Effects of single-dose antipurinergic therapy on behavioral and molecular alterations in the valproic acid-induced animal model of autism.

Autism spectrum disorder (ASD) is characterized by deficits in communication and social interaction, restricted interests, and stereotyped behavior. Environmental factors, such as prenatal exposure to valproic acid (VPA), may contribute to the increased risk of ASD. Since disturbed functioning of the purinergic signaling system has been associated with the onset of ASD and used as a potential therapeutic target for ASD in both clinical and preclinical studies, we analyzed the effects of suramin, a non-selective purinergic antagonist, on behavioral, molecular and immunological in an animal model of autism induced by prenatal exposure to VPA. Treatment with suramin (20 mg/kg, intraperitoneal) restored sociability in the three-chamber apparatus and decreased anxiety measured by elevated plus maze apparatus, but had no impact on decreased reciprocal social interactions or higher nociceptive threshold in VPA rats. Suramin treatment did not affect VPA-induced upregulation of P2X4 and P2Y2 receptor expression in the hippocampus, and P2X4 receptor expression in the medial prefrontal cortex, but normalized an increased level of interleukin 6 (IL-6). Our results suggest an important role of purinergic signaling modulation in behavioral, molecular, and immunological aberrations described in VPA model, and indicate that the purinergic signaling system might be a potential target for pharmacotherapy in preclinical studies of ASD.

Normal radial migration and lamination are maintained in dyslexia-susceptibility candidate gene homolog Kiaa0319 knockout mice.

Developmental dyslexia is a common disorder with a strong genetic component, but the underlying molecular mechanisms are still unknown. Several candidate dyslexia-susceptibility genes, including KIAA0319, DYX1C1, and DCDC2, have been identified in humans. RNA interference experiments targeting these genes in rat embryos have shown impairments in neuronal migration, suggesting that defects in radial cortical migration could be involved in the disease mechanism of dyslexia. Here we present the first characterisation of a Kiaa0319 knockout mouse line. Animals lacking KIAA0319 protein do not show anatomical abnormalities in any of the layered structures of the brain. Neurogenesis and radial migration of cortical projection neurons are not altered, and the intrinsic electrophysiological properties of Kiaa0319-deficient neurons do not differ from those of wild-type neurons. Kiaa0319 overexpression in cortex delays radial migration, but does not affect final neuronal position. However, knockout animals show subtle differences suggesting possible alterations in anxiety-related behaviour and in sensorimotor gating. Our results do not reveal a migration disorder in the mouse model, adding to the body of evidence available for Dcdc2 and Dyx1c1 that, unlike in the rat in utero knockdown models, the dyslexia-susceptibility candidate mouse homolog genes do not play an evident role in neuronal migration. However, KIAA0319 protein expression seems to be restricted to the brain, not only in early developmental stages but also in adult mice, indicative of a role of this protein in brain function. The constitutive and conditional knockout lines reported here will be useful tools for further functional analyses of Kiaa0319.

Comment on "Oxytocin-mediated GABA inhibition during delivery attenuates autism pathogenesis in rodent offspring".

Tyzio et al. (Reports, 7 February 2014, p. 675) reported that bumetanide restored the impaired oxytocin-mediated γ-aminobutyric acid (GABA) excitatory-inhibitory shift during delivery in animal models of autism, ameliorating some autistic-like characteristics in the offspring. However, standard practices in the study of these models, such as the use of sex-dimorphic or males-only analyses and implementation of tests measuring social behavior, are lacking to definitely associate their findings to autism.

Long-term effects of gestational nicotine exposure and food-restriction on gene expression in the striatum of adolescent rats.

Gestational exposure to environmental toxins such as nicotine may result in detectable gene expression changes in later life. To investigate the direct toxic effects of prenatal nicotine exposure on later brain development, we have used transcriptomic analysis of striatal samples to identify gene expression differences between adolescent Lister Hooded rats exposed to nicotine in utero and controls. Using an additional group of animals matched for the reduced food intake experienced in the nicotine group, we were also able to assess the impact of imposed food-restriction on gene expression profiles. We found little evidence for a role of gestational nicotine exposure on altered gene expression in the striatum of adolescent offspring at a significance level of p<0.01 and |log2 fold change >0.5|, although we cannot exclude the possibility of nicotine-induced changes in other brain regions, or at other time points. We did, however, find marked gene expression differences in response to imposed food-restriction. Food-restriction resulted in significant group differences for a number of immediate early genes (IEGs) including Fos, Fosb, Fosl2, Arc, Junb, Nr4a1 and Nr4a3. These genes are associated with stress response pathways and therefore may reflect long-term effects of nutritional deprivation on the development of the stress system.

