Biological correlates of altered circadian rhythms, autonomic functions and sleep problems in autism spectrum disorder

Abstract

Autism spectrum disorder (ASD) is a neurodevelopmental condition characterized by a complex and multifaceted neurobehavioral syndrome. In the last decades, several studies highlighted an increased prevalence of sleep problems in ASD, which would be associated with autonomic system and circadian rhythm disruption. The present review aimed to summarize the available literature about sleep problems in ASD subjects and about the possible biological factors implicated in circadian rhythm and autonomic system deregulation in this population, as well as possible therapeutic approaches. Shared biological underpinnings between ASD symptoms and altered circadian rhythms/autonomic functions are also discussed. Studies on sleep showed how ASD subjects typically report more problems regarding insufficient sleep time, bedtime resistance and reduced sleep pressure. A link between sleep difficulties and irritability, deficits in social skills and behavioral problems was also highlighted. Among the mechanisms implicated, alteration in genes related to circadian rhythms, such as CLOCK genes, and in melatonin levels were reported. ASD subjects also showed altered hypothalamic pituitary adrenal (HPA) axis and autonomic functions, generally with a tendency towards hyperarousal and hyper sympathetic state. Intriguingly, some of these biological alterations in ASD individuals were not associated only with sleep problems but also with more autism-specific clusters of symptoms, such as communication impairment or repetitive behaviors Although among the available treatments melatonin showed promising results, pharmacological studies for sleep problems in ASD need to follow more standardized protocols to reach more repeatable and reliable results. Further research should investigate the issue of sleep problems in ASD in a broader perspective, taking into account shared pathophysiological mechanisms for core and associated symptoms of ASD.

Introduction

The center of circadian rhythms regulation can be identified in the hypothalamus. It consists of several neuronal populations which produce neuropeptides and neurotransmitters with a crucial role for temperature, metabolic rate, thirst, hunger, sexual behavior, reproduction and emotional responses. The hypothalamus exerts its functions mainly through the productions of neuropeptides, such as the transcription factor orthopedia (OTP), crucial for proper development of diencephalic dopaminergic neurons or the steroidogenic factor 1 (SF-1), which is important for neuronal and structural connectivity. Another transcriptional factor, Sim-1, is implicated in hypothalamus differentiation [1]. Deregulation of autonomic system and of daily activity rhythms often precede underlying pathophysiological changes in the brain of subjects with neurodegenerative diseases, such as Alzheimer disease. Similarly, dysautonomias have been seen in metabolic syndromes, cardiovascular diseases, diabetes and cancer. In the last decades, several studies highlighted the importance of altered circadian rhythms, autonomic functions and sleep quality in ASD, while some authors reported associations between these alterations and specific autistic-like symptoms. Intriguingly, an abnormal connectivity between hypothalamus and amygdala has been associated with inappropriate responses to socially relevant stimuli in autism spectrum disorder (ASD) [2]. In particular, a dysfunction of the endogenous circadian system was hypothesized to be part of the neurodevelopmental alteration at the basis of the autistic condition [3]. Sleep problems are frequent among ASD individuals. Several hypotheses have been proposed for explaining this association, including a deficit in melatonin production, anxiety and arousal at the time of going to bed or specific gene mutations. Sleep disturbances have been related to the severity of ASD symptoms (restrictive and repetitive behaviors, difficulties in reciprocal and social interactions, irritability, etc.). It was recently reported an association between actigraphy-derived sleep efficiency, number of awakenings, anxiety and hyperactivity in ASD children. Other studies showed instead a positive correlation between sleep latency, affective problems and aggressive behavior in this population [4]. Some of the genes responsible of regulating circadian rhythms seem to be mutated or down-regulated in ASD, thus providing a molecular explanation for sleep problems: in this framework, deepening the knowledge about genes implicated in circadian rhythms deregulation could be crucial for the development of eventual biomarkers of sleep functions [5]. Circadian hormones such as melatonin and cortisol were also found to be altered in ASD. ASD individuals seem to show an altered production of melatonin, which, in turn, has been related to sleep difficulties, altered gastrointestinal motility, deficits in behavioral and emotional regulation, sensory protein dysfunction. As in the case of gene deregulation, alterations of melatonin/serotonin pathways could be promising biomarkers for ASD [6]. The increased cortisol levels of ASD people were related to hyperarousal and wake after sleep onset (WASO) duration in subjects with insomnia and in controls. On the other hand, a lower cortisol response was linked to poorer self-reported sleep-in healthy population [7]. Autonomic variables such as cardio-vagal activity and pupil size were also investigated in ASD, highlighting in ASD children a lower cardio-vagal activity as measured by heart rate variability (HRV) and increased sympathetic activity measured by means of urinary vanillylmandelic acid (VMA) [8]. It was pointed out that dysautonomias in ASD may be linked to the presence of repetitive behaviors [9]. Starting from these considerations, two important therapeutic strategies, such as melatonin supplementation and Early Start Denver Model (ESDM), a developmental and behavioral intervention, have been studied in ASD not only for sleep problems but also for improvements in social communications, stereotyped behaviors, rigidity, and anxiety [10]. In this review, we aimed to summarize previous reports about sleep problems in ASD children and adults, and about possible biological factors implicated in the alteration of circadian rhythms and autonomic function in this population, together with possible therapeutic approaches. Eventual shared biological underpinnings between ASD symptoms and altered circadian rhythms/autonomic functions will also be discussed.

Methods

To identify the studies focused on the relationship between ASD and sleep problems, a comprehensive literature review was performed using multiple databases (Pubmed, Web of Science, Scopus). Key words used were “autism spectrum disorder”, “autism”, “neurodevelopmental disorder”, “sleep problem”, “sleep disorder”, “circadian rhythm”, “autonomic nervous system”, “pharmacological therapy”, “pharmacological treatment”, “biological correlates”, “hormone”. Articles were considered eligible if written in English from 1980 to 2022.

Sleep problems in ASD: from childhood to adulthood

Several studies highlighted a link between ASD and sleep problems, reporting specific correlations with some symptoms. Besides questionnaires, methods used for evaluating sleep in children and adolescents with ASD include polysomnography and actigraphy. The first one measures multiple physiological sleep parameters, such as brain waves, eye movements, etc. However, in ASD children, due to sensory sensitivity related problem, systematic desensitization is often required. The actigraph is a portable accelerometer device that can be worn on the wrist or ankle and records different sleep variables such as time of sleep onset, total sleep time, morning waking time and frequency of night-time waking [11]. Actigraphy enables to distinguish ASD poor sleepers from good sleepers, because the first ones would show prolonged initial sleep latency, decreased sleep efficiency and increased number of night awakenings [12].

