Neurobehavioral changes in mice treated with methylmercury at two different stages of fetal development

Neurobehavioral changes in mice treated with methylmercury at two

different stages of fetal development

F.Y. Dorea,*, S. Gouleta, A. Gallaghera, P.-O. Harveya, J.-F. Cantina, T. D’Aigleb, M.-E. Miraultb

aCentre de Recherche Universite Laval Robert-Giffard and Ecole de Psychologie, Pavillon F.A. Savard, Universite Laval, Quebec, Quebec, Canada G1K 7P4bCentre de Recherche du CHUL-CHUQ and Department of Medicine, Universite Laval, Quebec, Quebec, Canada

Received 6 March 2001; received in revised form 9 July 2001; accepted 12 July 2001

Abstract

Pregnant C57BL/6 mice were orally given daily doses of 4 or 6 mg/kg of methylmercury chloride (MeHg) or vehicle during either

gestational days 7–9 (GD7–9) or days 12–14 (GD12–14). Their female offspring were tested between 6 and 16 weeks of age on a variety of

behavioral tasks. Motor coordination on the rotarod and visual discrimination learning in the Y maze were not affected by administration of

MeHg either at GD7–9 or at GD12–14. In the open field, the total number of square crossings was lower in mice treated with 4 and 6 mg/kg

of MeHg at GD12–14 than in control mice whether the environment was new or familiar, but prenatal administration of MeHg at GD7–9

had no effect on this measure. Administration of MeHg either at GD7–9 or at GD12–14 had no effect on the percentage of central square

crossings or on the frequency of rearings in the open field. On spatial alternation training in the T maze, both treated groups in Condition

GD7–9 and the group treated with 6 mg/kg at GD12–14 required more sessions to reach the learning criterion than their respective vehicle

groups. When spatial alternation was tested with delays, treated groups did not differ from their respective control groups. In the radial arm

maze, the performance of mice treated at GD7–9 was normal, but reference memory and working memory were impaired by administration

of MeHg at GD12–14. In mice treated with 4 mg/kg of MeHg, reference memory was impaired only on the first block of trials, whereas in

mice treated with 6 mg/kg, the deficit persisted on all blocks of trials. Overall, these results indicate that prenatal administration of MeHg at

GD12–14 had more detrimental effects on behavioral performance than administration at GD7–9. It reduced locomotor activity and

impaired reference memory for egocentric and allocentric spatial information as well as working memory for places. D 2001 Elsevier Science

Inc. All rights reserved.

Keywords: Methylmercury; Prenatal exposure; Activity; Learning; Memory

1. Introduction

The teratological and neurobehavioral effects of pre-

natal exposure to methylmercury (MeHg) on human and

animal development have been well documented for

decades (for reviews, see Refs. [3,7–9,21,29,38]). These

effects include reduced survival rate and weight gain,

sensory and motor dysfunctions as well as learning and

memory deficits. In mice, prenatal MeHg exposure tends

to retard reflexive behavior in early development (Ref.

[31], but see Ref. [36]), impairs motor coordination

[13,34,36] and decreases spontaneous locomotor activity

[17,35]. Early experiments [34,35,38] reported longer

latencies to explore the environment in the open field

test and decreased [35,38] or increased [36] frequencies

of rearings. In a recent experiment [19], less overall

locomotor activity and more locomotion directed toward

the center of the field were observed in this apparatus.

Studies of learning and memory in mice showed that

acquisition [16,36] and extinction [32,36] of avoidance

responses were impaired by prenatal MeHg exposure and

suggest that spatial long-term memory in the Morris water

maze was also impaired [19].

Only a few studies [17,34] have compared the neuro-

behavioral consequences of repeated administrations of

MeHg at different fetal stages of mice development. In the

following experiment, we investigated neurobehavioral

changes in female offspring of mice, which received vehicle

or methylmercury chloride (MeHg) at one of two daily

0892-0362/01/$ – see front matter D 2001 Elsevier Science Inc. All rights reserved.

PII: S0892 -0362 (01 )00167 -2

* Corresponding author. Tel.: +1-418-656-2376; fax: +1-418-

656-3646.

E-mail address: [email protected] (F.Y. Dore).

Neurotoxicology and Teratology 23 (2001) 463–472

doses (4 or 6 mg/kg) and at one of two developmental

windows, i.e., during days 7–9 or days 12–14 of gestation.

Doses and treatment windows were selected on the basis of

early [34,36] and more recent studies [17,19,38], as well as

on the basis of neurodevelopmental landmarks. Until gesta-

tional day 12 (GD12), interference with cell proliferation

can result in usual teratologies and gross defects of the

central nervous system [30]. In the case of MeHg, the eighth

or ninth day of gestation is especially important since it is

the beginning of maximum susceptibility of the developing

rodent brain to this neurotoxic agent [16,33]. From GD12 to

birth, cell proliferation is marked by bursts of activity in the

cerebellum, the thalamus, the striatum, the limbic structures

and the cerebral cortex [18,30]. Therefore, brain damage can

also occur after the critical period for malformations.

Five behavioral tasks were used in our experiment.

Acquisition of motor coordination and equilibrium was

measured on the rotarod. In mice with cerebellar lesions

[1,5,6] and in cerebellar mutant mice [14,22], fall latencies

on the rotarod task have been reported to be shorter than in

control mice. Activity and exploration were assessed in the

open field both when the environment was new, as in most

studies on developmental exposure to MeHg in rodents, and

when it was becoming familiar. Although the precise neuro-

behavioral significance of activity in the open field is not

well understood, it seems that limbic structures (amygdala,

hippocampal formation and prelimbic cortex) modulate the

behavioral response to novelty, whereas the nucleus accum-

bens mediates locomotor activity and exploration [4].

Performance was also examined in a variety of learning

and memory tasks. A visual discrimination learning task

was administered in the Y maze. Acquisition of a similar

task was shown to be impaired in rats by selective lesions of

the hippocampus [24]. Two tasks were used to test spatial

learning and memory. One task was spatial alternation in

the T maze, which requires the animal to choose the arm

opposite to the one selected on the previous trial. Since the

walls of the maze were opaque, no extramaze cues were

available and the spatial alternation response could be

learned by relying on egocentric information. Egocentric

localization memory deficits have been reported in rodents

after lesions of the caudate nucleus [10,28]. Training to

spatial alternation was followed by testing with delays.

Delayed spatial alternation and spatial working memory

have been repeatedly shown to be impaired by frontal

lesions [11,20,23,37]. The other spatial task involved place

learning and memory in the radial arm maze. In this task,

the maze is surrounded by a variety of extramaze stimuli,

which are visible from different arms. To solve the problem,

the animal relies on allocentric spatial information, i.e.,

information that is independent of the position of the

animal. In our experiment, we used the reference working

memory version of the radial arm maze task [26]. Short-

term and long-term retention for places are impaired on this

task by hippocampal lesions [26] and by caudate lesions

[27], respectively.