BDNF-TrkB signaling in striatopallidal neurons controls inhibition of locomotor behavior.

The physiology of brain-derived neurotrophic factor signaling in enkephalinergic striatopallidal neurons is poorly understood. Changes in cortical Bdnf expression levels, and/or impairment in brain-derived neurotrophic factor anterograde transport induced by mutant huntingtin (mHdh) are believed to cause striatopallidal neuron vulnerability in early-stage Huntington's disease. Although several studies have confirmed a link between altered cortical brain-derived neurotrophic factor signaling and striatal vulnerability, it is not known whether the effects are mediated via the brain-derived neurotrophic factor receptor TrkB, and whether they are direct or indirect. Using a novel genetic mouse model, here, we show that selective removal of brain-derived neurotrophic factor-TrkB signaling from enkephalinergic striatal targets unexpectedly leads to spontaneous and drug-induced hyperlocomotion. This is associated with dopamine D2 receptor-dependent increased striatal protein kinase C and MAP kinase activation, resulting in altered intrinsic activation of striatal enkephalinergic neurons. Therefore, brain-derived neurotrophic factor/TrkB signaling in striatopallidal neurons controls inhibition of locomotor behavior by modulating neuronal activity in response to excitatory input through the protein kinase C/MAP kinase pathway.

Behavioural and functional characterization of Kv10.1 (Eag1) knockout mice.

Kv10.1 (Eag1), member of the Kv10 family of voltage-gated potassium channels, is preferentially expressed in adult brain. The aim of the present study was to unravel the functional role of Kv10.1 in the brain by generating knockout mice, where the voltage sensor and pore region of Kv10.1 were removed to render non-functional proteins through deletion of exon 7 of the KCNH1 gene using the '3 Lox P strategy'. Kv10.1-deficient mice show no obvious alterations during embryogenesis and develop normally to adulthood; cortex, hippocampus and cerebellum appear anatomically normal. Other tests, including general health screen, sensorimotor functioning and gating, anxiety, social behaviour, learning and memory did not show any functional aberrations in Kv10.1 null mice. Kv10.1 null mice display mild hyperactivity and longer-lasting haloperidol-induced catalepsy, but there was no difference between genotypes in amphetamine sensitization and withdrawal, reactivity to apomorphine and haloperidol in the prepulse inhibition tests or to antidepressants in the haloperidol-induced catalepsy. Furthermore, electrical properties of Kv10.1 in cerebellar Purkinje cells did not show any difference between genotypes. Bearing in mind that Kv10.1 is overexpressed in over 70% of all human tumours and that its inhibition leads to a reduced tumour cell proliferation, the fact that deletion of Kv10.1 does not show a marked phenotype is a prerequisite for utilizing Kv10.1 blocking and/or reduction techniques, such as siRNA, to treat cancer.

Anxious, hypoactive phenotype combined with motor deficits in Gtf2ird1 null mouse model relevant to Williams syndrome.

Williams-Beuren syndrome (WBS) is a rare genetic disorder caused by a hemizygous deletion of around 28 genes on the long arm of chromosome 7 (7q11.23), characterized by a unique spectrum of behavioral impairments, including mental retardation, deficits in visuospatial constructive cognition, hypersociability, anxiety and simple phobias. Physical characteristics include dysmorphic faces, short stature, oculomotor deficits, gross and fine coordination impairments, diminished control of balance and mild extrapyramidal signs as well as gait abnormalities resembling gait hypokinesia. Genes near the distal deletion breakpoint appear to contribute most to the WBS cognitive and behavioral profile and include the GTF family of transcription factors: GTF2I, GTF2IRD1, GTF2IRD2. We have previously shown that heterozygous deletions of GTF2IRD1 in humans and homozygous deletion in mice contributes to craniofacial abnormalities. Here we show an important role of this gene in motor coordination and anxiety ascertained from extensive behavioral mouse phenotyping. Gtf2ird1 null mice showed lower body weight, decreased spontaneous and circadian locomotor activity, diminished motor coordination and strength, gait abnormalities, increased anxiety and an elevated endocrinological response to stress. Gtf2ird1 heterozygous mice displayed lower body weight and decreased circadian activity, but only minor motor coordination and anxiety-related behavioral dysfunctions. Our study strongly supports a role for GTF2IRD1 in the motoric and anxiety-related abnormalities seen in Williams-Beuren syndrome, and suggests basal ganglia and potentially cerebellar abnormalities in Gtf2ird1 mice.

