OBJECTIVES: To investigate the role of a neural pathway from oxytocin (OXT) neurons in the paraventricular hypothalamic nucleus (PVN) to γ-aminobutyric acid (GABA) neurons (GABAergic neurons) in the trigeminal nucleus caudalis (TNC) in regulating pain sensitization in a mouse model of chronic migraine and to explore the underlying mechanisms. METHODS: A chronic migraine mouse model was established by intraperitoneal injection of nitroglycerin (NTG, 1 mg/mL, 10 mg/kg) on days 1, 3, 5, 7, and 9. The study consisted of four parts: PartⅠ: 24 male wild-type C57BL/6J mice were divided into four groups (n=6 in each), receiving single or repeated injection of NTG or saline, respectively. Immunofluorescence was used to detect c-Fos and OXT expression in the PVN. Part Ⅱ: 6 male OXT-Cre transgenic C57BL/6J mice were used for anterograde monosynaptic tracing combined with RNAscope and immunofluorescence to identify neural projections from PVN OXT neurons to TNC GABAergic neurons. Part Ⅲ: 30 male OXT-Cre transgenic C57BL/6J mice were bilaterally injected Cre-dependent chemogenetic activation virus into the PVN. These mice were randomly divided into five groups, with six mice in each group. Mice in the clozapine N-oxide (CNO) group and the control group were intra-peritoneally injected with 0.1 mg/mL of CNO solution (1 mg/kg) and the same volume of isotonic normal saline, respectively. 3 hours after the injection, the brain tissues were harvest and c-Fos immunofluorescence staining was performed to verify the efficiency of chemogenetic activation virus. Mice in the model control group and the CNO activated model group were subjected to chronic migraine modeling, with bilateral TNC injection of isotonic normal saline and CNO, respectively, on day 10. The mice in the negative control group were bilaterally intra-TNC injected with isotonic normal saline. After 30 minutes, the Von-Frey filament and acetone tests were used to assess the mechanical pain threshold and cold pain response time in the periorbital region of the mice in these three groups. Part Ⅳ: 24 male OXT-Cre transgenic C57BL/6J mice were bilaterally injected with the Cre-dependent chemogenetic activation virus into the PVN. These mice were randomly divided into four groups, with six mice in each group. Mice in the model control group, the CNO activated model group and the atosiban group were subjected to chronic migraine modeling. On day 10, mice in the negative control group and the model control group were intraperitoneally injected with isotonic normal saline, while mice in the CNO activated model group and the atosiban group were intraperitoneally injected with CNO. After 15 minutes, mice in the atosiban group were bilaterally intra-TNC injected with atosiban, while mice in other three groups were bilaterally intra-TNC injected with isotonic normal saline containing 1% dimethyl sulfoxide. After 15 minutes, the Von-Frey filament and acetone tests were used to assess the mechanical pain threshold and cold pain response time in the periorbital region of the mice. The GABA content in the bilateral TNC was detected by high-performance liquid chromatography (HPLC). RESULTS: Mice with chronic migraine models exhibited reduced periorbital mechanical pain thresholds and increased periorbital cold pain reaction time, accompanied by an increase in both the number of c-Fos⁺ neurons and the percentage of c-Fos⁺ OXT neurons in the PVN (all P<0.05). The anterograde tracing virus and RNAscope combined with immunofluorescence staining showed that PVN OXT neurons projected to TNC GABAergic neurons. Immuno-fluorescence staining demonstrated that compared with the control group, the percentage of c-Fos⁺ OXT neurons in the PVN of CNO group increased (P<0.05). In bilateral intra-TNC drug administration experiments, compared with the model control group, the periorbital mechanical pain threshold increased, and the periorbital cold pain reaction time decreased in the CNO activated model group (both P<0.05). In intraperitoneal drug administration experiments, compared with the CNO activate model group, the periorbital mechanical pain threshold decreased, and the periorbital cold pain reaction time increased in the atosiban group (both P<0.05). HPLC analysis showed that, compared with the negative control group, the model control group and the atosiban group, GABA level of TNC in the CNO activated model group increased (all P<0.05). CONCLUSIONS PVN OXT neurons exert a descending facilitatory effect on GABAergic neurons in the TNC via OXT release, thereby ameliorating pain sensitization in chronic migraine.
Animals ; Paraventricular Hypothalamic Nucleus/physiopathology* ; Male ; Mice, Inbred C57BL ; Migraine Disorders/physiopathology* ; Mice ; GABAergic Neurons/physiology* ; Oxytocin/physiology* ; Disease Models, Animal ; Neurons/physiology* ; Mice, Transgenic ; Neural Pathways ; Chronic Disease