2. Materials and methods

2.1. Animals

Mice of the C57BL/6 strain were obtained from a local

breeder (Charles River, St. Constant, Quebec, Canada).

For mating, ninety-eight 11- to 12-week-old primigravid

females were placed two per cage with one male breeder.

GD1 was confirmed by the presence of a vaginal plug in

the morning. The 79 females with vaginal plugs were

placed into individual nesting cages and were assigned at

random to two conditions. In Condition GD7–9, they

were treated on GD7, GD8 and GD9, whereas in Con-

dition GD12–14, they were treated on GD12, GD13 and

GD14. In each condition, plugged females received a dose

of either 0 mg MeHg/kg body weight/day (GD7–9:

n = 10; GD12–14: n = 12), 4 mg/kg (GD7–9: n = 12;

GD12–14: n = 12) or 6 mg/kg (GD7–9: n = 12; GD12–14:

n = 21), for a cumulative dose of 0, 12 and 18 mg/kg,

respectively. MeHg (Laboratoire MAT, Beauport, Quebec,

Canada) was diluted with sterile phosphate-buffered saline

(PBS). MeHg or an equivalent volume of PBS was

administered by peroral injection to treated and vehicle

groups, respectively.

Three females treated with 3� 6 mg/kg of MeHg in

Condition GD12–14 were sacrificed at GD15 and two were

sacrificed at GD17; mercury levels were determined in

pools of livers and pools of brains of fetuses from each of

these females. The remaining females (n = 74) were checked

every morning for the presence of newborns. They gave

birth to 58 litters (GD7–9: 8, 10 and 9 litters treated with 0,

4 and 6 mg/kg of MeHg, respectively; GD12–14: 11, 9 and

11 litters treated with 0, 4 and 6 mg/kg of MeHg, respect-

ively). On the day of birth (GD19 or GD20), which was

defined as postnatal day 1 (PND1), three litters treated with

3� 6 mg/kg of MeHg and three vehicle litters in Condition

GD12–14 were randomly selected. Pups from these litters

were used for determinations of liver and brain levels of

mercury according to a procedure similar to the one used

with GD15 and GD17 fetuses, with the exception that

separate pools were made for females and males, and only

female pools were analyzed. In the remaining litters (n = 52),

the average number of pups per litter was very similar across

groups [Condition GD7–9: 8.6 ( ± S.E.M. = 0.6) in Group

3� 0 mg/kg; 9.0 ( ± S.E.M. = 0.4) in Group 3� 4 mg/kg;

and 8.1 ( ± S.E.M. = 0.7) in Group 3� 6 mg/kg; Condition

GD12–14: 8.9 ( ± S.E.M. = 1.0) in Group 3� 0 mg/kg; 9.3

( ± S.E.M. = 0.7) in Group 3 � 4 mg/kg; and 9.1

( ± S.E.M. = 0.4) in Group 3� 6 mg/kg]. These pups

remained with their biological mothers until weaned at

PND21. There was potential exposure to MeHg during

nursing, but a previous study [16], which used a fostering

procedure, showed that behavioral deficits were not sig-

nificantly influenced by the treatment to the mother, which

reared the pups, and were rather due to exposure of the mice

in utero.

F.Y. Dore et al. / Neurotoxicology and Teratology 23 (2001) 463–472464

At the age of 5 weeks, one female per litter was selected

for behavioral testing. Each female pup was individually

weighed and coded to make the experimenters blind to the

nature of their treatment. Mice were maintained on a 12:12 h

light–dark photoperiod and the tests were administered

during the light phase of the cycle. Because food reinforce-

ment (pieces of Fruit Loops cereals, Kellogg) was used in

three of the five behavioral tasks, the normal daily food

ration was restricted to reduce body weight to 85% of free-

feeding level and therefore, mice had to be housed indi-

vidually in standard plastic cages. In order to reduce body

weight to 85% of free-feeding level while allowing growth,

the weights of the mice under experiment were compared

and adjusted to the average weight of mice of the same age

(n = 15) receiving normal food ration. Water was provided

ad libitum.

Behavioral tests were administered 7 days/week. All mice

of Groups 3� 0 (n = 8), 3� 4 (n = 10) and 3� 6 mg/kg

(n = 9) in Condition GD7–9 and all mice of Groups 3� 0

(n = 8) and 3� 4 mg/kg (n = 9) in Condition GD12–14

completed the five tests. In Group 3� 6 mg/kg of Condition

GD12–14 (n = 8), three mice had to be replaced by another

female in the course of testing—one because of small

weight and weakness and two because of injuries.

2.2. Mercury determinations

For total mercury determinations, each tissue was

digested in concentrated nitric acid prior to reduction of

mercury to its metal state by stannous chloride and quan-

tification of mercury vapour by UV [12]. A certified

reference material (CRM) was used in the analytic process

for quality control purposes. The CRM used was a homo-

genised dogfish liver tissue sample (DOLT-2-) obtained

from the National Research Council of Canada. The

obtained value for mercury was 2.2 mg/g. The certified

value is 2.0 mg/g. The coefficient of variation for 50

determinations on different days was 4.9%.

2.3. Motor coordination on the rotarod

At 6 weeks of age, the female offspring were tested on

the rotarod. The balance rod was a 45-cm-long cylinder of

3 cm diameter wrapped with masking tape and suspended at

the top and center of a wooden enclosure (35� 45� 115 cm).

It was surrounded by three 30-cm-high black walls to

prevent animals from climbing off the rod and was divided

in the middle by a 30-cm-high vertical black screen, so that

two mice could be tested simultaneously. The cylinder was

connected to a DC motor (115 V, 33 A, 1/50 hp; Fisher

Scientific Canada), which supplied power to a gear box as a

function of electrical output from a variable transformer.

The floor of the enclosure was covered with a 40-cm-thick

cotton cushion to prevent any harm when the mice fell off

the rod. Each daily session included five trials during which

the mice were required to maintain balance on the rod for

120 s. A fall occurring in the first 10 s of a trial was defined

as a false start, which was a rare occurrence, and the trial

was resumed 30 s later. Each trial was administered on

squads of eight mice, so the intertrial interval was approx-

imately 8–10 min. Mice tested simultaneously were timed

on two separate chronometers and if one of the two mice

fell off the rotarod, only the chronometer for this mouse was

restarted. On the first session, the mice were placed on the

stationary rod (0 rpm) and on the second session, the rod

rotated at a constant speed of 3 rpm. On the following four

daily sessions, the rod rotated at a constant speed of 20 rpm.

Fall latencies were recorded and results from the four

sessions at 20 rpm were analyzed.

