Pierre L. Roubertoux

Mitochondrial DNA modifies cognition in interaction with the nuclear genome and age in mice.

Several lines of evidence indicate an association between mitochondrial DNA (mtDNA) and the functioning of the nervous system. As neuronal development and structure as well as axonal and synaptic activity involve mitochondrial genes, it is not surprising that most mtDNA diseases are associated with brain disorders. Only one study has suggested an association between mtDNA and cognition, however. Here we provide direct evidence of mtDNA involvement in cognitive functioning. Total substitution of mtDNA was achieved by 20 repeated backcrosses in NZB/BlNJ (N) and CBA/H (H) mice with different mtDNA origins. All 13 mitochondrial genes were expressed in the brains of the congenic quartet. In interaction with nuclear DNA (nDNA), mtDNA modified learning, exploration, sensory development and the anatomy of the brain. The effects of mtDNA substitution persisted with age, increasing in magnitude as the mice got older. We observed different effects with input of mtDNA from N versus H mice, varying according to the phenotypes. Exchanges of mtDNA may produce phenotypes outside the range of scores observed in the original mitochondrial and nuclear combinations. These findings show that mitochondrial polymorphisms are not as neutral as was previously believed.

Role of the serotonin transporter gene in the behavioral expression of autism

The promoter polymorphism of the serotonin transporter gene (HTT, locus SLC6A4) is of special interest in autism given the well-replicated platelet hyperserotonemia of autism, treatment effects of serotonin reuptake inhibitors, and the role of serotonin in limbic functioning and neurodevelopment. Parent-offspring transmission of the long (l) and short (s) alleles of the deletion/insertion polymorphism in the HTT promoter region was examined in families of 71 children with autism using the transmission test for linkage disequilibrium (TDT). Transmission of HTT promoter alleles did not differ between probands with autism and their unaffected siblings. However, allelic transmission in probands was dependent upon severity of impairments in the social and communication domains, with greater s allele transmission in severely impaired individuals and greater l transmission in mild/moderately impaired individuals. This relationship between HTT promoter alleles and severity of autistic impairment was also seen when ratings of social and communication behaviors were compared across genotypes. The data indicate that HTT promoter alleles by themselves do not convey risk for autism, but, rather, modify the severity of autistic behaviors in the social and communication domains. The results require replication and, given the size of the groups and subgroups examined, must be considered still preliminary. The results suggest that future research on the genetics of autism should carefully assess each of the major behavioral domains and seriously consider the possible role of modifying loci.

Comparative Diagnoses of Twin Zygosity by SSLP Variant Analysis, Questionnaire, and Dermatoglyphic Analysis

Zygosity diagnosis has been performed in 79 pairs of twins using three methods. Simple sequence repeat length polymorphism (SSLP) analysis allows an efficient classification (MZ or DZ) with only a few markers following a simplified technique of extraction and amplification. A method based on a full questionnaire completed by parents about twin similarity correctly classifies 97.46% of the pairs; 92.41% are correctly classified using only four questions as suggested by logistic regression analysis. The third method, using dermatoglyphic analyses, correctly classifies 86.76% of pairs. To lower the cost of DNA diagnosis we stress the possibility of limiting its use to pairs with scores in the overlap area between MZ and DZ twins with a validated questionnaire.

Serotonin transporter promoter variants in autism: functional effects and relationship to platelet hyperserotonemia