Noticeably, some studies also highlighted an association between sleep problems and other symptoms in this population. Considering children, Schreck et al. [13] reported a relationship between reduced sleep time and deficits in social skills in this population. Moreover, Sikora et al. [14] found that sleep problems were often associated with both externalizing (such as impulsivity and aggression) and internalizing symptoms (such as anxiety and depression). In a study conducted on hospitalized ASD children, higher scores on the Aberrant Behavior Checklist-Community scale (ABC-C) at admission were associated with fewer minutes slept during the last five nights of hospitalization. Meanwhile, sleep problems seem not to be influenced by the subtype of ASD, or by the degree of cognitive impairment. Goldman et al. [15] compared sleep problems of older children and adolescents with ASD with those of control toddlers and younger children. Participants were divided in four groups: less than 5 years, from 5 to 7, from 7 to 11, more than 11 years. While all groups reported sleep related problems, differences in the kind of problem were reported depending on age: younger children showed greater levels of sleep anxiety, bedtime resistance, parasomnias and night walking, while older children and adolescents more frequently reported short sleep duration, delays in sleep onset and daytime sleepiness [12, 15]. A recent study evaluated the association between sleep problems and autistic traits in toddlers (n = 426). The classification was based on the total Modified Checklist for Autism in toddlers (M-CHAT-JV) scores. The autistic group (n = 26) was composed mainly by males. In terms of sleep problems, three items were more frequently endorsed in the autistic group: “bedtime resistance” (time attended before going to bed), “abnormality in circadian rhythm” (the total sleep time varies from one night to another), “sleepiness outside the naptime”. It has emerged that children aged 3–5 years with more difficult temperaments showed substantially more bedtime resistance than children with less difficult temperaments. Bedtime resistance in children at around 18 months of age may be related to subclinical autistic traits. It has been suggested that children with ASD have a deregulation of sleep homeostasis, resulting in a reduction of sleep pressure [16]. Stressful events, such as home confinement during COVID-19 pandemic, were recently shown to exert an impact on sleep in ASD children. In particular, a study was conducted among Turkish ASD children (n = 46). Children were given different questionnaires, such as the children’s chronotype questionnaire (CCQ), autism behavior checklist (AuBC) and children’s sleep habits questionnaire (CSHQ). Results reported that ASD children had greater sleep problems and altered chronotype during home confinement period than during normal state. The severity of ASD symptoms increased together with the chronotype score and sleep score during home confinement. Interestingly, the level of sleep problems was considered a mediating factor in the relationship between CCQ score and autism symptom severity. Moreover, a significant correlation was found between increased CSHQ and CCQ score and ASD symptoms during the home confinement period [17]. Other authors also hypothesized that neonatal-irritable sleep–wake rhythm may be involved in ASD pathogenesis. Miike et al. [18] conducted an analysis among 177 ASD children and 203 controls. In the ASD group irritable/over-reactive types of sleep–wake rhythms were observed more frequently than in control group. These alterations may cause difficulties in raising the child, due to frequent waking up, difficulties in falling asleep, short sleep hours, continuous crying and grumpiness. Noticeably, the number of mothers who went to bed after midnight during pregnancy was higher in ASD group than in controls. Yavuz-Kodat et al. [19] investigated 52 ASD children aged 3–10 years. Sleep and circadian rest-activity rhythms were evaluated objectively with actigraphy and subjectively with the CSHQ, while the ABC-C was used for assessing behavioral difficulties. A shorter continuous sleep period was reported among children with high irritability compared to those with lower irritability. A similar trend emerged in children with high stereotyped behaviors compared to those with less stereotypies.

Other studies focused instead on ASD adolescents and adults, finding that insomnia, parasomnias and circadian rhythms disorders were common also in this population. Ballester et al. [20] led a study on 41 adults with ASD and intellectual disability (ID), and 51 typically developed adults. They used an ambulatory circadian monitoring recording wrist temperature, motor activity, body position, sleep and light intensity. ASD individuals showed several sleep difficulties, such as low sleep efficiency, prolonged sleep latency, increased number and length of night awakenings, with daily sedentary behavior and increased nocturnal activity. ASD adults had an advanced sleep–wake phase disorder. Sleep and markers of the circadian system were significantly different between adults with both ASD and ID and the control group. Sleep disturbances described for adults with ASD and ID are similar to those for ASD adults without ID. Even elderly ASD individuals seem to report more difficulties to fall asleep than other people. Jovevska et al. [21] enrolled in their studies subjects aged from 15 to 80 years (297 ASD participants and 233 controls). Sleep quality, sleep onset latency, total night sleep and sleep efficiency, as measured by Pittsburgh Sleep Quality Index, were analyzed. Moreover, the authors investigated autistic traits, mental health condition, medication, employment and sex as predictors of sleep quality. Results highlighted lower sleep quality and increased sleep onset latency in the whole ASD group rather than in the control one. ASD subjects also showed poorer sleep quality and longer sleep onset latency than controls of similar age, especially in adulthood and middle age, while no significant difference was found in adolescent and elderly ASD groups with respect to age-matched controls. Predictors seemed to account for the 21% of sleep quality variance in the ASD group, with sex as the strongest predictor (female sex more related to sleep problems). In the comparison group, the strongest predictor was mental health condition, with predictors accounting for 25% of sleep quality variance [21].

Some authors evaluated in adult samples the relationship between sleep problems and other features. Baker et al. [22] investigated sleep–wake rhythms in 36 ASD adults. Participants were assessed through a 14-day sleep–wake diary and 14-day actigraphy. ASD individuals seemed to show sleep patterns associated with circadian rhythm disturbances, although authors highlighted that the role of employment status, comorbid anxiety and depression should be taken into account. Another study from the same group evaluated employment status in ASD adults with sleep problems. A total of 72 individuals were enrolled in the study (36 ASD people, 36 controls). Subjects were assessed by questionnaires and 14-day actigraphy. 20 ASD individuals versus 4 controls met criteria for insomnia and/or circadian rhythm sleep–wake disorders. Adults with ASD who met the criteria for sleep disorders according to the International classification of Sleep-Disorders, Third edition, reported higher scores on the Pittsburgh Sleep Quality Index (indicating worse sleep quality) and were more frequently unemployed when compared with ASD adults without sleep problems. This result highlighted an association between sleep problems and unemployment [22]. Deserno et al. [23] also evaluated the impact of sleep on quality of life among 598 ASD adults. Autistic features, comorbid complaints, daily functioning and socio-demographic characteristics were evaluated by means of questionnaires. Results reported that sleep problems were an important predictor of subjective quality of life, while subjective experience of individual contribution in the society seemed to statistically predict the level of daily activities. The authors stressed how sleep problems may be considered an important treatment target for improving quality of life in ASD individuals.

In conclusion, while the literature seems to have reached a global agreement on the presence of sleep alterations in ASD subjects, during both childhood and adult life, results about the possible relationship between sleep problems and ASD symptoms are more limited and controversial, also due to high heterogeneity in the methodology and in the specific symptoms measured (see Table 1).

