Genomic imprinting, action, and interaction of maternal and fetal genomes.

Mammalian viviparity (intrauterine development of the fetus) introduced a new dimension to brain development, with the fetal hypothalamus and fetal placenta developing at a time when the fetal placenta engages hypothalamic structures of the maternal generation. Such transgenerational interactions provide a basis for ensuring optimal maternalism in the next generation. This success has depended on genomic imprinting and a biased role of the matriline. Maternal methylation imprints determine parent of origin expression of genes fundamental to both placental and hypothalamic development. The matriline takes a further leading role for transgenerational reprogramming of these imprints. Developmental errors are minimized by the tight control that imprinted genes have on regulation of downstream evolutionary expanded gene families important for placental and hypothalamic development. Imprinted genes themselves have undergone purifying selection, providing a framework of stability for in utero development with most growth variance occurring postnatally. Mothers, not fathers, take the lead in the endocrinological and behavior adaptations that nurture, feed, and protect the infant. In utero coadaptive development of the placenta and hypothalamus has thus required a concomitant development to ensure male masculinization. Only placental male mammals evolved the sex determining SRY, which activates Sox9 for testes formation. SRY is a hybrid gene of Dgcr8 expressed in the developing placenta and Sox3 expressed in hypothalamic development. This hybridization of genes that take their origin from the placenta and hypothalamus has enabled critical in utero timing for the development of fetal Leydig cells, and hence testosterone production for hypothalamic masculinization.

The post-natal chemosensory environment induces epigenetic changes in vomeronasal receptor gene expression and a bias in olfactory preference.

Vomeronasal stem cells are generated throughout the life of a mouse and differentiate into neurons that express one vomeronasal type 1 (V1r), one or two vomeronasal type 2 (V2r), or one olfactory receptor. Vomeronasal stem cells can be induced to differentiate into neurons by treatment with lipocalins from mouse urine or by epigenetic modification following treatment with histone deacetylase inhibitors. An important question is, do chemosensory signals, modify the detection capabilities of the vomeronasal organ and affect behaviour. Rearing mice in the presence of urine (and its pheromonal signals) derived from a different mouse strain, affected the behavioural preference for non-kin which were accompanied by changes in vomeronasal receptor expression. Significant changes in the expression of vomeronasal V1r, V2r and olfactory receptors, major urinary proteins, and a number of genes thought to be involved in transcriptional regulation were also observed following urine treatment. These results suggest that modification of a mouse's urinary environment may exert epigenetic effects on developing vomeronasal neurons, which modify the type of vomeronasal receptors that are expressed. This may provide a mechanism by which environmental changes are able to modify the detection capabilities of the vomeronasal organ to respond optimally to the most likely social environment that a mouse will encounter when mature.

Epigenetics and brain evolution.

Fundamental aspects of mammalian brain evolution occurred in the context of viviparity and placentation brought about by the epigenetic regulation of imprinted genes. Since the fetal placenta hormonally primes the maternal brain, two genomes in one individual are transgenerationally co-adapted to ensure maternal care and nurturing. Advanced aspects of neocortical brain evolution has shown very few genetic changes between monkeys and humans. Although these lineages diverged at approximately the same time as the rat and mouse (20 million years ago), synonymous sequence divergence between the rat and mouse is double that when comparing monkey with human sequences. Paradoxically, encephalization of rat and mouse are remarkably similar, while comparison of the human and monkey shows the human cortex to be three times the size of the monkey. This suggests an element of genetic stability between the brains of monkey and man with a greater emphasis on epigenetics providing adaptable variability.

Placental protection of the fetal brain during short-term food deprivation.

The fetal genome regulates maternal physiology and behavior via its placenta, which produces hormones that act on the maternal hypothalamus. At the same time, the fetus itself develops a hypothalamus. In this study we show that many of the genes that regulate placental development also regulate the developing hypothalamus, and in mouse the coexpression of these genes is particularly high on embryonic days 12 and 13 (days E12-13). Such synchronized expression is regulated, in part, by the maternally imprinted gene, paternally expressed gene 3 (Peg3), which also is developmentally coexpressed in the hypothalamus and placenta at days E12-13. We further show that challenging this genomic linkage of hypothalamus and placenta with 24-h food deprivation results in disruption to coexpressed genes, primarily by affecting placental gene expression. Food deprivation also produces a significant decrease in Peg3 gene expression in the placenta, with consequences similar to many of the placental gene changes induced by Peg3 mutation. Such genomic dysregulation does not occur in the hypothalamus, where Peg3 expression increases with food deprivation. Thus, changes in gene expression brought about by food deprivation are consistent with the fetal genome's maintaining hypothalamic development at a cost to its placenta. This biased change to gene dysregulation in the placenta is linked to autophagy and ribosomal turnover, which sustain, in the short term, nutrient supply for the developing hypothalamus. Thus, the fetus controls its own destiny in times of acute starvation by short-term sacrifice of the placenta to preserve brain development.