In modern society, people are increasingly exposed to chronic stress, leading to various mental disorders. However, the activities of brain regions, especially neural firing patterns related to specific behaviors, remain unclear. In this study, we introduce a novel approach, NeuroSync, which integrates open-field behavioral testing with electrophysiological recordings from emotion-related brain regions, specifically the central amygdala and the paraventricular nucleus of the hypothalamus, to explore the mechanisms of negative emotions induced by chronic stress in mice. By applying machine vision techniques, we quantified behaviors in the open field, and signal processing algorithms elucidated the neural underpinnings of the observed behaviors. Synchronizing behavioral and electrophysiological data revealed significant correlations between neural firing patterns and stress-related behaviors, providing insights into real-time brain activity underlying behavioral responses. This research combines deep learning and machine learning to synchronize high-resolution video and electrophysiological data, offering new insights into neural-behavioral dynamics under chronic stress conditions.
Animals ; Mice ; Male ; Emotions/physiology* ; Stress, Psychological/physiopathology* ; Action Potentials/physiology* ; Mice, Inbred C57BL ; Behavior, Animal/physiology* ; Machine Learning ; Amygdala/physiopathology* ; Neurons/physiology* ; Paraventricular Hypothalamic Nucleus/physiopathology* ; Brain/physiology*

Magnetic stimulation has made significant strides in the treatment of psychiatric disorders. Nonetheless, current magnetic stimulation techniques lack the precision to accurately modulate specific nuclei and cannot realize deep brain magnetic stimulation. To address this, we utilized superparamagnetic iron oxide nanoparticles as mediators to achieve precise targeting and penetration. We investigated the effects of magnetic fields with varying frequencies on neuronal activity and compared the activation effects on neurons using a 10-Hz precise magneto-stimulation system (pMSS) with repetitive transcranial magnetic stimulation in mice. Oxytocin levels, dendritic morphology and density, and mouse behavior were measured before and after pMSS intervention. Our findings suggest that pMSS can activate oxytocinergic neurons, leading to upregulation of oxytocin secretion and neurite outgrowth. As a result, sociability was rapidly improved after a one-week pMSS treatment regimen. These results demonstrate a promising magneto-stimulation method for regulating neuronal activity in deep brain nuclei and provide a promising therapeutic approach for autism spectrum disorder.
Animals ; Autism Spectrum Disorder/physiopathology* ; Paraventricular Hypothalamic Nucleus/physiology* ; Disease Models, Animal ; Transcranial Magnetic Stimulation/methods* ; Male ; Social Behavior ; Mice ; Oxytocin/metabolism* ; Mice, Inbred C57BL ; Neurons/physiology*

