TY - JOUR
AU - Tanimura, Takashi
AB - Abstract The effects of prenatal aflatoxin B1 (AFB) exposure on eight behavioral parameters in Jcl:Wistar rat offspring were assessed. Pregnant rats were injected subcutaneously with 0.3 mg/kg/day of AFB dissolved in dimethylsulfoxide on days 11–14 (Group A) or 15–18 (Group B) of gestation. Controls received the vehicle similarly on days 11–18 of gestation. Before weaning, the offspring were examined using the cliff avoidance response (5 days of age), the negative geotaxis reflex (7 days), and swimming development (6, 8, and 10 days). After weaning, animals were examined using the rotarod test (5 weeks of age), the open field test (6 weeks), a conditioned avoidance learning test (14 weeks), an underwater T-maze test (15 weeks), and a reproduction test (16 weeks). The preweaning offspring in the AFB-A group showed significantly lower success rates than controls in cliff avoidance responses. In swimming development, the offspring in the AFB-A group had significantly lower scores than controls for swimming direction. In the rotarod test, the AFB-A group remained on the rod for a significantly shorter time than the controls at 15 rpm on both the first and second trial days. The avoidance performance of the rats in AFB-A and AFB -B groups was significantly poorer than that of controls. These results indicate that prenatal exposure to AFB produced a delay of early response development, impaired locomotor coordination, and impaired learning ability in the offspring of rats exposed to AFB during middle pregnancy, and the early gestational exposure appears to produce more effects than latter exposure. aflatoxin B1, 2,3,6aα,9aα-tetrahydro-4-methoxycyclopenta[c]furo[3′,2′,: 4,5]furo[2, 3-h][1] benzopyran-1, 11-dione, mycotoxin, behavioral teratology, prenatal exposure, developmental toxicity, neurotoxicity Aflatoxin B1 (AFB), produced by several fungi such as Aspergillus flavus and A. parasiticus (Wilson et al., 1968) (Fig. 1), is one of the most important food borne mycotoxins and is one of the most potent hepatotoxins and carcinogens in many animal species (Eaton and Gallagher, 1994; Wogan and Newberne, 1967), and it has also been implicated in the etiology of human liver cancer in a number of epidemiologic investigations (IARC, 1993). The developmental toxicity of AFB has been studied in various animals (for review, see Hayes 1981; Hood, 1979; Hood and Szczech, 1983). AFB has been reported to be teratogenic and/or embryotoxic in rats (Elegbe et al., 1974; Grice et al., 1973; LeBreton et al., 1964; Panda et al 1970; Tanaka, 1975), in mice (Arora et al., 1981; DiPaolo et al., 1967; Roll et al., 1990; Tanimura et al., 1982), in hamsters (DiPaolo et al., 1967; Elis and DiPaolo, 1967), in chick embryos (Bassir and Adekunle,1970; Cilievici et al., 1980; Dietert et al., 1985), in tadpoles (Gabor et al., 1973), and Japanese medaka eggs (Llewellyn et al., 1977). Tranplacental carcinogenesis in rats (Goerttler et al., 1980; Grice et al., 1973; Tanaka, 1975), and selective immune depression in chick embryos (Dietert et al., 1985) have also been reported. A previous study in our laboratory demonstrated that AFB induced cleft palate, skeletal malformations, and intrauterine growth retardation in mouse fetuses of dams injected intraperitoneally with doses of 32 mg/kg/day for 2 days of days 6–7, 8–9, 10–11, 12–13 of gestation (Tanimura et al., 1982). In the past 20 years, the importance of postnatal evaluation for behavioral teratology has received increasing recognition, and the test battery system for assessment of behavioral teratogenic potential in reproductive and developmental toxicity study has been used widely (Riley and Vorhees, 1986; Tanimura, 1990, 1992; Ulbrich and Palmer, 1996). However, there is little published data on the behavioral teratogenic effects of AFB in animals or in humans. There has been only one study to date on the functional effects caused by prenatal AFB exposure. Chentanez et al. (1986) showed that AFB 2.0 mg/kg administered intravenously to Fisher rats on days 8–10 or 15–17 of gestation induced a decrease in some types of behaviors and in motor activity levels in 1-month-old offspring. The present study, therefore, was conducted to determine the behavioral