The Effects of Preoperative Intranasal Administration of Dexmedetomidine and Esketamine on Negative Postoperative Behavioral Changes in Children With Autism Spectrum Disorder
Phase: NA
Conditions: Autism Spectrum Disorder
Sponsor: Sun Fei

Version: V2.0 | Date: May 12,2025
Clinical Research Protocol
Title of the Study: Effects of Preoperative Nasal Sedation with
Dexmedetomidine and Eszopiclone on Postoperative
Behavioral Changes in Children with Autism
Spectrum Disorder
Source of the project:Horizontal Project
Project Leader: Zhang Li
Project contact: Sun Fei
Contact number: 18100619994
Study period: December 2025 to November 2027
Version: V2.0
Version date: May 12,2025

Version: V2.0 | Date: May 12,2025
1. Basis for the Proposal (This section primarily outlines the background of the
study, including domestic and international research advancements, the novelty of the
study, and the prominent clinical and scientific issues addressed. It proposes research
hypotheses and includes a list of key references. If the study involves drug research,
please specify whether the indicated indications are within the approved scope of use
by the National Medical Products Administration (NMPA).)
1.1 Background of the Study, Domestic and International Research Progress, and
Research Hypotheses
Autism Spectrum Disorder (ASD) is a complex neurodevelopmental disorder
with clinical phenotypes encompassing various subtypes, including typical autism,
pervasive developmental disorder (PDD), and Asperger syndrome. The core
symptoms of ASD primarily manifest as social interaction impairments, language
communication deficits, and restricted, repetitive patterns of behavior and interests.
ASD is also associated with numerous non-core symptoms, including anxiety,
irritability,
aggression,
attention
deficit
and
hyperactivity
disorder
(ADHD),
depression,
epilepsy,
gastrointestinal
and
immune
dysfunction,
metabolic
abnormalities, and sleep disorders [1].According to the latest epidemiological survey
data from the World Health Organization (WHO), the global prevalence of autism
spectrum disorder (ASD) in children exceeds 1%, with a sustained upward trend in
recent years [2]. This phenomenon may be associated with multiple factors, including
revisions in diagnostic criteria, increased public awareness, and changes in
environmental factors [3].
Compared to normally developing peers, children with ASD may face more
complex medical conditions and more frequent healthcare system visits [4]. Due to
their
inherent
social
communication
impairments
and
sensory
processing
abnormalities, they struggle to adapt to unfamiliar environments and exhibit
heightened sensitivity to external stimuli, posing unique challenges in seeking
medical services and undergoing surgical procedures [5]. Perioperative adverse
experiences can trigger significant physiological and psychological stress responses,
with the incidence of Negative Postoperative Behavioural Changes (NPOBCs) being

Version: V2.0 | Date: May 12,2025
markedly higher than that in healthy age-matched populations. This disparity holds
substantial clinical implications for postoperative recovery and long-term quality of
life [6].
NPOBCs (Non-Prominent Post-Anesthesia Complications) are one of the
significant
complications
in
pediatric
anesthesia,
exhibiting
distinct
biphasic
characteristics in their onset: the acute phase (post-anesthesia recovery period)
primarily manifests as postoperative delirium (ED), characterized by environmental
disorientation and time-space cognitive disturbances; the long-term phase (several
weeks
to months postoperatively)
presents
with
multidimensional
behavioral
abnormalities, including but not limited to sleep rhythm disorders, altered eating
patterns, emotional regulation disturbances (irritability/anxiety states), iatrogenic
phobias (e.g., white coat phobia/nightophobia), and reduced treatment compliance,
forming a composite behavioral phenotype.Epidemiological data indicate that
approximately 50% of pediatric patients undergoing general anesthesia experience
NPOBCs [7], with a significant positive correlation observed between this occurrence
and adverse perioperative outcomes such as prolonged hospital stays and increased
medical costs.More importantly, if these behavioral changes persist without timely
intervention, they may negatively impact children's emotional regulation and
cognitive development, leading to reduced medical compliance and increased
frequency of visits, thereby causing sustained harm to the physical and mental health
of affected children [8].
Studies have shown that mutations in ASD-related genes (e.g., SHANK3,
CNTNAP2) lead to abnormal synaptic pruning and excitatory/inhibitory imbalance,
which may enhance the impact of perioperative stress on neural circuits [9,10].
Additionally, alterations in the neurotransmitter system of ASD children, particularly
metabolic abnormalities in the dopaminergic and serotoninergic systems, have been
widely reported. These signaling disorders may be associated
with clinical
manifestations such as postoperative anxiety and aggressive behavior [11,12].
Furthermore, ASD children exhibit abnormal pain perception. Clinical evidence
indicates that developmental abnormalities in the dorsal root ganglion (DRG) alter the

