Long-lasting developmental effects in rat offspring after maternal exposure to acetamiprid in the drinking water during gestation.

Neonicotinoids (NNTs) are a class of insecticides proposed to be safe for pest control in urban, suburban, and agricultural applications. However, little is known about their developmental effects after repeated low-dose exposures during gestation. Here, we tested a dose considered subthreshold for maternal toxicity in rats (6 mg/kg/day) by assessing several morphological, biochemical, and neurobehavioral features in preterm fetuses and developing pups after maternal administration of the NTT acetamiprid (ACP) dissolved in the drinking water during gestational days (GD) 2-19. The exploratory evaluation included monitoring maternal body weight gain, fetal viability, body weight and sex ratio, cephalic length, neonatal body weight and sex ratio, metabolic enzymes in the placenta, maternal blood and fetal liver, and anogenital distance and surface righting response during infancy. We also used the circling training test to study the integrity of the associative-spatial-motor response in adolescence. Results showed no consistent findings indicating maternal, reproductive or developmental toxicity. However, we found ACP effects on maternal body weight gain, placental butyrylcholinesterase activity, and neurobehavioral responses, suggestive of a mild toxic action. Thus, our study showed a trend for developmental susceptibility at a dose so far considered subtoxic. Although the ACP concentration in environmental samples of surface water and groundwater has been mostly reported to be much lower than that used in our study, our results suggest that the ACP point of departure used in current guidelines aimed to prevent developmental effects may need to be verified by complementary sensitive multiple-endpoint testing in the offspring.

The gut microbiota as a biomarker for realistic exposures to pesticides: A critical consideration.

In mammals, the gut microbiota (GM) exerts important physiological functions. The abundance of bacterial, fungal and protozoan microorganisms colonizing the gut determines the nutrients that will be taken from the diet, the production and absorption rates of vitamins, and the ability of various metabolic pathways to activate or detoxify xenobiotics. Several studies have allowed characterizing the plastic changes occurring in the GM from birth to adulthood, the extent of intra- and inter-individual variability, and the influence of the housing/caging conditions and diet on the microbiome composition of the gut. The structural and functional integrity of the GM has been increasingly proposed as a biological determinant of the susceptibility to developmental disorders. More recently, some research articles have shown some potentiality of this microbial assembly as a possible biomarker for environmentally relevant exposure to chemical hazards. In this review, we performed a comprehensive survey on GM studies in rats and mice exposed to insecticides, herbicides and fungicides, exhaustively considering the influence of several biological and experimental factors, including the animal's age, diet, breeding type, housing and caging conditions and sampling protocol, on the results observed. For pesticides for which no reliable biomarker has yet been developed in mammals, the utility and predictive power of microbiome assays will strongly depend on the rigorous selection and application of the study design features.

Relationship Between the Dose Administered, Target Tissue Dose, and Toxicity Level After Acute Oral Exposure to Bifenthrin and Tefluthrin in Young Adult Rats.

Most pyrethroid insecticides (PYRs) share a similar primary target site in mammals. However, the potency estimates of the lethal and sublethal effects of these compounds differ up to 103-fold. The aim of this study was to evaluate the relationship between the dose administered, the target tissue dose, and the effect of 2 highly toxic PYRs, tefluthrin (TEF; 0.1-9 mg/kg) and bifenthrin (BIF; 0.5-12 mg/kg), by using the oral route, a corn oil vehicle (1 ml/kg) and subcutaneous temperature (Tsc) monitoring assays in adult rats. The Tsc was determined at 30-min intervals for 5 h (TEF) or 4.5 h (BIF) after dosing. Rats were sacrificed at 6 h after dosing, and BIF and TEF concentrations were determined in blood (Bd), liver (Lv), and cerebellum (Cb) by using a GC-ECD system. The minimal effective dose of BIF (3 mg/kg) affecting Tsc was similar to that found in prior studies using other testing paradigms. Regarding TEF, a very steep relationship between the dose administered and toxicity was observed, with a near-threshold to low-effective range for Tsc at 0.1-6 mg/kg, and a near lethal syndrome at ≥ 7.5 mg/kg. At 6-7.5 mg/kg TEF, the Cb/Bd and Cb/Lv concentration ratios were both > 1. Conversely, for BIF, the Cb concentration was barely over the Bd concentration and the Cb/Lv concentration ratio remained < 1. Our results and previous findings call for more comprehensive consideration to establish the relevance of the distribution into target tissues and the tissue dosimetry for health risks through the exposure to PYRs in humans.

Relationship between exposure, body burden and target tissue concentration after oral administration of a low-dose mixture of pyrethroid insecticides in young adult rats.

Pyrethroids (PYRs) are synthetic insecticides increasingly used in agricultural and household pest control. Little is known on how the toxicity of highly effective bolus doses of single compounds compares to more realistic scenarios of low-level exposure to PYR mixtures. In this study, we examined a quaternary mixture of two noncyano (tefluthrin, TEF; bifenthrin, BIF) and two cyano (α-cypermethrin, α-CPM; deltamethrin, DTM) PYRs in young adult rats. These compounds are mostly composed of PYR isomers ranking top ten in acute lethality in rats. Concurrently, we administered near-threshold levels of the four PYRs dissolved in corn oil by oral route. Six hours later blood was collected and the liver and cerebellum were dissected out to determine PYR concentrations in these tissues using Gas Chromatography with Electron Capture Detector (GC-ECD). The mixture caused mild-to-moderate changes in non-locomotor behaviors and subcutaneous body temperature (up to +1.2-1.5 °C increase at 2-4 h after dosing, respectively, compared to pre-dosing records). The most toxic PYRs BIF and TEF reached higher concentrations in the cerebellum than the cyano-compounds α-CPM and DTM. In addition, PYR concentrations in the cerebellum were correlated to single compound proportions in the dosing solution and changes in body temperature. Our results suggest that aggregate exposures resulting in a target tissue burden of ∼10-1 nmoles PYR/g may be toxicologically relevant, expanding the evidence on exposure-dose-effect relationships for PYRs, and serving to design convenient pharmacokinetic models for environmentally relevant exposures to PYR mixtures.