Table 1 Main findings about sleep problems in ASD

Biochemical correlates of altered circadian regulation in ASD

Genetic pathways

Several genes have been related to alterations of circadian rhythms. Among them, CLOCK gene was reported to be constitutive in the central nervous system (CNS). It was highlighted that the regulation of the cellular circadian clock would be the result of a particular loop including the transcriptional activators CLOCK and BMAL1, which stimulate the expression of cryptochrome (Cry) and period (Per) genes. The protein product of the genes, after having formed dimers in the cytoplasm represses their own transcription, via the inhibition of CLOCK-BMAL1, while on the other hand they activate BMAL1, by an unknown mechanism. As a result, they contribute to the constitution of a mammalian cell-autonomous oscillator. CLOCK genes are expressed not only in CNS, but also peripherally, in the oral mucosa, skin and blood. The co-expression of the gene in the blood, hippocampus and prefrontal cortex allow hypothesizing that its presence peripherally could be a good biomarker of the gene expression in the brain [24]. The importance of gene CLOCK and its epigenetic modulation in the circadian rhythmicity also emerges from evidences of hypo-methylation of CLOCK promoter, as well as of a higher CLOCK expression in blood cells of nightshift workers [25] CLOCK’s products play an important role in the dopaminergic outputs regulation, a pathway linked to many psychiatric disorders. CLOCK protein controls tyrosine hydroxylase (TH), cholecystokinin (CCK) and many other regulators of dopaminergic transmission. Meanwhile, acetylation of CLOCK is thought to regulate cortisol signaling responses thanks to both the reduction in the binding of glucocorticoid receptors to glucocorticoid response elements and the transcription repression of many glucocorticoid-responsive genes related to hypothalamic–pituitary–adrenal axis [26]. While circadian genes in ASD could be highly polymorphic, a dysfunction of circadian genes may be implied in ASD pathogenesis, or may contribute to its pathophysiology. In this framework, it is worth noting that a decreased expression of CLOCK seems to be present in individuals with ASD and some studies examined the specific role of CLOCK genes in this population. Period circadian regulator 1 (PER1), a CLOCK gene expressed in brain areas which affect sleep–wake transition, was also suggested to be mutated in ASD subjects. This alteration was hypothesized to be involved in the oscillations of the circadian clock in ASD, eventually also by affecting melatonin release [27]. Starting from the consideration that neuroanatomical and cellular changes potentially related to social impairment are particularly evident in the medial prefrontal cortex (mPFC) among ASD individuals, a study investigated the mPFC impaired molecular networks in a valproic acid (VPA) rat model of ASD. They found in the mPFC of this rat model 2 subsets of genes with a different expression: one was involved in circadian rhythm regulation, while the other featured collagen genes acting within the extracellular matrix [28].

Some human studies are also available in this field. Hu et al. [5] enrolled a group of ASD individuals who were divided on the basis of scores reported in the Autism Diagnostic Interview-Revised (ADIR) in three phenotypic groups. Their lymphoblastoid cell lines (LCL) were analyzed by means of microarray analyses, comparing the clinical groups with healthy controls. The authors identified a set of differentially expressed genes in each ASD group as well as a set of genes altered in all ASD groups. In the ASD group associated with higher severity, 15 genes regulating circadian rhythm were reported to be differentially expressed with respect to other groups, including the gene encoding for aralkylamine N-acetyltransferase (AANAT), an enzyme which catalyzes the first step of the biochemical conversion of 5-HT to melatonin, a key regulator hormone of circadian cycle. Noticeably, these genes were also linked to metabolic and neurological alteration typical of ASD. Among the genes reported to show an altered expression in all ASD groups, 20 novel genes associated with androgen sensitivity were revealed, leading authors to hypothesize that testosterone levels may be a risk factor for ASD [5]. More recently, a Japanese study investigated the mutations in CLOCK and related genes among ASD individuals versus controls. Many genetic variants located in the CLOCK gene locus, such as rs3762836, were found exclusively in ASD patients with sleep disorders [26, 29]. In a study on 28 ASD individuals and 23 controls, the coding regions of 18 canonical CLOCK genes and CLOCK-controlled genes were sequenced, reporting that mutations of circadian-relevant genes able to affect gene function were more frequent among ASD subjects than among controls [29]. Finally, a more recent work by Abel et al. [30] investigated the issue from another perspective, searching for possible genetic and biological mechanisms involved in the increased rate of sleep problems in this population. They found several overlapping genes between ASD and sleep problem gene sets, including CACNA1C gene, which is implicated in the regulation of calcium channels and has a circadian expression pattern, supporting circadian entrainment.

It should be noted that research in this field focused on different gene sets, with a lack of confirmation studies: thus it did not allow reaching a proper understanding, nor clarifying whether the association between ASD and altered gene expression related to circadian rhythms would be specific or possibly shared also by other mental disorders. In addition, very limited evidence is available on the association between gene variants and the specific presence of sleep problems in this population. In conclusion, while reports of altered CLOCK gene expression in ASD may suggest possible genetic underpinnings for circadian rhythmicity problems in these subjects, results in this field should be considered as preliminary (see Table 2).

Table 2 Genetic alterations in circadian rhythm of ASD individuals

Role of melatonin

Melatonin is a circadian hormone which crosses the placental barrier and exerts a crucial role in fetal neurodevelopment. It is implicated in circadian physiology, and specifically in the sleep wake and core body temperature rhythm. It is secreted by pineal gland cells thanks to synchronization of these cells with 24-h day/night cycle made by photosensitive ganglion cells of the retina, which perceive environmental light [31]. Melatonin is produced by pineal gland during the night, starting from serotonin (5-HT). The conversion of 5-HT in melatonin features two enzymatic steps and the intermediate N-acetylserotonin (NAS) [32]. From a pharmacokinetic point of view, melatonin is subject to hepatic metabolism: its poor adsorption and substantial hepatic first pass metabolism explain the low bioavailability of this molecule. Melatonin is also metabolized in 6-sulfatoxy-melatonin and then excreted in the urine [33,34,35]. While a low amplitude and a possible delayed melatonin rhythm have been associated with increased sleep problems, melatonin deregulation has been supposed to be one of the main responsible for alteration of circadian rhythms in ASD. On the other hand, aberrant phase cycle of melatonin has been related to seizures and electroencephalogram (EEG) discrepancies in ASD individuals [3].

