Objective: To investigate the role of methylation of placental glucocorticoid response gene in the association between pregnancy-related anxiety in the third trimester and birth outcomes. Methods: Based on a prospective cohort study, singleton live births and their mothers from the Ma'anshan Birth Cohort Study (MABC) were included as participants in this study. The maternal pregnancy-related anxiety symptoms in the third trimester of pregnancy were evaluated by using the Pregnancy-related Anxiety Questionnaire. The neonatal birth outcomes were collected from medical records. The placental tissues from 300 pregnant women with pregnancy-related anxiety and 300 without pregnancy-related anxiety were collected to detect the methylation of FKBP5, NR3C1 and HSD11B2 genes using the Methyl Target approach. The methylation factors were extracted by exploratory factor analysis. Linear regression or logistic regression models were used to analyze the association between pregnancy-related anxiety in the third trimester, methylation factor scores, and birth outcomes. The mediating role of methylation factors in the association between pregnancy-related anxiety in the third trimester and birth outcomes was analyzed by using the Process procedure. Results: The mean age of 2 833 pregnant women was (26.60±3.60) years old. After adjusting for confounding factors, pregnancy-related anxiety in the third trimester increased the risk of small-for-gestational-age (OR=1.32, 95%CI:1.00-1.74). A total of 5 methylation factors were extracted, and the factor 5 was loaded with FKBP5 CpGs 18-21. Pregnancy-related anxiety in the third trimester was negatively correlated with the factor 5 (β=-0.24,95%CI:-0.44--0.05). The factor 5 was positively correlated with the gestational age (β=0.17, 95%CI:0.06-0.27). In addition, the factor 2 (β=0.02,95%CI:0.00-0.04) and factor 3 (β=0.03,95%CI:0.01-0.05) were positively correlated with 5-min Apgar score after delivery. However, this study did not found the mediating role of the scores of the factor characterized by FKBP5 in the relationship between pregnancy-related anxiety and birth outcomes. Conclusion: Pregnancy-related anxiety in the third trimester may reduce the methylation level of FKBP5 CpGs 18-21 in placental tissues and is associated with the risk of small-for-gestational-age.
Infant, Newborn ; Pregnancy ; Female ; Humans ; Young Adult ; Adult ; Pregnancy Trimester, Third ; Placenta ; Glucocorticoids/metabolism* ; Cohort Studies ; Prospective Studies ; Methylation ; Factor V/metabolism* ; Anxiety/genetics*

OBJECTIVE: Anxiety is one of the most common symptoms associated with autistic spectrum disorder. The essential oil of Cananga odorata (Lam.) Hook. f. & Thomson, usually known as ylang-ylang oil (YYO), is often used in aromatherapy as a mood-regulating agent, sedative, or hypotensive agent. In the present study, the effects and mechanisms of YYO in alleviating anxiety, social and cognitive behaviors in autism-like rats were investigated. METHODS: The prenatal valproic acid (VPA) model was used to induce autism-like behaviors in offspring rats. The effectiveness of prenatal sodium valproate treatment (600 mg/kg) on offspring was shown by postnatal growth observation, and negative geotaxis, olfactory discrimination and Morris water maze (MWM) tests. Then three treatment groups were formed with varying exposure to atomized YYO to explore the effects of YYO on the anxiety, social and cognitive behaviors of the autistic-like offspring through the elevated plus-maze test, three-chamber social test, and MWM test. Finally, the monoamine neurotransmitters, including serotonin, dopamine and their metabolites, in the hippocampus and prefrontal cortex (PFC) of the rats were measured using a high-performance liquid chromatography. RESULTS: Offspring of VPA exposure rats showed autism-like behaviors. In the VPA offspring, medium-dose YYO exposure significantly elevated the time and entries into the open arms in the elevated plus-maze test, while low-dose YYO exposure significantly enhanced the social interaction time with the stranger rat in session 1 of the three-chamber social test. VPA offspring treated with YYO exposure used less time to reach the platform in the navigation test of the MWM test. YYO exposure significantly elevated the metabolism of serotonin and dopamine in the PFC of VPA offspring. CONCLUSION YYO exposure showed the effects in alleviating anxiety and improving cognitive and social abilities in the offspring of VPA exposure rats. The role of YYO was related to the regulation of the metabolism of serotonin and dopamine. Please cite this article as: Zhang N, Wang ST, Yao L. Inhalation of Cananga odorata essential oil relieves anxiety behaviors in autism-like rats via regulation of serotonin and dopamine metabolism. J Integr Med. 2023; 21(2): 205-214.
Pregnancy ; Female ; Rats ; Animals ; Autistic Disorder/drug therapy* ; Oils, Volatile/therapeutic use* ; Serotonin/metabolism* ; Cananga/metabolism* ; Dopamine ; Anxiety/drug therapy* ; Valproic Acid/pharmacology* ; Plant Oils ; Disease Models, Animal

