Neural pathways from hypothalamic orexin neurons to the ventrolateral preoptic area mediate sleep impairments induced by conditioned fear.

Introduction: Fear and sleep impairments common co-exist, but the underlying mechanisms remain unclear. Hypothalamic orexinergic neurons are involved in the regulation of sleep-wake and fear expression. The ventrolateral preoptic area (VLPO) is an essential brain region to promote sleep, and orexinergic axonal fibers projecting to the VLPO are involved in the maintenance of sleep-wake. Neural pathways from hypothalamic orexin neurons to the VLPO might mediate sleep impairments induced by conditioned fear. Methods: To verify above hypothesis, electroencephalogram (EEG) and electromyogram (EMG) were recorded for analysis of sleep-wake states before and 24 h after conditioned fear training. The retrograde tracing technique and immunofluorescence staining was used to identify the projections from the hypothalamic orexin neurons to the VLPO and to observe their activation in mice with conditioned fear. Moreover, optogenetic activation or inhibition of hypothalamic orexin-VLPO pathways was performed to observe whether the sleep-wake can be regulated in mice with conditioned fear. Finally, orexin-A and orexin receptor antagonist was administered into the VLPO to certify the function of hypothalamic orexin-VLPO pathways on mediating sleep impairments induced by conditioned fear. Results: It was found that there was a significant decrease in the non-rapid eye movement (NREM) and rapid eye movement (REM) sleep time and a significant increase in the wakefulness time in mice with conditioned fear. The results of retrograde tracing technique and immunofluorescence staining showed that hypothalamic orexin neurons projected to the VLPO and observed the CTB labeled orexin neurons were significantly activated (c-Fos+) in the hypothalamus in mice with conditioned fear. Optogenetic activation of hypothalamic orexin to the VLPO neural pathways significantly decreased NREM and REM sleep time and increased wakefulness time in mice with conditioned fear. A significant decrease in NREM and REM sleep time and an increase in wakefulness time were observed after the injection of orexin-A into the VLPO, and the effects of orexin-A in the VLPO were blocked by a pre-administrated dual orexin antagonist (DORA). Conclusion: These findings suggest that the neural pathways from hypothalamic orexinergic neurons to the VLPO mediate sleep impairments induced by conditioned fear.

Mechanism of Potassium Release from Feldspar by Mechanical Activation in Presence of Additives at Ordinary Temperatures.

To improve the potassium availability of feldspar at ordinary temperatures, the mechanical grinding and addition of sodium hydroxide/salts were employed to study the effects of mechanical activation and strong alkali addition on particle characteristics, water-soluble potassium, and the available potassium of feldspar. A laser particle size analyzer was utilized for the direct determination of particle size distribution (PSD) using ground samples. The Brunauer-Emmett-Teller (BET) method was employed for specific surface areas. X-ray diffraction (XRD) was employed for structural characterization, scanning electron microscopy (SEM) for morphology exploration, and energy dispersive spectroscopy (EDS) to determine the chemical composition of potassium feldspar powder. The results revealed that the mechanical activation of potassium feldspar could reduce the particle size and produce agglomerated nanoparticles in the later period. The addition of NaOH and sodium salt did not cause agglomeration, and NaOH dissolved the nanoparticles. The water-soluble potassium content of feldspar in each treatment increased during mechanical grinding, from 21.64 mg kg-1 to 1495.81 mg·kg-1, by adding NaOH 5% weight of potassium feldspar powder and to 3044.08 mg·kg-1 by adding NaOH 10% weight with effects different from those of mechanical shaking. By comparison, only 162.93 mg·kg-1 water-soluble potassium was obtained by adding NaOH 5% weight. The dissolved potassium in the former case was significantly higher than in the latter, and the addition of NaOH and sodium salts significantly enhanced the water-soluble potassium contents due to ion exchange. Furthermore, the addition of sodium hydroxide improved the water-soluble potassium due to its mechanochemical action on potassium feldspar. The mechanical energy changed the crystal structure of potassium feldspar, explaining the increase in available potassium. The addition of sodium salts did not promote change in the feldspar's structure, thereby did not raise the available potassium content. The reason for this was related to the mechanochemical action on sodium hydroxide and feldspar, which could promote the dissolution of fine particles, thereby incrementing the available potassium.