In a systematic review, Rossignol et al. [36] summarized results from 35 studies about melatonin in ASD. The authors concluded that levels of melatonin or melatonin derivatives were below average in individuals with ASD compared with controls. Melatonin pathway was reported to be altered in several ASD subjects, seeming also to correlate with ASD symptoms. In particular, abnormalities in genes involved in melatonin production and receptor function were reported, while some studies highlighted lower melatonin or melatonin metabolite concentrations, as well as altered melatonin circadian rhythm in ASD individuals when compared with healthy subjects or laboratory reference ranges. However, other studies reported instead daytime melatonin levels significantly higher in ASD than in controls [36]. On the basis of previously reported alterations of melatonin and 5-HT (hyperserotonemia) in ASD patients, more recently Pagan et al. [32] proposed that disruption of 5-HT-NAS-melatonin pathway may be a promising biomarker for ASD. To confirm their hypothesis, whole-blood 5-HT, platelets NAS and plasma melatonin were assessed in 278 patients with ASD, 506 first-degree relatives, 416 sex- and age-matched controls. Results confirmed the presence of hyperserotonemia and the deficit in melatonin production in the ASD group with respect to controls, together with increased platelet NAS levels. Relatives’ group reported intermediate levels between ASD and control groups. Interestingly, a strong correlation was found between platelet NAS and plasma melatonin levels in patients with ASD (more than in relatives’ group and controls), probably due to a common factor of deregulation. The melatonin deficit was significantly associated with insomnia. Other studies also focused on relatives of ASD patients, such as the study of Braam et al. [33], which found a significant lower 6-SM excretion rates in mothers of ASD children compared to controls [34, 35].

As reported above, some of the works reviewed by Rossignol et al. [36] investigated genes coding for melatonin receptors such as melatonin receptor 1A (MNTR1A), MTNR1B, GPR50, reporting, in a case, variants in genes MTNR1A and MTNR1B among 2.8% of ASD individuals [37]. Moreover, authors that investigated enzymes involved in hormone synthesis, such as Acetylserotonin O-methyltransferase (ASMT), showed a single-nucleotide polymorphisms in 2.6% of ASD individuals compared with healthy controls [38], while in another study two single-nucleotide polymorphisms in ASMT (rs4446909 and rs5989681) were reported to be more frequent among ASD individuals and were associated with a decrease in ASMT enzymatic activity and lower plasma melatonin levels [39]. Several other studies focused on genetic mutations of factors implied in the melatonin pathway in ASD. Wang et al. [40] also showed new coding mutations of ASMT in 6 ASD individuals, after having investigated the neighboring region in 398 subjects with ASD and in 437 controls. Veatch et al. [41] highlighted in ASD children treated with melatonin an association between sleep onset delay and dysfunctional variants in genes related to melatonin pathway, such as cytochrome (CYP) 1A2. The children who responded to treatment with melatonin showed a strong correlation between their genotypes in ASMT and CYP1A2. Another study focused on the genetic pathway of melatonin deregulation: they enrolled 295 ASD individuals, 362 European controls, and 284 subjects from human genome pluriethnic diversity panel, investigating the presence of rare variants of MTNR1A and MTNR1B. Their findings revealed MTNR1A and MTNR1B mutations linked to an alteration of the receptor functional properties: a significant association was detected between ASD and two variants in affected males [42, 43]. In ASD samples a reduction in pineal gland volume (PGV) was also highlighted [44, 45]. This volume reduction seemed related to primary insomnia, parasomnias and problems waking in the morning [45]. Pagan et al. [44] analyzed post-mortem pineal glands, gut samples and blood platelets of 239 individuals, respectively, for melatonin and 5-HT production and found a reduction of two enzymes related to melatonin synthesis (AANAT and ASMT) in blood platelets from individuals with ASD. However, the disruption of melatonin synthesis was not sufficient to induce 5-HT accumulation. The authors reported a significant correlation between ASMT deficits in blood platelets of ASD people and insomnia. Melatonin, ASMT, AANAT were significantly decreased in ASD individuals when compared with controls. By analyzing gastrointestinal tract of ASD people, no difference was highlighted between ASD individuals and controls with respect to 5-HT content. Melatonin content was instead lower in ASD patients than in controls. NAS was increased in blood platelets of ASD patients who had a disruption of melatonin synthesis. These authors also investigated the presence of ASMT mutations, finding that, in pineal glands of ASD patients, both degradation and synthesis of ASMT were impaired. Some interesting alterations in melatonin pathway were also observed by the same authors in first-degree relatives of ASD individuals. In particular, ASMT activity was significantly reduced in platelets of ASD subjects’ first-degree relatives when compared with controls, even if AANAT activity resulted diminished only in ASD participants and not in the group of relatives. Maruani et al. [45] evaluated PGV with magnetic resonance imaging (MRI) and early morning melatonin levels in 215 individuals (78 subjects with ASD, 90 unaffected relatives, 47 controls). Results highlighted that plasma melatonin levels were significantly different depending on the group and on the PGV. In another study, exons of 7 ASD patients and 17 controls were sequenced for ASMT and two rare variants were identified. 12 out of 290 individuals with ASD carried ASMT micro-duplication, versus 10 out of 324 of controls.

It should be noted that, in ASD subjects, a deregulation of melatonin production was related to sleep difficulties, but also to altered gastrointestinal motility, deficit in behavioral and emotional regulation as well as to sensory protein dysfunction [6]. In this framework, a positive correlation was reported between levels of 6-SM, severity of verbal communication impairment and daytime sleepiness in ASD subjects [46], whereas another work showed an association between lower mean serum melatonin level and abnormal EEG in ASD people [47]. However, other authors did not found instead correlations between Autism Diagnostic Interview and lower ASMT [39].

Finally, Rossignol et al. [36] also reported studies where administration of melatonin to ASD subjects had a beneficial effect in total sleep duration, number of night-time awakenings and sleep onset latency compared to placebo. In particular, some of these studies highlighted in ASD individuals treated with melatonin an improvement in daytime behaviors, such as less behavioral rigidity, ease of management for parents and teacher, better social interaction, fewer temper tantrums, less irritability, more playfulness, better academic performance and increased alertness. In a low number of subjects, minor side-effects related to melatonin treatment were also reported, such as mild morning tiredness, headache, dizziness, but also diarrhea, night-time awakening and excitement before going to bed [36].

Despite the majority of studies in this field seem to have reached an agreement about the presence in ASD population of altered melatonin levels and metabolism, it should be considered that altered melatonin was reported to be associated in this field not only with sleep problems but also with other symptoms. From this perspective, further studies are needed for clarifying if melatonin alteration may be involved in sleep problems directly or indirectly among ASD subjects, and if it should be considered as a causative factor, a consequence or a parallel process with respect to the neurodevelopmental condition (see Table 3).