Version: V2.0 | Date: May 12,2025
function of pain conduction pathways, resulting in atypical postoperative pain
expression and consequently affecting the timeliness of clinical interventions [13].
Finally, ASD patients demonstrate dysfunction of the hypothalamic-pituitary-adrenal
(HPA) axis, characterized by increased cortisol secretion and delayed ACTH response.
HPA axis dysfunction is also correlated with disease severity [14].As the severity of
autism spectrum disorder (ASD) increases, baseline cortisol levels significantly
decrease, ACTH levels rise, and the cortisol response to ACTH stimulation diminishes
[15]. Dysfunction of the HPA axis mediates neurodevelopmental abnormalities,
leading to impaired executive function and reduced emotional regulation, thereby
forming a vicious cycle of "medical stress-behavioral abnormalities-health damage,"
ultimately significantly reducing the quality of life in affected children. These
neurobiological abnormalities (including synaptic dysfunction, neurotransmitter
disorders, and abnormal pain perception) interact pathologically with the inherent
maladaptation to medical environments in ASD children, resulting in a higher
incidence of postoperative non-postoperative brain contusions (NPOBCs) and more
severe symptom manifestations in this population. Although postoperative NPOBCs
in ASD children have garnered widespread attention from clinicians worldwide,
current research remains limited, and systematic and effective prevention and
management strategies are still lacking [16].
Perioperative management for children with autism spectrum disorder (ASD)
aims to control preoperative anxiety, fear, and other emotions, reduce sympathetic
nervous system excitability, and minimize stress responses. Special emphasis is
placed on achieving comfortable care throughout all perioperative stages. Therefore,
effective preoperative sedation is of significant importance in improving the
perioperative experience for ASD children. Certain sedative agents, such as
dexmedetomidine and esketamine, have demonstrated protective effects in the
treatment of psychiatric disorders like depression, and are promising as potential
sedative therapies to reduce the risk of postoperative non-painful bowel movements
(NPOBCs) in ASD populations.

Version: V2.0 | Date: May 12,2025
Dextromethorphan is a highly selective α2-adrenergic receptor agonist that has
been widely used in pediatric anesthesia practice due to its excellent sedative,
analgesic, and anxiolytic effects without affecting respiration. In the ASD population,
Ahmend et al. [17] reported the use of dextromethorphan intravenously during
pediatric magnetic resonance imaging (MRI), demonstrating that ASD children
required the same dose of dextromethorphan as healthy children of the same age, with
only a slightly longer recovery period and no other complications observed. Lubitsch
et al. [18] similarly reported the successful use of dextromethorphan combined with
midazolam for sedation during pediatric MRI, with its safety and efficacy partially
validated. However, no existing studies have reported outcomes of non-postoperative
behavioral complications (NPOBCs) in ASD children. Lee-Archer et al. [8] found that
in the general pediatric population, dextromethorphan reduced the incidence of
postoperative delirium and agitation, and continuous intraoperative infusion of
dextromethorphan decreased the risk of NPOBCs within 28 days postoperatively. It
remains unclear whether dextromethorphan provides the same or even superior
protection against postoperative adverse behaviors in ASD children, which requires
further investigation through well-designed, large-sample, randomized controlled
clinical trials with higher evidence levels.
As a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist,
eszopiclone is characterized by rapid onset and significant analgesic effects. In recent
years, its application in the treatment of mental disorders has become increasingly
widespread, with preliminary studies demonstrating its efficacy in alleviating anxiety
symptoms and improving mood states. A prospective observational study conducted
by Yanling Liao et al. [19] revealed that among non-ASD children aged 2-5 years who
underwent
tonsillectomy,
the
combination
therapy
of
eszopiclone
and
dexmedetomidine showed significant advantages over monotherapy: compared to
eszopiclone alone, the combination therapy more effectively reduced the incidence of
postoperative
delirium;
compared
to
dexmedetomidine
alone,
it
significantly
decreased the number of postoperative NPOBCs on day 7. Animal experimental

Version: V2.0 | Date: May 12,2025
studies have shown that downregulation of methyl-CpG-binding protein 2 (MeCP2)
gene expression in the hippocampus can induce behavioral abnormalities associated
with autism-like traits in rats [20]. The MeCP2 gene is one of the few genes with a
clear causal relationship in the pathogenesis of ASD, and eszopiclone intervention can
effectively prevent the occurrence of these abnormal behaviors [21].In clinical case
studies, Miriam Olivola et al. [22] reported a successful case of nasal ethylchloramine
treatment for ASD combined with treatment-resistant depression, suggesting its
potential safety and efficacy. However, only a limited number of case reports have
indicated this potential, and large-scale randomized controlled trials are still lacking to
verify the impact of ethylchloramine on postoperative NPOBCs in ASD children.
Despite the relatively low prevalence of ASD, this population exhibits a higher
risk of postoperative NPOBCs due to their specific psychological-behavioral
disorders, making the exploration of perioperative intervention strategies for ASD
children particularly crucial. Dexmedetomidine and eszopiclone, as commonly used
sedatives in pediatric anesthesia, have been well-established in terms of their sedative
safety and efficacy. However, their impact on postoperative NPOBCs remains unclear.
Existing studies are limited by insufficient sample sizes and single-arm designs,
necessitating high-quality, multicenter randomized controlled clinical trials to provide
more reliable evidence-based medical data.
In conclusion, this study aims to systematically evaluate the effects of
dexmedetomidine and eszopiclone nasal spray on postoperative NPOBCs in children
with ASD through an international multicenter, prospective, randomized, controlled
trial. The study hypothesizes that dexmedetomidine and eszopiclone nasal spray can
reduce the incidence of NPOBCs in ASD children postoperatively. The findings will
provide critical reference for clinicians in decision-making within relevant fields,
while also contributing to the optimization of perioperative management strategies for
children with autism spectrum disorder (ASD), improving patients' perioperative
experience, enhancing postoperative recovery quality, and thereby alleviating the
burden on families and society. This study holds significant clinical and societal
value.