Selective activation of the hypothalamic orexinergic but not melanin-concentrating hormone neurons following pilocarpine-induced seizures in rats.

Introduction: Sleep disorders are common comorbidities in patients with temporal lobe epilepsy (TLE), but the underlying mechanisms remain poorly understood. Since the lateral hypothalamic (LH) and the perifornical orexinergic (ORX) and melanin-concentrating hormone (MCH) neurons are known to play opposing roles in the regulation of sleep and arousal, dysregulation of ORX and MCH neurons might contribute to the disturbance of sleep-wakefulness following epileptic seizures. Methods: To test this hypothesis, rats were treated with lithium chloride and pilocarpine to induce status epilepticus (SE). Electroencephalogram (EEG) and electromyograph (EMG) were recorded for analysis of sleep-wake states before and 24 h after SE. Double-labeling immunohistochemistry of c-Fos and ORX or MCH was performed on brain sections from the epileptic and control rats. In addition, anterograde and retrograde tracers in combination with c-Fos immunohistochemistry were used to analyze the possible activation of the amygdala to ORX neural pathways following seizures. Results: It was found that epileptic rats displayed prolonged wake phase and decreased non-rapid eye movement (NREM) and rapid eye movement (REM) phase compared to the control rats. Prominent neuronal activation was observed in the amygdala and the hypothalamus following seizures. Interestingly, in the LH and the perifornical nucleus, ORX but not MCH neurons were significantly activated (c-Fos+). Neural tracing showed that seizure-activated (c-Fos+) ORX neurons were closely contacted by axon terminals originating from neurons in the medial amygdala. Discussion: These findings suggest that the spread of epileptic activity from amygdala to the hypothalamus causes selective activation of the wake-promoting ORX neurons but not sleep-promoting MCH neurons, which might contribute to the disturbance of sleep-wakefulness in TLE.

Deep Neural Network for Point Sets Based on Local Feature Integration.

The research of object classification and part segmentation is a hot topic in computer vision, robotics, and virtual reality. With the emergence of depth cameras, point clouds have become easier to collect and increasingly important because of their simple and unified structures. Recently, a considerable number of studies have been carried out about deep learning on 3D point clouds. However, data captured directly by sensors from the real-world often encounters severe incomplete sampling problems. The classical network is able to learn deep point set features efficiently, but it is not robust enough when the method suffers from the lack of point clouds. In this work, a novel and general network was proposed, whose effect does not depend on a large amount of point cloud input data. The mutual learning of neighboring points and the fusion between high and low feature layers can better promote the integration of local features so that the network can be more robust. The specific experiments were conducted on the ScanNet and Modelnet40 datasets with 84.5% and 92.8% accuracy, respectively, which proved that our model is comparable or even better than most existing methods for classification and segmentation tasks, and has good local feature integration ability. Particularly, it can still maintain 87.4% accuracy when the number of input points is further reduced to 128. The model proposed has bridged the gap between classical networks and point cloud processing.

Characteristics of ultrafine-ground potassium-bearing shale and the effects of roasting pre-cracking.

Potassium-bearing shale is being developed as a potential alternative to potash for use in fertilisers. The first step in this process is to reduce its particle size by crushing. This paper explores whether roasting pre-cracked potassium-bearing shale can improve the quality of the resulting ultrafine product. Analysis of the particle size distribution of the ultrafine product and its fractal dimension found contradictory results: the minimum particle size distribution was obtained by roasting for 2.5 h, while the minimum fractal dimension was obtained by roasting for 1 h. Fuzzy comprehensive evaluation was conducted with three indicators-(1) the weight of the - 10 µm product, (2) the fractal dimension of the particle size distribution, and (3) d97-to obtain a unique combination of indicators that reflects the quality and quantity of the products. The weights of the three indicators were calculated by an analytic hierarchical process to be 0.69, 0.149 and 0.161, respectively. Roasting pre-cracked shale for 2-2.5 h was found to improve the mean values of the fuzzy comprehensive evaluation indicators by about 0.07. However, the cost increased from 2.82 RMB to ≥ 10.08 RMB, which is not feasible for widespread industrial implementation.

