Mood-related behavioral and neurochemical alterations in mice exposed to low chlorpyrifos levels during the brain growth spurt.

Organophosphates are among the most used pesticides. Particularly, chlorpyrifos (CPF) is responsible for a number of deleterious effects on brain development, which may program behavioral changes later in life. Here, we investigated whether a regimen of early low level CPF exposure that did not result in a significant inhibition of acetylcholinesterase (AChE) had deleterious effects on mood-related behaviors, as well as on cholinergic and serotonergic biomarkers in the mice brain. From the 3rd to 9th postnatal day (PN), male and female Swiss mice were subcutaneously injected with CPF. Mice were submitted to a battery of behavioral tests from PN60 to PN63: open field, elevated plus maze and forced swimming tests. The cholinergic and serotonergic biomarkers were assessed at PN10 and PN63. Our data indicated that early CPF exposure increased anxiety-like behavior in females and altered decision-making behavior in both sexes. Most biochemical alterations were sex-dependent and restricted to females. At PN10, CPF female mice showed increased serotonin and choline transporter binding in cerebral cortex. Distinctively, in adult females, the effects indicated a hypoactive state: CPF exposure reduced 5-HT1a receptor binding in cerebral cortex, as well as serotonin transporter binding and choline acetyltransferase activity in brainstem. Our results indicate that CPF exposure during the brain growth spurt deregulates serotonergic and cholinergic biomarkers. The effects are consistent with impaired synaptic function, may be related to long-term mood disorders and point out to higher female susceptibility.

Ethanol exposure during the brain growth spurt affects social behavior and increases susceptibility to acute ethanol effects during adolescence in male mice.

The brain is particularly vulnerable to ethanol effects during its growth spurt. Outcomes of early ethanol exposure such as hyperactivity have been extensively investigated; however, persons with fetal alcohol spectrum disorder frequently have social impairments and are heavy drinkers. Despite that, scant information is available regarding the neurobiological basis of these latter behavioral issues. Here, Swiss mice exposed to ethanol (Etoh, 5 g/kg i.p., alternate days) or saline during the brain growth spurt [postnatal day (PN) 2 to 8] were used to assess social behavior after an ethanol challenging during adolescence. At PN39, animals were administered with a single ethanol dose (1 g/Kg) or water by gavage and were then evaluated in the three-chamber sociability test. We also evaluated corticosterone serum levels and the frontal cerebral cortex serotoninergic system. Etoh males showed reductions in sociability. Ethanol challenging reverted these alterations in social behavior, reduced corticosterone levels, and increased the 5-HT2 receptor binding of male Etoh mice. No alterations were observed in 5-HT and 5-HIAA contents. These data support the idea that ethanol exposure during the brain growth spurt impacts social abilities during adolescence, alters ethanol reexposure effects, and suggests that stress response and serotoninergic system play roles in this phenomenon.

Maternal undernutrition during lactation alters nicotine reward and DOPAC/dopamine ratio in cerebral cortex in adolescent mice, but does not affect nicotine-induced nAChRs upregulation.

Early undernutrition causes long lasting alterations that affect the response to psychoactive drugs. Particularly, undernutrition during lactation affects the acute locomotor response to nicotine during adolescence, but the reward effect of continued exposure to nicotine remains unknown. The goal of this study was to investigate the effects of undernutrition during lactation on the nicotine susceptibility indexed via conditioned place preference (CPP), on dopamine content and turnover and on nicotine-induced nicotinic cholinergic receptor (nAChR) upregulation in the cerebral cortex, midbrain and hippocampus of adolescent mice. The impact of undernutrition and nicotine exposure on stress-related hormones and leptin was also investigated. From postnatal day 2 (PN2) to weaning (PN21), dams were randomly assigned to one of the following groups: Control (C) - free access to standard laboratory diet (23% protein); Protein Restricted (PR) - free access to isoenergenetic diet (8% protein); Calorie Restricted (CR) - access to standard laboratory diet in restricted quantities (mean ingestion of PR). PR and CR groups showed less mass gain and less visceral fat mass. While C and CR were equally susceptible to nicotine-induced place preference conditioning, PR failed to show a conditioning pattern. In contrast, all groups presented a nicotine-evoked nAChR upregulation in the cerebral cortex. While dopamine and DOPAC levels did not differ between groups, the DOPAC/dopamine ratio was increased in CR animals. No differences in endocrine parameters were observed. Taken together, our results indicate that undernutrition during lactation programs for brain alterations later in life. Our data also suggest that early undernutrition does not affect the rewarding associative properties of nicotine at adolescence.