Epigenetically regulated imprinted genes and foetal programming.

Genomic imprinting is a widespread epigenetic phenomenon in mammals and many imprinted genes are expressed in the developing hypothalamus and placenta. The placenta and brain are very different structures with very different roles, but in the pregnant mother they functionally interact coordinating and ensuring the provision of nutrients, timing of parturition and priming of hypothalamus for maternal care and nurturing. This interaction has been evolutionarily fine-tuned to optimise infant survival such that when resources are poor, the mother 'informs' this condition to the foetus producing a thrifty phenotype that is adapted to survive scarce resources after birth.

Paternal influence on female behavior: the role of Peg3 in exploration, olfaction, and neuroendocrine regulation of maternal behavior of female mice.

Genomic imprinting represents a mechanism through which parent-of-origin effects on offspring development may be mediated. However, investigation of the influence of imprinted genes on behavior has been limited. Here the authors investigate the role of the maternally imprinted/paternally expressed gene, Peg3, in several aspects of behavior using both 129Sv- and B6-Peg3 mutant female mice. Virgin Peg3 females on both genetic backgrounds were less exploratory and had higher rates of defecation with strain-dependent effects on activity levels and olfactory discrimination. Reproductive success, pup retrieval, and postnatal maternal care of pups were reduced in these females whereas indices of maternal aggression were higher among B6 Peg3-KO females. Differences in maternal care were apparent in females caring for biological or cross-fostered offspring and deficits in pup retrieval apparent beyond the immediate postpartum period. Oxytocin receptor binding in the MPOA and LS was reduced in Peg3-KO females. Thus, the authors demonstrate that disruptions to Peg3 influences aspects of female behavior that are critical for mediating maternal effects on offspring development, such as postpartum licking/grooming, and that effects of Peg3 are dependent on the maternal genetic background.

Increased apoptosis during neonatal brain development underlies the adult behavioral deficits seen in mice lacking a functional paternally expressed gene 3 (Peg3).

Inactivation of the maternally imprinted, paternally expressed gene 3 (Peg3) induces deficits in olfactory function, sexual and maternal behaviors, oxytocin neuron number, metabolic homeostasis and growth. Peg3 is expressed in a number of developing hypothalamic and basal forebrain structures and is a component of the P53 apoptosis pathway. Peg3 inactivation in neuronal cell culture lines inhibits P53 mediated apoptosis, which is important in the early postnatal development and sexual differentiation of the brain. In this study, we investigated the effect of inactivating the Peg3 gene on the incidence of caspase 3 positive cells (a marker of apoptosis) in 4- and 6-day postpartum mouse brain. Inactivating the Peg3 gene resulted in an increase in the incidence of total forebrain caspase 3 positive cells at 4 and 6 days postpartum. Increases in specific neuroanatomical regions including the bed nucleus of the stria terminalis, nucleus accumbens, caudate putamen, medial pre-optic area, arcuate nucleus, medial amygdala, anterior cortical and posteriodorsal amygdaloid nuclei, were also observed. In wild-type mice, sex differences in the incidence of caspase 3 positive cells in the medial amygdala, bed nucleus of the stria terminalis, nucleus accumbens, arcuate nucleus and the M2 motor cortex, were also observed. This neural sex difference was ameliorated in the Peg-3 mutant. These findings suggest that the neuronal and behavioral deficits seen in mice lacking a functional Peg3 gene are mediated by increases in the incidence of early neonatal apoptosis in neuroanatomical regions important for reproductive behavior, olfactory and pheromonal processing, thermoregulation and reward.

The evolution of pheromonal communication.

Small-brained rodents have been the principle focus for pheromonal research and have provided comprehensive insights into the chemosensory mechanisms that underpin pheromonal communication and the hugely important roles that pheromones play in behavioural regulation. However, pheromonal communication does not start or end with the mouse and the rat, and work in amphibians reveals much about the likely evolutionary origins of the chemosensory systems that mediate pheromonal effects. The dual olfactory organs (the main olfactory epithelium and the vomeronasal organ), their receptors and their separate projection pathways appear to have ancient evolutionary origins, appearing in the aquatic ancestors of all tetrapods during the Devonian period and so pre-dating the transition to land. While the vomeronasal organ has long been considered an exclusively pheromonal organ, accumulating evidence indicates that it is not the sole channel for the transduction of pheromonal information and that both olfactory systems have been co-opted for the detection of different pheromone signals over the course of evolution. This has also led to great diversity in the vomeronasal and olfactory receptor families, with enormous levels of gene diversity and inactivation of genes in different species. Finally, the evolution of trichromacy as well as huge increases in social complexity have minimised the role of pheromones in the lives of primates, leading to the total inactivation of the vomeronasal system in catarrhine primates while the brain increased in size and behaviour became emancipated from hormonal regulation.

