The results of this study demonstrate strikingly different timing and patterns of brain and behavior development in two neurodevelopmental disorders that share overlapping behavioral phenotypes. Notably, brain and behavior changes in FXS are present from the age of 6 months and persist until 24 months. In contrast, ASD characteristics change over the first 2 years of life, starting with a period of relatively typical brain and behavior development at 6 months (which coincides with data in this study as well as previous studies [
1–
4]), demonstrating relatively intact cognition and behavior at this young age in infants who later develop autism. The increased growth rate of the amygdala between 6 and 12 months occurs prior to the emergence of the social deficits that are diagnostic for autism (
2,
4), and well before the typical age of consolidation of the symptoms of ASD into a diagnosable syndrome. This gradual onset of brain and behavior changes in ASD, but not FXS, suggests an age- and disorder-specific pattern of cascading brain changes leading to autism. More specifically, in ASD, we observed increased early postnatal growth rate of the amygdala, a brain structure often implicated in the social aspects of ASD, occurring prior to the consolidation of the defining social deficits in ASD, whereas, in contrast, in FXS we noted an earlier onset of enlargement of the caudate (consistent with findings in other basal ganglia structures), which remained stable over infancy—paralleling the temporal pattern of cognitive deficits in FXS that are evident by 6 months of age and not showing the same degree of decline over time observed in infants who later meet criteria for ASD. The present findings add to the growing evidence that the onset of ASD occurs in the early postnatal period, following a presymptomatic period when the defining features are not yet consolidated into the full, clinically defined syndrome that is typically observed later.
Potential Cellular and Developmental Processes Underlying the Specific Timing of Early Amygdala Overgrowth and Its Relation to Emerging Social Deficits in ASD
Our findings identify the onset and define the subsequent early development of the long-standing observation of enlarged amygdala volume in children with ASD. The specific timing of increased amygdala volume growth rate from 6 to 12 months of age raises the question of what cellular and developmental processes could be occurring that underlie this volume overgrowth. There is postmortem histological evidence in ASD demonstrating an excess of amygdala neurons in early childhood (
23) (an age when amygdala enlargement has already been established in many children with autism), suggesting that early postnatal cellular growth may be dysregulated in individuals with ASD. Excess neuronal production could elicit a cascade of changes, such as increased dendritic arborization, leading to amygdala enlargement, dysfunctional communication between amygdala neurons and other brain regions, and clinical impairment (
50). In the first year of life, dramatic dendritic growth and synapse formation takes place as dendrites establish synaptic connections with neurons. Therefore, more neurons in the amygdala during the first year of life in ASD might lead to an even greater volume and density of dendrites. Indeed, postmortem studies in ASD have reported excessive amygdala neurons (
23) and increased dendritic spine density in the amygdala (
24).
Previous work has shown that neurons in the amygdala mature in an experience-dependent fashion (
52), raising the possibility that the excessive production of amygdala neurons (
23) and increased dendritic spine density (
24) in ASD may be related to altered activity-dependent maturation of amygdala neurons. During the first year of normative brain development, synapses compete for neural growth factors to survive: neural connections that are underactive are pruned in an activity-dependent manner, resulting in a functional network of efficient synaptic connections between neurons (
53). However, emerging evidence suggests that sensory (particularly visual) processing may differ in infants who develop autism (
1,
17,
54), during the early postnatal period when synaptic pruning is driven by sensory input. If there are excess neurons (
23), and dendrites grow in a dysregulated manner by not undergoing efficient activity-dependent synaptic pruning (
24), this could result in amygdala volume enlargement and aberrant signal transmission. This altered growth pattern would then be expected to lead to altered amygdala function and altered connectivity with brain regions that support sensory function, and thus contribute to social deficits. The amygdala is crucial for interpreting salient cues from the environment and coordinates multiple brain regions to detect threat and prepare an appropriate response: the visual system to detect a stimulus, the fusiform gyrus for face processing, and the orbitofrontal cortex for initiating goal-directed action. There are direct anatomical and functional connections between the amygdala, visual system, fusiform gyrus, and orbitofrontal cortex, all of which have been shown to be disrupted in young children with ASD (
55).
During this period of increased amygdala growth from 6 to 12 months of age, we have observed contemporaneous hyperexpansion of surface area in the occipital cortex (
9) and aberrant visual orienting (
54) in this same sample of infants who later developed ASD. Aberrant growth of the amygdala and visual cortex, which is also temporally related to abnormal visual orienting, may represent an aberrant feedback loop between visual and attention regions and the amygdala. It is possible that altered sensory experience leads to hypertrophy of the amygdala and altered amygdala circuitry affecting sensory and social processing systems. The stress model of the amygdala, proposed by McEwen (
56), posits that hypertrophy of the amygdala could be caused by stress. The amygdala stimulates the hypothalamic-pituitary-adrenal (HPA) system and stress response: sensory information enters the basolateral amygdala and is relayed to neurons in the central nucleus, and when neurons in the central nucleus are activated, the stress response ensues. The amygdala regulates the HPA axis by evoking the fight-or-flight response by increasing vigilance or alertness (
56). The amygdala is involved in encoding memories of emotional and painful events, and thus fearful, distressing experiences can form quickly and be long-lasting. Neurons in the amygdala learn to respond to stimuli associated with fear and distress, which aids in recognizing similar fearful stimuli in the future and then evoking a fear response (
57). For example, excessive activity of the amygdala is associated with anxiety disorders and autism (
58–
60). In autism, there is evidence that infants who later develop autism have more reactive temperament (
61), which has been shown to be a precursor to later anxiety (
62). Abnormal visual processing and atypical sensory experience (
54) could result in distress and anxiety in infants who develop autism as their sensory experiences are altered. This early stress could drive hyperresponsiveness of the amygdala, increased HPA activity, and hypertrophy of the amygdala.