teratogenic effects on the rat offspring of dams injected subcutaneously with a subteratogenic dose of AFB during mid or late organogenesis. The test battery system recently developed in our laboratory (Kihara, 1991; Kihara et al.,1995) was used for functional evaluations. MATERIALS AND METHODS Animals and Treatments Nine-week-old male (230–250 g) and female (170–190 g) Jcl:Wistar rats were purchased from Clea Japan, Osaka, Inc., and acclimated to the laboratory for 2 weeks prior to mating. Pellet diet OA-2 (Clea Japan, Osaka, Inc.) and tap water were available . Animals were maintained in a room with a controlled temperature of 23 ± 20C, a relative humidity of 50 ± 10%, and a 12-h light:dark cycle (lights on at 07:00 a.m.). Eleven-week-old females were mated overnight for 16 h with one male each. Mating was confirmed if sperm was found in a vaginal smear or if a plug was observed. The following morning (day 0 of gestation), the pregnant rats were randomly assigned to one of three groups, each consisting of 10 female rats at conception. They were individually housed in polypropylene cages with wooden shavings provided throughout gestation and lactation, and left undisturbed except for treatment and weighing until parturition. Pregnant rats were injected subcutaneously with 0.3 mg/kg/day of AFB (Makor Chemical Ltd., Jerusalem, Israel) dissolved in 0.9 mg/kg of dimethyl sulfoxide (Wako Pure Chemical Industries Ltd., Osaka) on days 11–14 (Group A) or 15–18 (Group B) of gestation. Control animals received subcutaneously the vehicle only on days 11–18 of gestation. The treatment volume was 1.0 ml/kg body weight. Fresh solutions of AFB were prepared on the day of use. The dams were allowed to deliver spontaneously and rear their offspring until weaning. At 4 days after birth, the litters were culled randomly to groups of eight offspring with the same number of males and females as far as possible. At 21 days, the offspring were weaned, separated by sex, and housed in hanging wire-mesh cages with four littermates of the same sex. All offspring were identified by marking with dry ink before weaning and with picric acid-ethanol solution after weaning. Body Weight and Physical Landmarks The dams were weighed on days 0, 7, 14, and 21 of gestation, daily during treatment, and on days 0, 4, 7, 14, and 21 after delivery. After weaning their litters (21 days after delivery), all the dams were killed by ether overdose and examined for numbers of implantation sites and any abnormalities of the reproductive organs. At birth, all live and dead offspring were counted. Live offspring were weighed, sexed, and examined for external malformations. The live offspring were again counted and weighed on 4, 7, 14, and 21 days after birth. After weaning, all the offspring were counted and weighed weekly until 20 weeks of age. The following physical landmarks were noted: bilateral pinna unfolding at 4 days of age, abdominal hair emergence at 7 days, low incisor eruption and bilateral eye opening at 14 days, descent of both testes in each male at 28 days, and vaginal opening at 42 days in females. Behavioral Test Battery The behavioral tests and ages at testing are listed in Table 1. All the behavioral testing procedures were conducted blind with regard to the treatment groups, and tests were performed between 09:00 a.m. and 05:00 p.m. Cliff avoidance. A rat was placed on a table edge with the forepaws and nose over the edge. The amount of time required to complete backing and turning away from the edge was recorded. Each offspring was tested in one trial. (Altman and Sudarshan, 1975; Brunner et al., 1978; Kihara, 1991). The number of rats with successful responses within 30 s was recorded. Negative geotaxis. The time taken to complete a 180-degree turn when placed in a head-down position on a 25-degree inclined plywood surface was measured. Each animal was given one trial. (Kihara, 1991; Vorhees et al., 1979a,b). The number of rats with successful responses within 30 s was recorded. Swimming development. This procedure has been described elsewhere (Kihara, 1991; Schapiro et al., 1970; Vorhees et al.,1979a,b). Each rat was individually placed in a tank of water (280C) for 5–10 s and direction, angle in the water (head position), and