Are preconceptional stressful experiences crucial elements for the aetiology of autism spectrum disorder? Insights from an animal model.

Autism spectrum disorders (ASD) are a group of neurodevelopmental disorders characterized by changes in social interactions, impaired language and communication, fear responses and presence of repetitive behaviours. Although the genetic bases of ASD are well documented, the recent increase in clinical cases of idiopathic ASD indicates that several environmental risk factors could play a role in ASD aetiology. Among these, maternal exposure to psychosocial stressors during pregnancy has been hypothesized to affect the risk for ASD in offspring. Here, we tested the hypothesis that preconceptional stressful experiences might also represent crucial elements in the aetiology of ASD. We previously showed that social isolation stress during adolescence results in a marked decrease in the brain and plasma concentrations of progesterone and in the quality of maternal care that these female rats later provide to their young. Here we report that male offspring of socially isolated parents showed decreased agonistic behaviour and social transmission of flavour preference, impairment in reversal learning, increased seizure susceptibility, reduced plasma oxytocin levels, and increased plasma and brain levels of BDNF, all features resembling an ASD-like phenotype. These alterations came with no change in spatial learning, aggression, anxiety and testosterone plasma levels, and were sex-dependent. Altogether, the results suggest that preconceptional stressful experiences should be considered as crucial elements for the aetiology of ASD, and indicate that male offspring of socially isolated parents may be a useful animal model to further study the neurobiological bases of ASD, avoiding the adaptations that may occur in other genetic or pharmacologic experimental models of these disorders.

Juvenile social isolation affects maternal care in rats: involvement of allopregnanolone.

RATIONALE: Social isolation of rats immediately after weaning is thought to represent an animal model of anxiety-like disorders. Socially isolated virgin females showed a significant decrease in allopregnanolone levels, associated with increased anxiety-related behavior compared with group-housed rats. OBJECTIVES: The present study investigates whether post-weaning social isolation affects maternal behavior and assesses neuroactive steroid levels in adult female rats during pregnancy and postpartum. RESULTS: Socially isolated dams displayed a reduction in the frequency of arched back nursing (ABN) behavior compared to group-housed dams. In addition, both total and active nursing were lower in socially isolated dams compared to group-housed dams. Compared to virgin females, pregnancy increases allopregnanolone levels in group-housed as well as isolated dams and such increase was greater in the latter group. Compared to pregnancy levels, allopregnanolone levels decreased after delivery and this decrease was more pronounced in isolated than group-housed dams. Moreover, the fluctuations in plasma corticosterone levels that occur in late pregnancy and during lactation follow a different pattern in socially isolated vs. group-housed rats. CONCLUSIONS: The present results show that social isolation in female rats decreases maternal behavior; this effect is associated with lower allopregnanolone concentrations at postpartum, which may account, at least in part, for the poor maternal care observed in socially isolated dams. In support of this conclusion is the finding that finasteride-treated dams, which display a decrease in plasma allopregnanolone levels, also showed a marked reduction in maternal care, suggesting that allopregnanolone may contribute to the quality of maternal care.

Altered stress responsiveness and hypothalamic-pituitary-adrenal axis function in male rat offspring of socially isolated parents.

Social isolation in male rats at weaning results in reduced basal levels of the neuroactive steroid 3α,5α-tetrahydroprogesterone (3α,5α-TH PROG) in the brain and plasma as well as increased anxiety-like behavior. We now show that socially isolated female rats also manifest a reduced basal cerebrocortical concentration of 3α,5α-TH PROG as well as an anxiety-like profile in the elevated plus-maze and Vogel conflict tests compared with group-housed controls. In contrast, despite the fact that they were raised under normal conditions, adult male offspring of male and female rats subjected to social isolation before mating exhibited an increased basal cerebrocortical level of 3α,5α-TH PROG but no difference in emotional reactivity compared with the offspring of group-housed parents. These animals also showed an increased basal activity of the hypothalamic-pituitary-adrenal axis as well as reduced abundance of corticotropin-releasing factor in the hypothalamus and of corticotropin-releasing factor receptor type 1 in the pituitary. Moreover, negative feedback regulation of hypothalamic-pituitary-adrenal axis activity by glucocorticoid was enhanced in association with up-regulation of glucocorticoid receptor expression in the hippocampus. There was also attenuation of corticosterone release induced by foot-shock stress in the offspring of socially isolated parents. The increase in the brain concentration of 3α,5α-TH PROG induced by acute stress was also blunted in these animals. Our results thus show that a stressful experience before mating can influence neuroendocrine signaling in the next generation.

Chlorpyrifos residues levels in fruits and vegetables after field treatment.

Chlorpyrifos (O,O-diethyl O-3,5,6-trichloro-2-pyridyl phosphorothioate) was applied with three different formulations on oranges, peaches, tomatoes, wine and table grapes, and its behaviour was evaluated after field treatment. The formulations applied were emulsifiable concentrates (EC), microencapsulates (ME), and wettable granules (WG). The residues were similar in all crops studied in the EC and WG experiments, except peaches with WG treatment, the residue amount was lower than EC values. Tomatoes which were grown in greenhouse showed similar residues in all treatments just after treatment. Wine and table grapes showed different decline curves in the EC experiments ascribable to the different growing technology. Instrumental limit of determination (LOD) and limit of quantification (LOQ) for all matrices were 0.01, and 0.03 mg kg⁻¹, respectively. Repeated treatments showed that Chlorpyrifos can accumulate leading to residue levels at the preharvest interval (PHI) over the maximum residue level (MRL), especially on oranges and peaches. Among the formulates used ME showed the higher risk of residues over the MRL at harvest.