Version: V2.0 | Date: May 12,2025
1.2 Off-label Use
1.2.1 Domestic Levometomidine Package Insert
Dextromethorphan Hydrochloride Injection, 2 mL: 0.2 mg, Nanjing Zhengda
Tianqing Pharmaceutical Co., Ltd.
Pediatric Medication
The safety and efficacy of this product in pediatric patients under 18 years
of age are not well established. Therefore, this product is not recommended for
use in this population.
1.2.2 Off-label Use in This Study
Dextromethorphan 2.0 μ g/kg administered via nasal route was selected for
preoperative sedation in children based on the "China Anesthesia Guidelines and
Expert Consensus" and the current clinical application status of dextromethorphan
both
domestically
and
internationally,
considering
the
applicable
population,
indications, route of administration, and dosage.
1.2.3 Expert Consensus on Off-Label Use
In Chapter 32 of the "China Anesthesia Guidelines and Expert Consensus" (2017
Edition) (the latest consensus), titled "Expert Consensus on Off-Label Use of
Common Pediatric Anesthetic Drugs," there is clear evidence that dexmedetomidine is
superior to midazolam as a preoperative medication for children. Dexmedetomidine
can effectively reduce the tension and anxiety of separated children and parents,
decrease delirium and post-anesthetic tremors, reduce postoperative agitation,
improve perioperative analgesia, and reduce the use of postoperative opioids.
Dexmedetomidine at 2.0 μg/kg has a better sedative effect than 1.0 μg/kg; intranasal
administration is more effective than oral administration (Evidence Level 1a,2a, or
2b). In this study, the use of dexmedetomidine falls within the recommended scope of
the "China Anesthesia Guidelines and Expert Consensus."
1.2.4 Current Research Status at Home and Abroad
Pediatric
patients
often
experience
anxiety
and
tension
before
surgery.

Version: V2.0 | Date: May 12,2025
Preoperative sedatives can provide sedation to alleviate preoperative anxiety, thereby
reducing pre-induction crying and improving participation and cooperation. In recent
years, preoperative sedation for pediatric patients has been widely recommended and
used. Although the prescribing information for dexmedetomidine does not include
specific dosing instructions for pediatric use, with the advancement of numerous
clinical drug trials, dexmedetomidine has increasingly been recommended as a
preoperative sedative for pediatric patients.
Anesthetic drugs, particularly those used in children during growth and
development,
exhibit
inconsistent
pharmacokinetics
and
pharmacodynamics,
necessitating specialized administration. Additionally, the inherent conflict between
pediatric populations and drug clinical trials has led to widespread delays in updating
drug labeling for pediatric anesthesia. As a preoperative sedative for pediatric patients,
dexmedetomidine has demonstrated a gradual increase in clinical use. Recent studies
on the sedative effects of dexmedetomidine in preoperative settings have effectively
validated its clinical efficacy and safety. Nasal administration of dexmedetomidine
provides sedative and anxiolytic effects without causing respiratory depression, and
the absence of nasal discomfort in pediatric patients ensures high patient acceptance.
In
recent
years,
numerous
clinical
studies
on
the
preoperative
nasal
administration
of
dexmedetomidine
have
been
reported
domestically
and
internationally. We have
systematically
reviewed
five
systematic
evaluations
published over the past decade to analyze literature on this topic [23-27]. Two of these
systematic reviews and meta-analyses concluded that nasal dexmedetomidine is a safe
and effective alternative to other drugs for pediatric preoperative sedation. In Kim et
al. 's systematic review, 11 randomized controlled trials (RCTs) were selected,
demonstrating superior sedative effects in children compared to oral benzodiazepines.
Jun et al.' s systematic review included 13 RCTs, which showed that nasal
dexmedetomidine provided effective preoperative sedation in children without serious
adverse events. Jun et al. also reported that nasal dexmedetomidine achieved better
sedation in children during separation from parents compared to other drugs and
reduced the need for postoperative analgesics. Extensive research indicates that

Version: V2.0 | Date: May 12,2025
preoperative
nasal
dexmedetomidine
administration
is
becoming
increasingly
common and appears to be a superior choice among preoperative sedatives. Nasal
administration eliminates the need for intravenous access, and the drug is absorbed
through the nasal mucosa, avoiding hepatic first-pass metabolism. Theoretically,
plasma concentrations are comparable to intravenous doses. As an α 2-adrenergic
agonist, dexmedetomidine is colorless, odorless, and does not cause respiratory
depression. It exerts its sedative effect by reducing norepinephrine release in the raphe
nucleus, producing a sedative state similar to natural non-REM sleep, thereby
ensuring high safety and efficacy.
In summary, based on the actual application of dexmedetomidine in clinical
practice, adhering to the principle of being most beneficial to patients, and
considering the current research status both domestically and internationally, the
clinical efficacy and safety of off-label use of dexmedetomidine are fully evaluated.
The
use
of
dexmedetomidine
is
reasonably
applied
without
exceeding
the
recommended scope of use in the "China Anesthesiology Guidelines and Expert
Consensus". A safety committee is established to monitor sedation-related adverse
reactions in real time during the study period, and the study may be terminated if
necessary.
[1].
[1].
Hodis B, Mughal S, Saadabadi A. Autism Spectrum Disorder. StatPearls. Treasure
Island (FL) ineligible companies. Disclosure: Saba Mughal declares no relevant financial
relationships with ineligible companies. Disclosure: Abdolreza Saadabadi declares no relevant
financial relationships with ineligible companies.: StatPearls Publishing
Copyright © 2025, StatPearls Publishing LLC., 2025
[2].
[2].
Issac A, Halemani K, Shetty AP, et al. The global prevalence of autism spectrum
disorder in children: a systematic review and meta-analysis. Osong public health and research
perspectives, 2025
[3].
[3].
Hirota T, King BH. Autism Spectrum Disorder: A Review. Jama, 2023, 329: 157-168
[4].
[4].
Zablotsky B, Rast J, Bramlett MD, et al. Health Care Transition Planning Among Youth
with ASD and Other Mental, Behavioral, and Developmental Disorders. Maternal and child
health journal, 2020, 24: 796-804
[5].
[5].
Chiri G, Warfield ME. Unmet need and problems accessing core health care services
for children with autism spectrum disorder. Maternal and child health journal, 2012, 16:
1081-1091

