Dexmedetomidine promotes NREM sleep by depressing the neuronal activity of OXT neurons in the PVN

doi:10.21203/rs.3.rs-4207174/v1

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Dexmedetomidine promotes NREM sleep by depressing the neuronal activity of OXT neurons in the PVN

https://doi.org/10.21203/rs.3.rs-4207174/v1

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published 30 Jul, 2024

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This study aimed to investigate the mechanism of Dexmedetomidine (DEX) on sleep homeostasis. We will focus on the effect of DEX on the activity and the signal transmission of oxytocin-expressing (Oxytocin, OXT) neurons in the paraventricular nucleus (PVN) of the hypothalamus (PVN^OXT) in the regulation of sleep-wakefulness cycle. In this study, the guide cannula was implanted to microinject DEX into PVN. PVN^OXT neurons were specifically activated by the chemogenetic method. Differences in sleep-wakefulness states were monitored by electroencephalogram (EEG)/ electromyogram (EMG) recording. In addition, the effect of DEX on the electrophysiological activity of PVN^OXT neurons was examined by whole-cell patch-clamp technique. EEG/EMG results showed that microinjection of DEX in PVN significantly increased the duration of nonrapid eye movement (NREM) sleep in mice. Chemogenetic activation of PVN^OXT neurons after internal cannula injection of DEX to PVN increased the amount of wake. Electrophysiological results show that DEX could inhibit the frequency of action potential (AP) and the spontaneous excitatory postsynaptic current (sEPSC) in PVN^OXT neurons through α₂-adrenoceptors. DEX maintained sleep homeostasis by inhibiting excitatory synaptic signaling in PVN^OXT neurons through α₂-adrenoceptors.

DEX

OXT neurons

PVN

EEG/EMG recording

patch clamp

Dexmedetomidine (DEX) is a highly specific α₂-adrenoreceptor agonist, with a 1620 times affinity of binding to α₁ adrenoreceptors [1]. α₂-adrenoceptors are a class of heterotrimeric guanine nucleotide-binding protein receptors (GPCR) [2], which play a wide role in regulating physiological functions, including heart rate, blood pressure, blood glucose, insulin homeostasis, and analgesia. They are involved in presynaptic feedback inhibition mediating the release of neurotransmitters from the terminal of norepinephrine synapses [3–5]. As an anesthetic drug, DEX has the characteristics of sedation and analgesia with few side effects and a wide application range[6–12]. The specific role of DEX in postoperative sleep and its related mechanisms are poorly understood.

As a sedative-hypnotic drug, DEX has a certain impact on the sleep-wake “switches” [13]. Intraperitoneal injection of DEX can increase the c-Fos expression in the overlapping areas of the preoptic area (POA) and diaphragm of the hypothalamus [14], ventral tegmental area (VTA) [15], and area postrema (AP*) [16] and decrease the c-Fos expression in the supraoptic nucleus (SON) [17], primary motor cortex (M1), and secondary motor cortex (M2) [18], and these brain regions are associated with the modulation of sleep-wakefulness [19]. Studies have shown that DEX can increase the calcium signal intensity of dopamine (DA) neurons in the VTA (VTA^DA). DEX can increase the excitability of VTA^DA neurons through α adrenoreceptors. It also increases the concentration of DA neurotransmitters in the medial prefrontal cortex (MPFC) and nucleus accumbens (NAc), an effect that may be related to the sedative arousal properties of DEX [15]. The activation of VTA^DA neurons can induce the state change from an unconscious anesthetic to wakefulness, triggering a strong awakening-promoting effect [20], which may be the mechanism of DEX-induced rapid awakening after sedation.