2.4. Spatial alternation training and delay testing in the

T maze

The mice were 7 weeks old at the beginning of the

T-maze task, which was administered in two identical

T mazes made of opaque acrylic with 30-cm-high walls

and 10-cm-wide corridors. The stem was divided into a start

box (30 cm) and a runway (50 cm). At the end of each

choice arm (30 cm), food reward could be concealed in an

opaque food cup. Black guillotine doors separated the start

box and the choice arms from the runway. On the first day,

the mice were familiarized to the apparatus. Several rein-

forcers were scattered on the stem, on the choice arms and in

the food cups. The mouse was placed in the start box and

was free to move in the maze while the experimenter

periodically opened and closed the guillotine doors. It was

allowed to explore the maze for a total of 10 min or until all

the reinforcers had been consumed, whichever occurred

first. Training to spatial alternation began the day after

familiarization. Each daily session consisted of 11 trials.

On the first trial (trial 0) of each session, both food cups in

the choice arms were baited with a food reinforcer and the

mouse was allowed to choose one arm. For the next 10

trials, the reinforcer was placed in the arm opposite to that

chosen by the mouse on the previous trial. Criterion for

ending training was eight successes out of 10 trials averaged

over two consecutive days. A maximum of 30 sessions was

administered. Once criterion was reached, delays of 30, 60

and 120 s were interposed between trials. Each delay was

used for two consecutive days and then was increased to the

next delay value.

2.5. Activity in the open field

The mice were 10 weeks old at the beginning of testing

in the open field. The floor of the apparatus measured

100� 100 cm and was divided into 25 equal squares. The

floor was surrounded by opaque acrylic 30-cm-high walls;

squares adjacent to walls were referred to as periphery, and

the nine remaining squares were referred to as center. On

each of five consecutive days, each mouse was moved from

its home cage and placed in the open field, facing the lower

F.Y. Dore et al. / Neurotoxicology and Teratology 23 (2001) 463–472 465

left corner. They were allowed to move freely for 15 min

but data were taken only in the first and in the last 5 min.

The following behavioral measures were recorded during

these 10 min: number of peripheral and central square

crossings and frequency of rearings. Total number of square

crossings, percentage of central square crossings and fre-

quency of rearings were averaged for the first session (new

environment) and for the following four daily sessions

(familiar environment).

2.6. Visual discrimination learning in the Y maze

The mice were 11 weeks old at the beginning of this

task. Two identical Y mazes made of opaque acrylic with

30-cm-high walls were used for visual discrimination learn-

ing. In each maze, the start box (10� 30 cm) led to a stem

choice area (20� 15 cm), which led itself to two parallel

runways (10� 30 cm) with opaque food cups placed at their

ends. Black guillotine doors separated the start box, the

stem choice area and the runways. Each runway and half of

the stem choice area leading to it were covered with a white

or a black interchangeable acrylic floor insert (10� 45 cm).

The floor inserts were thus visible to the mouse as soon as it

emerged from the start box. On each trial, the runway

covered with the black insert was baited (S+), whereas

the runway covered with the white insert was not (S� ).

The left and right positions of the inserts were determined

by a Gellerman pseudorandom sequence. Each daily session

consisted of 20 trials separated by a 10-s intertrial interval.

On the first two sessions, a correction procedure was used:

if the mouse chose the S� , an error was recorded and the

trial was immediately repeated with only the S+ runway

opened (forced successful choice). On the following ses-

sions, no correction procedure was used and each trial was

administered only once. Criterion for completion of the task

was 17 successes out of 20 trials averaged over two

consecutive days.

2.7. Spatial working reference memory in the radial

arm maze

The mice were 14 weeks old at the beginning of this

task. The eight-arm radial maze was made of wood and

painted flat gray. The maze was 60 cm from the floor. Each

arm was 60 cm long and 9 cm wide. A recessed food well

was located at the end of each arm. The center platform was

40 cm in diameter and was surrounded by a 40-cm-high

wooden wall. The entrance to each arm was blocked by a

gray acrylic guillotine door. The maze was centrally located

in a room providing a number of large objects (a window, a

poster, a vertical grid, etc.) and surrounded on two sides by

160-cm-high folding screens decorated with posters. The

animals’ behavior was monitored with a video camera that

was suspended over the maze. The camera was connected

to a video monitor that was located outside the folding

screens where the experimenter raised and lowered the

guillotine doors by pulling on strings in order to provide

access to the radial arms. The working reference memory

version of the radial arm maze task was used [26]. In this

version, half of the arms, always the same, is baited on each

daily trial and the other half is unbaited. As the animal

learns the rules of the task and/or features which are

constant from trial to trial (reference or long-term memory),

it becomes able to discriminate baited from unbaited arms

and avoids entering unbaited arms. It also becomes able to

avoid baited arms it has already visited within a trial

(working or short-term memory).

The first five daily trials served to familiarize the animals

with the apparatus. On the first day of familiarization,

several reinforcers were scattered on the central platform

and on four of the eight arms. The mouse was placed on the

central platform and was free to move in the maze for 5 min

while the experimenter periodically opened and closed the

guillotine doors. If the reinforcers in the four baited arms

were not found and consumed in the first 5 min, the mouse

was placed at the end of each baited arm and had to walk to

the central platform; then, from the central platform, it was

allowed to explore the maze for 5 min. On the following

4 days of familiarization, four baited arms were randomly

selected each day and the number of reinforcers was

gradually reduced until only the food wells of the four

selected arms were baited.

Training began the day after the last familiarization

session. There was one daily trial for 15 consecutive days.

On each trial, only four arms, always the same so they could

be maintained in reference memory, were baited. At the

beginning of a trial, the mouse was placed on the central

platform with all guillotine doors closed. After 10 s, the

doors were opened and the mouse was allowed to choose an

arm. When the mouse returned to the central platform, the

doors were closed and a 10-s waiting period began. After

the waiting period, all the doors were opened and the mouse

was allowed again to choose an arm. This procedure

continued until one of the following criteria was reached,

whichever occurred first: the four baited arms were visited,

16 choices were made or 10 min had elapsed. A reference

memory error was recorded each time the mouse visited for

the first time one of the four never baited arms and a

working memory error was recorded each time it revisited

an arm within a trial, whether this arm was originally baited

or not.

2.8. Statistical analysis

Within each condition (GD7–9 or GD12–14), results of

Groups 3� 0, 3� 4 and 3� 6 mg/kg were subjected to

ANOVAs with Group as a between-subject factor. On the

rotarod and delayed spatial alternation tasks, Session and

Delay were within-subject factors, respectively. Analyses of

simple main effects of the interaction (with Satterthwaite’s

correction for the error term and its degrees of freedom [15])

and Newman–Keuls tests served to locate specific signific-


ant effects. On the radial arm maze task, the total numbers of

correct choices, of reference memory errors and of working

memory errors made by mice in Condition GD7–9 were

also subjected to an ANOVA with Group as a between-

subject factor and Block of five daily trials as a within-

subject factor. As for Condition GD12–14, it appeared that

mice exposed to MeHg made fewer reference memory

errors, but also fewer choices, than control mice. Since the

total number of first choices made in 10 min differed in

treated and control mice, the measures in the radial arm

maze had to be subjected to special analyses. In each group

of Condition GD12–14, the numbers of visits to baited arms

(correct choices) and to unbaited arms (reference memory

errors) were compared by an ANOVAwith Type of arm and

Block of five trials as within-subject factors; the numbers of

working memory errors in the first and third blocks of five

trials were compared by a unilateral paired t test.