Version: V2.0 | Date: May 12,2025
[6].
[6].
Elliott AB, Holley AL, Ross AC, et al. A prospective study comparing perioperative
anxiety and posthospital behavior in children with autism spectrum disorder vs typically
developing children undergoing outpatient surgery. Paediatric anaesthesia, 2018, 28: 142-148
[7].
[7].
Yuki K, Daaboul DG. Postoperative maladaptive behavioral changes in children.
Middle East journal of anaesthesiology, 2011, 21: 183-189
[8].
[8].
Lee-Archer PF, von Ungern-Sternberg BS, Reade M, et al. The effect of
dexmedetomidine on postoperative behaviour change in children: a randomised controlled
trial. Anaesthesia, 2020, 75: 1461-1468
[9].
[9].
Gandhi T, Canepa CR, Adeyelu TT, et al. Neuroanatomical Alterations in the
CNTNAP2 Mouse Model of Autism Spectrum Disorder. Brain sciences, 2023, 13
[10].
[10]. Gao R, Penzes P. Common mechanisms of excitatory and inhibitory imbalance in
schizophrenia and autism spectrum disorders. Current molecular medicine, 2015, 15: 146-167
[11].
[11].
Antón-Galindo E, Cabana-Domínguez J, Torrico B, et al. The pleiotropic contribution
of genes in dopaminergic and serotonergic pathways to addiction and related behavioral traits.
Frontiers in psychiatry, 2023, 14: 1293663
[12].
[12]. Chavez B, Chavez-Brown M, Sopko MA, Jr., et al. Atypical antipsychotics in children
with pervasive developmental disorders. Paediatric drugs, 2007, 9: 249-266
[13].
[13]. Olena VB, Volodymyr BB, Adrien P, et al. The Current View on the Paradox of Pain in
Autism Spectrum Disorders. Frontiers in psychiatry, 2022
[14].
[14]. Hamza RT, Hewedi DH, Ismail MA. Basal and adrenocorticotropic hormone stimulated
plasma cortisol levels among Egyptian autistic children: relation to disease severity. Italian
journal of pediatrics, 2010, 36: 71
[15].
[15]. Spratt EG, Nicholas JS, Brady KT, et al. Enhanced cortisol response to stress in
children in autism. Journal of autism and developmental disorders, 2012, 42: 75-81
[16].
[16]. Snow SL, Smith IM, Bird SJ, et al. A call to solve the puzzle together by building an
evidence base for perioperative management of children with autism spectrum disorder.
Paediatric anaesthesia, 2016, 26: 772-773
[17].
[17]. Ahmed SS, Unland T, Slaven JE, et al. Successful use of intravenous dexmedetomidine
for magnetic resonance imaging sedation in autistic children. Southern medical journal, 2014,
107: 559-564
[18].
[18]. Lubisch N, Roskos R, Berkenbosch JW. Dexmedetomidine for procedural sedation in
children with autism and other behavior disorders. Pediatric neurology, 2009, 41: 88-94
[19].
[19]. Liao Y, Xie S, Zhuo Y, et al. Intranasal Dexmedetomidine-Esketamine Combination
Premedication versus Monotherapy for Reducing Emergence Delirium and Postoperative
Behavioral Changes in Pediatric Tonsillectomy and/or Adenoidectomy: A Randomized
Controlled Trial. Drug design, development and therapy, 2024, 18: 4693-4703
[20].
[20]. Orefice LL, Mosko JR, Morency DT, et al. Targeting Peripheral Somatosensory
Neurons to Improve Tactile-Related Phenotypes in ASD Models. Cell, 2019, 178:
867-886.e824
[21].
[21]. Choi M, Ko SY, Seo JY, et al. Autistic-like social deficits in hippocampal MeCP2
knockdown rat models are rescued by ketamine. BMB reports, 2022, 55: 238-243
[22].
[22]. Olivola M, Arienti V, Bassetti N, et al. Use of intranasal esketamine in a girl with
treatment-resistant depression and autism spectrum disorders: a case report. EXCLI journal,
2022, 21: 540-543

Version: V2.0 | Date: May 12,2025
[23].
Plambech MZ, Afshari A. Dexmedetomidine in the pediatric population: a review. Minerva
Anestesiol 2015;81:320–32.
[24].
AlSarheed MA. Intranasal sedatives in pediatric dentistry. Saudi Med J 2016;37:948–56.
[25].
Kim HJ, Shin WJ, Park S, Ahn HS, Oh JH. The sedative effects of the intranasal
administration of dexmedetomidine in children undergoing surgeries compared to other
sedation methods: A systematic review and meta-analysis. J Clin Anes-th 2017;38:33–9.
[26].
Jun JH, Kim KN, Kim JY, Song SM. The effects of intra-nasal dexmedetomidine
premedication in children: a systematic review and meta-analysis. Can J Anaesth 2017;64:947
–61.
[27].
Lewis, J. and C.R. Bailey, Intranasal dexmedetomidine for sedation in children; a review. J
Perioper Pract, 2020. 30(6): p. 170-175.
II. Research Content
2.1 Research Objectives and Significance
Primary objective: To compare the effects of right 美托咪啶and eszopiclone
nasal spray versus saline control on NPOBCs in children with ASD.
Secondary objective: To investigate the correlation between postoperative EEG
characteristics and NPOBCs in children with ASD.