Neocortex- and hippocampus-specific deletion of Gabrg2 causes temperature-dependent seizures in mice.

Mutations in the GABRG2 gene encoding the γ-aminobutyric acid (GABA) A receptor gamma 2 subunit are associated with genetic epilepsy with febrile seizures plus, febrile seizures plus, febrile seizures, and other symptoms of epilepsy. However, the mechanisms underlying Gabrg2-mediated febrile seizures are poorly understood. Here, we used the Cre/loxP system to generate conditional knockout (CKO) mice with deficient Gabrg2 in the hippocampus and neocortex. Heterozygous CKO mice (Gabrg2fl/wtCre+) exhibited temperature-dependent myoclonic jerks, generalised tonic-clonic seizures, increased anxiety-like symptoms, and a predisposition to induce seizures. Cortical electroencephalography showed the hyperexcitability in response to temperature elevation in Gabrg2fl/wtCre+ mice, but not in wild-type mice. Gabrg2fl/wtCre+ mice exhibited spontaneous seizures and susceptibility to temperature-induced seizures. Loss of neurons were observed in cortical layers V-VI and hippocampus of Gabrg2fl/wtCre+ mice. Furthermore, the latency of temperature- or pentylenetetrazol-induced seizures were significantly decreased in Gabrg2fl/wtCre+ mice compared with wild-type mice. In summary, Gabrg2fl/wtCre+ mice with Gabrg2 deletion in the neocortex and hippocampus reproduce many features of febrile seizures and therefore provide a novel model to further understand this syndrome at the cellular and molecular level.

Effects of Glucagon-Like Peptide-1 Receptor Agonist Exendin-4 on the Reinstatement of Cocaine-Mediated Conditioned Place Preference in Mice.

A high percentage of relapse to compulsive cocaine-taking and cocaine-seeking behaviors following abstinence constitutes a major obstacle to the clinical treatment of cocaine addiction. Thus, there is a substantial need to develop effective pharmacotherapies for the prevention of cocaine relapse. The reinstatement paradigm is known as the most commonly used animal model to study relapse in abstinent human addicts. The primary aim of this study is to investigate the potential effects of systemic administration of glucagon-like peptide-1 receptor agonist (GLP-1RA) exendin-4 (Ex4) on the cocaine- and stress-triggered reinstatement of cocaine-induced conditioned place preference (CPP) in male C57BL/6J mice. The biased CPP paradigm was induced by alternating administration of saline and cocaine (20 mg/kg), followed by extinction training and then reinstatement by either a cocaine prime (10 mg/kg) or exposure to swimming on the reinstatement test day. To examine the effects of Ex4 on the reinstatement, Ex4 was systemically administered 1 h after the daily extinction session. Additionally, we also explored the associated molecular basis of the behavioral effects of Ex4. The expression of nuclear factor κß (NF-κß) in the nucleus accumbens (NAc) was detected using Western blotting. As a result, all animals that were treated with cocaine during the conditioning period successfully acquired CPP, and their CPP response was extinguished after 8 extinction sessions. Furthermore, the animals that were exposed to cocaine or swimming on the reinstatement day showed a significant reinstatement of CPP. Interestingly, systemic pretreatment with Ex4 was sufficient to attenuate cocaine- and stress-primed reinstatement of cocaine-induced CPP. Additionally, the expression of NF-κß, which was upregulated by cocaine, was normalized by Ex4 in the cocaine-experienced mice. Altogether, our study reveals the novel effect of Ex4 on the reinstatement of cocaine-induced CPP and suggests that GLP-1R agonists appear to be highly promising drugs in the treatment of cocaine use disorder.

GPER-Deficient Rats Exhibit Lower Serum Corticosterone Level and Increased Anxiety-Like Behavior.