Efficacy of heat treatments with water and fludioxonil for postharvest control of blue and gray molds on inoculated pears and fludioxonil residues in fruit.

The residue levels of fludioxonil (FLU) were determined in pear cultivars Precoce di Fiorano, Coscia, and Spadona estiva after a 2-min dip in an aqueous mixture of FLU containing 300 or 100 mg/liter of active ingredient at 20 or 50 degrees C and after 12 days at 17 degrees C and 80% relative humidity (simulated shelf life conditions). The accumulation trend of FLU residues was determined in 'Precoce di Fiorano' pears after treatments with 25, 50, 100, or 200 mg/liter of active ingredient at 20 or 50 degrees C for 2 min or at 60 degrees C for 1 min. The efficacy of heat treatments with water and FLU was investigated on artificially inoculated 'Precoce di Fiorano', 'Coscia', and 'Spadona estiva' pears for the control of postharvest blue mold and gray mold caused by Penicillium expansum and Botrytis cinerea, respectively. Treatment with 300 mg/liter FLU at 20 degrees C resulted in residue levels similar to those from treatment with 100 mg/liter FLU at 50 degrees C in 'Coscia' fruit but in significantly lower residues in 'Precoce di Fiorano' and 'Spadona estiva' pears. Post-shelf life residues decreased in all cultivars, especially in 'Spadona estiva' pears treated with 300 mg/liter FLU at 20 degrees C. Residue levels of FLU in 'Precoce di Fiorano' pears treated at 20, 50, or 60 degrees C were correlated with fungicide dosage. When an equal rate was used, treatment at 50 degrees C resulted in a higher and a notably higher FLU deposition than that found under treatment at 60 and 20 degrees C, respectively. The in vitro tests showed that both pathogens were very sensitive to FLU, with MICs averaging 0.05 and 0.1 mg/liter for B. cinerea and P. expansum isolates, respectively. The 50% effective concentration ranged between 0.01 and 0.05 mg/liter for B. cinerea and between 0.05 and 0.1 mg/liter for P. expansum. In the in vivo trials, hot water treatment effectively reduced the incidence of both diseases during the first 4 to 8 days, depending on cultivar, dip temperature, and type of inoculum. However, as the incubation time proceeded, decay reduction was generally lower and the benefit of heat treatments was notably reduced or almost lost. In contrast, all treatments with FLU had a long-lasting effect. Treatments with heated FLU were more effective than those with unheated FLU; reduced concentrations of active ingredient were required to achieve a comparable control of blue and gray mold decay in these pears.

Residue levels and storage responses of nectarines, apricots, and peaches after dip treatments with fludioxonil fungicide mixtures.

Mature apricots (Prunus armeniaca), nectarines [Prunus persica var. nectarine (Ait.)], and peaches [P. persica (L.) Batsch.] were subjected to a 2 min dip treatment with warm water at 48 degrees C or with fludioxonil (FLU) at 100 mg L-1 and 20 degrees C or at 25 mg L-1 FLU and 48 degrees C and then stored at 5 degrees C and 90-95% relative humidity (RH) for 1 week plus 1 additional week at 18 degrees C and approximately 80% RH. Fruit residue uptake was determined as a function of fungicide concentration, dip temperature, treatment time (only on nectarines), and fruit storage conditions. FLU residue level was closely related to fungicide concentration and treatment temperatures and was dependent on fruit species. FLU residues showed great persistence over both storage and shelf life. Fruit dipping in water at 48 degrees C effectively reduced decay development in cvs. 'May Grand' nectarines and 'Pelese' apricots but was ineffective in cvs. 'Red Top' and 'Sun Crest' nectarines during 7 days of storage compared with nontreated fruit. Decay rates in cvs. 'Glo Haven' peaches and 'Fracasso' apricots were very low in fruit dipped in water at both 20 and 48 degrees C. Fungicide treatments at 20 and 48 degrees C resulted in the total or almost total suppression of decay in all cultivars. During shelf life, fruit became very prone to decay, averaging 25.7-100% depending on the cultivar. Fruit dipping in hot water effectively reduced decay in 'Pelese' and 'Fracasso' apricots, 'Sun Crest' peaches, and 'May Grand' nectarines as compared to control, but was ineffective in 'Glo Haven' and 'Red Top' peaches. Fungicide treatments at 20 degrees C were more effective than hot water in most cultivars. The combination of FLU with water at 48 degrees C further improved the fungicide performance. Indeed, reduced levels (a fourth) of active ingredient were required to achieve a control of decay comparable to that for treatment at 20 degrees C. Residue levels in fruit after treatment with 100 mg L-1 FLU at 20 degrees C or with 25 mg L-1 FLU at 48 degrees C averaged approximately 0.6-2 mg kg-1, which were notably lower than the maximum residue limit (5 mg kg-1) allowed in the United States for stone fruit.

Gas chromatographic ion trap mass spectrometry determination of zoxamide residues in grape, grape processing, and in the fermentation process.

A gas chromatographic ion-trap mass spectrometry (GC-ITMS) method was developed for the determination of the fungicide zoxamide in grape, must, wine, and spirits. Samples were extracted with hexane and analyzed without any clean up. The gas chromatograph was fitted with a carbofrit inserted into the glass liner to allow large volume injections. Analyses were carried out both in EI and CI mode. Recoveries from fortified samples ranged between 86 and 114% at four different fortification levels (n=6 each), ranging between 0.05 and 2.00 mg/kg. The relative standard deviation was below 19%. Both in EI and CI mode the calculated limit of detection (LOD) and quantification (LOQ) were 0.01 and 0.05 mg/kg (0.08 mg/kg in CI), respectively. Moreover the influence of yeasts and bacteria fermentation was evaluated.