limb usage was observed. Direction scores consisted of sinking (0 points), floating (1), circling (2), and swimming straight or nearly straight (3). Angle scores consisted of head submerged (0), nose at the surface (1), nose and top of head at or above the surface but ears still below the surface (2), ears half way above the surface (3), and ears completely above the surface (4). Limb usage scores consisted of no paddling (0), paddling with all four limbs (1), and paddling with hind limbs only with forelimbs stationary (2). Rotarod. The apparatus consisted of a rod 7 cm in diameter with a hard rubber surface, and linked to a variable-speed motor. The top of the rod was 34 cm above the base of the apparatus (Shinano Seisakusho Ltd., Tokyo, SN-498). Animals were placed individually on the rod for at least 5 s, and it was rotated at speeds of 5 or 15 rpm.. Rats were tested on two trials per day for 2 consecutive days. The maximum trial duration was 180 s, and the intertrial interval was about 30 min. The time each animal remained on the rod at each rotation speed was recorded (Kihara, 1991; Kihara et al., 1995). Open field. Rats were tested in a circular open field (100-cm diameter circular black polyvinyl chloride) on 3 consecutive days for 180 s per day. The time to leave the start area (latency), number of sections entered with all the four legs (ambulation), number of rearings, and number of fecal boluses were recorded (Kihara, 1991; Kihara, et al., 1995). Acquisition of conditioned avoidance. The apparatus was comprised of an operant chamber (O'hara and Co. Ltd., Tokyo, Model GT-7710), and a programming unit (Model GT-7715). As a visual and auditory warning device, a pilot lamp (24W) and a loud speaker (400 Hz) were provided over the lever. The chamber was placed inside a wooden sound-attenuating box (Model GT-7720). The conditioned avoidance schedule was as follows: a 20-s intertrial interval (ITI), a 5-s warning duration (conditioned stimuli; CS), and shock (approximately 85–90V, 0.5 mA, 60 Hz AC) for a maximum of 5 s. Stimuli were immediately terminated by the first lever-press elicited during the CS period, and foot shock was avoided. Both lever-pressing at times other than the CS presentation period and lever-holding were ineffective. Each session consisted of 1 h of training per day; animals were tested every other day for 15 sessions. The acquisition processes were considered to reflect discriminated avoidance learning. The index of conditioned avoidance learning was rates of avoidance. The number of avoidance responses, stimuli presented, and shocks delivered were recorded, and the mean percent of avoidance and the mean response rate per day were calculated (Kihara, 1991; Kihara et al., 1995). Underwater T-maze. The T-maze apparatus consisted of a grey polyvinyl chloride cylinder 20 cm in diameter; each arm was 50 cm long, and all the exits were curved upward. The maze was completely filled with water maintained at a temperature of 280C. Each animal was given 10 trials per day. A maximum swimming time of 60 s was allowed for each trial. An error was designated as entry into a blocked arm or re-entry into the stem. Swimming time and the number of errors in the maze were recorded for each trial (Kihara, 1991; Kihara et al., 1995). Reproduction test. One male and one female from each litter were selected at random for the reproduction test; sibling matings did not occur. They were mated monogamously for 2 weeks (about three estrus cycles) and were examined every morning for the presence of a vaginal plug. If a vaginal plug was observed (day 0 of gestation), the female was placed in an individual cage containing wooden chips for nesting. The dams were killed by ether overdose and necropsied on day 21 of gestation. These uteri were examined and the number of implantation sites, resorptions, and dead and live fetuses were recorded. The live fetuses were sexed, weighed, and examined for external malformations. Brain Weight One male offspring from each litter was killed at 20 weeks of age by ether overdose and autopsied. The brain was removed, weighed, and stored in 10% neutral formalin solution. Statistical Analysis The number of implantation sites and live fetuses and offspring as well as the