Neuropathic pain is a chronic condition caused by damage or dysfunction in the nervous system. While the spleen may influence neuropathic pain, its role has been poorly understood. This study demonstrates that the spleen plays a crucial role in regulating neuropathic pain through the bed nucleus of the stria terminalis (BNST) - paraventricular nucleus of the hypothalamus (PVN) neural circuit in a chronic constriction injury (CCI) mouse model. Splenectomy, splenic denervation, or splenic sympathectomy significantly increased the mechanical withdrawal threshold (MWT) and reduced macrophage infiltration in the dorsal root ganglia (DRG) of CCI mice. Pseudorabies virus injections into the spleen revealed connections to the BNST and PVN in the brain. Chemogenetic inhibition of the BNST-PVN circuit increased macrophage infiltration in the DRG and decreased the MWT; these effects were reversed by splenectomy, splenic denervation, or sympathectomy. These findings underscore the critical role of the spleen, regulated by the BNST-PVN circuit, in neuropathic pain.
Animals ; Neuralgia/pathology* ; Septal Nuclei/physiopathology* ; Male ; Spleen/physiopathology* ; Paraventricular Hypothalamic Nucleus/physiopathology* ; Mice, Inbred C57BL ; Splenectomy ; Mice ; Neural Pathways/physiopathology* ; Disease Models, Animal ; Ganglia, Spinal/physiopathology* ; Sympathectomy ; Macrophages

At present, the problem of drug addiction treatment mainly lies in the high relapse rate of drug addicts. Addictive drugs will bring users a strong sense of euphoria and promote drug seeking. Once the drug is withdrawn, there will be withdrawal symptoms such as strong negative emotions and uncomfortable physical reactions. The recurrence of context-induced withdrawal memory is an important reason for drug relapse. Our previous study has shown increased c-Fos expression in prelimbic cortex (PrL) neurons projecting to paraventricular nucleus of the thalamus (PVT) (PrL^-PVT) during conditioned context-induced retrieval of morphine withdrawal memory. However, whether PrL^-PVT neurons are involved in withdrawal memory retrieval and the underlying molecular mechanisms remain unknown. In this study, we used conditioned place aversion (CPA) model combined with in vivo calcium signal recording, chemogenetics and nucleus drug injection methods to investigate the role and molecular mechanism of PrL^-PVT neurons in retrieval of morphine withdrawal memory. The results showed that the calcium signals of PrL^-PVT neurons were significantly enhanced by withdrawal-related context; Inhibition of PrL^-PVT neurons blocked the conditioned context-induced morphine withdrawal memory retrieval; Activation of PrL^-PVT neurons caused animals to escape from the context; After the inhibition of NMDA receptors in the PrL, withdrawal-related context failed to increase c-Fos and Arc expressions in PrL^-PVT neurons. The above results suggest that NMDA receptors in PrL^-PVT neurons are associated with retrieval of morphine withdrawal memory. This study is of great significance for further understanding the neural circuit mechanism of withdrawal memory retrieval as well as the intervention and prevention of drug relapse.
Animals ; Substance Withdrawal Syndrome/physiopathology* ; Morphine/adverse effects* ; Neurons/physiology* ; Receptors, N-Methyl-D-Aspartate/metabolism* ; Male ; Rats ; Paraventricular Hypothalamic Nucleus/metabolism* ; Memory ; Rats, Sprague-Dawley ; Morphine Dependence/physiopathology* ; Midline Thalamic Nuclei/physiology* ; Neural Pathways/metabolism*