Table 3 Biological studies on melatonin in ASD

Cortisol levels and hyperarousal

ASD children and adolescents often experience anxiety symptoms like “butterflies”, nausea or sweating, which may cause delays in sleep onset and even insomnia. ASD people have more pre-sleep arousal than typically developed individuals: the main worry is that of have difficulties in falling asleep. Pre-sleep arousal and anxiety were hypothesized to be involved in the difficulty in falling asleep among ASD children and adults [22]. In this framework, it should be noted that a deregulation of hypothalamic pituitary adrenal (HPA) axis and of cortisol production was reported among ASD subjects and it was linked to a deregulation of circadian rhythms [48]. However, while more literature is available about altered HPA in ASD, few studies specifically evaluated the association between these alterations and the actual presence of sleep problems.

Some of the works in this field focused on measuring cortisol/corticosteroids levels in different specimen. In an investigation led by Priya et al. [49], the urinary level of free cortisol, corticosteroids, VMA and 5-hydroxyindole acetic acid were determined among children with low functioning (LFA), medium functioning (MFA) and high functioning (HFA) ASD and in controls. Corticosteroids excretion levels were higher in all the groups of children with ASD than in the control group. An alteration in the pattern of cortisol excretion was observed in children with LFA. The level of 5-hydroxyindole acetic acid was higher in children with LFA and MFA than in the control group. On the basis of these findings, the authors stressed that altered cortisol excretion patterns and high level of corticosteroids in urine may be a consequence of altered HPA function, which may eventually contribute to the pathogenesis of ASD. Cortisol diurnal rhythm seems also to be different in ASD children with respect to controls. Tomarken et al. [50] measured salivary cortisol at four time points during the day in a sample of 36 children with ASD and 27 typically developed peers. A decline in evening levels of cortisol was detected, whereas no difference was reported in the morning levels. In particular, 25% of ASD children had an attenuated linear decline in cortisol level, while the trajectory of the other ones was indistinguishable from that of TD children.

Other variables investigated in this field are the diurnal fluctuation of cortisol (DF) and cortisol after awakening response (CAR). Results about DF in ASD population are controversial. Inconsistencies in the reported findings may be related to the differences in age and ASD severity among the investigated samples. In particular, while some studies reported significant differences between ASD patients and controls, others failed to find an association of CAR or DF alterations with ASD [48, 51,52,53,54]. Some authors also evaluated if the presence of repetitive behaviors in ASD could be linked to cortisol regulation by measuring diurnal salivary cortisol levels in a sample of ASD children (n = 21). Results showed that participants with more severe repetitive behaviors had lower diurnal salivary cortisol than others. As stated by the authors, while repetitive behaviors may be considered a maladaptive strategy for mitigating stress, it is also possible that in this population a down regulation of glucocorticoid system would be associated with prolonged distress [51]. Sharpley et al. [53] specifically focused on ASD females, enrolling 39 girls with high-functioning ASD (diagnosed on the basis of DSM-IV criteria for ASD) for the evaluation of DF and CAR. More than a half of participants showed inverse CAR and more than 14% of the subjects reported inverted DF cortisol concentrations compared to previous models in general population [55]. Three potential sets of predictor factors (physiological, ASD-related, and mood-related) revealed that self-reported depressive symptoms were associated with CAR status, while the most important contributor to the CAR variance was suicidal ideation [53]. Other authors, focusing also on older patients, evaluated it age and puberty may exert a role on cortisol diurnal rhythm in ASD subjects, leading a study a study among ASD individuals of different age ranges (n = 113) and controls. Higher evening levels of cortisol, as well as a blunted diurnal slope, were reported in ASD individuals with respect to controls. According to these authors, evening cortisol levels seemed to increase through development in ASD individuals, being higher in adolescents than in children, whereas morning levels of the hormone were generally lower. The authors also suggested that the increase in cortisol level may be involved in the impaired sleep quality [48].

In a more recent work, Baker et al. [7] specifically investigated the relationship between sleep and cortisol levels, comparing ASD adults (n = 29) and controls (n = 29) by means of questionnaires, actigraphy and salivary samples. ASD participants reported greater reduction in evening cortisol concentrations when compared with controls participants. In the ASD group, poor sleep efficiency and increased wake duration was significantly correlated with cortisol levels measured 1 h before habitual sleep onset time. Moreover, increased sleep onset latency and poorer sleep efficiency was associated with higher subjective arousal in the ASD group. The authors further suggested that low cortisol levels in ASD adults may be linked to a deregulation of HPA axis. During the last years, another study reported alterations in HPA axis in children with ASD, comparing them with subjects with ADHD and typically developed children [54]. A total of 150 children, aged between 6 and 12 years old, were enrolled and distributed into four groups: ADHD, ASD, specific learning disorder (SLP), and typically developed group. Salivary samples were collected at three time points during the day, as well as before and 5 min after an academic performance test and a moral cognition task. ASD children showed lower diurnal salivary alpha-amylase (sAA) secretion, adjusted for age, compared to typically developed ones. Moreover, sAA evening levels resulted significantly higher in ADHD group compared to controls. Authors hypothesized that lower sAA daily output could be related to higher chronic stress in ADHD and ASD children. According to these authors, the academic performance task increased sAA levels in ASD children, while the moral cognition task did not activate the sympathetic nervous system in any group [54].

When considering results from these studies it should be noted that other researches did not find differences between ASD and controls with respect to these parameters. Kidd et al. [56] did not find significant differences between pre-schooling ASD subjects and controls (n = 52) with respect to cortisol and sAA levels at different time of the day. However, differences were reported in level variability, which may eventually exert an impact on circadian rhythmicity [56]. Similarly, Corbett et al. [52] comparing CAR in 46 ASD prepuberal males (aged 8–12 years) and 48 controls did not find any significant difference in CAR between the first and the second group. Further studies are needed for clarifying the actual role of HPA-related alterations in ASD (see Table 4).

Table 4 Cortisol level and hyperarousal in ASD

Autonomic nervous system (ANS) alterations in ASD

Several studies highlighted altered ANS responses in ASD, which may be related to altered circadian rhythms. Alteration of autonomic variables in ASD was reported since early infancy [57]. Noticeably, alterations of ANS have also been implicated in metabolic, sleep, gastrointestinal disorders, as well as in seizures and hormonal dysfunction [58]. On the other hand, the association between autonomic instability and psychiatric symptoms like depression, anxiety and sleep disorders was previously stressed in the literature [57, 58].