Version: V2.0 | Date: May 12,2025
2.2 Research Content and Key Issues to be Addressed in the Project
2.2.1 Research Content of the Project
This study planned to enroll 234 pediatric patients undergoing elective surgery
and randomly divide them into three groups: dexmedetomidine nasal spray group
(Group D): receiving dexmedetomidine 2.0 μg/kg nasal spray 30 minutes before
anesthesia induction; ketamine nasal spray group (Group L): receiving ketamine 1.0
mg/kg nasal spray 30 minutes before anesthesia induction; saline control group
(Group C): receiving an equal volume of saline nasal drops 30 minutes before
anesthesia induction. The incidence of NPOBCs at 3 days, 7 days, and 28 days
postoperatively in the three groups of ASD patients was observed, as well as the
correlation between the EEG characteristics of the patients at the time of awakening
and postoperative NPOBCs.
2.2.2 Key Issues to be Addressed
(1) This study aims to systematically evaluate the effects of dexmedetomidine and
eszopiclone nasal spray on postoperative NPOBCs in children with ASD.
The research findings will provide critical reference for clinicians 'decision-making in
relevant fields, while also contributing to the optimization of perioperative
management strategies for children with autism spectrum disorder (ASD), improving
patients' perioperative experience, enhancing postoperative recovery quality, thereby
alleviating the burden on families and society, demonstrating significant clinical and
societal value.
2.3 Study Design
2.3.1 Types of Research Designs and Research Centers
This study employed a dynamic stratified block randomization method with
central randomization. Stratification was performed based on center (8 centers), age
(0-6 years, 7-12 years), and surgical type (head and neck surgery, abdominal surgery,
limb surgery). Within each stratification level, block randomization was conducted by
pre-generating random block sequences of 6 or 9 units and placing them into a

Version: V2.0 | Date: May 12,2025
random sequence pool. When patients were enrolled in a given stratification level, a
block sequence was randomly selected from the central randomization pool to assign
to that level until the block sequence was exhausted. Recruitment was completed once
all block sequences in the pool had been allocated.
This study employed a double-blind design for both investigators and patients.
Approximately 30 minutes prior to surgery, the pediatric patient was admitted to the
anesthesia preparation room, where an anesthesia nurse not involved in the study
received the randomized allocation results from the central system and prepared the
corresponding formulations. The investigational drug was administered via the same
unmarked nasal spray pump, with all formulations diluted to 1 ml before being
injected into sterile blank nasal spray pumps. These pumps were then handed over to
the study intervention implementer, who was unaware of the administered drug. The
intervention was completed and the patient was removed from the study. Subsequent
investigators remained unaware of the group assignments. Postoperative follow-up
was conducted by another independent investigator, and the data analysis was
performed by a statistical expert not involved in the study.
This multicenter, prospective, randomized controlled trial enrolled 234 pediatric
patients undergoing elective surgery from June 2025 to January 2027 at the following
institutions: Children's Hospital of Nanjing Medical University, Zhongda Hospital of
Southeast University, Children's Hospital of Xuzhou Medical University, Wuxi
Children's Hospital of Jiangnan University, Changzhou Children's Hospital of
Nantong University, Pizhou Hospital of Xuzhou Medical University, Dongtai
Hospital of Nantong University (Dongtai People's Hospital), and Suqian Hospital of
Nanjing Drum Tower Hospital Group.
2.3.2 Randomization and Blinding
This study employed a dynamic stratified block randomization method with
central randomization. Stratification was performed based on center (8 centers), age
(0-6 years, 7-12 years), and surgical type (head and neck surgery, abdominal surgery,
limb surgery). Within each stratification level, block randomization was conducted by
pre-generating random block sequences of 6 or 9 units and placing them into a

Version: V2.0 | Date: May 12,2025
random sequence pool. When patients were enrolled in a given stratification level, a
block sequence was randomly selected from the central randomization pool to assign
to that level until the block sequence was exhausted. Recruitment was completed once
all block sequences in the pool had been allocated.
This study employed a double-blind design for both investigators and patients.
Approximately 30 minutes prior to surgery, the pediatric patient was admitted to the
anesthesia preparation room, where an anesthesia nurse not involved in the study
received the randomized allocation results from the central system and prepared the
corresponding formulations. The investigational drug was administered via the same
unmarked nasal spray pump, with all formulations diluted to 1 ml before being
injected into sterile blank nasal spray pumps. These pumps were then handed over to
the study intervention implementer, who was unaware of the administered drug. The
intervention was completed and the patient was removed from the study. Subsequent
investigators remained unaware of the group assignments. Postoperative follow-up
was conducted by another independent investigator, and the data analysis was
performed by a statistical expert not involved in the study.
2.3.3 Sample Size
Based on our preliminary study results, the incidence of adverse behavioral changes at
7 days postoperatively was 50% among autistic children undergoing day surgery. We
hypothesize that dexmedetomidine or eszopiclone can reduce the incidence of these
events by at least 50%, meaning the clinical minimum effective difference is 25%.
Using PASS15.0 software to calculate the sample size with a 1:1:1 ratio, 70 cases per
group would provide 80% power and a two-tailed significance level of 0.05.
Considering a 10% dropout rate, the actual sample size required per group is 78 cases,
with a minimum total sample size of 234 cases.
2.4 Study Subjects
Pediatric patients undergoing elective surgery at participating centers in this
study.
2.5 Research Procedures and Technical Approach
2.5.1 Research Steps