Ample evidence suggests that estrogens have strong influences on the occurrence of stress-related mood disorders, but the underlying mechanisms remain poorly understood. Through multiple approaches, we demonstrate that the G protein-coupled estrogen receptor (GPER) is widely distributed along the HPA axis and in brain structures critically involved in mood control. Genetic ablation of GPER in the rat resulted in significantly lower basal serum corticosterone level but enhanced ACTH release in response to acute restraint stress, especially in the female. GPER-/- rats of either sex displayed increased anxiety-like behaviors and deficits in learning and memory. Additionally, GPER deficiency led to aggravation of anxiety-like behaviors following single-prolonged stress (SPS). SPS caused significant decreases in serum corticosterone in WT but not in GPER-deficient rats. The results highlight an important role of GPER at multiple sites in regulation of the HPA axis and mood.

Postretrieval Microinjection of Baclofen Into the Agranular Insular Cortex Inhibits Morphine-Induced CPP by Disrupting Reconsolidation.

Environmental cues associated with drug abuse are powerful mediators of drug craving and relapse in substance-abuse disorders. Consequently, attenuating the strength of cue-drug memories could reduce the number of factors that cause drug craving and relapse. Interestingly, impairing cue-drug memory reconsolidation is a generally accepted strategy aimed at reducing the intensity of cues that trigger drug-seeking and drug-taking behaviors. In addition, the agranular insular cortex (AI) is an important component of the neural circuits underlying drug-related memory reconsolidation. GABAB receptors (GABABRs) are potential targets for the treatment of addiction, and baclofen (BLF) is the only prototypical GABAB agonist available for application in clinical addiction treatment. Furthermore, ΔFosB is considered a biomarker for the evaluation of potential therapeutic interventions for addiction. Here, we used the morphine-induced conditioned place preference (CPP) paradigm to investigate whether postretrieval microinjections of BLF into the AI could affect reconsolidation of drug-reward memory, reinstatement of CPP, and the level of ΔFosB in mice. Our results showed that BLF infused into the AI immediately following morphine CPP memory retrieval, but not 6 h postretrieval or following nonretrieval, could eliminate the expression of a morphine CPP memory. This effect persisted in a morphine-priming-induced reinstatement test, suggesting that BLF in the AI was capable of preventing the reconsolidation of the morphine CPP memory. Our results also showed that the elimination of morphine CPP memory was associated with reduced morphine-associated ΔFosB expression in the longer term. Taken together, the results of our research provide evidence to support that GABABRs in the AI have an important role in drug-cue memory reconsolidation and further our understanding of the role of the AI in drug-related learning and memory.

The novel estrogen receptor GPER1 decreases epilepsy severity and susceptivity in the hippocampus after status epilepticus.

The steroid hormone 17ß-estradiol (estrogen) exerts neuroprotective effects in several types of neurological disorders including epilepsy. The novel G protein-coupled estrogen receptor 1 (GPER1), also called GPR30, mediates the non-genomic effects of 17ß-estradiol. However, the specific role of GPER1 in status epilepticus (SE) remains unclear. In this report, we evaluated the effects of GPER1 on the hippocampus during SE and the underlying mechanism was studied. Our results revealed that pilocarpine-induced GPER1-KD epileptic rats exhibited a shorter latency to generalized convulsions and strikingly elevated seizure severity. Additionally, the electroencephalographic seizure activity also corresponded to these results. Fast-Fourier analysis indicated an enhancement of power in the theta and alpha bands during SE in GPER1-KD rats. In addition, epilepsy-induced pathological changes were dramatically exacerbated in GPER1-KD rats, including neuron damage and neuroinflammation in hippocampus. GPER1 might be associated with the susceptibility to and severity of epileptic seizures. In summary, our results suggested that GPER1 plays a neuroprotective role in SE, and might be a candidate target for epilepsy therapy.

GABAergic neurons in the insular cortex play an important role in cue-morphine reward memory reconsolidation.