In the field of ASD, differences in heart rate (HR) response between ASD children and controls were frequently highlighted, and both HR and respiratory sinus arrhythmia (RSA), which may be a good index of CNS regulation of HR, have been used as indicators of the integrity of the ANS [59]. In this framework, a recent study, evaluating HR and RSA among subjects from 1 to 72 months of age at 8 different time points, found a progressive reduction in HR and increases in RSA. However, the RSA increase was of smaller entity among those subjects who later received a diagnosis of ASD [59]. Bharath et al. [8] evaluated in a sample of ASD children (n = 40) and controls (n = 40) HRV together with urinary levels of VMA. They found that low frequency HRV was more represented in the ASD group, while high frequency HRV was less represented. In addition, urinary VMA concentrations were higher among ASD children. The authors hypothesized that ASD children may have a reduced cardio-vagal activity as measured by HRV, and increased sympathetic activity as measured by urinary VMA. Another study in a smaller sample further highlighted how HR variability (HRV) might be a promising parameter for differentiating ASD children from controls also in a very early stage. The authors enrolled 20 children (6 children with ASD, 14 controls), recording photo plethysmography (PPG) for 4 min in each child during resting state and color stimulus test condition. PPG was used for deriving HRV. Different responses between ASD and controls during resting state were highlighted for specific features of HRV [60]. Thapa et al. [61] compared ASD adults (n = 55) and controls from the general population (n = 55), and reported a significantly higher resting state HR among ASD patients, together with lower square root of mean squared differences of successive R–R intervals (RMSSD) and lower high frequency HRV. No significant difference was reported for low frequency HRV. These authors hypothesized that their results may suggest a general deregulation in resting autonomic activity among ASD adults, characterized by lower parasympathetic activity.

Among other factors linked to ANS alterations, pupil size was also considered as a potential biomarker for ASD. Anderson et al. [62] compared a group of ASD children with age matched controls, reporting that ASD individuals seemed to show a larger tonic pupil size and lower afternoon levels of sAA. The authors also highlighted a little diurnal variation of sAA among ASD children, while among controls a linear increase throughout the day was observed. Globally, they hypothesized that these features may be associated with an over-production of Norepinefrine (NE) [62]. In a recent work, Arora et al. [63] reviewed previous studies that evaluated autonomic arousal by measuring HR, pupillometry and electrodermal activity (EDA) in ASD individuals compared with controls. A general hyperarousal in ASD individuals with respect to controls was reported during resting states, eventually linked to the difficulties showed by ASD people in regulating their arousal.

Considering the issue of sleep problems, some studies specifically focused on the relationship between ANS and sleep alterations in ASD. In particular, Pace et al. [64] investigated HRV during sleep in high-functioning ASD children (n = 19) and controls (n = 19). Two indices of cardiac activity, the high frequency (HF) spectrum and RMSSD, were significantly higher in ASD individuals when compared with other ones. In both groups, lower mean HR values were found during sleep with respect to those registered during wakefulness. However, the ASD group showed a lower decrease in HR during deep sleep despite the presence of a higher parasympathetic tone. Another study analyzed HR and HRV during sleep in ASD. A group of ASD children (n = 21) and controls (n = 23) was evaluated by means of overnight polysomnography. HR was found higher in ASD subjects than in controls during stages N2 and N3 of non-REM sleep and during REM sleep. High frequency oscillations of HRV, which reflect vagal modulation, resulted lower in N3 stage of non-REM sleep and in REM sleep among ASD children. Low frequency to high frequency ratio (LF/HF) was instead higher during REM sleep. These data, together with the higher HR, lead to hypothesize a sympathetic dominance during sleep among ASD children, which would be associated with a decreased vagal influence [65]. Tessier et al. [66] investigated HRV in ASD subjects (17 adults and 13 children) and typically developed ones (16 adults and 13 children) before and after sleep. They divided the sample into four groups: ASD and control adults and ASD and control children. Each group was evaluated for two nights in a sleep laboratory. Normalized values of low frequency, normalized values of high frequency and LF/HF ratio were evaluated during sleep and vigilance state. Morning high frequency values resulted to be lower among ASD adults when compared with controls. LF/HF ratio during REM sleep was instead higher in both ASD adults and controls than in children. These results are in line with a lower parasympathetic activity in ASD individuals in the morning. However, it should be noted that while this study provided a good insight on possible differences in the investigated parameters depending on age, the presence of multiple group comparison analyses in a limited sample size limited the extensibility of their results. More recently, Chong et al. [67] investigated the relationship between EDA, another parameter used as a marker of sympathetic nervous system activation, and sleep deregulation in ASD children (n = 13). Two EDA indices, non-specific skin conductance responses (NSSCR) and tonic skin conductance levels (SCL) were measured. The group of children with deregulated sleep showed fewer NSSCRs and lower SCL in the afternoon.

Finally, other confirmations of the presence of altered ANS response in ASD may came from pharmacological studies: some authors proposed propranolol, a beta-blocker drug, as a possible therapeutic option for dysautonomias and emotional behaviors in ASD. According to the available literature, this drug seems to significantly improve cognitive performances, such as verbal problem solving, social skills, mouth fixation and conversation reciprocity. Moreover, it seems to improve anxiety, behavioral and autonomic deregulation, aggressive, self-injurious, and hypersexual behaviors, including among subjects with acquired brain injury. However, the specific benefits with respect to sleep still need to be further investigated [68].

The above described literature seems to suggest the presence of hyperarousal states in ASD individuals, possibly with a hyper-sympathetic state not properly counterbalanced by the vagal parasympathetic influences [62,63,64,65,66,67]. However, also in this case the available studies are still scant in number and highly heterogeneous in methodology. Moreover, studies specifically focused on the possible relationship between altered ANS function and sleep alteration are even fewer and did not allow reaching a conclusive agreement on this topic (see Table 5).

Table 5 Autonomic nervous system deregulation in ASD

Current pharmacological perspectives for sleep disorders in ASD

To date, the first line treatment for ASD sleep problems, especially in children, are sleep hygiene practices and behavioral interventions [34, 69]. There are some extinction measures which parents of ASD children with sleep problems may follow, such as decreasing afternoon naps, or placing the child in his/her own bed without extraneous stimuli [70]. Associated behavioral manifestations, such as aggressive behaviors, may also be controlled by atypical antipsychotic agents such as risperidone or aripiprazole [71].

Among sleep related pharmacological interventions, administration of melatonin and its agonist drug agomelatine was reported to reduce latency of sleep and the number of awakenings in ASD children. As reported in the previous chapters, in ASD populations a reduction or an alteration in melatonin secretion was highlighted, as well as a reduction in gamma-Aminobutyric acid (GABA)_B receptors in the anterior and posterior cingulate cortex and in the fusiform gyrus cortex [12].