Locomotor response to acute nicotine in adolescent mice is altered by maternal undernutrition during lactation.

Undernutrition during brain development causes long lasting alterations in different neurotransmitter systems that may alter responses to psychoactive drugs. Despite the recognized effects of early undernutrition on the cholinergic system, no evidence that demonstrates the influence of this insult on nicotine susceptibility has been reported. We investigated the effects of protein/calorie restriction during lactation on the susceptibility to nicotine in adolescent mice. Dams were randomly assigned to one of the following groups: Control (C, 20 litters)--free access to standard laboratory diet (23% protein); Protein Restricted (PR, 12 litters)--free access to a isoenergetic, 8% protein diet; Calorie Restricted (CR, 12 litters)--access to standard laboratory diet in restricted quantities (mean ingestion of PR: pair-fed group). Undernutrition extended from postnatal day 2 (PN2) to weaning (PN21). At PN30, animals either received an i.p. injection of nicotine (0.5mg/Kg) or saline and were immediately placed in open field (OF). After the OF, adrenal glands and serum were collected for the analyses of stress-related endocrine parameters and leptin concentration. PR and CR offspring showed less body mass gain and visceral fat mass. PR offspring presented reduced serum leptin concentration. In the OF, nicotine increased locomotor activity of C and PR, but not of CR. CR and PR offspring showed decreased adrenal catecholamine content, which was not dependent on nicotine exposure. Our results indicate that early undernutrition interferes with nicotine-elicited locomotor effects in adolescent mice and suggest that endocrine parameters alterations in malnourished animals do not influence the behavioral response to nicotine.

Methamidophos exposure during the early postnatal period of mice: immediate and late-emergent effects on the cholinergic and serotonergic systems and behavior.

Organophosphates (OPs) are among the most used pesticides. Although some OPs have had their use progressively more restricted, other OPs are being used without sufficient investigation of their effects. Here, we investigated the immediate neurochemical and delayed neurochemical and behavioral actions of the OP methamidophos to verify whether there are concerns regarding exposure during early postnatal development. From the third to the nineth postnatal day (PN), Swiss mice were sc injected with methamidophos (1mg/kg). At PN10, we assessed cholinergic and serotonergic biomarkers in the cerebral cortex and brainstem. From PN60 to PN63, mice were submitted to a battery of behavioral tests and subsequently to biochemical analyses. At PN10, the effects were restricted to females and to the cholinergic system: Methamidophos promoted increased choline transporter binding in the brainstem. At PN63, in the brainstem, there was a decrease in choline transporter, a female-only decrease in 5HT1A and a male-only increase in 5HT2 receptor binding. In the cortex, choline acetyltransferase activity was decreased and 5HT2 receptor binding was increased both in males and females. Methamidophos elicited behavioral alterations, suggestive of increased depressive-like behavior and impaired decision making. There were no significant alterations on anxiety-related measures and on memory/learning. Methamidophos elicited cholinergic and serotonergic alterations that depended on brain region, sex, and age of the animals. These outcomes, together with the behavioral effects, indicate that this OP is deleterious to the developing brain and that alterations are indeed identified long after the end of exposure.

Exposure to methamidophos at adulthood adversely affects serotonergic biomarkers in the mouse brain.