3. Results

3.1. Liver and brain levels of Hg and percentages of

survival at the age of 5 weeks

At GD15, GD17 and at birth, liver levels of mercury

in mice treated with 6 mg/kg of MeHg at GD12–14

were 12.70 ( ± S.E.M. = 1.79), 15.67 ( ± S.E.M. = 0.27) and

15.76 mg/g ( ± S.E.M. = 0.54), respectively, whereas brain

levels were 16.74 ( ± S.E.M.=.1.56), 13.26 ( ± S.E.M. = 3.76)

and 12.09 mg/g ( ± S.E.M. = 0.89). In control mice, liver and

brain levels of mercury at birth were 0.17 ( ± S.E.M. = 0.003)

and 0.13 mg/g ( ± S.E.M. = 0.01), respectively. Although liver

and brain levels of mercury were measured in a small

number of mice and only in Condition GD12–14, it is clear

that they were very high shortly after treatment (GD15 and

GD17) as well as at birth [3].

In each litter, the number of offspring which survived to

the age of 5 weeks, i.e., 1 week before the beginning of

behavioral testing, was divided by the number of offspring at

birth and the result was multiplied by 100 to give the

percentage of survival. In Condition GD7–9, the percentages

of survival in Groups 3� 0, 3� 4 and 3� 6 mg/kg were

90.3% ( ± S.E.M. = 3.4), 86.0% ( ± S.E.M. = 3.8) and 61.8%

( ± S.E.M. = 11.9), respectively. These percentages signific-

antly differed [F(2, 24) = 4.12, P < .05] and the Newman–

Keuls test (P < .05) showed that the percentage of survival in

Group 3� 6 mg/kg was significantly lower than in Groups

3� 0 and 3� 4 mg/kg. In Condition GD12–14, the percen-

tages of survival were 86.5% ( ± S.E.M. = 5.6) for Group

3� 0mg/kg, 68.7% ( ± S.E.M. = 11.2) for Group 3� 4mg/kg

and 60.5% ( ± S.E.M. = 7.4) for Group 3� 6 mg/kg. These

percentages also significantly differed [F(2, 22) = 11.34,

P < .01] and the Newman–Keuls test (P < .01) showed that

the percentage of survival in Group 3� 6 mg/kg was sig-

nificantly lower than in Group 3� 0 mg/kg. Thus, prenatal

exposure to MeHg reduced the percentage of survival at the

age of 5 weeks in mice treated with 3� 6 mg/kg of MeHg at

GD7–9 and at GD12–14.

3.2. Motor coordination on the rotarod task

In the first 2 days of testing, all mice were able to

maintain balance on the rod in at least four of the five trials

when the rod was stationary or rotated at a speed of 3 rpm.

In the following four daily sessions, when the rod rotated at

a constant speed of 20 rpm (Fig. 1), fall latencies increased

across sessions in all groups and thus, there was some

learning of motor coordination. However, prenatal exposure

to MeHg had no effect on motor learning. In Condition

GD7–9 [F(3, 72) = 14.73, P < .0001] and in Condition

GD12–14 [F(3, 66) = 19.49, P < .0001], the factor Session

was significant but the factor Group [Condition GD7–9:

F(2, 24) = 1.09; Condition GD12–14: F(2, 22) = 1.25] and

the interaction were not significant [Condition GD7–9:

F(6, 72) = 0.59; Condition GD12–14: F(6, 66) = 0.58].

3.3. Spatial alternation training and delay testing in the

T maze

Prenatal exposure to MeHg impaired training to spatial

alternation in the T maze (Fig. 2A). The number of sessions

to reach the learning criterion [F(2, 24) = 3.93, P < .05]

significantly differed in the groups of Condition GD7–9.

The Newman–Keuls test (P < .05) showed that mice treated

with 4 and 6 mg/kg of MeHg required more sessions than

control mice. In Condition GD12–14, there was also a

Fig. 1. Fall latencies on the rotarod task. Data points are group means

± S.E.M. for four consecutive daily sessions. Number of mice in each

group: Condition GD7–9: 3� 0 (n= 8); 3� 4 (n= 10); 3� 6 mg (n= 9);

Condition GD12–14: 3� 0 (n= 8); 3� 4 (n= 9); 3� 6 mg (n= 8). Data of

each condition were analyzed by ANOVAwith Group as a between-subject

factor and Sessions (1–4) as a within-subject factor. The factor Group

[Condition GD7–9: F(2, 24) = 1.09; Condition GD12–14: F(2, 22) = 1.25]

and its interaction with the factor Session [Condition GD7–9: F(6,

72) = 0.59; Condition GD12–14: F(6, 66) = 0.58] were not significant, but

the factor Session was significant [Condition GD7–9: F(3, 72) = 14.73,

P < .0001; Condition GD12–14: F(3, 66) = 19.49, P< .0001].


significant difference between groups in the number of

sessions to reach criterion [F(2, 22) = 4.17, P < .05] and

the Newman–Keuls test (P < .05) revealed that Group 3� 6

mg/kg required more sessions to reach criterion than

Groups 3� 0 and 3� 4 mg/kg. In Condition GD12–14,

all mice were able to produce 80% correct responses, but in

Condition GD7–9, 4/10 mice treated with 3� 4 mg/kg of

MeHg and 2/9 mice treated with 3� 6 mg/kg failed to attain

this level of performance within the maximum of 30

sessions. Those mice were not tested in the delayed spatial

alternation test.

When delays of 30, 60 and 120 s were interposed between

trials, spatial alternation performance deteriorated compared

to the last two sessions of training (Fig. 2B), but it was not

impaired by prenatal administration of MeHg at GD7–9

or GD12–14. In both conditions, the factor Delay had

a significant effect [Condition GD7–9: F(3, 54) = 22.77,

P < .0001; Condition GD12 – 14: F(3, 66) = 18.33,

P < .001], whereas the factor Group [Condition GD7–9:

F(2, 18) = 0.10; Condition GD12–14: F(2, 22) = 0.16] and

the interaction [Condition GD7–9: F(6, 54) = 0.55; Con-

dition GD12–14: F(6, 66) = 0.52] were not significant.

However, differences between treated and control mice

might have been masked by a floor effect because at a 30-s

delay, the number of correct choices was already between

55% and 70%.