The paternally expressed gene Peg3 regulates sexual experience-dependent preferences for estrous odors.

Sexual experience has marked and long-lasting effects on male behavior in mammals, regulating traits such as the anticipation and display of sexual behavior, aggression, and olfaction. The authors conducted urine preference, habituation-dishabituation, and partner choice tests with sexually experienced and naive male mice and found that wild-type males acquire adaptively significant preferences for the odors of receptive, estrous females with sexual experience, and that these preferences are matched by changes in main olfactory system responses involving the piriform cortex, as indicated by c-Fos expression. The authors also report that these experiential effects are disrupted in male mice carrying a knockout of the imprinted gene Peg3. This paternally expressed gene regulates maternal care and offspring development, but the authors here report that Peg3 mutant males suffer a complex olfactory deficit that affects estrous odor preferences and the responses of the main olfactory system to such odors. Peg3 appears to have evolved to regulate the experience-dependent preference for receptive females, an adaptive trait that would enhance male reproductive success and so potentially increase paternal transmission of this paternally expressed gene.

Visualisation of the vomeronasal pheromone response system.

Mammalian vomeronasal receptors respond to pheromones conveying information on gender, reproductive status and individual recognition. The question arises as to how this information is coded, which parts of the code require combinatorial activity and whether or not there are specific receptor neurons committed to sex discrimination. Are there receptor neurons that are committed to responding for female or male pheromones? Is there a sex difference for the proportion of these receptors, bearing in mind that it is very much in the male's interest to distinguish the restricted oestrous phase of the female's cycle in order to successfully mate? Perhaps more intriguing is the complexity of individual recognition and whether or not the vomeronasal receptors actually possess this capacity. A recent paper in Science by Ron Yu and colleagues addresses these issues by literally visualising patterns of activity in VNO slices and determining what information is common across different individuals and what distinguishes them.

Epigenetics, brain evolution and behaviour.

Molecular modifications to the structure of histone proteins and DNA (chromatin) play a significant role in regulating the transcription of genes without altering their nucleotide sequence. Certain epigenetic modifications to DNA are heritable in the form of genomic imprinting, whereby subsets of genes are silenced according to parent-of-origin. This form of gene regulation is primarily under matrilineal control and has evolved partly to co-ordinate in-utero development with maternal resource availability. Changes to epigenetic mechanisms in post-mitotic neurons may also be activated during development in response to environmental stimuli such as maternal care and social interactions. This results in long-lasting stable, or short-term dynamic, changes to the neuronal phenotype producing long-term behavioural consequences. Use of evolutionary conserved mechanisms have thus been adapted to modify the control of gene expression and embryonic growth of the brain as well as allowing for plastic changes in the post-natal brain in response to external environmental and social cues.

Natural variations in postpartum maternal care in inbred and outbred mice.

The role of maternal care in mediating variation in offspring phenotype has been examined in the rat and demonstrates that mother-infant interactions are critical for inducing long-term changes in behavior. Though phenotypic differences between mice strains are often attributed to genetic factors, the influence of early maternal environment has not been extensively explored. To understand maternal influence on phenotype in mice, we must first explore the nature of differences in behavior. In the present study, we examine aspects of maternal care differentiating mice strains and explore the relationship between postpartum behavior and measures obtained by a standard test of maternal responsivity (Retrieval Test). We compared inbred 129Sv (n=25), C57BL/6J (n=23), and outbred Swiss (n=23) lactating female mice. Swiss females had shorter latencies to retrieve and crouch over pups (P<.01), whereas 129Sv females had shorter latencies to nestbuild (P<.05). Conversely, observations of homecage behavior indicate that 129Sv females nestbuild less frequently. 129Sv females also engaged in very low levels of pup licking/grooming (P<.001) and long periods of nursing/contact (P<.05) with pups compared to C57BL/6J and Swiss females. Temporal analysis suggests that the magnitude of these differences varies both within and between days. No significant correlations were found between any aspect of maternal responsivity and postpartum behavior. These results illustrate that through detailed analysis of maternal behavior in mice, variations between strains can be observed. These variations represent strain specific strategies for promoting growth and survival of offspring during infancy that may also mediate "epigenetic" differences in phenotype in adulthood.

Genomic imprinting mediates sexual experience-dependent olfactory learning in male mice.