AIMS: There have been recent reports that reconsolidation-based interventions attenuate drug reward memories in rodents. The insular cortex (IC) is an essential part of neural circuits that underlie cue-drug memory reconsolidation. GABAergic interneurons in the IC are a potent control on network excitability and play an important role in the inhibitory mediation of reward circuits. However, the function of GABAergic neurons in the IC for memory reconsolidation remains unclear; therefore, we conducted this study to clarify this. MAIN METHODS: We applied morphine-induced conditioned place preference (mCPP) paradigm and pharmacogenetic techniques to study the mediation effect of GABAergic neurons in the IC on mCPP reconsolidation. Moreover, we preliminarily explored the possible mechanisms of mediating GABAergic neurons in the IC involved in mCPP reconsolidation by assessing Arc and Erg-1 protein levels in the IC. KEY FINDINGS: We found that post-retrieval immediate activation of GABAergic neurons in the IC impaired mCPP reconsolidation. In addition, this effect was not reversed by a priming morphine injection. Further, post-retrieval inhibition and non-retrieval excitation of GABAergic neurons in the IC had no effect on mCPP. SIGNIFICANCE: Taken together, our findings suggest that GABAergic neurons in the IC are closely involved in mCPP reconsolidation. Specifically, their excitation could eliminate established mCPP and prevent the relapse risk by disruption of the reconsolidation. The underlying molecular biological mechanisms could involve reduced Arc and Erg-1 levels.

Amentoflavone protects hippocampal neurons: anti-inflammatory, antioxidative, and antiapoptotic effects.

Amentoflavone is a natural biflavone compound with many biological properties, including anti-inflammatory, antioxidative, and neuroprotective effects. We presumed that amentoflavone exerts a neuroprotective effect in epilepsy models. Prior to model establishment, mice were intragastrically administered 25 mg/kg amentoflavone for 3 consecutive days. Amentoflavone effectively prevented pilocarpine-induced epilepsy in a mouse kindling model, suppressed nuclear factor-κB activation and expression, inhibited excessive discharge of hippocampal neurons resulting in a reduction in epileptic seizures, shortened attack time, and diminished loss and apoptosis of hippocampal neurons. Results suggested that amentoflavone protected hippocampal neurons in epilepsy mice via anti-inflammation, antioxidation, and antiapoptosis, and then effectively prevented the occurrence of seizures.

Proteomic response to acupuncture treatment in spontaneously hypertensive rats.

Previous animal and clinical studies have shown that acupuncture is an effective alternative treatment in the management of hypertension, but the mechanism is unclear. This study investigated the proteomic response in the nervous system to treatment at the Taichong (LR3) acupoint in spontaneously hypertensive rats (SHRs). Unanesthetized rats were subject to 5-min daily acupuncture treatment for 7 days. Blood pressure was monitored over 7 days. After euthanasia on the 7(th) day, rat medullas were dissected, homogenized, and subject to 2D gel electrophoresis and MALDI-TOF analysis. The results indicate that blood pressure stabilized after the 5th day of acupuncture, and compared with non-acupoint treatment, Taichong-acupunctured rat's systolic pressure was reduced significantly (P<0.01), though not enough to bring blood pressure down to normal levels. The different treatment groups also showed differential protein expression: the 2D images revealed 571 ± 15 proteins in normal SD rats' medulla, 576 ± 31 proteins in SHR's medulla, 597 ± 44 proteins in medulla of SHR after acupuncturing Taichong, and 616 ± 18 proteins in medulla of SHR after acupuncturing non-acupoint. In the medulla of Taichong group, compared with non-acupoint group, seven proteins were down-regulated: heat shock protein-90, synapsin-1, pyruvate kinase isozyme, NAD-dependent deacetylase sirtuin-2, protein kinase C inhibitor protein 1, ubiquitin hydrolase isozyme L1, and myelin basic protein. Six proteins were up-regulated: glutamate dehydrogenase 1, aldehyde dehydrogenase 2, glutathione S-transferase M5, Rho GDP dissociation inhibitor 1, DJ-1 protein and superoxide dismutase. The altered expression of several proteins by acupuncture has been confirmed by ELISA, Western blot and qRT-PCR assays. The results indicate an increase in antioxidant enzymes in the medulla of the SHRs subject to acupuncture, which may provide partial explanation for the antihypertensive effect of acupuncture. Further studies are warranted to investigate the role of oxidative stress modulation by acupuncture in the treatment of hypertension.

[Effect of moderate acupuncture-stimulation of "Taichong" (LR 3) on blood pressure and plasma endothelin-1 levels in spontaneous hypertension rats].