body weight of dams and offspring were analyzed by one-way analysis of variance (ANOVA), followed by Dunnett's test if differences were found. The survival index data were analyzed by the chi-square test. The behavioral measures and physical developmental observations were evaluated by using the Mann-Whitney U-test for nonparametric comparison of group means (Siegel, 1956; Sokal and Rohlf, 1973). Before weaning, the data from individual subjects were averaged, and the litter was used as the unit of analysis. After weaning, the analysis was performed on the basis of individual animals from each litter. The accepted level of significance was p < 0.05. RESULTS Maternal Effects Neither death nor noticeable symptoms were observed in the dams of any group throughout gestation and lactation. AFB had no significant effect on either the length of gestation or maternal weight during the gestation and lactation periods (data not shown). Growth and Physical Landmarks of the Offspring At birth, the number of live offspring was significantly lower in the AFB-B group than those of controls, and body weights of male and female offspring in both the AFB-A and -B groups were significantly lower than the controls. However, there were no significant differences between the AFB groups and the control group with regard to the number of implants, sex ratio of offspring, offspring with external malformations, live birth index, survival rates at 4, 21 days of age (at weaning), or 20 weeks of age, or body weights at 4 and 7 days of age (data not shown), 21 days, and 20 weeks of age (Table 2). With regard to physical landmarks, there were no statistically significant effects of AFB-exposure on pinna unfolding, hair emergence, incisor eruption, testis descent, or vaginal opening (data not shown). However, the mean percentage of offspring with eyes opening at 14 days of age were 57.9, 17.9, and 30.0 for the control, AFB-A, and AFB-B groups, respectively. The AFB-A group showed significantly delayed eye opening compared to the controls. Behavioral Testing There were no sex differences on any of preweaning tests; therefore, males and females were combined for data analysis. The values presented the mean of the litter means in Table 3 and 4. Reflex behavior. The results of reflex testing are presented in Table 3. The AFB-A offspring were less successful in cliff avoidance than control offspring, and the AFB-A group also had significantly slower response times. Cliff avoidance was not affected in the AFB-B group. On the negative geotaxis reflex, there were no significant differences between the AFB-exposed groups and the control group. Swimming development. The results are shown in Table 4. With regard to swimming direction, the AFB-A group scored significantly lower than the control group at 6 days of age; however, there were no significant treatment effects at any other age observed. There were no significant differences between the AFB groups and the control group for swimming angle and limb usage on 6, 8, and 10 days of age. Rotarod performance. The rats in the AFB-A group remained on the rotarod for a significantly shorter time than the controls at 15 rpm on both the first and second test days. However, there were no statistically significant differences between the AFB-B and control groups (Table 5). Open field activity. There were no significant differences between the AFB-exposed groups and the control group for ambulation counts, rearing, and defecation frequencies, or the start latency time on any of the test days. (The mean control values [mean ± SE] were the ambulation 1st day 29.8 ± 7.2, 2nd 22.0 ± 4.7, 3rd 22.1 ± 5.7; the rearing 1st 9.4 ± 3.4, 2nd 3.6 ± 1.3, 3rd 2.5 ± 0.8; the defecation frequencies 1st 1.7 ± 0.5, 2nd 4.0 ± 0.8, 3rd 4.5 ± 0.6). Conditioned avoidance. Acquisition rates of the conditioned avoidance test are presented in Figure 2. The mean avoidance rates in both AFB-exposed groups were lower than those in the control group during 15 training sessions. Both the AFB-A group (5th–15th sessions) and the AFB-B group (12th, 14th, and 15th sessions) showed significantly lower avoidance rates than the control group. Furthermore, rates of AFB-A group during 5–11 sessions were also lower than avoidance rates of