In this framework, Wasdell et al. [72] led a study on 51 children with neurodevelopmental disabilities (age range 2–18 years), who were administered melatonin 20–30 min before the desired bedtime and were invited not to eat during the 2–3 h before the moment of drug assumption. Melatonin resulted to be effective in 47/51 children in improving sleep quality and reducing family stress. Malow et al. [73] performed instead a dose escalation study on 24 children with ASD, who were free of psychotropic medications. The dose response, tolerability and safety were studied during a 14-week open label design. The supplementation of melatonin (1–3 g) showed to improve sleep latency and was proved to be tolerated and safe. A work by Maras et al. [74] has proved 3-month efficacy and safety of a novel pediatric-appropriate prolonged-release melatonin (PedPRM), an easily swallowed formulation shown to be efficacious versus placebo, for long-term treatment (that is up to 52 weeks) of children with ASD who suffered from insomnia. The study also reported an improvement in caregivers’ quality of life. While several studies were conducted in this field, also review and meta-analyses are available: in particular, a meta-synthesis by Cuomo et al. [75], collected data from eight previous systematic reviews based on a total of 38 studies about sleep intervention in ASD. Interventions were categorized in four groups: melatonin supplementation, other pharmacological therapies (such as risperidone, clonidine, benzodiazepines), behavioral interventions, parent education/education programs, alternative therapies. Melatonin supplementation was showed to be the most successful intervention for sleep initiation and maintenance of sleep, while for other parameters, such as night wakings or self-settlings, a strong effect of behavioral intervention and education was reported. Despite the general agreement on the efficacy of melatonin for improving sleep parameters and daytime behaviors in ASD [36], it should be noted that studies which evaluated melatonin efficacy are affected by several limitations, such as small sample sizes, comorbid neurodevelopmental disabilities, heterogeneity in methodologies and in dosages, and that not all the studies clearly confirmed melatonin beneficial effects on all the outcomes [76]. Adverse effects linked to melatonin use were also reported, although generally mild, such as morning tiredness, headache, irritability, diarrhea or night-time awakenings [35, 36].

More limited evidence is available for other pharmacological interventions. Ramelteon, a melatonin agonist and a sleep-cycle regulator used for people who have difficulties in falling asleep, was also hypothesized to have a preventive effect on the onset of sleep disorder after general anesthesia in patients with ASD [77]. A case series is also available, which reported, during a trial on 3 ASD children with Ramalteon, a decrease in sleep problems such as insomnia as well as an improvement of ASD behavioral symptoms [78, 79]. The use of agomelatine was also evaluated in this population. Ballester et al. [80] highlighted among ASD subjects (n = 23) with sleep problems a significant increase of night total sleep time, together with phase correction and improvement of sleep stability, after 3-months of treatment with agomelatine. Other studies reported that donepezil, a reversible inhibitor of acetyl cholinesterase, may have beneficial effects on ASD individuals for improving sleep, communication, eye contact, hyperactivity, expressive and receptive speech [81, 82]. Donepezil combined with choline supplement also showed a sustainable effect on language skills in children with ASD for 6 months after treatment, particularly in subject aged below 10 years [83]. Finally, benzodiazepines, which are commonly used in other neuropsychiatric conditions, were also reported to improve some sleep problems in ASD, even if paradoxical reactions with agitation and hyperactivity were reported [84, 85]. The use of benzodiazepines in ASD might be supported by evidences of a GABA-A alpha 5 receptor deficit in this population [85], while an impairment in GABAergic functions were also highlighted in ASD murine models [86,87,88]. In a case series of 11 ASD children, the administration of 0.5–1 mg of clonazepam showed efficacy in 75% of the subjects [84]. However, the possibility that children with neurodisabilities could show an increased risk of paradoxical reaction prevents many clinicians from prescribing benzodiazepines in ASD children [88]. Another drug which has been object of investigation in this field is clonidine. In a case series of six children with neurodevelopmental disorders, clonidine was proven effective for reducing sleep problems and sleep initiation latency but showed a small impact on recurrent night time and early morning awakenings, mood instability and aggressiveness [89]. Among antidepressants, trazodone was suggested to be useful for treating the advanced sleep phase in ASD individuals [90]. Mirtazapine was reported to be effective on insomnia and symptoms related to sleep deprivation such as irritability and anxiety. In a study led by Posey et al. [91], 25 ASD participants were investigated to test the efficacy and tolerability of mirtazapine. An improvement of several symptoms, including aggressive behaviors, self-injury and irritability, was observed in nine subjects. A further research analyzed the effects of gabapentin administration to ASD children (n = 23). Gabapentin administered 30–45 min before the bedtime resulted to be effective in improving sleep in 18 participants [92, 93]. On the basis of the lower ferritin levels frequently reported among ASD children, an 8-week open-label treatment with oral iron supplementation was also conducted in 33 ASD children with restless sleep. A significant improvement in sleep quality was reported [11, 94].

Globally, it should be noted that while melatonin studies should be regarded in light of the previously mentioned limitations, it appears clear how studies on pharmacological interventions besides melatonin are still very limited in number, featuring also small sample sizes or, for some drugs, being limited to case series, thus increasing the risk of biases and placebo effects. As a result, the above reported research should be considered cautiously until confirmed by further investigations. In addition, another issue in this field lies in the fact that in most of the studies, including those on melatonin, the pharmacological treatment was reported to improve also other symptoms besides sleep problems. Although these data may further support the beneficial effect of the treatment object of investigation, it may be also considered a confounding factor: further studies should clarify if these drugs would exert a specific effect on sleep, or the sleep improvement should be considered a consequence of the improvement of other symptoms (see Table 6).

Table 6 Pharmacological therapies for sleep problems in ASD

Biological correlates of altered circadian rhythm and sleep problems in ASD: broadening the perspective

Studies on sleep showed how ASD subjects typically report more problems regarding bedtime resistance and reduced sleep pressure [16]. A link between sleep difficulties and irritability, deficits in social skills and behavioral problems was also highlighted in ASD children [13, 14, 19]. Generally, an insufficient sleep time seems to affect the quality of life of ASD individuals [23], including an important impact on employment status [22]. As reported in the previous chapters, several mechanisms may be responsible of this feature. The molecular explanation of circadian problems in ASD individuals may be found in mutation, variants or different expression of specific genes which regulate circadian rhythms, such as CLOCK [5, 26, 29], PER1 [27] and CACNA1C [30]. On the other hand, these problems may also be related to an alteration of melatonin levels (and, eventually, of PGV) as reported by several studies [3, 6, 32, 44, 45]. In addition, an impaired circadian regulation of cortisol [48, 56], as well as increased urinary corticosteroids levels were reported in ASD, possibly due to an altered HPA axis function [49]. Besides the development of anxiety and depressive symptoms, the chronic activation of HPA axis seems to exert a negative impact on ASD individuals’ daily life [48, 95] and to inversely correlate with gravity of repetitive behaviors [51]. Studies focused on autonomic functions in ASD globally pointed out the presence of hyperarousal state in ASD individuals [63]. ASD was hypothesized to be associated with the presence of a hyper-sympathetic state, which would be not adequately compensated by vagal parasympathetic influences. The analysis of autonomic indices such as pupil size [62], electrodermal activity [67] and HR highlighted a tendency towards a lower parasympathetic activity during daytime and a sympathetic dominance during sleep [64,65,66].