Recently, it was found that vesicular glutamate transporter 2 (Vglut2), oxytocin (OXT), prodynorphin (PDYN), corticotropin-releasing hormone (CRH), and vasopressin (AVP) neurons in the paraventricular nucleus of the hypothalamus (PVN) could induce different levels of arousal through chemogenetic activation [21]. PVN is composed of three types of neurons: the large cells (Magno), the small cells (Parvocellular), and the long projection neurons, which express OXT, PDYN, CRH, et.al. Almost one-third of neurons in PVN are expressing OXT (PVN^OXT) mainly distributed in the middle part of PVN. [22]. AVP and OXT neurons are involved in the regulation of physiology and behavior, including stress response and social behavior. Sleep-wakefulness has an essential attribution to these adaptive behaviors. Our previous study showed that DEX could depress the c-Fos expression of AVP neurons in PVN and reduce the concentration of AVP in serum [23]. It has been found that photogenetic and chemogenetic activation of PVN^AVP neurons significantly increases the amount of wake in mice [21, 24]. PVN^OXT neurons are an important component in the regulation of sleep and awakening [25]. Studies have shown that chemogenetic activation of PVN^OXT neurons could increase the amount of wake and decrease the amount of NREM sleep and rapid eye movement (REM) sleep [21]. Here, we aim to investigate the roles of PVN^OXT neurons in the regulation of sleep homeostasis by DEX. We used the chemogenetic methods to activate PVN^OXT neurons after canula administration of DEX to PVN, we found that activation of PVN^OXT neurons could counteract the increase of NREM sleep time induced by DEX. In addition, we found that DEX inhibited the frequency of action potential (AP) and spontaneous excitatory postsynaptic current (sEPSC) of PVN^OXT neurons through α₂-adrenoceptors. It is suggested that DEX might maintain sleep homeostasis by inhibiting excitatory synaptic signaling in PVN^OXT neurons.

2.1 Mice

The animal experimental procedures underwent rigorous scrutiny and approval by the Laboratory Animal Management Office of Laboratory Animals at Anhui Medical University, adhering strictly to the pertinent ethical directives established by the Institutional Animal Care Unit Committee of Anhui Medical University, under project identifier LLSC20190763. OXT-IRES-Cre mice (stock No. 024234; Jackson Laboratory, Bar Harbor, ME) and Ai9 mice (stock No. 007909; Jackson Laboratory, Bar Harbor, ME) were generously provided by Professor Tian Xue from the University of Science and Technology of China. Housing conditions for all mice were maintained under a 12-hour light-dark cycle (lights on at 08:00, illuminance ranging from 150–200 lux), with ad libitum access to food and water, and a room temperature (RT) set at 25°C.

2.2 Surgery and virus injection

Mice were deeply anesthetized with 50 mg/kg sodium pentobarbital and positioned in a stereotaxic frame (RWD Life Science, Shenzhen, China) with sterilized instruments. Utilizing coordinates from the mouse brain atlas[26], a microinjection needle delivered 200 nL of rAAV-EF1α-DIO-hM3D(Gq)-mCherry-WPREs (≥ 5.00E + 12 viral particles/mL, Brainvta, Wuhan, China) bilaterally into the PVN at a rate of 40 nL/min (M/L: ± 0.30 mm, A/P: -0.6 mm, D/V: -4.6). The guide cannula (RWD Life Science, Shenzhen, China) was implanted in the PVN with sagittal plane tilting 5°(M/L: 1.0 mm, A/P: -0.6 mm, D/V: -4.6) and secured with light-curable resin (BJBCJB, Sino-Dentex, Jilin, China). The length of internal cannulas (RWD Life Science, Shenzhen, China), is equal to guide cannulas. EEG electrodes were inserted into the skull (+ 1.5 mm Bregma, -1.5 mm midline – first recording electrode; +1.5 mm Lambda, + 1.5 mm midline – second recording electrode), while EMG electrodes were placed in neck musculature. All electrodes were affixed with light-cured resin and quick self-curing acrylic resin (SC01, Yamahahi Dental, Japan). Surgical sites were sutured and disinfected. Post-surgery, mice were housed individually.

2.3 EEG/EMG recordings and analysis

The animals were housed individually in an EEG/EMG recording cage under a 12-hour light:12-hour dark cycle (with light onset at 08:00 and offset at 20:00) for 3 days of acclimation. Electrical signals were recorded at 1 kHz, amplified with the MP150 system (BIOPAC, California, USA), and digitized using Acknowledge 4.2 software (BIOPAC, California, USA). Automated analysis categorized EEG/EMG signals into Wake, NREM sleep, and REM sleep states every 4 seconds using the Lunion Stage AI engine (LunionData, Shanghai, China), with manual verification and correction. EEG signals were filtered between 0.3–35 Hz and EMG signals between 10–200 Hz.