Evidence for effects on thermoregulation after acute oral exposure to type I and type II pyrethroids in infant rats.

Most pyrethroid (PYR) insecticides may be classified either as type-I compounds, which produce whole body tremors and hyperthermia, or type-II compounds, which produce salivation, choreoathetosis, and hypothermia (i.e., producing T and CS neurobehavioral syndromes, respectively). This classification is based on clinical observations in adult rats and mice after intracerebroventricular or intravascular administration of highly effective acute (bolus) doses. PYR neurotoxicity in infant animals is not characterized as much as in adult animals. Endpoints informing on vital determinants of mammal's maturation, such as body temperature may help recognizing age-related differences in susceptibility to PYRs. In this work, body temperature (Tb) was monitored at 30-min intervals after acute oral exposure to T-syndrome PYR bifenthrin (BIF), CS-syndrome PYR cypermethrin (CYPM), and a BIF­CYPM mixture in weanling rats by using a subcutaneous temperature monitoring system. In both single-compound assays, a time- and dose-related decline of Tb was the most evident impact on thermoregulation observed starting at ~2­3 h after dosing.Moreover, 15­18 mg/kg BIF induced a mild increase in Tb before the hypothermic action was apparent. The lowest effective dose for temperature perturbation was 15mg/kg for BIF and 10mg/kg for CYPM, and moderate neurobehavioral alterations were evident at 12 and 10mg/kg, respectively. When low effective doses of BIF and CYPM were co-administered mild behavioral effects and a transient increase in Tb (p=0.02) were observed at 1­2 h, and no Tb decline was apparent afterwards compared to control animals. Noteworthy, the hypothermic action of BIF in infant rats was quite different from the hyperthermia consistently reported in studies using mature animals. Our results suggest that body temperature monitoring may be useful as a complementary assessment to reveal qualitative age-specific pesticide effects in rats.

Permanent motor activity and learning disorders induced by exposure to phenytoin during gestation and early infancy in the rat.

Experimental models and clinical data indicate that the incidence of motor and learning disorders may be increased in children of epileptic mothers taking phenytoin (PHT) during pregnancy. There is little data on the vulnerability of infants to PHT-induced long-term behavioral toxicity after gestational or early life exposure (i.e., infantile convulsion therapy). We examined the persistence of alterations in circling behavior induced by exposure to PHT during gestation, infancy, or both. Pregnant Sprague-Dawley rats were injected i.p. with saline (SAL) or PHT (30 mg/kg/day) during gestational days (GD) 10-18. The offspring were then administered (i.p.) SAL or PHT (60 mg/kg/day) during postnatal days (PD) 13-23. Afterward, Circling Training tests were performed at three time points. At PD40 and PD80, the clockwise direction of circling was reinforced. At PD150, counterclockwise circling was rewarded instead. At PD40, all PHT-treated groups demonstrated increased circling velocities compared to saline-treated controls. Higher spatial error rates for direction of circling were also observed in gestation-only and infancy-only exposures. At PD80, groups exposed during gestation had higher circling velocities than control or infancy-only exposed groups. At PD150, increases in circling velocity were apparent for the reverse learning task in groups exposed during gestation. These results indicate that early postnatal exposure to PHT may exacerbate the known long-term behavioral effects of gestational exposure.

Postnatal haloperidol eliminates the deficit in circling behavior produced by prenatal exposure to the same drug.

Up to 35% of pregnant women take psychotropic drugs at least once during gestation [Austin and Mitchell, 1998]. From concurrent animal and human evidence, it has been proposed that exposure to several psychoactive medications in utero or during lactation increases the risk for permanent brain disorders. Present preventive or therapy practices applied on humans for this type of long-lasting behavioral alterations are mainly based on empirical results. Here, we test an experimental approach designed to counteract a circling performance deficit that appears in Sprague-Dawley rats at puberty on exposure to the dopaminergic blocker haloperidol (HAL) during gestation [J.L. Brusés, J.M. Azcurra, The circling training: A behavioral paradigm for functional teratology testing, in: P.M. Conn (Ed.), Paradigms for the study of behavior, Acad. Press, New York, 1993, pp. 166-179. Method Neurosci. 14]. Gestational exposure to HAL (GD 5-18, 2.5 mg/kg/day ip) induced the expected circling activity decrease in the offspring at the fifth week of life. When prenatal exposure to HAL was continued through lactation (PD5-21, 1.5 mg/kg/day ip), rats otherwise showed a control-like circling performance. No difference was yet found between lactation-only, HAL-exposed pups and saline (SAL)-treated controls (n=8 each group). We further performed saturating (3H)-spiroperidol (SPI) binding assays on striatal P2 membrane fractions 2 months later. The dopamine-type D2-specific binding results suggested that above circling behavior findings could be partially explained by enduring HAL-induced neurochemical changes. The role of critical periods of sensitivity as transient windows for opportunistic therapies for behavioral teratology is discussed.