It should be taken into account that some of the above reported alterations in ASD were not associated only with sleep problems but also with more ASD-specific clusters of symptoms, such as communication impairment or repetitive behaviors [51, 96]. These data suggest that the altered sleep patterns in ASD should be regarded in a broader perspective, considering the possible bidirectional interactions between ASD core symptoms and sleep problems in promoting and maintaining each other. Noticeably, the importance of rhythmicity and synchrony of motor, emotional and interpersonal rhythms for the development of social communication was recently stressed [97]. In addition, ASD children were reported to have difficulties in adapting their changes to the internal or external environment [98]. It would be worth mentioning that epileptic seizures, which are often comorbid with ASD, may be influenced by rapid rhythms of sensory stimuli in the external environment [99]. It was also hypothesized that melatonin may be implicated in the synchronization of the circadian clock network. As a result, interventions which combine melatonin treatment with behavioral measures may be useful for investigating the alterations of this clock [98]. The contribution of melatonin in the ontogenetic establishment of circadian rhythms and in the synchronization of circadian clocks suggests that melatonin may also be implicated in motor, emotional and interpersonal rhythms. An altered melatonin excretion or activity has been related to both timing problems in biological clocks and to the severity of social communication impairment [96, 100, 101]. Maternal variations in cardiac rhythm and in hormone levels were reported to influence the child’s ability to adjust to the environment, while parent–infant synchrony and the construction of shared timing were supposed to be important for social communication development [102]. Some authors also hypothesized that repetitive behaviors may be interpreted as a coping strategy for controlling anxiety and stress response as well as for experiencing more continuity and stabilizing circadian rhythms [51, 103]. In this framework, it should be remembered that melatonin seems to exert an effect also on the improvement of stereotyped behaviors [73]. Both melatonin supplementation and Early Start Denver Model (ESDM) have been studied in ASD not only for sleep problems but also for improving social communication, stereotyped behavior, rigidity, and anxiety. A significant improvement in global communication (including verbal and non-verbal scores) was reported [10], while improvements in rigidity were highlighted on the basis of parents’ and teachers’ comments [104]. Wasdell et al. [72] led an investigation to test the efficacy of melatonin in treatment of delayed sleep phase syndrome and sleep maintenance problems in children with neurodevelopmental disabilities and ASD, highlighting an anxiety reduction based on caregivers’ comment. ANS dysfunctions were also hypothesized to be involved in different ASD symptoms. Condy et al. [9] reported that repetitive behavior severity seems to be predicted by base line RSA and by its reactivity. Low baseline cardiac vagal control predicted less adaptive behaviors in children with and without ASD. ANS studies which analyzed HRV, pupil size and sAA, highlighted that ASD subjects differ from typically developed ones, showing a general reduction in parasympathetic function, while a tendency towards hypoarousal was reported in ASD by studies which measured electrodermal responsiveness [63]. Kaartinen et al. [105] investigated the relationship between autonomic arousal to direct gaze (measured by means of skin conductance responses) and social impairment in ASD children (n = 15) and controls (n = 16). Among ASD children, but not among controls, a positive association was found between impairment in social skills and increased arousal enhancement to direct gaze. In another study, RSA and HR were instead correlated with the accuracy and latency of recognition of facial emotions. Children with ASD were slower in recognizing facial emotions and made more errors in recognizing anger. While ASD subjects showed lower amplitude RSA and higher HR than controls, ASD children with more amplitude of RSA were faster at recognizing emotions. It was proposed that ASD children may live in a hyper-sympathetic state with a reduced ability to calm down. This asset may contribute to their impaired ability to engage in social interactions and to the increased levels of anxiety [106]. Bazelmans et al. [107] highlighted significant correlations between language skills and HR in non-ASD children and between language skills and HRV in ASD ones. These authors also stressed the possible use of autonomic control for detecting individual differences among ASD subjects. On the other hand, another study did not find specific associations between social functioning and autonomic/cortisol response in ASD and non-ASD subjects [108]. Recently, the role of gut microbiota in ASD physiopathology is also acquiring more attention. In particular, gut microbiota and CNS were supposed to influence each other through the “gut brain axis”, which would also involve mucosal immune system, enteric and autonomic nervous system (in particular the vagal branch) [109, 110]. In this framework, a recent investigation further broadened the perspective, stressing the importance of evaluating both microbiota and ANS alterations to properly differentiate specific ASD presentations. The authors evaluated autonomic functions and microbiota composition among ASD individuals comparing them with their non-affected first-degree relatives. A lower interbeat interval was reported in ASD patients, which might be eventually associated with higher sympathetic arousal and lower vagal tone. ASD individuals with sleep disturbances showed more gastrointestinal dysfunctions as well as altered interbeat interval and blood volume pulse, suggesting a possible role of these indices for detecting occult sleep disturbances among ASD subjects. Moreover, ASD subjects with greater microbial richness (increased alpha diversity of the gut flora) reported milder sensory/cognitive and language/communication impairment [110]. However, it should be noted that the use of first-degree relatives as controls should be considered as a major limitation for this study, considering the increasing interest about sub-threshold forms of autism [111,112,113]. Increasing literature is pointing out that autistic traits would be distributed in a continuum in the general population, being highly represented in clinical populations of psychiatric patients with other disorders as well as in first-degree relatives of ASD probands [114,115,116,117,118]. In this latter population, the presence of sub-threshold autistic traits also contributed in shaping the concept of “broad autism phenotype”. In this framework, biological studies in ASD field should include the category of patients’ relatives as an intermediate group and not as a control group [113, 118].

Conclusions

Globally, literature about the issue of sleep disturbance in ASD did not reach a sufficient agreement, especially when considering possible biochemical correlates. Further studies should investigate the link between ASD and sleep disturbances in a broader perspective, which would allow evaluating the shared biological underpinnings between ASD symptomatology and altered circadian rhythms. Moreover, an integrative model would allow clarifying the possible bidirectional interaction between sleep problems and other ASD symptoms: while disrupted sleep may worsen ASD clinical picture, exerting a detrimental effect on cognitive and behavioral features, on the other hand the typical core of ASD symptoms may facilitate the presence and maintenance of sleep problems in this population. In addition, pharmacological studies for sleep problems in ASD need to follow more standardized protocols to reach more repeatable and reliable results. As circadian rhythm alterations are known to exert a detrimental effect on psychopathological conditions, including ASD, improving sleep patterns in ASD subjects may lead to a significant improvement of the clinical outcome and of the psychopathological trajectory, eventually reducing also the risk of developing further disorders in comorbidity [47, 114]. Improve our knowledge in this field may lead to detect potential pathognomonic alterations or even biological markers for ASD, with a consequent improvement of diagnostic strategies. On the other hand, a better understanding of biological bases of ASD symptoms may pave the way for the investigation of further therapeutic targets, eventually improving actual treatment strategies for a population which is still poorly responsive to the currently approved treatments.