Version: V2.0 | Date: May 12,2025
On the day before surgery, investigators reviewed the surgical application form
to determine eligibility based on the inclusion criteria. During the preoperative visit,
basic patient information was collected, along with any risk factors that might cause
perioperative respiratory adverse events. Informed consent was obtained from the
patient's family members or guardians. All patients were required to fast and abstain
from liquids before surgery: 6-8 hours for solid foods, 4 hours for breast milk, 4 hours
for formula milk, and 2 hours for clear liquids. The patient's basic vital signs were
recorded.
Thirty minutes prior to surgery, the child was escorted by parents into the
anesthesia preparation room. A non-participating anesthesia nurse received the
randomized allocation results from the central system and prepared the corresponding
formulations. The study drug was administered via the same unmarked nasal spray
pump, with all formulations diluted to 1 ml before being injected into sterile blank
nasal spray pumps. These pumps were then handed over to the study intervention
implementer, who was unaware of the administered drugs. The child was withdrawn
from the study immediately after the intervention. Subsequently, continuous
monitoring of the child's non-invasive blood pressure, heart rate, and pulse oximetry
saturation was performed, with records taken every 5 minutes. Additionally, the
child's sedation alertness (OAA/S) score was assessed every 5 minutes. Prior to
entering the room, if the child exhibited severe anxiety or fear, the parents were first
provided with verbal reassurance, along with behavioral interventions such as playing
with cartoons or toys. If symptoms persisted, midazolam was administered for
sedation, with relevant records kept.
Upon admission, routine monitoring of heart rate, pulse oximetry, and
non-invasive blood pressure was performed every 5 minutes. The method of
anesthesia induction was selected by the lead anesthesiologist based on clinical
practice, either via inhalation anesthesia or simple intravenous induction. For
intravenous anesthesia induction, after establishing an open intravenous line, propofol
2-3 mg/kg and fentanyl 2 μg/kg were administered. For inhalation anesthesia

Version: V2.0 | Date: May 12,2025
induction, 8% sevoflurane was inhaled, and after loss of consciousness, the
intravenous line was established, followed by fentanyl 2 μg/kg or sufentanil 0.3 μ
g/kg. All patients received mivaclopramide 0.15 mg/kg or cisatracurium 0.15 mg/kg,
after which a tracheal tube was inserted and connected to an anesthesia machine for
mechanical ventilation. Mechanical ventilation was performed with either
pressure-controlled or volume-controlled settings, with an intraoperative
oxygen-to-air ratio of 1:1 and a flow rate of 2 L/min.
Anesthesia maintenance was achieved through continuous inhalation of 1%
sevoflurane via anesthesia machine, with concurrent intravenous continuous infusion
of propofol at a rate of 4-6 mg·kg⁻ ¹·h⁻ ¹, adjusted according to depth of anesthesia
monitoring. Intraoperative analgesia was determined by the anesthesiologist, with
optional addition of fentanyl or continuous infusion of remifentanil. The tidal volume
was adjusted to maintain PetCO₂
between 35-45 mmHg, with intraoperative AI
monitoring to sustain anesthesia depth within the range of 40-60. Records included
anesthesia and operative duration, fluid intake, and respiratory adverse events during
the induction and maintenance phases.
Postoperative inhalation anesthesia is administered, and the patient is intubated
and transferred to the Post-Anesthesia Care Unit (PACU) by another independent
anesthesiologist. The endotracheal tube is removed after the child regains
consciousness. The PACU stay time, tube removal time, and respiratory adverse
events are recorded. Postoperative pain scores and PAED scores are documented after
the child awakens. If additional sedative or analgesic medications are administered,
the type and dosage are recorded. The patient is returned to the ward once the Steward
score exceeds 4.
2.5.2 Technical Roadmap

Version: V2.0 | Date: May 12,2025
2.6 Evaluation Indicators
①Primary outcome indicators
The incidence of adverse behavioral changes (PHBQ-AS scale) at 7 days
postoperatively.
Secondary outcome indicators
The incidence of postoperative adverse behavioral changes at 3 days and 28 days
after surgery;
③Other outcome indicators
(1) 30-minute post-administration sedation score (MOAA/S scale);
(2) Post-sedation pain score (FLACC score);
(3) Postawake Delirium and Agitation Scale (PAED score);
(4) Electroencephalogram (EEG) characteristics of pediatric patients during surgery
and postoperative recovery;
(5) incidence of other perioperative adverse events such as nausea, vomiting, and
Data collection, follow-up
Group C: 1ml saline
nasal spray
Data organization and statistical
Enrollment of patients meeting the trial
randomized grouping
Declaration of Ethics,
According to the trial
Group L:
Eszopiclone 1.0
Group D:
Dexmedetomidine

Version: V2.0 | Date: May 12,2025
allergic reactions;
(6) Awakening time and postoperative hospitalization duration;
(7) Parental satisfaction.
④General data collection
The child's age, gender, height, weight, ASA classification, surgical type,
operative time, anesthesia duration, as well as the parents' educational level, the
child's temperament type (measured using the Emotion, Activity, and Social
Temperament Inventory), the parents' baseline anxiety level (measured using the
State-Trait Anxiety Scale), and the child's Modified Yale Preoperative Anxiety Scale
score.
2.7 Data Management and Statistical Analysis Plan, Confidentiality Plan
2.7.1 Data Management
Clinical research will establish corresponding data safety monitoring plans based
on the level of risk. During the implementation phase of clinical research, all adverse
events will be meticulously documented, properly managed, and tracked until they are
adequately resolved or the condition stabilizes. Serious adverse events and
unexpected incidents will be promptly reported to the ethics committee and the
competent authority. The principal investigator will conduct regular cumulative
reviews of all adverse events and convene investigator meetings when necessary to
assess the risks and benefits of the study. This trial is a double-blind trial, with
unblinding performed when necessary to ensure the safety and legitimate rights of
participants. Independent data monitors will be assigned to oversee the study data, and
an independent data safety monitoring committee will be established to monitor
accumulated safety and efficacy data, determining whether to continue the study.
2.7.2 Statistical Analysis Plan
（1）Statistical analysis was performed using SPSS 26.0, with GraphPad Prism 8.0.2
for data visualization and MATLAB for EEG data processing.
（2）The Kolmogorov-Smirnov test was used to determine whether the continuous
data followed a normal distribution.
（3）Continuous data with normal distribution within the group were expressed as
mean ±
standard deviation (x̄ ±
s), while non-normal data were presented as

