Two Brains Are Better Than One To summarize the information that was presented in the previous sections, we would expect that brain development requires the activity of at least two interacting epigenetic factors to coordinate during developmental processes. In nature, several different epigenetic factors work in concert to regulate both developmental plasticity and the proper maturation of tissue-specific neurons. It seems likely that in human brains, multiple epigenetic factors cooperate to establish cell types, in particular neurons that have unique activity. The coordinated action of many different epigenetic factors should ensure that all of the proper cell types are fully developed and fully functional. One way to understand how cooperation of several epigenetic factors might contribute to brain development is to examine the way that these different epigenetic factors cooperate during other developmental processes. For example, the developmental maturation of flowers depends on a network of synergistic interactions of a number of different epigenetic factors (Chandler and Dean [@CR21]). As they mature, flowers change shape from having radial symmetry in an open bud to having bilateral symmetry in a closed bud. This spatial rearrangement of the shape requires a network of interactive signals between several different epigenetic factors. Similar patterns are seen in animal development. For example, developmental remodeling of the *Drosophila* embryo depends on an interaction network between several different epigenetic factors, including Bicoid, Kruppel, Giant, Fushi-Tarazu and Pipsqueak (Struhl [@CR104]). It is likely that the proper activity of several different epigenetic factors is needed to specify mature neuronal types in mammalian brains. While each of these epigenetic factors would be able to specify neurons independently, the interactions between several different epigenetic factors is required to ensure that appropriate neuronal types are generated. The activity of many of these different epigenetic factors is regulated by a network of chemical and mechanical cues. In the case of Bicoid, for example, the pattern of gene expression specified by the maternal gene product *bicoid* is modified by other genes and mechanical cues produced by the eggshell (Struhl [@CR104]). The final result is that the Bicoid pattern of gene expression that has been established during oogenesis is altered during development in response to the cell's surrounding environment. A similar process could also occur during brain development, where different neuronal types are specified in response to a combination of epigenetic factors and environmental cues. It is important to point out that although many of the epigenetic factors that regulate developmental processes require the environment to fully execute their proper function, some epigenetic factors (e.g., nucleosomes and DNA methylation) are independent of the environment. A network of epigenetic factors can, therefore, be established in advance during development. Some epigenetic factors, therefore, might establish the initial pattern of neuronal connections before the neural circuits are exposed to a chemical or mechanical environment. Once an initial neuronal circuit is established, however, this initial pattern of connections could be modified by environmental signals. One obvious candidate for environmental regulation would be neural activity, which is necessary for proper brain development. It will be particularly interesting to determine how epigenetic factors cooperate to direct the activity-dependent maturation of brain circuitry and how their function is regulated by environmental signals. ### The brain has a unique epigenetic factor network that coordinates the development of neuronal connectivity in relation to environmental signals {#Sec8} Epigenetic factors may, therefore, play a unique and important role in determining neuronal morphology and synaptic connectivity by coordinating the activity of several different epigenetic factors in relation to the chemical and mechanical environments in which neural circuits develop. This concept may help to explain how neuronal types that are generated in the brain are able to reconfigure their connections in response to environmental stimuli. For example, as we will discuss later, certain epigenetic factors are capable of coordinating the activity of neural circuits and directing the process of neural circuit formation. These epigenetic factors might also have the unique ability to establish connections that depend on environmental stimuli. Although we know little about how epigenetic factors such as H3K4 methylation and acetyltransferases regulate neuronal connectivity, there is evidence that epigenetic factors could regulate gene expression and, hence, neuronal connectivity in response to an external environmental signal. For example, the activity of the transcriptional coactivator P300 was shown to be controlled by an extracellular stimulus (i.e., acetyl-CoA production) and to in turn regulate gene expression that controls neuronal fate and connectivity in a target cell (i.e., dopaminergic neurons in the ventral tegmental area of the midbrain; Park et al. [@CR77]). This study demonstrated that this epigenetic factor can transduce an extracellular stimulus into a gene expression response that regulates synaptic development. In addition, although we currently know little about how epigenetic factors influence the development of neuronal connectivity, an understanding of epigenetic regulation of neuron gene expression and neuronal connections is likely to be essential for understanding how neural circuits are able to sense and respond to their environment. To understand how epigenetic factors might be involved in neural circuit formation, we must first understand