The amygdala works in concert with the parietal and occipital association cortices to regulate visual attention, such that disrupted development of this attentional network (
54) and associated visual regions (e.g., middle occipital cortex [
9]) could affect the normal development of important social behaviors (e.g., social eye gaze, interpreting another person’s intentions and movements, and perceiving the spatial relations between oneself and the surrounding environment [
63]). This could contribute to the dysfunction of pivotal skills that are characteristic of social deficits in autism, including eye contact, response to name, and joint attention. For instance, joint attention comprises an important set of skills that support the development of language and social communication (
64): the child needs to make eye contact, follow the eye gaze of another person, and direct and coordinate attention to share a visual experience (
65). Joint attention impairment could relate to deficits in lower-level perceptual processes such as face processing, visual orienting and attention, and interpreting others’ actions (
21). Indeed, Shen et al. (
55) reported that preschool-age children with ASD have altered functional connectivity between the amygdala and areas important for social communication, including the medial prefrontal cortex (mPFC); this altered amygdala-mPFC connectivity was correlated with worse social deficits in ASD (
55). The mPFC regulates emotional responses triggered by the amygdala by providing contextual and experiential input to the amygdala, which in turn uses this information to interpret social stimuli and prepare behavioral and emotional responses (
55). Abnormal function of the amygdala and mPFC in children with ASD has been related to an exaggerated response of the amygdala to faces (
66), alterations in social reward and social motivation (
67), and increased ASD severity (
55). The present study demonstrated that increased amygdala growth in the first year of life in ASD was associated with later social deficits. While this is consistent with the above-mentioned literature on the amygdala’s role in social behavior, this specific brain-behavior finding warrants replication in an independent sample of HL infants, which is currently under way.
Increased amygdala growth in ASD could also be related to neuroinflammation in the first years of life (
68–
70). Microglia play an important role in responding to neuroinflammation and neuronal injury. Resting microglia are characterized by small cell bodies and long thin processes; but when microglia are activated in response to immune challenges, they readily increase in number, their processes thicken, and the cell body swells to as much as two to four times its normal volume, while quickly moving to the site of infection, where microglia interact with neurons to fight infection (
50,
68). Postmortem studies in autism have shown excessive microglia activation, both in number and size of microglia, in the amygdala (
71). It is possible that increased amygdala size in the first year of life reflects 1) microglia becoming activated (and increasing in number and size) in an inflammatory response to overproliferation of amygdala neurons (
23), both of which would contribute to early amygdala enlargement; or 2) increased number and size of microglial cells in a secondary response to some other postnatal neuroinflammatory insult (
68–
70).
What Could Underlie Caudate Enlargement in FXS and Its Relationship to Repetitive Behaviors?
The
FMR1 mutation responsible for FXS causes diminished production of the protein FMRP expressed in neurons. FMRP binds to various mRNAs and plays a vital role in neural development and synaptic plasticity (
27–
30). Stereological analyses have shown that FMRP is highly expressed in the caudate and other basal ganglia structures and involves multiple neuronal processes: the nucleus, cytoplasm, dendrites, and dendritic spines (
31). Disruption of FMRP could result in any number of prenatal and early postnatal events leading to increased caudate volume: increased neuron number, increased neuron size, decreased cell packing density, increased neuropil, reduced synaptic pruning, and greater volume and density of dendritic spines.
Previous research has identified a circuit-specific mechanism for repetitive behavior (
72) whereby the D
1 receptor regulates a specific loop between the caudate, globus pallidus, thalamus, and cortex that can increase repetitive behaviors. Excitatory/inhibitory activity imbalance in these regions (i.e., increased activity of excitatory neurons, decreased activity of inhibitory neurons) can lead to sensory dysfunction. For example, imbalance in activity between the direct and indirect pathways of the basal ganglia system—either reduced D
2 activity (leading to reduced inhibition) or increased D
1 activity (leading to increased excitation)—leads to increased excitability and greater repetitive behaviors (
73). Cortico-striatal-thalamo-cortical circuits underlie behavioral features of many neurodevelopmental disorders, including motor stereotypies, compulsive and ritualistic behaviors, atypical reward processing, and sensory dysfunction (
74,
75). The work of Ting and Feng (
76) demonstrates that specific circuits and behavior domains have different critical periods, and therefore specific windows for interventions during periods of plasticity. For example, the striatal circuit described above has greater plasticity and a longer and later therapeutic window for treating repetitive behaviors, compared to social behaviors. This raises the potential that identifying the pathophysiological mechanisms underlying caudate enlargement could lead to targeted treatments for repetitive behaviors in FXS.