2.4 Electrophysiology

2.4.1 Slice preparation

OXT::Ai9 mice (8–12 weeks old) were anesthetized with 1% isoflurane (RWD Life Science, Shenzhen, China) and decapitated. The brain was swiftly excised and immersed in a freezing oxygenated (95% O₂ and 5% CO₂) cutting solution containing N-methyl-D-glucamine, 1-deoxy-1-(methylamine)-D-sorbito, glucamine (NMDG) cutting solution (in mM: 92 NMDG, 1.2 potassium chloride (KCl), 1.2 potassium dihydrogen phosphate (KH₂PO₄), 30 sodium bicarbonate (NaHCO₃), 25 D-glucose, 20 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES), 5 L-ascorbic acid, 3 sodium pyruvate, 2 thiourea, 10 magnesium sulfate (MgSO₄) and 0.5 calcium chloride (CaCl₂); the pH adjusted to 7.2 ± 0.1 with hydrogen chloride (HCl), the osmolarity adjusted to 310 ± 5 mOsmol/L with NMDG). Horizontal brain slices (300 µm) were then obtained using a vibrating microtome (VT1200S, Leica, Germany). These slices were transferred to oxygenated (95% O₂ and 5% CO₂) aCSF (mM: 125 NaCl, 1.25 KCl, 2 CaCl₂, 1 MgCl₂, 1.25 KH₂PO₄, 25 NaHCO₃, 0.4 Na-pyruvate, 0.08 L-ascorbic acid, 25 D-glucose), and allowed incubating for 40–45 minutes at 36.6 ℃ before recording.

2.4.2 Whole-cell recordings

Whole-cell recordings were performed at RT in oxygenated (95% O₂ and 5% CO₂) aCSF solution with constant perfusion at a rate of 1–2 mL/min. For action potential (AP) recording, the electrodes were filled with a potassium-based internal solution (in mM: 125 K-gluconate, 10 KCl, 10 HEPES, 0.1EGTA, 5 ATP-Mg, 0.5GTP-Na; the pH adjusted to 7.2 ± 0.1 with potassium hydroxide (KOH); the osmolarity adjusted to 310 ± 5 mOsmol/kg with K-gluconate). For sEPSCs recording, the electrodes were filled with a CsMethane sulfonate-based internal solution containing (in mM: 135 CsMethane sulfonate, 10 cesium chloride (CsCl), 10 HEPES, 1EGTA, 5 ATP-Mg, 0.5 GTP-Na, the pH was adjusted to 7.2 ± 0.1 using cesium hydroxide (CsOH), and the osmolarity was adjusted to 310 ± 5 mOsmol/L using CsMethane sulfonate). The aCSF with 50 µM picrotoxin (PTX, HY-101391, MedChemExpress, USA) was recorded sEPSCs. For sIPSCs recording, the electrodes were filled with a cesium-based internal solution containing (in mM: 140 CsCl, 10 HEPES, 1EGTA, 5 ATP-Mg, 0.5 GTP-Na; the pH was adjusted to 7.2 ± 0.1 using CsOH, and the osmolarity was adjusted to 310 ± 5 mOsmol/L using CsCl). The aCSF with 10 µM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, ab120017, Abcam, UK) and 20 µM 5-phospho-D-norvaline (D-AP5, ab120003, Abcam, UK) were recorded sIPSCs. sEPSCs were recorded in the presence of the GABA_A receptor antagonist Picrotoxin (PTX) and sIPSCs were recorded in the presence of AMPA and NMDA receptor antagonists CNQX and D-AP5 in the aCSF. The neurons were held at -70 mV. The resistance of the patch pipette was 5–7 MΩ. A 40× water microscope (TH4-200 and U-RFL-T microscope, Olympus, Japan) was used to visualize the cells. Data were collected by using an Axopatch 700B patch clamp amplifier, Digidatal 1550B data acquisition system, and pClamp10.7 software (Molecular Devices, USA). Data were sampled at 20 kHz, and low-pass filtered with a cutoff frequency of 1 kHz in the current recording. Clampfit 10.7 (Molecular Devices, USA) was used to analyze the electrophysiological properties of the cells. If the hold current of the patch neuron exceeds 30 pA during whole-cell recording or the change of Access resistance (Ra) exceeds 20% during recording, statistics will not be included in the statistical analysis. Igor Pro (WaveMetrics, USA) was used to make statistical charts and representative charts.