Epidemiologic studies describe a potential risk of depression and suicide in farm workers exposed to organophosphates (OPs). In a previous study we observed an increase in depressive-like behavior in adult mice exposed to the OP pesticide methamidophos. Considering the association between depression and the serotonergic (5HT) system, in the present study we investigated whether a subchronic exposure to methamidophos affects the serotonergic system of adult mice. From postnatal day 60 to 89 (PN60 to PN89), one of two concentrations of methamidophos (higher dose: 5.25 µg/ml; lower dose: 1.31 µg/ml) or vehicle was administered in the drinking water of male Swiss mice. We evaluated three serotonergic biomarkers during (PN89) and after (PN100) the exposure period: 5HT(1A) receptor binding with [(3)H]OH-DPAT, 5HT(2) receptor binding with [(3)H]ketanserin and 5HT transporter binding with [(3)H]paroxetine. Methamidophos elicited robust decreases in binding for all 5HT markers. These decreases were evident in brain regions containing 5HT cell bodies and dendritic arbors (midbrain, brainstem) as well as in the cerebral cortex, which contains 5HT projections. In the cerebral cortex, effects were identified in mice exposed to the higher dose of methamidophos while in the midbrain and brainstem, both doses elicited significant effects. Overall, effects were present both during and after exposure, even though there were some regional disparities regarding the persistence of effects. Our results indicate that exposure to methamidophos affects synaptic transmission promoting decreases of specific serotonergic biomarkers. These data suggest a mechanism of action of this pesticide that might explain the increased depressive-like behavior in adult mice.

Exposure to nicotine and ethanol in adolescent mice: effects on depressive-like behavior during exposure and withdrawal.

Depression and use of addictive substances are two of the most frequent public health problems of adolescents. However, little is known about the association between depression and drug use. Considering that ethanol and nicotine are the most widely used and abused drugs by adolescents, here, we evaluated the depressive-like behavior of C57BL/6 male and female mice exposed to nicotine (NIC) and/or ethanol (ETOH) from the 30th to the 45th (PN30-45) postnatal day. Four groups were analyzed: 1) concomitant NIC (50µg/ml in 2% saccharin to drink) and ETOH (25%, 2g/kg i.p. injected every other day) exposure; 2) NIC exposure; 3) ETOH exposure; 4) vehicle. Immobile behavior, an animal model of depressive behavior, was assessed in the forced swimming test (FST) while the anhedonic state was assessed in the sucrose preference test (SPT) by the end of exposure (PN45-47) as well as during short- (PN50-52) and long-term (PN75-77) withdrawal. In the FST, ETOH female mice showed a reduction in immobility time by the end of exposure while, during long-term withdrawal, immobility time was increased. Short-term withdrawal elicited an increase in immobility time only in female NIC mice. In the SPT, males from both NIC and NIC+ETOH groups showed increased sucrose consumption, suggesting a reward-craving effect during short-term withdrawal. During long-term withdrawal, NIC male mice showed an anhedonic effect. Adolescent nicotine, ethanol and nicotine+ethanol combined exposures during adolescence thus elicit gender-selective effects both during exposure and withdrawal that may contribute to the increased prevalence of depression among drug users.

Neonatal nicotine exposure causes insulin and leptin resistance and inhibits hypothalamic leptin signaling in adult rat offspring.

Maternal nicotine (NIC) exposure during lactation leads to overweight, hyperleptinemia, and hypothyroidism in adult rat offspring. In this model, we analyzed adipocyte morphology, glucose homeostasis (serum insulin and adiponectin; liver and muscle glycogen), serum lipid, and the leptin signaling pathway. After birth, osmotic minipumps were implanted in lactating rats, which were divided into the groups NIC (6 mg/kg per day s.c. for 14 days) and control (C, saline). NIC and C offspring were killed at the age of 180 days. Adult NIC rats showed higher total body fat (+10%, P<0.05), visceral fat mass (+12%, P<0.05), and cross-sectional area of adipocytes (epididymal: +12% and inguinal: +43%, P<0.05). Serum lipid profile showed no alteration except for apolipoprotein AI, which was lower. We detected a lower adiponectin:fat mass ratio (-24%, P<0.05) and higher insulinemia (+56%, P<0.05), insulin resistance index (+43%, P<0.05), leptinemia (+113%, P<0.05), and leptin:adiponectin ratio (+98%, P<0.05) in the adult NIC group. These rats presented lower hypothalamic contents of the proteins of the leptin signaling pathway (leptin receptor (OB-R): -61%, janus tyrosine kinase 2: -41%, and p-signal transducer and activator of transcription 3: -56%, P<0.05), but higher suppressor of cytokine signaling 3 (+81%, P<0.05). Therefore, NIC exposure only during lactation programs rats for adipocyte hypertrophy in adult life, as well as for leptin and insulin resistance. Through the effects of NIC, perinatal maternal cigarette smoking may be responsible for the future development of some components of the metabolic syndrome in the offspring.