OBJECTIVE: To observe the effect of different strength of acupuncture stimulation on blood pressure and plasma endothelin (E)-1 in spontaneous hypertension rats (SHR) ,so as to seek a better acupuncture parameter for clinical treatment of hypertension. METHODS: Twenty-eight 9-week-old SHRs were randomized into mild-stimulation group, moderate-stimulation group, strong-stimulation group and model group (n = 7 in each group). Seven normotensive SD rats served as a normal control group. Acupuncture stimulation with mild, moderate and strong stimulation was applied to bilateral "Taichong" (LR 3) for 5 min, once daily for 7 days. Blood pressure (BP) was determined by using a non-invasive BP-6 detection system. Plasma ET-1 was assayed by radioimmunoassay. RESULTS: Compared with the model group, the systolic pressure of the moderate-stimulation group on the 6th and 7th day was decreased significantly after acupuncture of "Taichong" (LR 3) (P<0. 01), being significantly lower than that of the mild- and strong-stimulation groups (P<0. 01). In comparison with the normal control group, plasma ET-1 level in the model group was increased significantly (P<0.01), while compared with the model group, only that in the moderate-stimulation group was down-regulated considerably (P<0. 01). No significant differences were found between the mild-stimulation and model groups, between the strong-stimulation and model groups, and between the mild-stimulation and strong-stimulation groups in plasma ET-1 level (P>0. 05). CONCLUSION: Moderate-stimulation of "Taichong" (LR 3) can lower blood pressure and plasma EA-1 level in spontaneous hypertension rats. The reduced level of plasma ET-1 may be one of its mechanisms underlying improving hypertension.

Effects of phytoestrogen on mitochondrial structure and function of hippocampal CA1 region of ovariectomized rats.

The present study was undertaken to evaluate whether estrogen deprivation might lead to mitochondrial alteration of hippocampal neurons of ovariectomized (OVX) rats, and to evaluate the protective effect of estrogen and phytoestrogen on the mitochondrial alteration. First, OVX rats were used to mimic the pathologic changes of neurodegeneration of postmenopausal female, and we looked into the alteration of the mitochondrial ultrastructure and ATP content of hippocampal CA1 region after ovariectomy on different phase by transmission electron microscope (TEM) and reversed-phase high-performance liquid chromatography (HPLC), and found the best phase points of the alteration of the mitochondrial ultrastructure and ATP content. Next, estrogen and phytoestrogen were administered to the OVX rats for the protective effects on the mitochondrial ultrastructure and ATP content. Meanwhile, the density, size, shape, and distribution parameters of mitochondrial ultrastructure were analyzed according to the morphometry principle. The experimental results presented that (1) The alteration of mitochondrial ultrastructure elicited by ovariectomy worsened with the days going on, and the changes were the most noteworthy in volume density (Vv), average surface area (S), specific surface area (delta), and particle dispersity (Clambdaz) on 12th day (P < 0.05 or P < 0.01). Moreover, there was no statistical significance of the numerical density (Nv) among the five groups in the first step experiment. (2) The treatment with estrogen, genistein (Gs), and ipriflavone (Ip) significantly reversed the effect elicited by ovariectomy on Vv, S, delta, Clambdaz, Nv, and particle average diameter (D) of mitochondria of hippocampal CA1 region (P < 0.05). (3) Furthermore, ATP content of hippocampal CA1 region after ovariectomy declined significantly on 7th day (P < 0.05), and estrogen and phytoestrogen could reverse the alteration (P < 0.05). Taken together, these results revealed that phytoestrogen may have a protective role against the neurodegeneration after menopause via protecting mitochondrial structure and functions. Phytoestrogen may be a good alternative as a novel therapeutic strategy for menopausal syndrome.

[Study on the meridians and acupoints based on fasciaology: an elicitation of the study on digital human being].

OBJECTIVE: To explore the correlativity of meridians and acupoints with fascial system. METHODS: The lines and beads-like structures seen by fasciaology and scanning connective tissue, were combined with traditional meridians and collaterals, and acupoints to investigate channels, collaterals and acupoints. RESULTS: The high correlativity of the meridians and acupoints with the fascial system was found. CONCLUSION: The concept, functions, clinical application and mechanisms of meridians and acupoints can be preliminarily explained.