2.5 Drug administration

In the EEG/EMG recording, OXT-hM3D(Gq)-mCherry mice were recorded from Zeitgeber time (ZT) 14 to ZT0, after the cannula administration of saline or 2 µM DEX to investigate the DEX-induced effect of sleep-wakefulness states (Fig. 1b). OXT-hM3D(Gq)-mCherry mice were injected 1µL 2 µM DEX into internal cannulas at ZT14(22:00) and i.p injected saline or CNO at ZT15.5(23:30) to investigate the effect of sleep-wakefulness states in PVN^OXT neurons (Fig. 2c) [27, 28].

Electrophysiological characteristics of PVN^OXT neurons of APs were captured using the whole-cell patch-clamp method in brain slices immersed in the artificial cerebrospinal fluid (aCSF) with or without 2 µM DEX. RS79948, an α₂-adrenergic receptor antagonist, was utilized to block adrenergic signaling [29]. Following a 2-minute stabilization, PVN^OXT neurons were microinjected with 1 mM RS79948 for 30 seconds, followed by a 3-minute recording period. Subsequently, neurons received a combined microinjection of 1 mM RS79948 and 2 µM DEX for 30 seconds, with a further 3-minute recording. (Fig. 4a). To assess the impact of DEX on neuronal signal transmission, sEPSCs and spontaneous inhibitory postsynaptic currents (sIPSCs) were recorded. Initially, neurons were stabilized for 2 minutes, then microinjected with aCSF for 30 seconds, and data were recorded for 2 minutes. This was followed by 30-second microinjection of 2 µM DEX, with a 2-minute recording period (Fig. 5a). To determine the factors of the observed sEPSCs reduction, neurons were stabilized for 2 minutes, microinjected with 1 mM RS79948 for 30 seconds, and recorded for 1 minute, followed by a co-injection of 1 mM RS79948 and 2 µM DEX for 30 seconds, with a final 1-minute recording period (Fig. 5b).

2.6 Statistical Analysis

Data are presented as mean ± standard error of the mean (SEM). The normality of data was assessed using Shapiro-Wilk analysis prior to t-tests. Statistical comparisons between two groups were conducted using paired or unpaired Student's t-tests, Wilcoxon matched-pairs signed rank test, or Mann-Whitney test as appropriate. Multiple parameter comparisons were performed using two-way repeated-measures ANOVA, followed by Bonferroni's multiple comparison test. Prism 10.0 software (GraphPad Software, San Diego, CA, USA) was utilized for all statistical analyses. P < 0.05 was considered statistically significant.

3.1 DEX increased NREM sleep time and decreased the average duration of wakefulness.

For further exploration of the mechanism of DEX in the sleep-wakefulness cycle, EEG/EMG was divided into three states: NREM sleep, REM sleep, and wake. A guide cannula for drug administration was implanted in the PVN. The mice were cannula injected 1µL saline or 2 µM DEX into PVN with a speed of 1µL/min (Fig. 1a). We used the EEG/EMG recording to analyze the sleep-wakefulness phases in mice between the saline group and the DEX group (Fig. 1c-d). The results showed that the amount of wake during ZT14-18 was decreased (Fig. 1e), the amount of NREM sleep during ZT14-18 was increased (Fig. 1f), and the amount of REM sleep during ZT14-18 had no differences (Fig. 1g) in the DEX group, compared with the saline group., EEG Power spectrum density (PSD) in the DEX group decreased significantly from 1.0-3.9Hz within 4 hours, compared with the saline group (Fig. 1h). DEX resulted in a decrease in the mean duration of wake (Fig. 1i), but the number of sleep transitions had no differences (Fig. 1j).