how epigenetic factors could coordinate with the activity of the neurons that form a circuit to establish specific neuronal connections. The following section outlines possible molecular mechanisms by which activity-dependent cues and epigenetic factors coordinate the formation of neuronal circuits in the brain (Fig. [4](#Fig4){ref-type="fig"}). Although the development of neuronal connectivity requires both the coordinated activity of epigenetic factors and neural activity, understanding the activity-dependent development of neuronal connectivity is not the same as explaining how different circuits are generated in the brain. In the brain, the formation of different circuits relies on a combination of epigenetic factors and neural activity, as well as several other molecular and cellular mechanisms, to coordinately regulate the formation of the same circuit. For example, epigenetic factors and neural activity coordinately control the formation of the motor neuron circuit in both vertebrates and invertebrates (Wickramasinghe et al. [@CR125]; Gupta et al. [@CR37]; Gebhart et al. [@CR31]; Wark et al. [@CR122]). Motor neurons in *C. elegans* are located in different segments of the worm, and different motor neuron clusters are differentially specified by the action of a number of different epigenetic factors (Winn et al. [@CR127]). However, the correct formation of motor neuron connectivity also requires neural activity. Neuronal activity in the form of synaptic activity is essential for specifying the same motor neuron clusters (Chiang and Jorgensen [@CR22]). The activity-dependent specification of neuronal connections is not limited to invertebrate species. For example, a related study performed in rat brain explants showed that synaptic activity could also regulate synapse formation in mammals (Burette et al. [@CR12]). Thus, the study of activity-dependent neuronal connectivity formation in model organisms is likely to provide the first hints about how neuronal connectivity is formed in human brains.Fig. 4**Epigenetic factors could coordinate with the activity of neurons to specify neuronal connections**. **a** A schematic diagram of the relationship between epigenetic factors, neural activity and the formation of neuronal connections. Neurons establish neuronal circuits in response to environmental signals. Each neuron must sense its surrounding environment to determine whether it should be part of the neural circuit. One possible mechanism that establishes this network of neurons is the cooperation between epigenetic factors and neural activity. The epigenetic factors would specify the correct number and kind of neurons by regulating the expression of transcription factors that specify neuronal fate. Neurons could, then, respond to environmental signals by modulating the activity of epigenetic factors to create a pattern of gene expression that affects neuronal morphology and connectivity. For example, an extracellular chemical cue might stimulate or inhibit specific receptor proteins in neurons to alter gene expression. The activity-dependent regulation of gene expression could also be used to guide neuronal connectivity formation. For example, epigenetic factors may control the expression of a transcription factor that controls the synaptic development of neurons. Neurons that receive synaptic input from other neurons would express an active form of this transcription factor. Although neuronal activity is important for establishing the correct neuronal connections, cells in the brain also respond to a number of external stimuli. For example, experience-dependent changes in the environment regulate gene expression and neuronal connectivity. These experiences could trigger changes in gene expression in the brain to adapt to new external stimuli. **b** A schematic diagram of the relationship between epigenetic factors and synaptic activity. The regulation of synaptic activity is important for establishing proper neuronal connectivity. Specific epigenetic factors can influence neuronal synaptic activity. For example, epigenetic factors could regulate the expression of presynaptic proteins that are involved in the maintenance of synaptic vesicle pools. Synaptic activity could also regulate the expression of epigenetic factors. For example, synaptic activity could directly influence the transcription of epigenetics factors to help establish proper neuronal connections. Neuronal activity is also involved in learning and memory processes by altering the activity of neurotransmitters. Neurons and other neurons in the brain also produce a number of neurotransmitters to influence other neurons. The regulation of neural activity can also be important in memory processes (Rekart et al. [@CR89]) The study of epigenetic factors that regulate neuronal connectivity also provides a framework for understanding how chemical and mechanical signals regulate activity-dependent neuronal specification and connectivity formation. We know that epigenetic factors must be expressed early in order to specify the correct number and kind of neurons. Thus, changes in the amount or type of specific neurotransmitters or hormones may be able to regulate the expression of different epigenetic factors that are responsible for establishing the proper neuronal connectivity. In addition, a number of environmental factors may also alter gene expression in the brain to guide neuronal connectivity formation. For example, an interesting hypothesis is that specific environmental conditions during pregnancy and early development in utero are able to alter gene expression patterns in the brain to program the later activity-dependent differentiation of neurons (Roper and Renault [@CR92]). Although these environmental factors regulate gene expression to establish the correct developmental patterns