OBJECTIVE: This study investigates the sleep-modulating effects of ginsenoside Rg1 (Rg1, C₄₂H₇₂O₁₄), a key bioactive component of ginseng, and elucidates its underlying mechanisms. METHODS: C57BL/6J mice were intraperitoneally administered doses of Rg1 ranging from 12.5 to 100 mg/kg. Sleep parameters were assessed to determine the average duration of each sleep stage by monitoring the electrical activity of the brain and muscles. Further, orexin neurons in the lateral hypothalamus (LH) and corticotropin-releasing hormone (CRH) neurons in the paraventricular hypothalamic nucleus (PVH) were ablated using viral vector surgery and electrode embedding. The excitability of LH^orexin and PVH^CRH neurons was evaluated through the measurement of cellular Finkel-Biskis-Jinkins murine osteosarcoma viral oncogene homolog (c-Fos) expression. RESULTS: Rg1 (12.5-100 mg/kg) augmented the duration of non-rapid eye movement (NREM) sleep phases, while reducing the duration of wakefulness, in a dose dependent manner. The reduced latency from wakefulness to NREM sleep indicates an accelerated sleep initiation time. We found that these sleep-promoting effects were weakened in the LH^orexin and PVH^CRH neuron ablation groups, and disappeared in the orexin and CRH double-ablation group. Decreased c-Fos protein expression in the LH and PVH confirmed that Rg1 promoted NREM sleep by inhibiting orexin and CRH neurons. CONCLUSION Rg1 increases the duration of NREM sleep, underscoring the essential roles of LH^orexin and PVH^CRH neurons in facilitating the sleep-promoting effects of Rg1. Please cite this article as: Wang YY, Wu Y, Yu KW, Xie HY, Gui Y, Chen CR, Wang NH. Ginsenoside Rg1 promotes non-rapid eye movement sleep via inhibition of orexin neurons of the lateral hypothalamus and corticotropin-releasing hormone neurons of the paraventricular hypothalamic nucleus. J Integr Med. 2024; 22(6): 721-730.
Animals ; Ginsenosides/pharmacology* ; Orexins/metabolism* ; Mice, Inbred C57BL ; Neurons/metabolism* ; Paraventricular Hypothalamic Nucleus/metabolism* ; Male ; Hypothalamic Area, Lateral/metabolism* ; Corticotropin-Releasing Hormone/metabolism* ; Mice ; Sleep/drug effects*

The nucleus tractus solitarii (NTS) is one of the morphologically and functionally defined centers that engage in the autonomic regulation of cardiovascular activity. Phenotypically-characterized NTS neurons have been implicated in the differential regulation of blood pressure (BP). Here, we investigated whether phenylethanolamine N-methyltransferase (PNMT)-expressing NTS (NTS^PNMT) neurons contribute to the control of BP. We demonstrate that photostimulation of NTS^PNMT neurons has variable effects on BP. A depressor response was produced during optogenetic stimulation of NTS^PNMT neurons projecting to the paraventricular nucleus of the hypothalamus, lateral parabrachial nucleus, and caudal ventrolateral medulla. Conversely, photostimulation of NTS^PNMT neurons projecting to the rostral ventrolateral medulla produced a robust pressor response and bradycardia. In addition, genetic ablation of both NTS^PNMT neurons and those projecting to the rostral ventrolateral medulla impaired the arterial baroreflex. Overall, we revealed the neuronal phenotype- and circuit-specific mechanisms underlying the contribution of NTS^PNMT neurons to the regulation of BP.
Solitary Nucleus/metabolism* ; Blood Pressure/physiology* ; Phenylethanolamine N-Methyltransferase/metabolism* ; Neurons/metabolism* ; Paraventricular Hypothalamic Nucleus/metabolism*