3.4. Activity in the open field

Fig. 3A presents the total number of square crossings in

the open field during the 10 min of recording in the first

session and in the following four sessions. Only mice

treated with MeHg at GD12–14 were significantly hypo-

active. When the environment was new (Session 1), the

groups in Condition GD7–9 did not differ [F(2, 24) = 0.60],

but the groups in Condition GD12–14 did [F(2, 22) = 5.98,

P < .01]: Groups 3� 6 and Group 3� 4 mg/kg crossed

significantly fewer squares than Group 3� 0 mg/kg (New-

man–Keuls at P < .01). In Sessions 2–5, when the envir-

onment was becoming familiar, similar results were

observed. The groups in Condition GD7–9 did not differ

[F(2, 24) = 0.16], but the groups in Condition GD12–14 did

[F(2, 22) = 9.01, P < .005]: Groups 3� 4 and 3� 6 mg/kg

crossed significantly fewer squares than Group 3� 0 mg/kg

(Newman–Keuls at P < .01).

Fig. 3B presents the percentage of central square cross-

ings in 10 min. If this behavioral measure is an index of

fear or emotionality as it has frequently been described

[19], there was no evidence of altered function in mice

with prenatal exposure to MeHg. In both conditions, the

groups did not differ whether the environment was new

[Session 1: Condition GD7–9: F(2, 24) = 0.51; Condition

GD12–14: F(2, 22) = 0.11] or familiar [Sessions 2–5:

Condition GD7–9: F(2, 24) = 0.57; Condition GD12–14:

F(2, 22) = 0.97]. Finally, Fig. 3C presents the frequency of

rearings in the open field. Again, in both conditions, the

groups did not differ whether the environment was new

[Session 1: Condition GD7–9: F(2, 24) = 0.55; Condition

GD12–14: F(2, 22) = 0.02] or familiar [Sessions 2–5:

Condition GD7–9: F(2, 24) = 0.39; Condition GD12–14:

F(2, 22) = 0.82].

Fig. 2. Training and delay testing on spatial alternation in the T maze. (A)

Number of sessions to reach criterion. Number of mice in each group:

Condition GD7–9: 3� 0 (n = 8); 3� 4 (n = 10); 3� 6 mg (n = 9);

Condition GD12–14: 3� 0 (n= 8); 3� 4 (n= 9); 3� 6 mg (n= 8). Each

histogram represents the mean ± S.E.M. Data of each condition were

analyzed by a one-way ANOVA followed by a posteriori comparisons with

the Newman–Keuls test. In Condition GD7–9, the factor Group was

significant [ F(2, 24) = 3.93, P < .05]: mice treated with 4 and 6 mg/kg of

MeHg required more sessions to reach criterion than control mice ( P< .05).

In Condition GD12–14, the factor Group was also significant [ F(2,

22) = 4.17, P< .05] and Group 3� 6 mg/kg required more sessions to reach

criterion than Groups 3� 0 and 3� 4 mg/kg ( P< .05). (B) Frequency of

correct choices in delay testing. Number of mice in each group: Condition

GD7–9: 3� 0 (n= 8); 3� 4 (n= 6); 3� 6 mg (n= 9); Condition GD12–14:

3� 0 (n= 8); 3� 4 (n= 9); 3� 6 mg (n= 8). Data points are group means

± S.E.M. for each delay (0-s delay: data from the two criterion sessions).

Data of each condition were analyzed by ANOVAwith Group as a between-

subject factor and Delay as a within-subject factor. The factor Group


and the interaction [Condition GD7–9: F(6, 54) = 0.55; Condition GD12–

14: F(6, 66) = 0.52] were not significant, but the factor Delay was

significant [Condition GD7–9: F(3, 54) = 22.77, P < .0001; Condition

GD12–14: F(3, 66) = 18.33, P < .001].


3.5. Visual discrimination learning in the Y maze

Prenatal exposure to MeHg had no effect on learning the

visual discrimination in the Y maze (Fig. 4). The number of

sessions to reach criterion did not significantly differ in the

three groups of mice either in Condition GD7–9 [F(2,

24) = 1.29] or in Condition GD12–14 [F(2, 22) = 2.33].

For an unknown reason, all groups in Condition GD12–

14 seemed to require more sessions than groups in Con-

dition GD7–9.

3.6. Spatial working-reference memory in the radial

arm maze

In Condition GD7–9 (Fig. 5A), the performance in the

radial maze was similar in treated and control mice.

The frequency of correct choices was high and there

was no significant difference between groups [Group:

F(2. 24) = 0.02; Group�Block: F(4, 48) = 0.07] or across

blocks of trials [ F(2, 48) = 0.00]. The frequency of

reference memory errors significantly decreased across

blocks of trials [ F(2, 48) = 5.01, P < .01], but there


F(2, 24) = 0.13; Group�Block: F(4, 48) = 0.29]. Similarly,

the frequency of working memory errors significantly

decreased across blocks [F(2, 48) = 3.85, P < .05] and there


F(2, 24) = 1.57; Group�Block: F(4, 48) = 0.32].

In Condition GD12–14, the results on the radial arm

maze task were more complex. In the course of testing, it

became clear that mice of Groups 3� 4 and 3� 6 mg/kg

behaved differently than control mice of the same condition

(Fig. 5B): a trial had frequently to be ended after the 10-min

criterion was reached, before the four baited arms were

Fig. 4. Number of sessions to reach criterion on the visual discrimination in

the Y maze. Each histogram represents the mean ± S.E.M. Number of mice

in each group: Condition GD7–9: 3� 0 (n= 8); 3� 4 (n= 10); 3� 6 mg

(n= 9); Condition GD12–14: 3� 0 (n= 8); 3� 4 (n= 9); 3� 6 mg (n= 8).

Data of each condition were analyzed by a one-way ANOVA. In Conditions

GD7–9 [ F(2, 24) = 1.29] and GD12–14 [ F(2, 22) = 2.33], the groups did

not differ.

