1. Background—Behavioral Epigenetics
Epigenetics is the term that is widely used to describe the DNA-related molecular processes that may alter gene expression without changing the DNA sequence. In general, epigenetic modifications can be inherited both mitotically and meiotically. These epigenetic modifications play essential roles in gene regulation in which case epigenetic marks such as DNA methylation and histone modifications are often described as switches that regulate gene expression [1
]. Aberrant changes in the epigenetic profile might induce abnormal gene silencing. Subsequently, physiological regulation, such as cellular differentiation, genomic imprinting, genome stability, and even behavior can be potentially affected. As studies have shown, similar physiological regulatory mechanisms are also observed in the offspring of affected organisms, proving that aberrant changes in the epigenetic profile are heritable.
In fact, several comprehensive studies have noted that environmental factors are closely associated with epigenetic regulations. Diet [2
], toxicants [3
], pollutants [4
], and irradiation [5
] are such factors that trigger alterations in epigenetic profiles. Interestingly, environment enrichments such as adverse experiences have also been proposed as factors that may affect the epigenetic profile [6
]. For instance, childhood abuse and maternal separation are linked to aberrant DNA methylation profiles in the hypothalamic-pituitary-adrenal (HPA) stress response axis [7
]. HPA axis is a complex set of direct influences and feedback interactions among the hypothalamus, pituitary gland, and adrenal glands, and it plays an important role in the reaction to various kind of stress. As a result, aberrant epigenetic regulation can disrupt the HPA axis activity, leading to an increased risk of suicide [9
]. This type of epigenetic regulation induced by experience or enriched environment has attracted the attention of many epigenetic researchers.
To address this interesting phenomenon, researchers have recently worked on the concept of “behavioral epigenetics”. Behavioral epigenetics is defined as the study of how epigenetic alterations induced by experience and environmental stress may affect animal behavior, which describes the epigenetic alterations caused by environmental enrichments [5
]. Environmental enrichment generally relates to the provision of environmental stimuli that promotes the expression of species-appropriate behavior and mental activities [10
]. Furthermore, there is accumulating evidence of the presence of metastable epialleles in both plants and animals, which have been proven to be able to affect the activity of adjacent alleles [11
]. Furthermore, some of these metastable epialleles are found to be associated with parent-of-origin effects and transgenesis in epigenetic inheritance [14
]. However, the underlying mechanism of the transmission of epigenetic traits remains poorly understood. Therefore, herein, we study behavioral epigenetics by broadly surveying the research on epigenetic regulation induced by enriched environment and identifying the possible epigenetic marks involved. Moreover, the potentially transmissible pathways underlying behavioral epigenetics are highlighted in this review as well. We also suggest considerations in the establishment of a transgenerational epigenetic inheritance model in order to narrow the outlook for future behavioral epigenetic studies.
4. Considerations in the Establishment of Epigenetic Transgenerational Inheritance
are well-known examples of metastable epialleles in mammals and epigenetically regulated phenotypes such as agouti coat color and kinked tail are observed to be transmitted across generations [46
]. While there are stably inherited epialleles found in both animals and plants, there remains much uncertainty regarding transgenerational inheritance in behavioral epigenetics.
First, a precise understanding of the definition of epigenetic transgenerational inheritance is essential. Epigenetic transgenerational inheritance is generally described as the transmission of nongenetic information across at least two generations in the absence of direct environmental exposure [48
]. For transgenerational inheritance, patrilineal and matrilineal lines are considered to be different; in some previous research, epigenetic transgenerational inheritance was misidentified as an effect of multigenerational exposure. For instance, when the gestating female (F0) was exposed to environmental stress, the primordial germ cell (F1 germ cell) carrying the gestating female was considered to be exposed as well. Thus, examination of transgenerational inheritance effects in matrilineal lines should start from the F3 generation. In contrast, assessment of transgenerational inheritance in patrilineal lines can start from the F2 generation because there is no exposure in the gestating period during patrilineal transmission. Furthermore, even though postnatal experience is also an important concern in experience-dependent epigenetics, there are many interfering factors, such as in utero effects that should be taken into consideration when examining the postnatal effects [49
]. Thus, to address the difficulties associated with behavioral epigenetic research, a well-designed experiment is especially important. For instance, cross-fostering is crucial in determining the contribution of postnatal experience in epigenetic inheritance [51
]. In short, an understanding of the fundamental concept of epigenetic inheritance and precise experimental design are necessary to study epigenetic transgenerational inheritance mechanisms in behavioral epigenetics.
Second, an understanding of how epigenetic profiles can be inherited meiotically and sustained in subsequent generations is required. As mentioned above, the epigenetic profile in the genome is subjected to two dynamic reprogramming events during the formation of germ cells and before embryo implantation [52
]. Most biologists have linked epigenetic transgenerational inheritance to the incomplete erasure of epigenetic markers during the occurrence of reprogramming events before embryo implantation in the uterus. To address this issue, scientists have focused on the differentially methylated region (DMR) and differentially methylated gene (DMG) that often escape demethylation during reprogramming waves and hence get expressed in subsequent generations. Recently, one comprehensive review suggested that the escape of methylated regions from reprogramming waves is not a stochastic event, and DNMT variants have exhibited significant roles in the regulation of the methylation pattern [53
]. Hence, an underlying pattern of epigenetic regulation may be observed via the examination of epigenetic profiles across a variety of species. For instance, the methylation profile in humans in the previously mentioned CGIs is usually retained after the reprogramming event. Therefore, CGIs are selectively subjected to imprinting.