3.2 Chemogenetic activation of PVN^OXT neurons counteracted DEX-induced NREM sleep and increased arousal.

We injected rAAV-EF1α-DIO-hM3D(Gq)-mCherry-WPREs into PVN to infect OXT neurons (Fig. 2a). Patch clamp recording was used to certify the effectiveness of the virus. The OXT-hM3D(Gq)-expressing neurons were puff with the aCSF and 5µm clozapine-N-oxide (CNO, Brainvta, Wuhan, China) for 5 minutes respectively, indicating that neurons could be successfully activated by CNO (Fig. 2b). The results of i.p. injection of saline or 2 mg/kg CNO into ZT2 showed that activation of PVN^OXT neurons in the CNO group could increase the amount of wake and decrease NREM sleep in 1 hour, while REM sleep time had no differences (Fig. 2d-f). EEG/EMG was recorded for 10 hours from ZT14 (Fig. 2g, h). After 30 minutes of CNO injection, the time of three sleep states of 0.5, 1, and 1.5 hours were counted. Compared with the saline group, after chemogenetic inhibition of PVN^OXT neurons, the amount of wake during ZT16-17 in the CNO group was increased, while the amount of NREM time was decreased and the amount of REM had no differences (Fig. 2i-m). In conclusion, the DEX-induced increase in NREM sleep was counteracted by the activation of PVN^OXT neurons.

3.3 DEX inhibited the AP frequency of PVN^OXT neurons.

To further investigate the mechanism of the effect of DEX on PVN^OXT neurons on sleep homeostasis, we prepared brain slices containing PVN from OXT::Ai9 mice during ZT13 and ZT14. During electrophysiological recording, fluorescent images of recorded PVN^OXT neurons were collected (Fig. 3a). We examined the firing properties of PVN^OXT neurons before and after drug administration, such as the AP duration were measured at 50% repolarization (APD₅₀), AP amplitude, spike max, afterhyperpolarization (AHP) amplitudes, thresholds, resting cell membrane potential (RMP), membrane input resistance (R_in) (Fig. 3b). The results showed that the AP frequency of OXT neurons in the DEX group was significantly reduced, compared to the aCSF group (Fig. 3c). However, there were no significant differences in APD₅₀, AP amplitude, spike max, AHP, threshold, RMP, and R_in between the aCSF group and the DEX group (Fig. 3d-j). When the neurons received stimulation from − 40 pA to 0 pA, the DEX group showed a tendency to decrease the potential difference, compared with the aCSF group (Fig. 3k). DEX might reduce the rate of ion influx. Further analysis of the changes in membrane potential and resistance will help to reveal the finer regulatory mechanism of DEX on the function of OXT neurons.

3.4 DEX decreased the AP and sEPSC frequencies of PVN^OXT neurons through α₂-adrenoceptors.

As mentioned above, DEX is a highly potent α₂-adrenoceptors agonist. We recorded the effect of blocking the receptor-mediated signaling pathway on the electrophysiological characteristics of PVN^OXT neurons by patch-clamp experiments. The results showed that the AP frequency, APD₅₀, AP amplitude, spike max, AHP amplitude, threshold, RMP, and R_in have not changed in the RS79948 + DEX group, compared with the RS79948 group (Fig. 4b-j). This suggests that DEX might inhibited the AP frequency in PVN^OXT neurons through α₂-adrenoceptors. Subsequently, sEPSCs and sIPSCs were recorded to explore whether DEX could influence the neuronal signal transmission of PVN^OXT neurons. We observed a significant decrease in the frequency of sEPSC in the PVN^OXT neurons between the aCSF group and the DEX group (Fig. 5c), and no significant difference between the mean amplitude of sEPSC (Fig. 5d). DEX did not show significant effects on both the frequency and amplitude of sIPSC (Fig. 5e, f). Last, we puffed RS79948 to both the aCSF group and the DEX group and recorded sEPSCs and sIPSCs in PVN^OXT neurons. We found that RS79948 blocked the decreased frequency of sEPSC induced by DEX, with no difference in the amplitude of sEPSC (Fig. 5g, h). Combined with the above results, our study shows that DEX has a significant effect on modulating AP and excitatory synaptic transmission in PVN^OXT neurons through α₂-adrenoceptors.