the AFB-B group. There appears to be some increase in variance that may be treatment related. No significant differences in the mean lever-pressing responses were detected among the three groups throughout the 15 sessions of training (data not shown). Underwater T-maze. There were no significant differences between the AFB-treated groups and the control group for swimming time and number of errors on any of the trial days (The mean control values of swimming time (s ± SE) were the 1st trial day 6.2 ± 0.5, 2nd 4.6 ± 0.3, 3rd 4.3 ± 0.3, 4th 8.1 ± 0.7, 5th 5.1 ± 0.6, and 6th 6.1 ± 0.5; the number of errors ranged between 0.18 and 0.8). Reproductive performance. No significant differences were seen between the AFB-exposed and control groups in any reproductive parameters including copulation and pregnancy rates, litter size, sex ratio of offspring, fetal weight, or incidence of external malformations (data not shown). Brain Weight There were no statistically significant differences in brain weight between the AFB-exposed and the control groups. The values (mean: g ± SE) were 1.88 ± 0.05, 1.83 ± 0.04, and 1.85 ± 0.04 for the control group, the AFB-A and the AFB-B groups, respectively. DISCUSSION The present study is the first postnatal physical and behavioral evaluation of offspring development using a test battery system following prenatal exposure to AFB during mid or late organogenesis. The main effects observed were a smaller number of live births, lower mean birth weights, a delayed physical development, delayed behavioral developments in the preweaning period, and a impaired locomotor coordination and deficits in avoidance performance in the postweaning period. On the other hand, no effects on maternal mortality, maternal weight gains, gestation length, the offspring with malformations, and the brain weight were observed in the prenatal exposure to AFB groups. AFB treatment during late organogenesis (days 15–18) resulted in decreased numbers of live pups; however, no significant reduction in the live birth index was seen because the mean number of implants in group AFB-B was smaller than that the control group (Table 2). AFB exposure during both mid-organogenesis (days 11–14) and late organogenesis (days 15–18) resulted in reduced birth weights in both male and female offspring, but body weight differences were completely recovered after 4 days of age. Thus, growth rates of offspring in the AFB groups were equivalent to the control group during the remainder of the study. There was also a delay in eye opening in the prenatally exposed animals, but it was significant only when AFB was administered in the mid-organogenesis period (days 11–14). The present study is the first to show that prenatal exposure to AFB affects behavioral performance in the preweaning offspring and alters the rate of acquisition of a conditioned avoidance task, motor coordination, and body balancing in the postweaning offspring. Treatment with AFB during mid-organogenesis (days 11–14) but not during late organogenesis (days 15–18) delayed the development of the cliff avoidance response. Swimming ontogeny is a measure of the development of neuromotor coordination and swimming ability (Kihara, 1991; Schapiro et al., 1970; Vorhees et al., 1979a,b). In the present study, prenatal AFB exposure during mid-organogenesis (days 11–14) also results in a delay in the direction of swimming but not during late organogenesis (days 15–18). On the other hand, the negative geotaxis test results in the present study did not distinguish exposed from control rats. These result suggest that AFB has a significant toxic action on developmental patterns of behavior in newborn rats, particularly on development of motor coordination. The rotarod test has been used during the postweaning period to determine forced coordinated motor and balancing abilities (Altman and Sudarshan, 1975; Kaplan and Murphy, 1972; Kinnard and Carr, 1957). The results of the present study show a deficit in motor skills in the offspring of mothers treated with AFB during mid-organogenesis (days 11–14) but not during late organogenesis (days 15–18). As rotarod performance has been related to cerebellar function (Pellegrino and Altman, 1979), prenatal AFB exposure may produce