Prenatal exposure to nicotine impairs performance of the 5-choice serial reaction time task in adult rats.

Cigarette smoking is associated with a wide variety of adverse reproductive outcomes, including increased infant mortality and decreased birth weight. Prenatal exposure to tobacco smoke, of which nicotine is a major teratogenic component, has also been linked to the acceleration of the risk for different psychiatric disorders, including conduct disorder and attention deficit hyperactivity disorder (ADHD). Whether this increased risk is influenced by the direct effects of gestational nicotine exposure on the developing fetus remains uncertain. In this study we provide experimental evidence for the effects of prenatal nicotine exposure on measures of attention and impulsivity in adult male rats. Offspring of females exposed during pregnancy to 0.06 mg/ml nicotine solution as the only source of water (daily consumption: 69.6±1.4 ml/kg; nicotine blood level: 96.0±31.9 ng/ml) had lower birth weight and delayed sensorimotor development measured by negative geotaxis, righting reflex, and grip strength. In the 5-choice serial reaction time test, adult rats showed increased numbers of anticipatory responses and omissions errors, more variable response times, and lower accuracy with evidence of delayed learning of the task demands when the 1 s stimulus duration was introduced. In contrast, prenatal nicotine exposure had no effect on exploratory locomotion or delay-discounting test. Prenatal nicotine exposure increased expression of the D5 dopamine receptor gene in the striatum, but did not change expression of other dopamine-related genes (DRD4, DAT1, NR4A2, and TH) in either the striatum or the prefrontal cortex. These data suggest a direct effect of prenatal nicotine exposure on important aspects of attention, inhibitory control, or learning later in life.

Gestational exposure to nicotine in drinking water: teratogenic effects and methodological issues.

Individual differences in nicotine effects lead to questions about appropriate experimental procedures for prenatal nicotine exposure in rodent models. The objective of this study was to develop a method for gestational studies in rats based on oral nicotine exposure, and to evaluate the neurodevelopmental effects. Female Lister hooded rats were exposed to nicotine solutions both before and during pregnancy. These female rats were divided into groups consuming solutions of different concentrations such that animals that initially consumed the solutions most readily were exposed to progressively higher concentrations. Offspring of these female rats were evaluated in a test battery measuring maturational and developmental milestones. Female rats ingested nicotine solutions at levels that provided blood nicotine concentrations of 10-60 ng/ml, at daily dose levels of 2.9-6.2 mg/kg. Solutions with concentrations below 0.06 mg/ml were well tolerated with some moderate adverse effects at the highest dose. Concentrations above 0.08 mg/ml led to a large drop in fluid consumption and in body weight. Strong teratogenic effects of prenatal nicotine exposure were observed at concentrations above 0.04 mg/ml, including developmental and maturational delays shown by measures of pinnae detachment, fur appearance, incisor eruption, eye opening and righting reflex. Negative geotaxis, grip strength and weight gain were impaired and postnatal mortality was increased. This study design provides a model for the impact of prenatal exposure to nicotine at blood levels comparable with those in medium and heavy smokers. There were marked developmental and behavioural deficits induced in the offspring of nicotine-exposed female rats.

An animal model of premenstrual dysphoric disorder sensitive to antidepressants.

Premenstrual dysphoric disorder (PMDD) is characterized by the recurrence of a cluster of physical and negative mood symptoms, especially irritability, appearing when estrogen and progesterone levels decrease during the late luteal phase of the menstrual cycle. This unit describes a new animal model of PMDD that shows differentiation between female rats expressing and not expressing ovarian cycle-dependent irritability measured by the behavior of burying harmless objects. Burying behavior is enhanced in a subgroup of female rats at metestrus and decreased at the proestrus phase of the estrous cycle. Increased marble burying at metestrus does not habituate and can be reversed by acute treatment with several antidepressants, which mimic the pharmacology of PMDD in humans. This model may be used to study the neuroendocrine mechanisms triggering premenstrual irritability, and the mode of action of antidepressants used for the treatment of PMDD.

Gender-specific behavioral and immunological alterations in an animal model of autism induced by prenatal exposure to valproic acid.