Fig. 3. Behavior in the open field. Number of mice in each group:

Condition GD7–9: 3� 0 (n = 8); 3� 4 (n = 10); 3� 6 mg (n = 9);

Condition GD12–14: 3� 0 (n= 8); 3� 4 (n= 9); 3� 6 mg (n= 8). Each

histogram represents the mean ± S.E.M. Data of each condition were

analyzed by a one-way ANOVA followed by a Newman–Keuls test. (A)

Total number of square crossings during the 10-min of recording when the

environment was new (Session 1) and familiar (Session 2–5). In Session 1,

the groups did not differ in Condition GD7–9 [ F(2, 24) = 0.60], but they

significantly differed in Condition GD12–14 [ F(2, 22) = 5.98, P< .01],

with Group 3� 0 mg/kg crossing more squares than Group 3� 6 mg/kg

( P< .01). In Sessions 2–5, the groups did not differ in Condition GD7–9

[ F(2, 24) = 0.16], but they significantly differed in Condition GD12–14

[ F(2, 22) = 9.01, P< .005], with Group 3� 0 mg/kg crossing more squares

than Groups 3� 4 and 3� 6 mg/kg ( P< .01). (B) Percentage of central

square crossings when the environment was new (Session 1) and familiar

(Session 2–5). The groups in both conditions did not differ in Session 1


or in Sessions 2–5 [Condition GD7–9: F(2, 24) = 0.57; Condition GD12–

14: F(2, 22) = 0.97]. (C) Frequency of rearings when the environment was

new (Session 1) and familiar (Session 2–5). The groups in both conditions

did not differ in Session 1 [Condition GD7–9: F(2, 24) = 0.55; Condition

GD12–14: F(2, 22) = 0.02] or in Sessions 2–5 [Condition GD7–9: F(2,

24) = 0.39; Condition GD12–14: F(2, 22) = 0.82].


visited and before 16 choices were made. ANOVAs made

on the overall results of the 15 trials revealed that although

the groups did not differ in terms of working memory errors

[F(2, 22) = 1.72], they significantly differed in terms of

correct choices [F(2, 22) = 4.45, P < .05] and reference

memory errors [F(2, 22) = 5. 29, P < .05]. Newman–Keuls

tests (P < .05) showed that Groups 3� 4 and 3� 6 mg/kg

made fewer reference memory errors but they also made

fewer correct choices. Because the number of first entries

into arms was lower in prenatally exposed groups than in

control mice, direct intergroup comparisons were not pos-

sible. An intragroup strategy of analysis was adopted.

Long-term retention or reference memory of the rules and/

or constant features of the task implies that the animal

gradually learned to discriminate baited and unbaited arms.

Thus, reference memory was evaluated, within each group,

by comparing the number of visits to baited arms (correct

choices) and the number of visits to unbaited arms (reference

memory errors). An ANOVAwith Type of arm and Block of

five trials as within-subject factors was used for these

comparisons. In Group 3� 0 mg/kg, the factor Type of arm

was significant [F(1, 7) = 34.97, P < .001], and the factor

Block [F(2, 14) = 1.43] and the interaction [F(2, 14) = 2.78]

were not significant. Thus, control mice visited more baited

arms than unbaited arms on all three blocks of trials. Treat-

ment with MeHg in Condition GD12–14 impaired discrim-

ination of baited and unbaited arms and this impairment

was more severe at the highest dose. In Group 3� 4 mg/kg,

the factor Type of arm [F(1, 8) = 29.07, P < .001] and the

interaction [F(2, 16) = 5.61, P < .02] were significant, but

the factor Block [F(2, 16) = 0.19] was not significant. The

analysis of simple main effects showed that in Group

3� 4 mg/kg, more visits were made to baited arms than to

unbaited arms on the second [F(1, 16) = 15.11, P < .002] and

on the third blocks of trials [F(1, 16) = 44.20, P < .0001], but

not on the first block [F(1, 16) = 3.02]. In Group 3� 6mg/kg,

the frequency of visits to baited and unbaited arms did not

differ on any block: the factors Type of arm [F(1, 7) = 3.25]

and Block [F(2, 14) = 1.85], as well as the interaction [F(2,

14) = 0.11], were not significant. Whereas Group 3� 4 mg/

kg was initially slower to learn the rules and/or constant

features of the task than the vehicle group, Group 3� 6 mg/

kg did not discriminate baited and unbaited arms on any of the

three blocks of five trials.

In Condition GD12–14, working memory errors were

analyzed by comparing, within each group, the numbers of

reentries on the first and third blocks of trials. In normal

Fig. 5. Frequency of correct choices (visits to baited arms), of reference

memory errors (first visits to unbaited arms) and working memory errors

(reentries in baited or unbaited arms) in the radial arm maze. Data points

are group means ± S.E.M. for each block of five trials. (A) Data of

Condition GD7–9. Number of mice in each group: 3� 0 (n= 8); 3� 4

(n= 10); 3� 6 mg (n= 9). Data were analyzed by ANOVAs with Group as

a between-subject factor and Blocks of five trials as a within-subject factor.

There was no significant difference between groups or across blocks of

trials in the number of correct choices [Group: F(2, 24) = 0.02; Block: F(2,

48) = 0.00; Group�Block: F(4, 48) = 0.07]. The frequency of reference

memory errors significantly decreased [ F(2, 48) = 5.01, P< .01] and there

was no significant difference between groups [Group: F(2, 24) = 0.13;

Group� Block: F(4, 48) = 0.29]. The frequency of working memory errors

significantly decreased [ F(2, 48) = 3.85.P < .05] and there was no significant

difference between groups [Group: F(2, 24) = 1.57; Group�Block: F(4,

48) = 0.32]. (B) Data of Condition GD12–14. Number of mice in each

group: 3� 0 (n= 8); 3� 4 (n= 9); 3� 6 mg (n= 8). In order to compare

correct choices and reference memory errors, data of each group were

analyzed by ANOVAs with Type of arm and Block of five trials as a

within-subject factors. In Group 3� 0 mg/kg, the factor Type of arm was

significant [ F(1, 7) = 34.97, P < .001], and the factor Block [ F(2.

14) = 1.43] and the interaction [ F(2, 14) = 2.78] were not significant. In

Group 3� 4 mg/kg, the factor Type of arm [ F(1, 8) = 29.07, P < .001] and

the interaction [ F(2, 16) = 5.61, P < .02] were significant, but the factor

Block [ F(2, 16) = 0.19] was not significant. The analysis of simple main

effects showed that in Group 3� 4 mg/kg, more visits were made to

baited arms than to unbaited arms on the second [ F(1, 16) = 15.11.

P < .002] and third blocks [ F(1, 18) = 44.20, P < .0001], but not on the

first block [ F(1, 16) = 3.02]. Finally, in Group 3� 6 mg/kg, the frequency

of visits to baited and unbaited arms did not differ on any block because

the factor Type of arm [ F(1, 7) = 3.25], Block [ F(2, 14) = 1.85] and the

interaction [ F(2, 14) = 0.11] were not significant. For each group, the

frequencies of working memory errors on the first and third blocks of

trials were compared by a unilateral Student’s t test for paired samples.

The frequency of working memory errors significantly decreased in Group

3� 0 mg/kg [t (7) = 1.91, P < .05], but not in Group 3� 4 mg/kg

[t(8) = 0.46] or in Group 3� 6 mg/kg [t(7) = 0.40].


animals, these errors usually decrease across trials as they

learn to avoid revisiting an arm. In fact, the number of

working memory errors significantly decreased in Group

3� 0 mg/kg [t (7) = 1.91, P < .05], but not in Groups 3� 4

[t (8) = 0.46] and 3� 6 mg/kg [t (7) = 0.40].