alterations in the morphology and physiology of this area. With regard to the avoidance learning task, it is of considerable interest to note that rat offspring exposed to AFB during both mid and late organogenesis showed impaired learning ability, and that the impaired avoidance learning in the prenatal AFB exposure during mid-organogenesis was more severe than that during late organogenesis. Therefore, mid-organogenesis (rat, days 11–14 of gestation) appears to be a more vulnerable period for AFB disruption of the acquisition of the conditioned avoidance learning than is late organogenesis (days 15–18). Generally, learning ability is a very important parameter for assessment of developmental neurotoxicity (Riley and Vorhees, 1986, Tanimura, 1992). In this respect, the most important finding in the present study is that prenatal exposure of rats to AFB impaired performance in the conditioned avoidance test. The open field test is a measure of emotional reactivity and exploratory behavior in the rat (Hall, 1934), and has commonly been used in developmental toxicology studies. It may also be viewed as a measure of activity level (Adams, 1986; Walsh and Cummins, 1976). In the present study, no effects of 0.3 mg/kg/day AFB were observed in the open field test in Wistar rats. However, Chentanez et al. (1986) found a decrease in some types of behavior (sniffing, grooming, exploring, rearing licking, and gnawing) and motor activity in the open field box in 1-month-old offspring of dams treated intraperitoneally with AFB, at 2.0 mg/kg/day, during both days 7–9 or days 14–16 of gestation in Fisher rats but not in 2- and 3-month-old offspring. Thus, the results of the present study differ from those of Chentanez et al. (1986) with regard to the effect of the AFB on the open field activity. This may be due to dose differences, rat strain differences, treatment route differences, or observer differences. The most important feature of the present study is that prenatal exposure of rats to AFB, at a dose lower than that which causes other gross malformations or growth retardation, altered neurobehavioral performance in offspring in the preweaning and postweaning periods. Therefore, the behavioral changes observed here were the result of persistent effects of AFB on the fetal nervous system. Furthermore, the behavioral teratogenic effect of AFB is greater when administered during mid-organogenesis than during late organogenesis. It was widely believed that late organogenesis or the early postnatal period might be the most sensitive stage for disrupting behavioral development, as brain maturation actively takes place within those periods (Hutchings, 1983; Leonard, 1982). More recently, several studies in addition to this one have demonstrated that mid-organogenesis in rats is a more sensitive period for some behavioral teratogens (Kihara, 1991; Rodier et al., 1979).Vorhees (1983, 1987) also suggested that mid-organogenesis is the most vulnerable period. Mid-organogenesis, approximately days 11–15 in the rat, is an extremely active phase of neurogenesis for the visual areas, cerebral cortices, basal ganglia and forebrain, and for thalamic, hypothalamic, and limbic regions (Vorhees, 1987). The period of administration for the control group was twice as long as that of the AFB treatment groups. This additional handling stress of the control dams could influence some behavioral parameters, especially basic values in the vehicle control. The mechanisms of AFB-induced behavioral teratogenesis are unknown at present. However, AFB produces central nervous system malformations (Geissler and Faustman, 1988; Tanaka, 1975), inhibition of DNA replication and binding to DNA (Benasutti et al 1988; Jacobson et al., 1987; Sporn et al., 1966), mutagenicity (Ong, 1975; Wong and Hsieh, 1976; Yourtee and Kirk-Yourtee, 1987), chromosome aberrations (Adgigitov et al.,1984), and transplacental carcinogenesis (Goerttler et al., 1980; Grice et al., 1973; Tanaka, 1975). It is surmised that some of the actions of AFB mentioned above are developmental toxic actions, including behavioral teratogenicity. However, further studies are required to characterize any behavioral effects of developmental AFB exposure in postweaning female rats, behavioral effects in progeny of dams treated during the lactation period, neurobiochemical effects of AFB, and the mechanisms of its effects on developmental neurotoxicology. TABLE 1 Schedule of Behavioral Test Battery Procedure  Age of testing   Note. Age of offspring for preweaning tests in days; age of offspring for postweaning tests in weeks.  