This study investigated the effect of Suanzaoren Decoction on the expression of N-methyl-D-aspartate receptors(NMDAR) and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptors(AMPAR) in the hippocampus and synaptic plasticity in rats with conditioned fear-induced anxiety. The effect of Suanzaoren Decoction on rat behaviors were evaluated through open field experiment, elevated plus maze experiment, and light/dark box experiment. Enzyme-linked immunosorbent assay(ELISA) was used to measure the levels of glutamate(Glu) and γ-aminobutyric acid(GABA) in the rat hippocampus. Real-time fluorescence quantitative PCR(qRT-PCR) and Western blot were employed to assess the gene and protein expression of ionotropic glutamate receptors in the hippocampal region. Transmission electron microscopy was utilized to observe the changes in the ultrastructure of synaptic neurons in the hippocampal region. Long-term potentiation(LTP) detection technique was employed to record the changes in population spike(PS) amplitude in the hippocampal region of mice in each group. The behavioral results showed that compared with the model group, the Suanzaoren Decoction group effectively increased the number of entries into open arms, time spent in open arms, percentage of time spent in open arms out of total movement time, number of entries into open arms out of total entries into both arms(P<0.01), and significantly increased the time spent in the light box and the number of shuttle crossings(P<0.01). There was an increasing trend in the number of grid crossings, entries into the center grid, and time spent in the center grid, indicating a significant anxiolytic effect. ELISA results showed that compared with the model group, the Suanzaoren Decoction group exhibited significantly reduced levels of Glu, Glu/GABA ratio(P<0.01), and significantly increased levels of GABA(P<0.01) in the rat hippocampus. Furthermore, Suanzaoren Decoction significantly decreased the gene and protein expression of NMDAR(GluN2B and GluN2A) and AMPAR(GluA1 and GluA2) compared with the model group. Transmission electron microscopy results demonstrated improvements in synapses, neuronal cells, and organelles in the hippocampal region of the Suanzaoren Decoction group compared with the model group. LTP detection results showed a significant increase in the PS amplitude changes in the hippocampal region of Suanzaoren Decoction group from 5 to 35 min compared with the model group(P<0.05, P<0.01). In conclusion, Suanzaoren Decoction exhibits significant anxiolytic effects, which may be attributed to the reduction in NMDAR and AMPAR expression levels and the improvement of synaptic plasticity.
Rats ; Mice ; Animals ; Receptors, Ionotropic Glutamate/metabolism* ; Hippocampus ; Neuronal Plasticity ; Receptors, N-Methyl-D-Aspartate/genetics* ; Anxiety/genetics* ; gamma-Aminobutyric Acid