Autism is a severe behavioral disorder characterized by pervasive impairments in social interactions, deficits in verbal and non-verbal communication, and stereotyped behaviors, with a four times higher incidence in boys than in girls. The core symptoms are frequently accompanied by a spectrum of neurobehavioral and immunological derangements, including: aberrant sensitivity to sensory stimulation, anxiety, and decreased cellular immune capacity. Recently, a new potential rodent model of autism induced by prenatal exposure to valproic acid (VPA rats) has been proposed. In order to determine if gender has an influence on alterations observed in VPA rats, male and female rats have been evaluated in a battery of behavioral, immunological, and endocrinological tests. A plethora of aberrations has been found in male VPA rats: lower sensitivity to pain, increased repetitive/stereotypic-like activity, higher anxiety, decreased level of social interaction, increased basal level of corticosterone, decreased weight of the thymus, decreased splenocytes proliferative response to concanavaline A, lower IFN-gamma/IL-10 ratio, and increased production of NO by peritoneal macrophages. Female VPA rats exhibited only increased repetitive/stereotypic-like activity and decreased IFN-gamma/IL-10 ratio. Sexual dimorphism characteristics for measured parameters have been observed in both groups of animals, except social interaction in VPA rats. Our results confirm existence of similarities between the observed pattern of aberrations in VPA rats and features of disturbed behavior and immune function in autistic patients, and suggest that they are gender-specific, which is intriguing in light of disproportion in boys to girls ratio in autism.

Attenuation of estrous cycle-dependent marble burying in female rats by acute treatment with progesterone and antidepressants.

Premenstrual dysphoric disorder (PMDD) is characterized by the recurrence of a cluster of physical and negative mood symptoms, especially irritability, appearing when estrogen and progesterone levels decrease during the late luteal phase of the menstrual cycle. The aim of the present study was to explore a new potential model of premenstrual irritability. It has been suggested that burying of harmless objects by rodents may reflect a form of impulsive or anxiety-like behavior. This study demonstrates changes in burying behavior during various phases of the estrous cycle in some but not all female rats. Burying behavior was found to be enhanced at metestrus and decreased at proestrus, characterized by low and high ovarian hormone levels, respectively. No habituation of the cycle-dependent burying was observed. Enhanced burying was not observed in reproductive senescent and ovariectomized females characterized by stable, low levels of ovarian hormones. Increased marble burying at metestrus was reversed by acute treatment with antidepressants fluoxetine, desipramine, nomifensine, the benzodiazepine agonist diazepam, and progesterone, while the neuroleptic chlorpromazine was without effect. Reversal of cycle-dependent burying was unrelated to the drugs' effects on locomotor activity. These results indicate that estrous cycle-dependent marble-burying behavior displayed by a subgroup of female rats might be a manifestation of ovarian hormone-dependent irritability. This manifestation may be used to elucidate the neuroendocrine mechanisms triggering premenstrual irritability, and the detailed mode of action of antidepressants when used for PMDD.

Prenatal exposure to valproic acid disturbs the enkephalinergic system functioning, basal hedonic tone, and emotional responses in an animal model of autism.

RATIONALE: It has been suggested that behavioral aberrations observed in autism could be the result of dysfunction of the neuroregulatory role performed by the endogenous opioid peptides. Many of those aberrations have been recently modeled in rats exposed to valproic acid (VPA) on the 12th day of gestation (VPA rats). OBJECTIVES: The aim of the present study was to elucidate functioning of the enkephalinergic system, one of the endogenous opioid peptide systems strongly involved in emotional responses, in VPA rats using both biochemical and behavioral methods. MATERIALS AND METHODS: In situ hybridization was used to measure proenkephalin mRNA expression in adult VPA rats' central nucleus of the amygdala, the dorsal striatum, and the nucleus accumbens. Additional groups of animals were examined in a conditioned place aversion to naloxone, the elevated plus maze, and object recognition tests to assess their basal hedonic tone, anxiety, learning and memory, respectively. RESULTS: Prenatal exposure to VPA decreased proenkephalin mRNA expression in the dorsal striatum and the nucleus accumbens but not in the central nucleus of the amygdala. It also increased anxiety and attenuated conditioned place aversion to naloxone but had no impact on learning and memory. CONCLUSIONS: The present results suggest that prenatal exposure to VPA may lead to the decreased activity of the striatal enkephalinergic system and in consequence to increased anxiety and disregulated basal hedonic tone observed in VPA rats. Presented results are discussed in light of interactions between enkephalinergic, GABAergic, and dopaminergic systems in the striatum and mesolimbic areas of the brain.