The well-replicated platelet hyperserotonemia of autism has stimulated interest in serotonin (5-HT) in autism. We have examined the effects of the serotonin transporter gene (5-HTT, locus SLC6A4) promoter polymorphism (5-HTTLPR) on platelet 5-HT physiology in autism. Platelet 5-HT uptake rates and affinities (Vmax and Km), uptake site densities (Bmax) and 5-HT levels were examined in 31 French individuals with autism genotyped with respect to the 5-HTTLPR. Platelet 5-HT uptake and 5-HT levels were measured using HPLC; uptake sites were determined by radioligand binding. A 1.5-fold increased rate (Vmax) of platelet 5-HT uptake was observed in ll genotype individuals compared to those with ls and ss genotypes (Mann– Whitney U-test, P = 0.022). However, no significant relationship was observed between genotype and uptake site density (U-test, P = 0.51). Although median levels of platelet 5-HT in platelet-rich plasma were higher in the ll group, only trend level significance was observed (U-test, P= 0.069); platelet 5-HT content measured in whole blood was similar across genotypes. Uptake rates were well correlated with Bmax values (r = 0.66, P = 0.002); correlations between uptake and platelet 5-HT levels and between Bmax values and 5-HT levels were somewhat lower. While 5-HTTLPR alleles had an appreciable effect on platelet 5-HT uptake rates, effects on 5-HT levels and uptake site density were smaller or absent. Based on these preliminary data and prior studies of allele frequencies, we conclude that the 5-HTTLPR is not a major determinant of the group mean platelet serotonin elevation seen in autism. However, a role for increased uptake in the hyperserotonemia of autism can not be ruled out. In addition, it appears that studies of platelet 5-HT measures in autism and other disorders should take account of the effects of 5-HTTLPR genotype on 5-HT uptake

Differences in patterns of pup care in mice. V--Pup ultrasonic emissions and pup care behavior.

Newborn mice, like all newborn rodents, are able to emit high frequency signals, in particular when they are put out of the nest. Moreover, it is known that in this situation retrieving behaviors are induced in the foster mother, which are likely to reveal stable differences across inbred strains of mice. The question that arises is whether these differences are causally linked to differences in the pup rate of signalling and/or to the capacity of the females of these strains to perceive them. To provide insights into this question, the behavior of 8 inbred strains of mice was observed: A/J, BALB/c, CBA/H, C57BL/6, C57Br, DBA, NZB and XLII. Pup ultrasonic calls of each of these strains, emitted in the same conditions as a retrieving test, were recorded and tabulated. Auditory sensitivities of females belonging to these strains were determined by auditory evoked potentials recorded in the inferior colliculus. These two variables were analysed in relation to scores of females of these strains on three variables of a retrieving test. Results show that the presence of other factors than auditory cues must be taken into account to describe differences across strains in retrieving performances. This conclusion has been confirmed by results obtained using cross-fostering procedure. Female mice unable to utilize ultrasonic information may use other sensory channels. Furthermore, female mice capable of perceiving ultrasounds may also be able to use different sensory modalities in different situations.

Pain Reactivity and Plasma β-Endorphin in Children and Adolescents with Autistic Disorder

Background Reports of reduced pain sensitivity in autism have prompted opioid theories of autism and have practical care ramifications. Our objective was to examine behavioral and physiological pain responses, plasma β-endorphin levels and their relationship in a large group of individuals with autism. Methodology/Principal Findings The study was conducted on 73 children and adolescents with autism and 115 normal individuals matched for age, sex and pubertal stage. Behavioral pain reactivity of individuals with autism was assessed in three observational situations (parents at home, two caregivers at day-care, a nurse and child psychiatrist during blood drawing), and compared to controls during venepuncture. Plasma β-endorphin concentrations were measured by radioimmunoassay. A high proportion of individuals with autism displayed absent or reduced behavioral pain reactivity at home (68.6%), at day-care (34.2%) and during venepuncture (55.6%). Despite their high rate of absent behavioral pain reactivity during venepuncture (41.3 vs. 8.7% of controls, P<0.0001), individuals with autism displayed a significantly increased heart rate in response to venepuncture (P<0.05). Moreover, this response (Δ heart rate) was significantly greater than for controls (mean±SEM; 6.4±2.5 vs. 1.3±0.8 beats/min, P<0.05). Plasma β-endorphin levels were higher in the autistic group (P<0.001) and were positively associated with autism severity (P<0.001) and heart rate before or after venepuncture (P<0.05), but not with behavioral pain reactivity. Conclusions/Significance The greater heart rate response to venepuncture and the elevated plasma β-endorphin found in individuals with autism reflect enhanced physiological and biological stress responses that are dissociated from observable emotional and behavioral reactions. The results suggest strongly that prior reports of reduced pain sensitivity in autism are related to a different mode of pain expression rather than to an insensitivity or endogenous analgesia, and do not support opioid theories of autism. Clinical care practice and hypotheses regarding underlying mechanisms need to assume that children with autism are sensitive to pain.