Another consideration in epigenetic transgenerational inheritance is the transmission of epigenetic information in only matrilineal or patrilineal lines, indicating a sex-specific epigenetic inheritance model [54
]. The sex-specific inheritance model indicates a complicated regulatory network underlying the incomplete erasure of epigenetic markers on either parental side. A previous review proposed that the specialized chromatin structure is required to protect CGIs from demethylation [56
]. Furthermore, clustering of imprinted genes is associated with the existence of imprinting control regions, whereby clusters of methylated imprinted genes can have a higher probability of survival across reprogramming events. In this scenario, both paternal and maternal effects in behavioral epigenetic inheritance should be considered, as most research in the field of behavioral epigenetics is associated with only the exposure of adverse experiences during the gestation period. Perhaps the specific escape pattern during epigenetic reprogramming events in behavioral epigenetic cases will be revealed in future research, as the complex mechanisms underlying these events are substantial obstacles for the establishment of epigenetic inheritance mechanisms in mammals.
Recently, epigenetic marks associated with environmental enrichment have been predicted to be reversible by several studies. One of the studies suggests that treatment with the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) can reverse the hypermethylation status induced by maternal behavior in the exon 17
GR promoter [57
]. According to mapping results, TSA treatment decreased the methylation level of the maternal care group by increasing the K9 acetylation of H3 histones in the exon 17
GR promoter. In short, the DNA methylation profile in the hippocampus can be reversed through exposure to other aggressive stimuli that can alter chromatin structure. Therefore, in order to be transgenerationally inherited, the epigenetic markers should at least be sustained before conception.
Furthermore, accumulating evidence supports a possible role for in vitro fertilization (IVF) treatments in alternating the stability of epigenetic markers such as DNA methylation and histone modifications [58
]. In a mouse study, in vitro embryo cultures exhibited decreased growth rates and organ mass of offspring. Similar phenomena were observed in the subsequent generation as well [59
]. As indicated above, IVF treatments can disrupt epigenetic regulation. Therefore, it is crucial to understand the implications of IVF in behavioral epigenetics as some of the offspring in behavioral epigenetics research is conceived via IVF.
The interact epigenetic inheritance of somatic cells through germline also led to the consideration of the epigenomic impact of epigenetic information carrier circulated between somatic cells and the germline during the inheritance process. However, there is more than one regulatory factor that is suggested to be involved in the circulation of epigenetic information. To be transgenerationally inherited, interplay between epigenetic markers is especially important. Several studies have suggested that epigenetic markers that are transmissible to subsequent generations will be affected if one of these markers is absent or loses its function due to interference. For instance, regulation of DNMT variants is often linked to histone modifications in the induction of gene silencing [41
]. Without a clear understanding of the epigenetic inheritance mechanism, it is difficult to firmly state that epigenetic material can be transmitted via only one epigenetic mechanism. Further research is thus needed to study the cooperation between epigenetic markers.
Lastly, genetic effects should not be ignored in epigenetic studies. A study on the contribution of genetic variation to epigenetic inheritance in humans suggested that up to 20% of variation in DNA methylation profiles is due to the sequence-based DNA variants [60
]. Thus, in addition to environmental enrichment as the source of stimuli in behavioral epigenetic regulation, genetic factors may play a particularly important role in epigenetic inheritance.
Behavioral epigenetics is an interdisciplinary field involving neurobiology, psychology, reproductive biology, etc. Research in behavioral epigenetics over the past few years has revealed that adverse experiences associated with environmental enrichment are able to induce alteration in epigenetic marks. Hence, further investigation is required to answer important questions concerning epigenetic regulation upon exposure to an enriched environment, and the impact of such epigenetic regulation. Epigenetic alterations such as DNA methylation and histone modification are among the most discussed mechanisms mediated by adverse environmental enrichment, which includes childhood abuse, maternal separation, chronic stress, etc. Moreover, the regulation of noncoding RNAs, such as miRNA, has emerged as a potential mediator of epigenetic effects that may change the transcriptional activity of genes associated with the nervous system after adverse stress exposure. In some cases, the epigenetic marks mentioned might interact with each other to regulate the transcriptional activity. For instance, low levels of histone acetylation may induce DNA methylation, which are likely to result in permanent gene silencing. However, despite our current understandings on behavioral epigenetics, epigenetic communication research merits further investigation.
Furthermore, germline-mediated epigenetic inheritance, subsequent effects on somatic cells, and the germline inheritance model are proposed and explained in the review. For instance, our review concludes that epigenetic inheritance models provide a potential pathway for these epigenetic marks to be passed down to the offspring and that there is a possible restriction of epigenetic inheritance due to reprogramming events. In addition, current observations of behavioral epigenetics are specific to certain stress response loci, making it unclear whether there is a direct impact of enriched environment on genome-wide epigenetic regulation. Overall, researchers are far from deciphering the model of epigenetic inheritance because the current scope of epigenetic research is not comprehensive enough.
Another promising area of behavioral epigenetic is the establishment of transgenerational epigenetic inheritance. Epigenetic transgenerational inheritance in behavior occurs when similar behaviors shown in both parents and offspring sustain two generations and beyond in absence of direct environmental exposure. However, there are some limitations in conducting such interdisciplinary research. For instance, standardized guidelines for experimental design is needed to determine the mechanism underlying epigenetic inheritance, as most of the inducing stimuli lack consistency, leading to incoherent results. In addition, sex-specific epigenetic inheritance models and the potential reversibility of epigenetic information need further investigation to narrow or bridge the gap in our understanding of epigenetic transgenerational inheritance. Understanding the communication between epigenetic marks and genetic effects is also crucial as most of epigenetic alterations occur differently depending on the DNA sequences. In conclusion, further work such as long-term experiments and the development of advanced technologies is required to understand this topic. Hopefully in-depth research can lead to breakthroughs in the epigenetic field.