lead transfer to suckling offspring, Toxicol. Appl. Pharmacol., 55 (1980) 220-228. 26 Kiraly, E. and Jones, D.G., Dendritic spine changes in rat hippocampal pyramidal cells after postnatal lead treatment: a Golgi study, Exp. NeuroL, 77 (1982) 236-239. 27 Kirk, R.E., Experimental Design. Procedures for the Behavioral Sciences, Brooks/Cole, Belmont, 1982, pp. 83-84. 28 Kober, T.E. and Cooper, G.P., Lead competitively inhibits calcium-dependent synaptic transmission in the bullfrog sympathetic ganglion, Nature (London), 262 (1976) 704-705. 29 Kostas, J., McFarland, D.J. and Drew, W.G., Lead-induced behavioral disorders in the rat: effects of ampheta-

mine, Pharmacology, 16 (1978) 226-236. 30 Krigman, M.R., Druse, M.J., Traylor, T.D., Wilson, M.H., Newell, L.R. and Hogan, E.L., Lead encephalopathy in the developing rat: effect on cortical ontogenesis, J. Neuropathol. Exp. Neurol., 33 (1974) 671-686. 31 Leaton, R.N., Habituation of startle response, lick suppression and exploratory behavior in rats with lesions, J. Comp. Physiol. Psychol., 95 (1981) 813-826. 32 Louis-Ferdinand, R.T., Brown, D.R., Fiddler, S.F., Daughtrey, W.C. and Klein, A.W., Morphometric and enzymatic effects of neonatal lead exposure in the rat brain, Toxicol. Appl. PharmacoL, 43 (1978) 351-360. 33 Manalis, R.S. and Cooper, G.P., Presynaptic and postsynaptic effects of lead at the frog neuromuscular junction, Nature (London), 243 (1973) 354-356. 34 McCarren, M., Short- and long-term effects of neonatal lead exposure in rats: behavior and neurophysiology, Diss. Abstr., (1982). 35 McCauley, P.T., Bull, R.J. and Lutkenhoff, S.D., Association of alterations in energy and metabolism with lead-induced delays in rat cerebral cortical development, Neuropharmacology, 18 (1979) 93-101. 36 McClain, R.M. and Becker, B.A., Effects of organolead compounds on rat embryonic and fetal development, Toxicol. Appl. Pharmacol,, 21 (1972) 265-274. 37 McClain, R.M. and Becker, B.A., Teratogenicity, fetal toxicity and placental transfer of lead nitrate in rats, Toxicol. Appl. PharmacoL, 31 (1975) 72-82. 38 Michaelson, I.A. and Sauerhoff, M.W., Animal models of human disease: severe and mild lead encephalopathy in the neonatal rat, Environ. Health Perspect., 7 (1974) 201-225. 39 Nadel, L., Dorsal and ventral hippocampal lesions and behavior, Physiol. Behav., 3 (1968) 891-900. 40 O'Keefe, J. and Dostrovsky, J., The hippocampus as a spatial map. Preliminary evidence from unit activity in the freely moving rat, Brain Res., 34 (1971) 171-175. 41 Petit, T.L., Alfano, D.P. and LeBoutillier, J.C., Early lead exposure and the hippocampus: a review and recent advances, Neurotoxicology, 4 (1983) 79-94. 42 Petit, T.L. and LeBoutillier, J.C., Effects of lead exposure during development on neocortical dendritic and synaptic structure, Exp. Neurol., 64 (1979) 482-492. 43 Plunkett, R.P., Faulds, B.D. and Albino, R.C., Place learning in hippocampectomized rats, Bull. Psychon. Soc., 2 (1973) 79-80. 44 Press, M.F., Lead encephalopathy in neonatal LongEvans rats: morphologic studies, J. NeuropathoL Exp. Neurol., 36 (1977) 169-193. 45 Roberts, W.W., Dember, W.N. and Brodwick, M., Alternation and exploration in rats with hippocampal lesions, J. Comp. Physiol. Psychol., 55 (1962) 695-700. 46 Robinson, T.E., Hippocampal rhythmic slow activity (RSA:THETA): a critical analysis of selected studies and discussion of possible species-differences, Brain Res. Rev., 69 (1980) 69-101. 47 Ruppert, P.H., Walsh, T.J., Reiter, L.W. and Dyer, R.S., Trimethyltin-induced hyperactivity: time course and pattern, Neurobehav. Toxicol. Teratol., 4 (1982) 135-139. 48 Schlessinger, A.R., Cowan, W.M. and Gottlieb, D.I., An autoradiographic study of the time of origin and the pattern of granule cell migration in the dentate gyrus of the rat, J. Comp. Neurol., 159 (1975) 149-176. 49 Stanfield, B.B. and Cowan, W.M., The development of the hippocampus and dentate gyrus in normal and reeler

II0 mouse, J. Comp. Neurol.. 185 (1979) 423-459. 50 Suess, W.M. and Berlyne, D.E., Exploratory behavior as a function of hippocampal damage, stimulus complexity and stimulus novelty in the hooded rat, Behav. Biol., 23 (1978) 487-499. 51 Toraason, M.A., Barbe, J.S. and Knecht, E.A., Maternal lead exposure inhibits intestinal calcium absorption in rat

pups. Toxicol. Appl, PharmacvA, v,¢~i [~;i',i ) ¢C -~~; 52 Vanderwotf, ('.H., Kramis, R. and R¢~binson, "f.[!.., t]JF~ pocampal electrical activih during '.aaking behavior an~.~ sleep: analyses using centrally actin~z drugs [n CIBA l:ou~ dation Symposium. 58 (new ,~'trlc~i ,I:'u~ctions o[ the Sept~,Hippocampal,S'vstem. Elsevier, ,'~m~i 'rd~m, 107N