The present study was aimed to explore the involvement of dopamine D1 receptor of the anterior cingulate cortex (ACC) in the regulation of chronic inflammatory pain-related emotion. On the first day, the rats were acclimated to the environment and the baseline indices were measured. On the second day, the rats were administered with the dopamine D1 receptor antagonist SCH-23390 or agonist SKF38393 in the ACC, and then they were subcutaneously injected with complete Freund's adjuvant (CFA, 0.08 mL) in the left hind paw to establish conditioned place avoidance (CPA) response after pairing with specific environment. On the third day, the CPA response and the firing frequency of ACC neurons were observed synchronously, and the open-field behavior, mechanical pain behavior and paw withdrawal latency (PWL) tests were also observed subsequently. In other experiments, rats were given subcutaneous injection of normal saline (NS) on the left hind paw after SCH-23390 or SKF-38393 was administered in the ACC, and then the same observations were performed. The results showed that: (1) Compared with the control group, the PWL and mechanical pain thresholds of rats injected with CFA on the left hind paw were significantly decreased (P < 0.05); (2) The residence time of rats injected with CFA in the "pain environment" and open field center was significantly shortened (P < 0.05); (3) Pre-injection of antagonist SCH-23390 in ACC (10 μg) alleviated the anxiety-like negative behavior response induced by CFA (P < 0.05) and reversed CFA-induced increases of discharge frequency of ACC neurons (P < 0.05); (4) Pre-injection of agonist SKF-38393 in the ACC (10 μg) induced CPA-like behavioral response in rats injected with NS in the left hind paw, and increased the firing frequency of ACC neurons (P < 0.05); (5) Immunofluorescence detection showed that dopamine D1 receptor and NMDA receptor were co-expressed in the same neuron. These results suggest that inhibition of dopamine D1 receptor in ACC can alleviate the negative emotional response induced by persistent pain.
2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine/adverse effects* ; Animals ; Anxiety ; Chronic Pain ; Gyrus Cinguli ; Hyperalgesia ; Rats ; Receptors, Dopamine D1/metabolism*

Central sensitization is essential in maintaining chronic pain induced by chronic pancreatitis (CP), but cortical modulation of painful CP remains elusive. Here, we examined the role of the anterior cingulate cortex (ACC) in the pathogenesis of abdominal hyperalgesia in a rat model of CP induced by intraductal administration of trinitrobenzene sulfonic acid (TNBS). TNBS treatment resulted in long-term abdominal hyperalgesia and anxiety in rats. Morphological data indicated that painful CP induced a significant increase in FOS-expressing neurons in the nucleus tractus solitarii (NTS) and ACC, and some FOS-expressing neurons in the NTS projected to the ACC. In addition, a larger portion of ascending fibers from the NTS innervated pyramidal neurons, the neural subpopulation primarily expressing FOS under the condition of painful CP, rather than GABAergic neurons within the ACC. CP rats showed increased expression of vesicular glutamate transporter 1, and increased membrane trafficking and phosphorylation of the N-methyl-D-aspartate receptor (NMDAR) subunit NR2B and the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) subunit GluR1 within the ACC. Microinjection of NMDAR and AMPAR antagonists into the ACC to block excitatory synaptic transmission significantly attenuated abdominal hyperalgesia in CP rats, which was similar to the analgesic effect of endomorphins injected into the ACC. Specifically inhibiting the excitability of ACC pyramidal cells via chemogenetics reduced both hyperalgesia and comorbid anxiety, whereas activating these neurons via optogenetics failed to aggravate hyperalgesia and anxiety in CP rats. Taken together, these findings provide neurocircuit, biochemical, and behavioral evidence for involvement of the ACC in hyperalgesia and anxiety in CP rats, as well as novel insights into the cortical modulation of painful CP, and highlights the ACC as a potential target for neuromodulatory interventions in the treatment of painful CP.
Animals ; Anxiety/etiology* ; Chronic Pain/etiology* ; GABAergic Neurons ; Gyrus Cinguli/metabolism* ; Hyperalgesia/metabolism* ; Pancreatitis, Chronic/pathology* ; Rats ; Rats, Sprague-Dawley ; Receptors, N-Methyl-D-Aspartate/metabolism* ; Trinitrobenzenesulfonic Acid/toxicity*