Version: V2.0 | Date: May 12,2025
median (M) and interquartile range (IQR). Categorical variables were expressed as
rates.
（4）For continuous data with normal distribution within each time point across
three groups, repeated measures ANOVA was employed. For continuous data with
normal distribution at the same time point across three groups, one-way ANOVA was
used. P-values for pairwise comparisons were adjusted using the Bonferroni
correction. For continuous data with non-normal distribution across three groups, the
Kruskal-Wallis H-sum test was applied, while Mann-Whitney U test was used for
pairwise comparisons of non-normal data.
（5）The chi-square test of difference analysis was used to compare the data of
ordinal categorical variables among the three groups. According to the sample size
and the minimum theoretical frequency, Pearson's chi-square test or Fisher's exact
probability method was selected to determine whether there was an association
between the grouping and the outcome variable.
（6）The significance level was set at α=0.05, with P<0.05 indicating statistically
significant differences.
2.7.3 Data Confidentiality Plan
To ensure the confidentiality, security, and integrity of clinical research data and
related information, all researchers and other relevant personnel at the centers have
signed
confidentiality
agreements
and
understood
their
obligations
and
responsibilities. Data is recorded in paper-based collection forms and ultimately
registered in Excel by members of the Data Security Committee for storage.
Electronic data is encrypted, and only authorized personnel are permitted to access
and process the data. Access rights for researchers and other relevant personnel are
strictly restricted, with data disclosure permitted only when necessary. The protocol is
scheduled for regular review and updates to ensure compliance with the latest
technologies, regulations, and requirements. Upon completion of all enrollment, an
unblinded statistician conducts the analysis, reports the data results, and drafts the
conclusions. This process does not disclose any personal patient information, and all
patients have no access rights.
III. Case Selection

Version: V2.0 | Date: May 12,2025
3.1 Eligibility Criteria
（1）American Society of Anesthesiology (ASA) classification
Ⅰ~Ⅲlevel ；
（2）2-12 years of age;
（3）diagnosed with autism by a psychiatrist;
（4）The patient was scheduled for elective surgery under general anesthesia.
3.2 Exclusion Criteria
（1）cardiac, thoracic and neurosurgical procedures;
（2）Congenital diseases or severe hepatic/renal dysfunction;
（3）Allergy to the investigational drug;
（4）Neuromuscular disorders, cerebral palsy, epilepsy;
（5）Other mental or neurological disorders;
（6）BMI ≥30 kg/m²;
（7）Severe upper respiratory tract infection prior to surgery;
（8）Administration of sedatives or analgesics within 48 hours prior to surgery;
（9）Exposure
to
significant
life
stressors
(e.g.
changes
in
family/school
environment, parental divorce or death) within one month before surgery.
(10) Refusal of participation by family members or guardians.
3.3 Exit and Termination Criteria
The withdrawal of the subject by the guardian terminates the patient's
participation in the study.
3.4 Duration of Study Participation
Participated in the whole process from April 2025 to March 2028.
3.5 Recruitment Process
No recruitment.
IV. Risk/Benefit Assessment of the Study
4.1 Research Benefits (Individual and Social Benefits)
(1) This study is expected to reduce the incidence of NPOBCs in children with ASD

Version: V2.0 | Date: May 12,2025
after surgery through the administration of dexmedetomidine and eszopiclone
nasal sprays.
(2) The findings of this study will provide clinicians with critical reference for
decision-making in relevant fields, while also contributing to the optimization of
perioperative management strategies for children with autism spectrum disorder
(ASD), improving patients' perioperative experience, enhancing postoperative
recovery quality, and thereby alleviating the burden on families and society,
demonstrating significant clinical and societal value.
4.2 Research Risks
The subjects were patients undergoing elective surgery under general anesthesia,
without additional study risks.
4.3 Risks and Protection for Special Populations
The subjects were patients undergoing elective surgery under general anesthesia,
without additional study risks.
5、Multicenter clinical study: Please specify the lead institution and task
allocation within this center
The Children's Hospital of Nanjing Medical University served as the primary
responsible institution, with Director Zhang Li as the principal investigator and Sun
Fei as the project liaison.
VI. Ethics in Clinical Research
Clinical research will comply with the World Medical Assembly's "Declaration
of Helsinki" and other relevant regulations. Prior to the initiation of the study, the trial
protocol must be approved by the ethics committee before the clinical research is
conducted. Before enrolling each participant in the study, the investigator is
responsible for providing a complete and comprehensive explanation of the study's
purpose, procedures, and potential risks to the participant or their legal representative,
and obtaining a written informed consent (the version approved by the ethics
committee). Participants should be informed of their right to withdraw from the study
at any time, and the informed consent should be retained as part of the clinical
research documentation for future reference. The privacy and data confidentiality of