Clinical studies show that severe sleep disorders and delirium often occur after surgery [30]. Patients exhibit sleep deprivation, sleep fragmentation, and a reduction in the duration of slow-wave sleep and REM sleep [31]. The most significant features of postoperative delirium include cognitive deficits, dysfunction of circadian rhythm, and emotional disorders. Studies have shown that the DEX has characteristics of low side effects, which could improve postoperative sleep quality, and reduce the incidence of postoperative delirium after surgery [32, 33]. Disturbance of sleep structure is a serious problem for postoperative patients, which may lead to prolonged postoperative recovery time [34] and increase the incidence of cardiovascular diseases, cognitive dysfunctions [35], and impaired immune functions [36]. Therefore, we hope to find out the mechanism of DEX on the modulation of sleep homeostasis, further reducing the incidence of delirium.

Studies showed that oral administration of DEX can increase the duration of NREM sleep within 6 hours, and decrease the duration of REM sleep within 6 hours[13]. In a sleep deprivation experiment, the polysomnography results show that the delta wave spectrum in NREM sleep would increase during the recovery period after sleep deprivation [37]. DEX-induced sedation is similar to deep restorative sleep after sleep deprivation. Unlike the classical hypnotic drug diazepam, oral administration of DEX could increase the power of the delta spectrum in NREM sleep EEG, especially at high-frequency delta waves of 1.75–3.25 Hz, while the power of low-frequency delta waves of 0.5-1.0 Hz was reduced. The same phenomenon appeared in our study, the cannula injection of DEX to PVN could increase the amount of NREM sleep time and decrease the mean duration of wakefulness, and DEX decreased the delta spectrum at 1.0-3.9Hz. If subsequent studies can find the effect of DEX on different delta wave frequencies, it will be helpful to further analyze the regulatory effect of DEX on NREM sleep [13].

OXT-expressing neurons are predominantly located in the PVN and supraoptic nucleus (SON) of the mammalian hypothalamus. OXT plays a crucial role in modulates responses to stress, fear, and social behaviors[38–42]. Little research has investigated the influence of OXT on sleep-wakefulness. Interestingly, OXT might have a dual mechanism of action in dependence of the physiological state on sleep regulation. Long-term intranasal administration of OXT reduced sleep latencies and increased the sleep efficiency and percentage of REM sleep episodes in human [43]. A study in mice showed that acute lateral ventricle (ICV) infusion of OXT delayed sleep onset latency, which resulted in a transient reduction of NREM sleep and REM sleep, and augmented high-frequency activity in NREM sleep [44]. Optogenetic activation of OXT neurons in PVN can promote arousal [45]. The same phenomenon was found in our study, the chemogenetic activation of PVN^OXT neurons could increase the amount of wake. We found that the activation of wakeful-promoting PVN^OXT neurons through the chemogenetic method could decrease the one-hour amount of NREM sleep induced by DEX. DEX-induced sedation has the property of deep restorative sleep after sleep deprivation [14] and is easy to wake up [46]. Whether this process was modulated by working on OXT neurons needs further studies.

Different firing patterns of neurons dictate the amount and timing of neuropeptide release, and the activity of the network affects the neurons' integrative properties. The previous study indicates DEX can increase or decrease delta and theta wave percentage through chemogenetic inhibition or activation of VTA^DA neurons. DEX increases the firing rate of VTA^DA neurons through α₂ adrenoceptors [15]. DEX is widely recognized for its sedative properties and is predominantly believed to exert its effects by engaging the norepinephrine system. Extensive research indicates that DEX interacts with α₂ adrenoceptors located on the norepinephrine-producing neurons of the LC. This interaction triggers a hyperpolarization of the neuronal cell membranes via a Gi protein-mediated pathway. As a result, the release of the stimulating neurotransmitter norepinephrine is diminished across the brain, culminating in a state of sedation[47]. Here, we examined the influence of DEX on the membrane characteristics and synaptic properties of PVN^OXT neurons. We found DEX decreases the frequency of AP in PVN^OXT neurons. The chemogenetic activation of PVN^Vglut2 neurons can increase arousal indicate glutamate signaling pathway participates in modulating the sleep-wakefulness cycle [21], and in our study, we also found DEX could decrease the frequency of sEPSC of PVN^OXT neurons. Therefore, DEX might reduce the rate of ion influx by bounding α₂ adrenoceptors, resulting in decreased excitability of the PVN^OXT neurons.