OBJECTIVE: To investigate the effects of diethylhexyl phthalate (DEHP) exposure on anxiety-like behaviors and learning and memory ability in mice and explore the underlying mechanism. METHODS: Forty male ICR mice were randomized equally into control group (0 mg/kg) and 10, 50 and 100 mg/kg DEHP exposure groups, in which the mice were exposed to DEHP at the indicated doses by gavage for 4 weeks. After the treatments, the mice were assessed for behavioral changes using open filed test, elevated plus-maze and Morris water maze test. Brain tissues were collected from the mice for determination of malondialdehyde (MDA) content, pathologies and expressions of ZO-1 and occludin in the hippocampus. RESULTS: Compared with the control group, the mice with DEHP exposure for 4 weeks exhibited no significant body weight change (P>0.05) but presented with obvious behavioral changes, manifested by reduced movement distance (P < 0.05) and time spent in the center of the open field (P < 0.05), reduced movement distance (P < 0.05) and time spent in the open arm of the elevated maze (P < 0.05), significantly increased latency of searching for the platform (P < 0.05), and decreased frequency of crossing the platform (P < 0.05). HE staining showed obvious vertebral cell death in the hippocampal CA1 to CA3 regions of the mice with DEHP exposure. The exposed mice showed significantly increased MDA content and decreased expressions of ZO-1 and occludin at both the mRNA and protein levels in the hippocampus (P < 0.05 or 0.01). Multivariate linear regression analysis suggested a close correlation between anxiety-like behaviors and learning and memory abilities in DEHP-exposed mice. CONCLUSION DEHP exposure may cause damages of the blood-brain barrier and the pyramidal cells in the hippocampus of mice, thereby inducing anxiety-like behaviors and learning and memory impairment.
Animals ; Anxiety/chemically induced* ; Blood-Brain Barrier/metabolism* ; Diethylhexyl Phthalate/toxicity* ; Male ; Maze Learning ; Mice ; Mice, Inbred ICR ; Occludin/pharmacology*

Danzhi Xiaoyao Powder is a classical prescription for anxiety. This study aims to analyze the effect of this medicine on mitochondrial morphology and function of anxiety rats and explore the mechanism of it against anxiety. Specifically, uncertain empty bottle drinking water stimulation(21 days) was employed to induce anxiety in rats. The elevated plus-maze test and open field test were respectively performed on the 7 th, the 14 th, and the 21 st days of the stimulation, so as to detect the anxiety-related protein index brain-derived neurotrophic factor(BDNF) and evaluate the anxiety level of animals. On this basis, the effect of this prescription on anxiety rats was preliminarily evaluated. After the behavioral test on the 21 st day, rats were killed and the brain tissues were separated for the observation of the mitochondrial morphology and the determination of mitochondrial function-related indicators and the adenosine 5'-monophosphate-activated protein kinase(AMPK) level. The results showed that Danzhi Xiaoxiao Powder could alleviate the anxiety-like behavior of rats, significantly increase the percentage of time in open arm in elevated plus-maze test and the ration of activity time in the central area of the field, dose-dependently raise the activity levels of respiratory chain complex Ⅰ,Ⅱ,Ⅲ and Ⅳ and the adenosine triphosphate(ATP) content, and elevate the levels of BDNF and phosphorylated AMPK(p-AMPK). Clear structure and intact morphology of mitochondrial cristae in medial prefrontal cortex cells and amygdala were observed in the Danzhi Xiaoyao Powder group. In summary, Danzhi Xiaoyao Powder exerts therapeutic effect on anxiety, and the mechanism is the likelihood that p-AMPK protects the structure and maintains the function of mitochondria.
Rats ; Animals ; Brain-Derived Neurotrophic Factor/metabolism* ; Powders ; AMP-Activated Protein Kinases ; Anxiety/drug therapy* ; Mitochondria

OBJECTIVE: To explore the pathogenic role of changes of Wnt/β-catenin signaling pathway in the hippocampus in depression- and anxiety-like behaviors caused by prenatal stress (PS) in offspring rats. METHODS: Twelve female SpragueDawley rats weighing 240-260 g were randomly divided into control and restraint stress groups. The rats in the control group received no interventions, and those in restraint stress group were subjected to restraint stress (three times a day, 45 min each time) at the gestational age of 14-20 days. The 1-month-old offspring rats underwent open field test and forced swimming test to assess the anxiety- and depression-like behaviors, and the expressions of Wnt1, Gsk-3β and β-catenin in the hippocampus were detected using Western blotting. RESULTS: In open field test, the offspring rats with PS showed significantly decreased crossings of the center ( < 0.01) with reduced time spent in the center ( < 0.05) compared with control offspring rats. In forced swimming test, the offspring rats in PS group exhibited a significantly longer immobility time than in the control rats, and showed obvious depression- and anxiety-like behaviors. Compared with those in the control offspring rats, Gsk-3β expression increased significantly while the expressions of β-catenin and Wnt1 were significantly lowered in the hippocampus of the offspring rats in PS group ( < 0.01). CONCLUSIONS PS causes changes in Wnt/β-catenin signaling pathway in the hippocampus to contribute to the occurrence of depression-and anxiety-like behaviors in rats.
Animals ; Anxiety ; etiology ; metabolism ; Behavior, Animal ; Depression ; etiology ; metabolism ; Female ; Glycogen Synthase Kinase 3 beta ; Hippocampus ; metabolism ; Pregnancy ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Restraint, Physical ; psychology ; Stress, Psychological ; complications ; Swimming ; psychology ; Wnt Signaling Pathway