a All offspring in each litter were tested.  b One male from each litter was randomly assigned to each of the four postweaning tests, except the reproductive test, for which one male and one female from each litter were used.  Preweaninga     Cliff avoidance  5   Negative geotaxis  7   Swimming development  6, 8, 10  Postweaningb     Rotarod  5   Open field  6   Conditioned avoidance learning  14   Underwater T-maze  15   Reproduction  16  Procedure  Age of testing   Note. Age of offspring for preweaning tests in days; age of offspring for postweaning tests in weeks.  a All offspring in each litter were tested.  b One male from each litter was randomly assigned to each of the four postweaning tests, except the reproductive test, for which one male and one female from each litter were used.  Preweaninga     Cliff avoidance  5   Negative geotaxis  7   Swimming development  6, 8, 10  Postweaningb     Rotarod  5   Open field  6   Conditioned avoidance learning  14   Underwater T-maze  15   Reproduction  16  View Large TABLE 2 Reproductive Performance of the Dams Exposed to Aflatoxin B1, and Survival and Body Weight in the Offspring     0.3 mg/kg   Group  Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   Note. M, male; F, female.  a Gestational days exposed.  b Mean ± SE.  c Percentage of implants.  d Percentage of offspring at birth.  e Percentage of offspring at 4 days of age.  f Percentage of offspring at weaning.  * Significantly different from the control (p < 0.05).  No. of dams  10  10  10   No. of implantsb  16.2±0.4  15.8±0.7  15.2±0.9   No. of live birthsb  15.2±0.5  14.4±0.4  12.9±0.9*   No. of offspring malformed at birth  0  0  0   Live birth indexc  93.9  93.8  85.6   Survival index at 4 days of aged  98.8  96.0  95.5   Weaning index at 21 days of agee  93.8  97.5  98.8   Survival index at 20 weeks of agef  93.4  90.7  97.3   Body weight of offspring (g)b         At birth          M  5.7±0.1  5.1±0.1*  4.7±0.3*     F  5.4±0.1  4.9±0.1*  4.7±0.2*    At weaning          M  49.1±1.1  46.1±1.6  46.7±2.8     F  46.6±1.4  42.7±2.0  46.0±2.3    At 20 weeks          M  371±4.9  362±6.4  355±9.3     F  234±4.3  223±4.3  220±7.0      0.3 mg/kg   Group  Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   Note. M, male; F, female.  a Gestational days exposed.  b Mean ± SE.  c Percentage of implants.  d Percentage of offspring at birth.  e Percentage of offspring at 4 days of age.  f Percentage of offspring at weaning.  * Significantly different from the control (p < 0.05).  No. of dams  10  10  10   No. of implantsb  16.2±0.4  15.8±0.7  15.2±0.9   No. of live birthsb  15.2±0.5  14.4±0.4  12.9±0.9*   No. of offspring malformed at birth  0  0  0   Live birth indexc  93.9  93.8  85.6   Survival index at 4 days of aged  98.8  96.0  95.5   Weaning index at 21 days of agee  93.8  97.5  98.8   Survival index at 20 weeks of agef  93.4  90.7  97.3   Body weight of offspring (g)b         At birth          M  5.7±0.1  5.1±0.1*  4.7±0.3*     F  5.4±0.1  4.9±0.1*  4.7±0.2*    At weaning          M  49.1±1.1  46.1±1.6  46.7±2.8     F  46.6±1.4  42.7±2.0  46.0±2.3    At 20 weeks          M  371±4.9  362±6.4  355±9.3     F  234±4.3  223±4.3  220±7.0  View Large TABLE 3 Reflex Development of Preweaning Rat Offspring of Dams Exposed to Aflatoxin B1     0.3 mg/kg   Group  Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   a Gestational days exposed.  b Percentage of offspring achieving criterion.  c Mean positive response time (s) ± SE.  * Significantly different from the control (p < 0.05).  No. of offspring tested  80  80  77   Cliff avoidance (5 days of age)         Success rateb  97.5  76.0*  91.2    Response timec  6.4±0.7  16.7±2.3*  7.6±1.9   Negative geotaxis (7 days of age)         Success rateb  95.8  98.7  97.5    Response timec  17.7±2.6  13.9±1.1  20.2±2.6      0.3 mg/kg   Group  Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   a Gestational days exposed.  