In conclusion, these findings highlighted DEX increased NREM sleep by inhibiting PVN^OXT neurons, which promoted arousal and decreased the frequency of AP and sEPSC of PVN^OXT neurons. The results reveal the potential role of modulation of DEX in PVN^OXT neurons and provide new ideas for the treatment of postoperative delirium prevention. However, further research is still needed to resolve the molecular mechanisms of DEX and its more far-reaching effects on neuronal networks.

Acknowledgments

The authors thank Dandan Zang, Cheng Qian, and Guanjun Chen of the Center for Scientific Research of Anhui Medical University for their valuable help in our experiments.

Author Contributions

Juan Cheng and Liecheng Wang supervised and designed the work; Ying Zhang and Juan Cheng wrote and revised the manuscript; Ying Zhang and Jiaxin Li completed the patch-clamp experiment, EEG/EMG recording and gene identification. Ying Zhang, Jiaxin Li, Wei Wang, Daming Wang, and Junli Ding analyzed the experiment data and bred the mice.

Data Availability Statement:

Raw data will be made available on request to the corresponding author.

Declaration of Conflicting Interests

The author(s) declared no potential conflicts of interest concerning the research, authorship, and/or publication of this article.

Funding

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: Our work was supported by the National Natural Science Foundation of China (31800997 to Juan Cheng, 81971236, 81571293 to Liecheng Wang), the Natural Science Foundation of Universities of Anhui Province (2022AH050783 to Juan Cheng), Anhui Provincial Natural Science Foundation (2308085MC83 to Juan Cheng), Research Fund of Anhui Institute of translational medicine (2023zhyx-C71 to Daming Wang) , Basic and Clinical Cooperative Research Promotion Program of Anhui Medical University (2020xkjT027 to Junli Ding).

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Dexmedetomidine promotes NREM sleep by depressing the neuronal activity of OXT neurons in the PVN

Status:

Journal Publication

Version 1

Abstract

Figures

1 Introduction

2 Materials and Methods

2.1 Mice

2.2 Surgery and virus injection

2.3 EEG/EMG recordings and analysis

2.4 Electrophysiology

2.4.1 Slice preparation

2.4.2 Whole-cell recordings

2.5 Drug administration

2.6 Statistical Analysis

3 Result

3.1 DEX increased NREM sleep time and decreased the average duration of wakefulness.

3.2 Chemogenetic activation of PVN^OXT neurons counteracted DEX-induced NREM sleep and increased arousal.

3.3 DEX inhibited the AP frequency of PVN^OXT neurons.

3.4 DEX decreased the AP and sEPSC frequencies of PVN^OXT neurons through α₂-adrenoceptors.

4 Discussion

Declarations

Acknowledgments

Author Contributions

Data Availability Statement:

Declaration of Conflicting Interests

Funding

References

Additional Declarations

Status:

Journal Publication

Version 1

Dexmedetomidine promotes NREM sleep by depressing the neuronal activity of OXT neurons in the PVN

Status:

Journal Publication

Version 1

Abstract

Figures

1 Introduction

2 Materials and Methods

2.1 Mice

2.2 Surgery and virus injection

2.3 EEG/EMG recordings and analysis

2.4 Electrophysiology

2.4.1 Slice preparation

2.4.2 Whole-cell recordings

2.5 Drug administration

2.6 Statistical Analysis

3 Result

3.1 DEX increased NREM sleep time and decreased the average duration of wakefulness.

3.2 Chemogenetic activation of PVNOXT neurons counteracted DEX-induced NREM sleep and increased arousal.

3.3 DEX inhibited the AP frequency of PVNOXT neurons.

3.4 DEX decreased the AP and sEPSC frequencies of PVNOXT neurons through α2-adrenoceptors.

4 Discussion

Declarations

Acknowledgments

Author Contributions

Data Availability Statement:

Declaration of Conflicting Interests

Funding

References

Additional Declarations

Status:

Journal Publication

Version 1

3.2 Chemogenetic activation of PVN^OXT neurons counteracted DEX-induced NREM sleep and increased arousal.

3.3 DEX inhibited the AP frequency of PVN^OXT neurons.

3.4 DEX decreased the AP and sEPSC frequencies of PVN^OXT neurons through α₂-adrenoceptors.