Intermittent hypoxia (IH) has preventive and therapeutic effects on hypertension, myocardial infarction, cerebral ischemia and depression, but its effect on post-traumatic stress disorder (PTSD) has not been known. In this study, we used inescapable electric foot shock combined with context recapture to build PTSD mouse model. The levels of fear and anxiety were valued by the open field, the elevated plus maze (EPM) and the fear conditioning tests; the level of spatial memory was valued by Y maze test; the number of Fos positive neurons in hippocampus, amygdala and medial prefrontal cortex was valued by immunohistochemical staining; and the protein expressions of hypoxia inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) and brain derived neurotrophic factor (BDNF) in these brain area were valued by Western blot. The results showed that IH and model (foot shock) had an interaction on percentage of entering open arms (OE%) in EPM and freezing time and the number of fecal pellets in fear conditioning test. IH increased OE% in EPM and reduced the freezing time and the number of fecal pellets in fear conditioning test in PTSD model mice. At the same time, IH reduced the number of Fos positive neurons in the hippocampus, amygdala and medial prefrontal cortex of PTSD model mice, and increased the protein expression levels of HIF-1α, VEGF and BDNF in these brain tissues. In conclusion, IH pretreatment can relieve fear and anxiety behavior in post-traumatic stress model mice, suggesting that IH may be an effective means of preventing PTSD.
Animals ; Anxiety ; therapy ; Brain-Derived Neurotrophic Factor ; metabolism ; Fear ; Hypoxia ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Mice ; Stress Disorders, Post-Traumatic ; therapy ; Vascular Endothelial Growth Factor A ; metabolism

Stress and emotion are associated with several illnesses from headaches to heart diseases and immune deficiencies to central nervous system. Terminalia arjuna has been referred as traditional Indian medicine for several ailments. The present study aimed to elucidate the effect of T. arjuna bark extract (TA) against picrotoxin-induced anxiety. Forty two male Balb/c mice were randomly divided into six experimental groups (n = 7): control, diazepam (1.5 mg·kg), picrotoxin (1 mg·kg) and three TA treatemt groups (25, 50, and 100 mg/kg). Behavioral paradigms and PCR studies were performed to determine the effect of TA against picrotoxin-induced anxiety. The results showed that TA supplementation increased locomotion towards open arm (EPM) and illuminated area (light-dark box test), and increased rearing frequency (open field test) in a dose dependent manner, compared to picrotoxin (P < 0.05). Furthermore, TA increased number of licks and shocks in Vogel's conflict. PCR studies showed an up-regulation of several genes, such as BDNF, IP, DL, CREB, GABA, SOD, GPx, and GR in TA administered groups. In conclusion, alcoholic extract of TA bark showed protective activity against picrotoxin in mice by modulation of genes related to synaptic plasticity, neurotransmitters, and antioxidant enzymes.
Animals ; Antioxidants ; metabolism ; Anxiety Disorders ; drug therapy ; genetics ; metabolism ; psychology ; Brain-Derived Neurotrophic Factor ; genetics ; metabolism ; Dopamine Agents ; administration & dosage ; GABA Agents ; administration & dosage ; Glutathione Peroxidase ; genetics ; metabolism ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Neuronal Plasticity ; drug effects ; Neurotransmitter Agents ; metabolism ; Phytotherapy ; Picrotoxin ; adverse effects ; Plant Bark ; chemistry ; Plant Extracts ; administration & dosage ; Serotonin Agents ; administration & dosage ; Superoxide Dismutase-1 ; genetics ; metabolism ; Terminalia ; chemistry