b Percentage of offspring achieving criterion.  c Mean positive response time (s) ± SE.  * Significantly different from the control (p < 0.05).  No. of offspring tested  80  80  77   Cliff avoidance (5 days of age)         Success rateb  97.5  76.0*  91.2    Response timec  6.4±0.7  16.7±2.3*  7.6±1.9   Negative geotaxis (7 days of age)         Success rateb  95.8  98.7  97.5    Response timec  17.7±2.6  13.9±1.1  20.2±2.6  View Large TABLE 4 Swimming Development in Preweaning Rat Offspring of Dams Exposed to Aflatoxin B1     0.3 mg/kg   Group  Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   Note. Values are expressed using rating scales; mean ± SE.  a Gestational days exposed.  * Significantly different from the control (p < 0.05).  No. of offspring  80  80  77   6 days of age         Direction  2.3±0.1  1.8±0.1*  2.2±0.2    Angle  2.3±0.1  2.5±0.2  2.4±0.2    Limb usage  1.0±0  1.0±0  1.1±0.1   8 days of age         Direction  2.1±0.1  2.0±0.1  2.2±0.2    Angle  2.8±0.1  2.8±0.1  2.5±0.2    Limb usage  1.0±0  1.0±0  1.0±0   10 days of age         Direction  2.3±0.1  2.2±0.1  2.2±0.1    Angle  3.1±0.1  2.9±0.1  2.9±0.1    Limb usage  1.0±0  1.0±0  1.0±0      0.3 mg/kg   Group  Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   Note. Values are expressed using rating scales; mean ± SE.  a Gestational days exposed.  * Significantly different from the control (p < 0.05).  No. of offspring  80  80  77   6 days of age         Direction  2.3±0.1  1.8±0.1*  2.2±0.2    Angle  2.3±0.1  2.5±0.2  2.4±0.2    Limb usage  1.0±0  1.0±0  1.1±0.1   8 days of age         Direction  2.1±0.1  2.0±0.1  2.2±0.2    Angle  2.8±0.1  2.8±0.1  2.5±0.2    Limb usage  1.0±0  1.0±0  1.0±0   10 days of age         Direction  2.3±0.1  2.2±0.1  2.2±0.1    Angle  3.1±0.1  2.9±0.1  2.9±0.1    Limb usage  1.0±0  1.0±0  1.0±0  View Large TABLE 5 Rotarod Performance in Male Rat Offspring of Dams Exposed to Aflatoxin B1       0.3 mg/kg   Group    Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   Note. Values are means of time (s) spent on the rod ± SE. Maximum time was 180 s per trial. Ten male offspring were tested per group.  a Gestational days exposed.  b Revolutions per min.  * Significantly different from the control (p < 0.05).    rpmb        1st day  5  136.0±22.6  84.5±26.6  70.5±29.6     15  30.4±10.5  6.9±2.0*  46.7±25.3   2nd day  5  152.7±18.4  110.8±20.9  176.6±3.4     15  91.6±25.6  16.5±4.9*  87.8±31.7        0.3 mg/kg   Group    Control Days 11–18a  Days 11–14 (A)  Days 15–18 (B)   Note. Values are means of time (s) spent on the rod ± SE. Maximum time was 180 s per trial. Ten male offspring were tested per group.  a Gestational days exposed.  b Revolutions per min.  * Significantly different from the control (p < 0.05).    rpmb        1st day  5  136.0±22.6  84.5±26.6  70.5±29.6     15  30.4±10.5  6.9±2.0*  46.7±25.3   2nd day  5  152.7±18.4  110.8±20.9  176.6±3.4     15  91.6±25.6  16.5±4.9*  87.8±31.7  View Large FIG. 1. View largeDownload slide Structural formula of aflatoxin B1. FIG. 1. View largeDownload slide Structural formula of aflatoxin B1. FIG. 2. View largeDownload slide Acquisition of conditioned avoidance responses in male rat offspring of dams exposed to aflatoxin B1. Changes in the mean avoidance rates are shown. AFB-A: period of exposure of aflatoxin B1 (0.3 mg/kg/day) on days 11–14 of gestation; AFB-B: days 15–18; control (vehicle): days 11–18. Ten male offspring were tested per group. Vertical bars represent the mean standard error. *Significantly different from control (p < 0.05). FIG. 2. View largeDownload slide Acquisition of conditioned avoidance responses in male rat offspring of dams exposed to aflatoxin B1. Changes in the mean avoidance rates are shown. AFB-A: period of exposure of aflatoxin B1 (0.3 mg/kg/day) on days 11–14 of gestation; AFB-B: days 15–18; control (vehicle): days 11–18. Ten male offspring were tested per group. 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TI - Effects of Prenatal Aflatoxin B1 Exposure on Behaviors of Rat Offspring
JF - Toxicological Sciences
DO - 10.1093/toxsci/53.2.392
DA - 2000-02-01
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SP - 392
EP - 399
VL - 53
IS - 2
DP - DeepDyve
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