Early-onset behavioral and neurochemical deficits in the genetic mouse model of phenylketonuria.

Phenylketonuria (PKU) is one of the most common human inborn errors of metabolism, caused by phenylalanine hydroxylase deficiency, leading to high phenylalanine and low tyrosine levels in blood and brain causing profound cognitive disability, if untreated. Since 1960, population is screened for hyperphenylalaninemia shortly after birth and submitted to early treatment in order to prevent the major manifestations of the disease. However, the dietetic regimen (phenylalanine free diet) is difficult to maintain, and despite the recommendation to a strict and lifelong compliance, up to 60% of adolescents partially or totally abandons the treatment. The development and the study of new treatments continue to be sought, taking advantage of preclinical models, the most used of which is the PAHenu2 (BTBR ENU2), the genetic murine model of PKU. To date, adult behavioral and neurochemical alterations have been mainly investigated in ENU2 mice, whereas there are no clear indications about the onset of these deficiencies. Here we investigated and report, for the first time, a comprehensive behavioral and neurochemical assay of the developing ENU2 mice. Overall, our findings demonstrate that ENU2 mice are significantly smaller than WT until pnd 24, present a significant delay in the acquisition of tested developmental reflexes, impaired communicative, motor and social skills, and have early reduced biogenic amine levels in several brain areas. Our results extend the understanding of behavioral and cerebral abnormalities in PKU mice, providing instruments to an early preclinical evaluation of the effects of new treatments.

Age-specific autistic-like behaviors in heterozygous Fmr1-KO female mice.

Fragile X syndrome (FXS) is a major developmental disorder and the most frequent monogenic cause of autism. Surprisingly, most existing studies on the Fmr1-KO mouse model for FXS have focused on males, although FX women, who are mostly heterozygous for the Fmr1 mutation, are known to exhibit several behavioral deficits, including autistic-like features. Furthermore, most animal research has been carried out on adults only; so that little is known about the age progression of the behavioral phenotype of Fmr1 mutants, which is a crucial issue to optimize the impact of therapeutic interventions. Here, we performed an extensive analysis of autistic-like social behaviors in heterozygous (HET) Fmr1-KO females and their WT littermates at different ages. No behavioral difference between HET and WT mice was observed at infancy, but some abnormalities in social interaction and communication were first detected at juvenile age. At adulthood some of these alterations disappeared, but avoidance of social novelty appeared, together with other FXS-relevant behavioral deficits, such as hyperactivity and reduced contextual fear response. Our data provide for the first time evidence for the presence of autistic-relevant behavioral abnormalities in Fmr1-HET female mice, demonstrating the utility of this mouse line to model autistic-like behaviors in both sexes. These results also highlight the importance of taking into account age differences when using the Fmr1-KO mouse model, suggesting that the early post-natal phases are the most promising target for preventive interventions and the adult age is the most appropriate to investigate the behavioral impact of potential therapies. Autism Res 2017. © 2017 International Society for Autism Research, Wiley Periodicals, Inc. Autism Res 2017, 10: 1067-1078. © 2017 International Society for Autism Research, Wiley Periodicals, Inc.

Social threat exposure in juvenile mice promotes cocaine-seeking by altering blood clotting and brain vasculature.

Childhood maltreatment is associated with increased severity of substance use disorder and frequent relapse to drug use following abstinence. However, the molecular and neurobiological substrates that are engaged during early traumatic events and mediate the greater risk of relapse are poorly understood and knowledge of risk factors is to date extremely limited. In this study, we modeled childhood maltreatment by exposing juvenile mice to a threatening social experience (social stressed, S-S). We showed that S-S experience influenced the propensity to reinstate cocaine-seeking after periods of withdrawal in adulthood. By exploring global gene expression in blood leukocytes we found that this behavioral phenotype was associated with greater blood coagulation. In parallel, impairments in brain microvasculature were observed in S-S mice. Furthermore, treatment with an anticoagulant agent during withdrawal abolished the susceptibility to reinstate cocaine-seeking in S-S mice. These findings provide novel insights into a possible molecular mechanism by which childhood maltreatment heightens the risk for relapse in cocaine-dependent individuals.

Early handling and repeated cross-fostering have opposite effect on mouse emotionality.

Early life events have a crucial role in programming the individual phenotype and exposure to traumatic experiences during infancy can increase later risk for a variety of neuropsychiatric conditions, including mood and anxiety disorders. Animal models of postnatal stress have been developed in rodents to explore molecular mechanisms responsible for the observed short and long lasting neurobiological effects of such manipulations. The main aim of this study was to compare the behavioral and hormonal phenotype of young and adult animals exposed to different postnatal treatments. Outbred mice were exposed to (i) the classical Handling protocol (H: 15 min-day of separation from the mother from day 1 to 14 of life) or to (ii) a Repeated Cross-Fostering protocol (RCF: adoption of litters from day 1 to 4 of life by different dams). Handled mice received more maternal care in infancy and showed the already described reduced emotionality at adulthood. Repeated cross fostered animals did not differ for maternal care received, but showed enhanced sensitivity to separation from the mother in infancy and altered respiratory response to 6% CO2 in breathing air in comparison with controls. Abnormal respiratory responses to hypercapnia are commonly found among humans with panic disorders (PD), and point to RCF-induced instability of the early environment as a valid developmental model for PD. The comparisons between short- and long-term effects of postnatal handling vs. RCF indicate that different types of early adversities are associated with different behavioral profiles, and evoke psychopathologies that can be distinguished according to the neurobiological systems disrupted by early-life manipulations.