4. Discussion

The mercury determinations in brain and liver of mice

treated at GD12–14 indicate that high exposure to MeHg

lasted from very shortly after treatment until birth. A

weakness of the present experiment is that mercury levels

were not measured in mice treated at GD7–9. However, an

indication of the effects of administration of MeHg in this

condition is that, as in Condition GD12–14, survival to the

postnatal age of 5 weeks was significantly decreased by

administration of 6 mg/kg of MeHg.

Observations made during the experiment did not reveal

any obvious sign of motor deficit and testing on the

rotarod did not show any impairment in learning motor

coordination, even in mice treated with 3� 6 mg/kg.

Previous studies on rodents did find gross impairments

of motor function after prenatal exposure to MeHg, but

these studies examined reflexive behavior in early devel-

opment [25] or swimming ability [13,34,36]. On the other

hand, our results in the open field are consistent with early

[34,35,38] and with more recent [19] reports, which

showed that prenatal exposure to MeHg significantly

decreases locomotor activity. In our experiment, this effect

was observed only in mice treated at GD12–14 and

frequency of rearings was not affected. Contrary to the

results of Kim et al., there was no evidence that hypo-

activity in the open field was related to fear or to changes

in emotional status: the percentage of central square cross-

ings in treated mice and control mice did not differ.

Hypoactivity in mice treated with MeHg at GD12–14

was not specific to novelty and also appeared when the

open field was familiar. This result suggests that the

decrease in locomotor activity was related to damage to

the nucleus accumbens rather than to limbic structures [4].

In order to discriminate the black and white floors in the

Y maze, to acquire the spatial alternation response in the T

maze and to discriminate baited and unbaited arms in the

radial maze, mice had to learn the basic rules and/or

constant features of the task. Thus, the three tasks required

intact reference memory or long-term retention of either an

individual cue (black floor in the visual discrimination task)

or of spatial information (T maze and radial maze). In all

mice treated with MeHg, learning on the visual discrimina-

tion task was normal and this result is consistent with

previous experiments on rodents [2,13]. On spatial tasks,

reference memory was impaired by prenatal exposure to

MeHg whether the task involved egocentric or allocentric

information. However, the effects of treatment windows on

the two spatial tasks differed.

On spatial alternation training, both treated groups of

Condition GD7–9 required more sessions to reach the

learning criterion and in Condition GD12–14, only Group

6 mg/kg was impaired. This result suggests that reference

memory for egocentric spatial information was affected by a

lower dose ofMeHgwhen treatment occurred at GD7–9 than

at GD12–14. In the radial arm maze, reference memory for

allocentric spatial information was impaired only in Con-

dition GD12–14 and this impairment was more severe after

prenatal exposure to the highest dose.Whereas discrimination

of baited and unbaited arms was impaired on all three blocks

after prenatal exposure to 6mg/kg, it was impaired only on the

first block of trials in mice exposed to 4 mg/kg. Reference

memory for egocentric spatial information [10,28] and ref-

erence memory for allocentric spatial information [27] have

been both associated with the function of the caudate nucleus

and yet, exposure to MeHg at two fetal developmental stages

had different effects on spatial alternation training and on

the radial arm maze task. One possible explanation is that

treatment at GD7–9 and GD12–14 altered the function of

different neurotransmitter systems in the caudate nucleus.

Delay testing of spatial alternation in the T maze suggests

that working memory for egocentric spatial information and

frontal function were not affected by prenatal exposure to

MeHg. However, this conclusion can only be tentative.

First, working memory errors on this task were already

low at the shortest delay and the possibility of a floor effect

cannot be definitely excluded. Second, in Condition GD7–9,

some treated mice did not reach criterion and could not be

tested with delays although their performance on this task

was probably more affected by exposure to MeHg than in

Condition GD12–14. In contrast, the analysis of reentries in

the radial arm maze revealed that in Condition GD12–14,

but not in Condition GD7–9, working memory errors did not

decrease between the first and the third blocks of trials in

mice prenatally exposed to either dose of MeHg. Thus,

working memory for places was impaired and this deficit

is usually associated with dysfunctions of the hippocampus

and/or adjacent entorhinal cortex.

In summary, treatment with MeHg at two different stages

of fetal development had different effects. In mice treated

with MeHg at GD7–9, reference memory for egocentric

spatial information was impaired, whereas in mice treated

with MeHg at GD12–14, a variety of neurobehavioral

functions were altered. Damages to the ventral striatum,

the dorsal striatum and the hippocampal formation are

suggested by the hypoactivity observed in the open field,

by reference memory deficits on spatial alternation training

and on the radial maze task, and by working memory

deficits in the radial maze, respectively.

Acknowledgments

The authors thank Mr. Alain Tremblay and Ms.

Micheline Noel for their excellent technical assistance. This


research was supported by funds from Fonds de Recherche

en Sante du Quebec (FRSQ)/Hydro-Quebec (Programme de

recherche en sante de l’enfant) and by Health Canada

(Programme Saint-Laurent Vision 2000). The research

received approval from the Comite de protection des

animaux de laboratoire de l’Universite Laval, which is

responsible for the application and enforcement of the rules

of the Canadian Council on Animal Care.

References

[1] N. Auvray, J. Caston, A. Reber, T. Stelz, Role of the cerebellum in the

ontogenesis of the equilibrium behavior in the young rat: A behavioral

study, Brain Res. 505 (1989) 291–301.

[2] J. Buelke-Sam, C.A. Kimmel, J. Adams, C.J. Nelson, C.V. Vorhees,

D.C. Wright, V. St. Omer, B.A. Korol, R.E. Butcher, M.A. Geyer,

J.F. Holson, C.L. Kutscher, M.J. Wayner, Collaborative behavioral

teratology study: Results, Neurobehav. Toxicol. Teratol. 7 (1985)

591–624.

[3] T.M. Burbacher, P.M. Rodier, B. Weiss, Methymercury developmental

neurotoxicity: A comparison of effects in humans and animals, Neuro-

toxicol. Teratol. 12 (1990) 191–202.

[4] L.H. Burns, L. Annett, A.E. Kelley, B.J. Everitt, T.W. Robbins, Ef-

fects of lesions to amygdala, ventral subiculum, medial prefrontal

cortex, and nucleus accumbens on the reaction to novelty: Implication

for limbic–striatal interactions, Behav. Neurosci. 110 (1996) 60–73.

[5] J. Caston, N. Jones, T. Stelz, Role of preoperative and postoperative

sensorimotor training on restoration of the equilibrium behavior in

adult mice following cerebellectomy, Neurobiol. Learn. Mem. 64

(1995) 195–202.

[6] J. Caston, F. Vasseur, T. Stelz, C. Chianale, N. Delhaye-Bouchaud,

J. Mariani, Differential roles of cerebellar cortex and deep cerebellar

nuclei in the learning of the equilibrium behavior: Studies in intact and

cerebellectomized lurcher mutant mice, Dev. Brain Res. 86 (1995)

311–316.