Gene × Environment Interactions in Autism Spectrum Disorders: Role of Epigenetic Mechanisms

Several studies support currently the hypothesis that autism etiology is based on a polygenic and epistatic model. However, despite advances in epidemiological, molecular and clinical genetics, the genetic risk factors remain difficult to identify, with the exception of a few chromosomal disorders and several single gene disorders associated with an increased risk for autism. Furthermore, several studies suggest a role of environmental factors in autism spectrum disorders (ASD). First, arguments for a genetic contribution to autism, based on updated family and twin studies, are examined. Second, a review of possible prenatal, perinatal, and postnatal environmental risk factors for ASD are presented. Then, the hypotheses are discussed concerning the underlying mechanisms related to a role of environmental factors in the development of ASD in association with genetic factors. In particular, epigenetics as a candidate biological mechanism for gene × environment interactions is considered and the possible role of epigenetic mechanisms reported in genetic disorders associated with ASD is discussed. Furthermore, the example of in utero exposure to valproate provides a good illustration of epigenetic mechanisms involved in ASD and innovative therapeutic strategies. Epigenetic remodeling by environmental factors opens new perspectives for a better understanding, prevention, and early therapeutic intervention of ASD.

Additive and interactive effects of genotype and maternal environment

Publisher Summary The interaction between genotype and environment is one of the most widely employed concepts in genetics as it is in the behavioral sciences. This chapter discusses on maternal environments and their additive or interactive effects with the offspring genotype. It presents the thesis: that there are nonlinear relationships between gene and the central nervous system (CNS) functioning, including behavior. Most of the genetic information involved in higher nervous activity is susceptible to modification by the effect of components of the environment, particularly of the maternal one. The concepts of interactive and additive effects are discussed from a genetic standpoint. The components of the maternal environment are defined. Their interactive and additive effects with offspring genotype are presented. The physiological mechanisms underlying these effects are also analyzed.

Hippocampal mossy fiber distributions and intermale aggression in seven inbred mouse strains

The capacity to initiate attack behavior against a passive standard opponent was measured in 140 male mice belonging to seven different inbred mouse strains. Large strain differences were found, which strongly correlated with the size of the hippocampal intra- and infrapyramidal mossy fibers terminal fields. These results, combined with those obtained from earlier experiments, point to a possible modulating role of the hippocampus in the regulation of attack behavior in male mice.

Vocalizations in newborn mice: Genetic analysis

Two kinds of vocalizations are produced by newborn mice: whistles (between 50 and 150 ms in length), having a narrow bandwidth in each strain that ranges from 30 to 90 kHz; and clicks, which are shorter (about 1 ms) and have a larger bandwidth. These vocalizations were individually recorded in 1-day-old pups from seven inbred strains of laboratory mice, at two temperatures (23±0.5 and 15±0.5°C). The numbers of clicks and whistles were counted under these two conditions. Moreover, the length and frequencies at the beginning, apex, and end of the whistles were measured during the 15°C condition. Correlations, including several components—additivity, epistasis (between homozygous loci), and maternal environment—were calculated between the characteristics of the whistles during the 15°C condition. Clicks and whistles were also counted from 1 to 8 days of age during the 15°C condition. The numbers of clicks and whistles were age dependent, with a decrease from day 1 to day 8 for the clicks and a consistent production of whistles. A quantitative genetic analysis was also performed on the 1-day-old pups from the mendelian generations produced by the inbred strains most contrasting for the number of whistles produced in the cold condition: NZB/BINJ and CBA/H. The heterozygous genotype of the mother induced an increment of the number of whistles. Moreover, a significant part of the additive variance was suspected from the first design, and found with the second one, for this variable. Quantitative genetic analysis showed significant dominance and epistasis between homozygous loci and homozygous and heterozygous loci. This points to multigenic correlates for the number of whistles in this population. The significant additive values for all the variables recorded during the 15±0.5°C condition and for the number of whistles produced during the 23±0.5°C condition are compatible with an effect the indicates neither directional nor stabilizing selection. This results is examined in the light of the multichannel sensorial process implicated in maternal behavior in mice.