Mammalian imprinted genes are generally thought to have evolved as a result of conflict between parents; however, recent knockout studies suggest that coadaptation between mother and offspring may have been a significant factor. We present evidence that the same imprinted gene that regulates mammalian maternal care and offspring development also regulates male sexual behavior and olfaction. We have shown that the behavior of male mice carrying a knockout of the imprinted gene Peg3 does not change with sexual experience and that the mice are consequently unable to improve their copulatory abilities or olfactory interest in female odor cues after mating experience. Forebrain activation, as indexed by female odor-induced c-Fos protein induction, fails to increase with sexual experience, providing a neural basis for the behavioral deficits that the male mice display. The behavioral and neural effects of the Peg3 knockout show that this imprinted gene has evolved to regulate multiple and varied aspects of reproduction, from male sexual behavior to female maternal care, and the development of offspring. Moreover, sexual experience-driven behavioral changes may represent an adaptive response that enables males to increase their reproductive potential over their lifespan, and the effects we have found suggest that the evolution of genomic imprinting has been influenced by coadaptation between males and females as well as between females and offspring.

Suppression of prolactin does not reduce infant care by parentally experienced male common marmosets (Callithrix jacchus).

High levels of prolactin have been found to correlate with the expression of paternal care in a variety of taxa. However, in mammals, there is little experimental evidence that prolactin is causally involved in the stimulation or maintenance of paternal care. Here, we suppressed prolactin production in paternally experienced common marmoset fathers in their family groups during the first 2 weeks after their infants were born. Circulating prolactin levels were suppressed using cabergoline (Dostinex: Pfizer), a long acting dopamine (D2) agonist with minimal behavioural side-effects. A within-subject design was used to compare behavioural and hormonal data on 5 paternally experienced fathers during two consecutive births. Cabergoline reduced prolactin to negligible levels in all fathers without effecting testosterone, DHT and cortisol and without adverse side-effects. However, lowering prolactin had no significant effect on the expression of majority of the behaviour patterns associated with paternal care. These included infant carrying, infant grooming and the frequency with which fathers retrieved and rejected infants. The only infant-related behaviour to be affected was the frequency with which fathers touched, licked and investigated infants. We noted a marginally significant increase in this behaviour during cabergoline treatment. Despite the lack of effect on paternal care, cabergoline did exert an effect on the affiliative/sexual behaviour of fathers as there was a significant increase in the grooming behaviour fathers directed at and received from their mates during drug treatment. This study showed that experienced male marmosets can express paternal behaviour in the absence of the high prolactin levels normally seen after infants are born.

Urinary pheromones promote ERK/Akt phosphorylation, regeneration and survival of vomeronasal (V2R) neurons.

The G protein-coupled pheromone receptor neurons (V1R and V2R) of the vomeronasal organ (VNO) are continually replaced throughout the lifetime of the mouse. Moreover, active signalling of V2Rs via the transient receptor potential 2(TRPC2) channel is necessary for regeneration of receptors, as the TRPC2 null mutant mouse showed a 75% reduction of V2Rs by the age of two months. Here we describe V2R mediated signalling in a neuronal line established from vomeronasal stem cells taken from postnatal female mice. Cells were immunoreactive for Galpha(o) and V2R, whereas V1R and Galpha(i) immunoreactivity could not be detected. Biological ligands (dilute urine and its protein fractions) were found to increase proliferation and survival of these neurons. Dilute mouse urine but not artificial urine also induced ERK, Akt and CREB signalling in a dose dependent way. The volatile fraction of male mouse urine alone was without effect while the fraction containing peptides (> 5 kDa) also stimulated ERK and Akt phosphorylation. The ERK, Akt and CREB phosphorylation response was sensitive to pertussis toxin, confirming the involvement of V2R linked Galpha(o). Dilute mouse urine or its high molecular weight protein fraction increased survival and proliferation of these neurons. Hence, urinary pheromones, which signal important social information via mature neurons, also promote survival and proliferation of their regenerating precursors. These data show that regenerating V2Rs respond to urine and the urinary peptides by activation of the Ras-ERK and PI3-Akt pathways, which appear to be important for vomeronasal neural survival and proliferation.

Genes, brains and mammalian social bonds.

Recent studies of monogamous species have revealed the role of the neuropeptides oxytocin and vasopressin in activating reward mechanisms of the brain that are involved in establishing partner recognition and selective 'bonding'. The evolutionary history of these findings resides, at a mechanistic level, in the reciprocal bonding between mother and infant that is common to all mammals. However, in Old World primates, where mother and infant alone would not survive, living in large social groups brings extended family relationships and provides for alloparenting. This has required the emancipation of parenting behaviour from the constraints of hormonal state and the evolution of large brains for decision making that was previously restricted and determined by hormonal state. How this has been achieved, what conserved mechanisms underpin social bonding, and what genetic and mechanistic changes have occurred in the evolution of social bonds are the issues addressed here.