Increased depressive-like traits in an animal model of premenstrual irritability.

Women are at higher risk of anxiety and mood disorders, especially at transitions across the reproductive life cycle (premenstruum, postpartum, menopause). Premenstrual dysphoric disorder (PMDD) is one of female mood disorders associated with changing ovarian hormone levels. Because anxiety and depression frequently occur in women with PMDD, premenstrual dysphoria might be a manifestation of certain vulnerability traits increasing the risk of those disorders. The present study was conducted to elucidate a potential association between estrous cycle-dependent aggression, the rodent model of "premenstrual irritability" (resident-intruder test), and anxiety (elevated plus maze), depressive-like traits (forced swim test) as well as carbohydrate craving in female Wistar rats. Some aggressive and nonaggressive females were restraint-stressed before testing to determine their sensitivity to stress at different hormonal stages. The results revealed that females expressing the estrous cycle-dependent aggression but not those not expressing cycle-dependent aggression spent longer time immobile and shorter time swimming in the forced swim test at metestrus compared to proestrus phase of the estrous cycle. There was no difference between aggressive and nonaggressive females in anxiety, locomotor activity and sensitivity to restraint stress and sucrose consumption. The present study suggests a common neurobiological background for the estrous cycle-dependent aggression and depressive-like traits in rodents. This phenomenon could potentially aid the elucidation of premenstrual emotional dysfunctions and might be used as an ethological model to study a biochemical and genetic proneness to depression.

Environmental enrichment reverses behavioral alterations in rats prenatally exposed to valproic acid: issues for a therapeutic approach in autism.

Environmental enrichment has been repeatedly shown to affect multiple aspects of brain function, and is known to improve cognitive, behavioral, and histopathological outcome after brain injuries. The purpose of the present experiments was to determine the effect of an enriched environment on behavioral aberrations observed in male rats exposed to valproic acid on day 12.5 of gestation (VPA rats), and proposed on the basis of etiological, anatomical, and behavioral data as an animal model of autism. Environmental enrichment reversed almost all behavioral alterations observed in the model. VPA rats after environmental enrichment (VPA-E) compared to VPA rats reared in standard conditions have higher sensitivity to pain and lower sensitivity to nonpainful stimuli; stronger acoustic prepulse inhibition; lower locomotor, repetitive/stereotypic-like activity, and enhanced exploratory activity; decreased anxiety; increased number of social behaviors; and shorter latency to social explorations. In comparison with control animals (Con), VPA-E rats exhibited increased number of pinnings in adolescence and social explorations in adulthood, and were less anxious in the elevated plus maze. Similar differences in social behavior and anxiety were observed between control rats exposed to environmental enrichment (Con-E) and control group reared in standard conditions. These results suggest that postnatal environmental manipulations can counteract the behavioral alterations in VPA rats. We propose environmental enrichment as an important tool for the treatment of autism spectrum disorders.

Behavioral alterations in rats prenatally exposed to valproic acid: animal model of autism.

Autism is a severe behavioral disorder characterized by pervasive impairments in social interactions, deficits in verbal and nonverbal communication, and stereotyped, repetitive patterns of behaviors and interests. Recently, a new rodent model of autism was created by exposure of rat fetuses to valproic acid (VPA) on the 12.5th day of gestation (VPA rats). The model has striking anatomical, pathological, and etiological similarities to human data; however, it has not been characterized behaviorally. In order to determine if VPA rats present behavioral aberrations observed in autism, their behavior was extensively evaluated in a battery of tests. The results of the present experiments demonstrate that VPA rats exhibit: (1) lower sensitivity to pain and higher sensitivity to nonpainful stimuli, (2) diminished acoustic prepulse inhibition, (3) locomotor and repetitive/stereotypic-like hyperactivity combined with lower exploratory activity, and (4) decreased number of social behaviors and increased latency to social behaviors. In addition, VPA rats showed delayed maturation, lower body weight, delayed motor development, and attenuated integration of a coordinated series of reflexes, delayed nest-seeking response mediated by olfactory system, and normal negative geotaxis. Interestingly, all behavioral aberrations described in this paper appear before puberty, which could distinguish the VPA rat model of autism from other animal models of neurodevelopmental disorders, especially rodent models of schizophrenia. Our results bring further support to validity of the proposed VPA animal model of autism, suggesting similarities between the observed pattern of behavioral alterations in VPA rats and features of disturbed behavior in autistic patients.