It is well established that increased excitability of the presympathetic neurons in the hypothalamic paraventricular nucleus (PVN) during hypertension leads to heightened sympathetic outflow and hypertension. However, the mechanism underlying the overactivation of PVN presympathetic neurons remains unclear. This study aimed to investigate the role of endogenous corticotropin-releasing factor (CRF) on the excitability of presympathetic neurons in PVN using Western blot, arterial blood pressure (ABP) and renal sympathetic nerve activity (RSNA) recording, CRISPR/Cas9 technique and patch-clamp technique. The results showed that CRF protein expression in PVN was significantly upregulated in spontaneously hypertensive rats (SHRs) compared with normotensive Wistar-Kyoto (WKY) rats. Besides, PVN administration of exogenous CRF significantly increased RSNA, heart rate and ABP in WKY rats. In contrast, knockdown of upregulated CRF in PVN of SHRs inhibited CRF expression, led to membrane potential hyperpolarization, and decreased the frequency of current-evoked firings of PVN presympathetic neurons, which were reversed by incubation of exogenous CRF. Perfusion of rat brain slices with artificial cerebrospinal fluid containing CRF receptor 1 (CRFR1) blocker, NBI-35965, or CRF receptor 2 (CRFR2) blocker, Antisauvagine-30, showed that blocking CRFR1, but not CRFR2, hyperpolarized the membrane potential and inhibited the current-evoked firing of PVN presympathetic neurons in SHRs. However, blocking CRFR1 or CRFR2 did not affect the membrane potential and current-evoked firing of presympathetic neurons in WKY rats. Overall, these findings indicate that increased endogenous CRF release from PVN CRF neurons enhances the excitability of presympathetic neurons via activation of CRFR1 in SHRs.
Rats ; Animals ; Rats, Inbred SHR ; Paraventricular Hypothalamic Nucleus/physiology* ; Receptors, Corticotropin-Releasing Hormone/metabolism* ; Rats, Inbred WKY ; Corticotropin-Releasing Hormone/metabolism* ; Neurons/physiology* ; Hypertension ; Sympathetic Nervous System

Olfaction and food intake are interrelated and regulated. In the process of feeding, the metabolic signals in the body and the feeding signals produced by food stimulation are first sensed by the arcuate nucleus of hypothalamus and the nucleus tractus solitarius of brain stem, and then these neurons project to the paraventricular nucleus of hypothalamus. The paraventricular nucleus transmits the signals to other brain regions related to feeding and regulates feeding behavior. In this process, olfactory signals can be transmitted to hypothalamus through olfactory bulb and olfactory cortex to regulate feeding behavior. At the same time, gastrointestinal hormones (ghrelin, insulin, leptin, etc.) and some neurotransmitters (acetylcholine, norepinephrine, serotonin, endocannabinoid, etc.) produced in the process of feeding act on the olfactory system to regulate olfactory function, which in turn affects the feeding itself. This review summaries the research progress of the interaction between olfaction and food intake and its internal mechanism from the aspects of neuronal and hormonal regulation.
Arcuate Nucleus of Hypothalamus/metabolism* ; Feeding Behavior/physiology* ; Hypothalamus ; Paraventricular Hypothalamic Nucleus ; Smell

Acupuncture has remarkable effects on treating functional gastrointestinal diseases, but its central mechanism is not clear. At present, the research has mainly focused on several central nuclei, such as the dorsal vagus complex (DVC), nucleus raphe magnus (NRM), locus coeruleus (LC), subnucleus reticularis dorsalis (SRD), hypothalamic paraventricular nucleus (PVN), cerebellar fastigial nucleus (FN), central amygdala (CeA), etc. It is not clear whether the nuclei are involved in acupuncture regulation of gastric function through certain interrelation. A further summary of related literature indicates that many brain regions or nuclei in the central nervous system are closely related to gastric function, such as DVC, NRM, parabrachial nuclei (PBN), LC, periaqueductal gray (PAG), cerebellum, PVN, arcuate nucleus (Arc), hippocampus, CeA, etc. Most of these nuclei have certain fiber connections with each other, in which DVC is the basic center, and other nuclei are directly or indirectly involved in the regulation of gastric function through DVC. Is DVC the key target in acupuncture regulation of gastric function? Does other nuclei have direct or indirect neural circuit with DVC to participate in the regulation of gastric function by acupuncture, such as the possibility of CeA-DVC neural loop in acupuncture regulating gastric function. Therefore, more advanced techniques such as photogenetics, chemical genetics should be introduced and the central mechanism of acupuncture on regulating gastric function with DVC as center, from the view of nerve loop, will become the focus of further research, which could explain the central integration mechanism of acupoint compatibility by modern neuroscience technology.
Acupuncture Therapy ; Locus Coeruleus ; Paraventricular Hypothalamic Nucleus ; Vagus Nerve