Early social enrichment rescues adult behavioral and brain abnormalities in a mouse model of fragile X syndrome.

Converging lines of evidence support the use of environmental stimulation to ameliorate the symptoms of a variety of neurodevelopmental disorders. Applying these interventions at very early ages is critical to achieve a marked reduction of the pathological phenotypes. Here we evaluated the impact of early social enrichment in Fmr1-KO mice, a genetic mouse model of fragile X syndrome (FXS), a major developmental disorder and the most frequent monogenic cause of autism. Enrichment was achieved by providing male KO pups and their WT littermates with enhanced social stimulation, housing them from birth until weaning with the mother and an additional nonlactating female. At adulthood they were tested for locomotor, social, and cognitive abilities; furthermore, dendritic alterations were assessed in the hippocampus and amygdala, two brain regions known to be involved in the control of the examined behaviors and affected by spine pathology in Fmr1-KOs. Enrichment rescued the behavioral FXS-like deficits displayed in adulthood by Fmr1-KO mice, that is, hyperactivity, reduced social interactions, and cognitive deficits. Early social enrichment also eliminated the abnormalities shown by adult KO mice in the morphology of hippocampal and amygdala dendritic spines, namely an enhanced density of immature vs mature types. Importantly, enrichment did not induce neurobehavioral changes in WT mice, thus supporting specific effects on FXS-like pathology. These findings show that early environmental stimulation has profound and long-term beneficial effects on the pathological FXS phenotype, thereby encouraging the use of nonpharmacological interventions for the treatment of this and perhaps other neurodevelopmental diseases.

Neuroprotective effects of donepezil against cholinergic depletion.

INTRODUCTION: Intraparenchymal injections of the immunotoxin 192-IgG-saporin into medial septum and nucleus basalis magnocellularis causes a selective depletion of basal forebrain cholinergic neurons. Thus, it represents a valid model to mimic a key component of the cognitive deficits associated with aging and dementia. Here we administered donepezil, a potent acetylcholinesterase inhibitor developed for treating Alzheimer's disease, 15 days before 192-IgG-saporin injection, and thus we examined donepezil effects on neurodegeneration and cognitive deficits. METHODS: Caspase-3 activity and cognitive performances of lesioned rats pre-treated with donepezil or saline were analyzed and compared to the outcomes obtained in pre-treated sham-lesioned rats. RESULTS: Cholinergic depletion increased hippocampal and neocortical caspase-3 activity and impaired working memory, spatial discrimination, social novelty preference, and ultrasonic vocalizations, without affecting anxiety levels and fear conditioning. In lesioned animals, donepezil pre-treatment reduced hippocampal and neocortical caspase-3 activity and improved working memory and spatial discrimination performances and partially rescued ultrasonic vocalizations, without preventing social novelty alterations. CONCLUSIONS: Present data indicate that donepezil pre-treatment exerts beneficial effects on behavioral deficits induced by cholinergic depletion, attenuating the concomitant hippocampal and neocortical neurodegeneration.

Unstable maternal environment, separation anxiety, and heightened CO2 sensitivity induced by gene-by-environment interplay.

BACKGROUND: In man, many different events implying childhood separation from caregivers/unstable parental environment are associated with heightened risk for panic disorder in adulthood. Twin data show that the occurrence of such events in childhood contributes to explaining the covariation between separation anxiety disorder, panic, and the related psychobiological trait of CO(2) hypersensitivity. We hypothesized that early interference with infant-mother interaction could moderate the interspecific trait of response to CO(2) through genetic control of sensitivity to the environment. METHODOLOGY: Having spent the first 24 hours after birth with their biological mother, outbred NMRI mice were cross-fostered to adoptive mothers for the following 4 post-natal days. They were successively compared to normally-reared individuals for: number of ultrasonic vocalizations during isolation, respiratory physiology responses to normal air (20%O(2)), CO(2)-enriched air (6% CO(2)), hypoxic air (10%O(2)), and avoidance of CO(2)-enriched environments. RESULTS: Cross-fostered pups showed significantly more ultrasonic vocalizations, more pronounced hyperventilatory responses (larger tidal volume and minute volume increments) to CO(2)-enriched air and heightened aversion towards CO(2)-enriched environments, than normally-reared individuals. Enhanced tidal volume increment response to 6%CO(2) was present at 16-20, and 75-90 postnatal days, implying the trait's stability. Quantitative genetic analyses of unrelated individuals, sibs and half-sibs, showed that the genetic variance for tidal volume increment during 6%CO(2) breathing was significantly higher (Bartlett χ = 8.3, p = 0.004) among the cross-fostered than the normally-reared individuals, yielding heritability of 0.37 and 0.21 respectively. These results support a stress-diathesis model whereby the genetic influences underlying the response to 6%CO(2) increase their contribution in the presence of an environmental adversity. Maternal grooming/licking behaviour, and corticosterone basal levels were similar among cross-fostered and normally-reared individuals. CONCLUSIONS: A mechanism of gene-by-environment interplay connects this form of early perturbation of infant-mother interaction, heightened CO(2) sensitivity and anxiety. Some non-inferential physiological measurements can enhance animal models of human neurodevelopmental anxiety disorders.