[7] L.W. Chang, Neurotoxic effects of mercury: A review, Environ. Res.

14 (1977) 329–373.

[8] L.W. Chang, Mercury in experimental and clinical neurotoxicology,

in: P.S. Spencer, H.H. Schaumburg (Eds.), Experimental and Clinical

Neurotoxicology, Williams and Wilkins, London, 1980, pp. 508–526.

[9] L.W. Chang, G.L. Guo, Fetal Minamata disease: Congenital methyl-

mercury poisoning, in: W.S. Slikker, L.W. Chang (Eds.), Handbook of

Developmental Neurotoxicology, Academic Press, San Diego, 1998,

pp. 507–515.

[10] D. Cook, R.P. Kesner, Caudate nucleus and memory for egocentric

localization, Behav. Neural Biol. 49 (1988) 332–343.

[11] J.M. De Brabander, J.P.C. De Bruin, C.G. van Enden, Comparison of

the effects of neonatal and adult medial prefrontal cortex lesions on

food hoarding and spatial delayed alternation, Behav. Brain Res. 42

(1991) 67–75.

[12] V. Ebbestadt, N. Gunderson, T.A. Torgrimsen, Simple method for the

determination of inorganic mercury and methylmercury in biological

samples by flameless atomic absorption, At. Absorpt. Newsl. 14

(1975) 142–143.

[13] J. Elsner, B. Hodel, K.E. Suter, D. Oelke, B. Ulbrich, G. Shreiner,

V. Cuomo, R. Cagiano, R.E. Rosengren, J.E. Karlsson, K.G. Halid,

Detection limits of different approaches in behavioral teratology,

and correlation of effects with neurochemical parameters, Neuro-

toxicol. Teratol. 10 (1988) 155–167.

[14] R. Gerlai, K.J. Millen, K. Herrup, K. Fabien, A.L. Joyner, J. Roder,

Impaired motor learning performance in cerebellar En-2 mutant mice,

Behav. Neurosci. 110 (1996) 126–133.

[15] D.C. Howell, Statistical Methods for Psychology, third ed., PWS-Kent

Publishing, Boston, 1992.

[16] J.A. Hughes, Z. Annau, Postnatal behavioral effects in mice after

prenatal exposure to methylmercury, Pharmacol. Biochem. Behav. 8

(1976) 385–391.

[17] M. Inouye, K. Murao, Y. Kajiwara, Behavioral and neuropathological

effects of prenatal methylmercury exposure in mice, Neurobehav.

Toxicol. Teratol. 7 (1985) 227–232.

[18] M.H. Kaufman, The Atlas of Mouse Development, Academic Press,

London, 1992.

[19] C.-Y. Kim, K. Nakai, Y. Kasanuma, H. Satoh, Comparison of neuro-

behavioral changes in three inbred strains of mice prenatally exposed

to methylmercury, Neurotoxicol. Teratol. 22 (2000) 397–403.

[20] B. Kolb, A.J. Nonneman, R.K. Singh, Double dissociation of spatial

impairment and perseveration following selective prefrontal lesions in

rats, J. Comp. Physiol. Psychol. 87 (1974) 772–780.

[21] M.R. Krigman, Neuropathology of heavy metal intoxication, Environ.

Health Perspect. 26 (1978) 117–120.

[22] R. Lalonde, A.N. Bensoula, M. Filali, Rotarod sensorimotor learning

in cerebellar mutant mice, Neurosci. Res. 22 (1995) 423–426.

[23] J.K. Larsen, I. Divac, Selective ablations within the prefrontal cortex

of the rat and performance of delayed alternation, Physiol. Psychol. 6

(1978) 15–17.

[24] T. Myhrer R.J., T.S. Johannesen, Learning and retention of a visual

discrimination task in rats with various combinations of lesions in the

temporal–hippocampal region, Brain Res. Bull. 36 (1995) 499–503.

[25] K. Olson, G.M. Boush, Decreased learning capacity in rats exposed

prenatally and postnatally to low doses of mercury, Bull. Environ.

Contam. Toxicol. 13 (1975) 73–79.

[26] D.S. Olton, B.C. Papas, Spatial memory and hippocampal function,

Neuropsychologia 17 (1979) 669–682.

[27] M.G. Packard, N.W. White, Lesions of the caudate nucleus selectively

impair ‘‘reference memory’’ acquisition in the radial maze, Behav.

Neural Biol. 53 (1990) 39–50.

[28] M. Potegal, The caudate nucleus egocentric localization system, Acta

Neurobiol. Exp. 32 (1969) 479–494.

[29] D.C. Rice, Sensory and cognitive effects of developmental methyl-

mercury exposure in monkeys, and a comparison to effects in rodents,

Neurotoxicology 17 (1996) 139–154.

[30] P.M. Rodier, Chronology of neuron development: Animal studies

and their clinical implications, Dev. Med. Child Neurol. 22 (1980)

525–545.

[31] H. Satoh, N. Yasuda, S. Shimai, Development of reflexes in neonatal

mice prenatally exposed to methylmercury and selenium, Toxicol.

Lett. 25 (1985) 199–203.

[32] S. Shimai, H. Satoh, N. Yasuda, Taste aversion learning and perinatal

methylmercury exposure in mice, Ind. Health 22 (1984) 41–44.

[33] J.M. Spyker, M. Smithberg, Effects of methylmercury on prenatal

development in mice, Teratology 5 (1972) 181–190.

[34] J.M. Spyker, S.B. Sparber, A.M. Goldberg, Subtle consequences of

methylmercury exposure: Behavioral deviations in offspring of treated

mothers, Science 177 (1972) 621–623.

[35] M.Q. Su, G.T. Okita, Behavioral effects on the progeny of mice

treated with methylmercury, Toxicol. Appl. Phamacol. 38 (1976)

195–205.

[36] T. Tanimura, E. Ema, T. Kihara, Effects of combined treatment with

methylmercury and polychlorinated biphenyls (PCSs) on the develop-

ment of mouse offspring, in: P.V.N. Persaud (Ed.), Neural and Behav-

ioral Teratology, MTP Press, Lancaster, 1980, pp. 163–198.

[37] F. Van Haaren, J.P.C. De Bruin, R.P.W. Heinsbroek, N.E. Van de Poll,

Delayed spatial response alternation: Effects of delay interval duration

and lesions of the medial prefrontal cortex on response accuracy of

male and female Wistar rats, Behav. Brain Res. 19 (1985) 41–49.

[38] C. Watanabe, H. Satoh, Evolution of our understanding of methyl-

mercury as a health threat, Environ. Health Perspect. 104 (1996)

367–379 (Supplement).


Documents

Neurobehavioral changes in mice treated with methylmercury at two different stages of fetal development