Version: V2.0 | Date: May 12,2025
participants will be protected throughout the study.
VII. Annual Plan
April 2025-May 2025: Ethics review and trial registration
June 2025-December 2026: Data collection and statistical analysis
January 2027-June 2027: Thesis writing, submission, and project completion
VIII. Evaluation Indicators
(1)
Case data collection: Complete the collection of all case numbers from all centers to obtain
relatively comprehensive pediatric case data.
(2)
Data analysis: Complete data organization and statistical analysis based on the primary and
secondary research objectives.
(3)
Research Publication: Authored and published 2 papers in SCI-indexed journals.
IX. Preliminary Research Foundation and Working Conditions
(1) Theoretical Feasibility: Dexmedetomidine inhibits excessive activation of the
sympathetic-adrenal cortex axis by activating α 2-adrenergic receptors, reduces the
release of neuroinflammatory factors (e.g., IL-6, TNF-α), and enhances the activity of
prefrontal GABAergic neurons to regulate dysfunction in the amygdala-cortical
circuit. The induced increase in δ-wave power may reduce postoperative behavioral
abnormalities by stabilizing brain network synchrony. In contrast, eszopiclone
antagonizes NMDA receptors to block glutamatergic excitotoxicity and activates the
BDNF-mTOR
pathway
to
promote
synaptic
plasticity
repair.
The
induced
enhancement of γ
oscillations improves default mode network dissociation. Both
agents target the core pathological mechanisms of neurotransmitter imbalance
(GABA/glutamate)
and
synaptic
dysfunction
in ASD
children
through
dual
anti-inflammatory and anti-excitotoxic pathways during the perioperative period.
Preliminary clinical evidence shows that dexmedetomidine reduces postoperative
agitation in children by 30%, while low-dose ketamine nasal spray has been
demonstrated to decrease aggressive behavior in ASD children by 42%. Combined
with the predictive model of theta/ β
wave power ratio established in our team's

Version: V2.0 | Date: May 12,2025
preclinical trial, this provides theoretical and practical support for the correlation
between intervention mechanisms and behavioral outcomes.
(2) Technical Feasibility: Nanjing Children's Hospital affiliated with Nanjing
Medical University is a Grade III, Class A hospital with long-term experience in
pediatric surgery, demonstrating exceptional technical expertise to provide the
required surgical technical support for this study. The Department of Anesthesiology
possesses extensive theoretical
and
clinical practical
experience in pediatric
anesthesia, with medical technical capabilities that are far ahead of the rest in China.
(3) This study was conducted at the Children's Hospital affiliated to Nanjing
Medical University, Zhongda Hospital affiliated to Southeast University, Children's
Hospital affiliated to Xuzhou Medical University, Wuxi Children's Hospital affiliated
to Jiangnan University, Changzhou Children's Hospital affiliated to Nantong
University, Pizhou Hospital affiliated to Xuzhou Medical University, Dongtai
Hospital affiliated to Nantong University (Dongtai People's Hospital), and Suqian
Hospital of Nanjing Drum Tower Hospital Group. These institutions boast superior
geographical locations, advanced medical expertise, extensive regional patient
distribution, a high annual volume of elective surgeries, and a rich pediatric case
source, which can meet the required sample size for the study.
(4) The research team members have extensive prior research experience.
Professor Zhang Li, the principal investigator of the project, serves as the Director of
the Department of Anesthesiology at Nanjing Children's Hospital, a member of the
Biomedical Committee of the Nanjing Association of Chinese Students in Europe and
America, a high-level talent under Jiangsu Province's "Six Major Talent Peaks"
program, and a visiting scholar at Cincinnati Children's Hospital and Dallas Children's
Hospital in the United States. With years of clinical research experience, she
possesses profound expertise in pediatric anesthesia.
(5) This study incorporated quality control feedback from experts in related fields such as
pediatric surgery, anesthesiology, and the Department of Statistics. The research was conducted
collaboratively by senior-title physicians, intermediate-title physicians, junior-title physicians, and
master's/doctoral students, ensuring quality control at each level.

Version: V2.0 | Date: May 12,2025
X. Investigator Information Form
1. Principal Investigator
surname
and
persona
l name
unit
Degree/Title
Ethical and
GCP
training
Phone/Email
Zhang Li
Children's
Hospital
of
Nanjing
Medical
University
Doctor/Chief
Physician
yes
13815406629/
drzhangli@njmu.edu.cn
2. Project team members
surname
and
persona
l name
unit
Degree/Title
Project task
division
Ethical and
GCP
training
Sun Fei
Children's
Hospital
of
Nanjing
Medical
University
Master/Attending
Physician
Data
acquisition
and analysis
yes
Shen
Fangming
Zhongda
Hospital
Affiliated
to
Southeast
University
Doctor/Resident
Data
acquisition
and analysis
XI. Principal Investigator's Commitment (Submission of the research protocol to
the ethics committee shall be deemed as compliance with this commitment):
I hereby certify the authenticity of the application content. I will fulfill the
responsibilities of a principal investigator, strictly comply with national
regulations pertaining to clinical research, ensure the timely completion of
research tasks, diligently perform my duties, and submit relevant materials on

Version: V2.0 | Date: May 12,2025
schedule. Should any inaccuracies or violations of regulations occur in the
submission, I shall bear full responsibility.
I agree to comply with the review comments of the ethics committee and
commence the clinical trial after approval, promptly report any changes in
clinical trial activities to the ethics committee, as well as any unforeseen issues
involving risks to subjects or other personnel, and obtain re-approval from the
ethics review before proceeding. During the study, I will conscientiously submit
relevant reports in accordance with the requirements specified in the ethics
committee's approval.