STED Imaging of Vesicular Endocytosis in the Synapse.

Endocytosis is a fundamental biological process that couples exocytosis to maintain the homeostasis of the plasma membrane and sustained neurotransmission. Super-resolution microscopy enables optical imaging of exocytosis and endocytosis in live cells and makes an essential contribution to understanding molecular mechanisms of endocytosis in neuronal somata and other types of cells. However, visualization of exo-endocytic events at the single vesicular level in a synapse with optical imaging remains a great challenge to reveal mechanisms governing the synaptic exo-endocytotic coupling. In this protocol, we describe the technical details of stimulated emission depletion (STED) imaging of synaptic endocytosis at the single-vesicle level, from sample preparation and microscopy calibration to data acquisition and analysis.

The thalamic reticular nucleus orchestrates social memory.

Social memory has been developed in humans and other animals to recognize familiar conspecifics and is essential for their survival and reproduction. Here, we demonstrated that parvalbumin-positive neurons in the sensory thalamic reticular nucleus (sTRNPvalb) are necessary and sufficient for mice to memorize conspecifics. sTRNPvalb neurons receiving glutamatergic projections from the posterior parietal cortex (PPC) transmit individual information by inhibiting the parafascicular thalamic nucleus (PF). Mice in which the PPCCaMKII→sTRNPvalb→PF circuit was inhibited exhibited a disrupted ability to discriminate familiar conspecifics from novel ones. More strikingly, a subset of sTRNPvalb neurons with high electrophysiological excitability and complex dendritic arborizations is involved in the above corticothalamic pathway and stores social memory. Single-cell RNA sequencing revealed the biochemical basis of these subset cells as a robust activation of protein synthesis. These findings elucidate that sTRNPvalb neurons modulate social memory by coordinating a hitherto unknown corticothalamic circuit and inhibitory memory engram.

10 W super-wideband ultra-low-intensity-noise single-frequency fiber laser at 1 µm.

A 10 W super-wideband ultra-low-intensity-noise single-frequency fiber laser (SFFL) at 1 µm is experimentally demonstrated, based on dual gain saturation effects from semiconductors and optical fibers, together with an analog-digital hybrid optoelectronic feedback loop. Three intensity-noise-inhibited units synergistically work, which actualizes a connection of effective bandwidth and enhancement of noise-suppressing amplitude. With the cascade action of the semiconductor optical amplifier and optical fiber amplifier, the laser power is remarkably boosted. Eventually, an SFFL with an output power of 10.8 W and a relative intensity noise (RIN) below -150 dB/Hz at the frequency range over 1 Hz is realized. More meaningfully, within the total frequency range of 10 Hz to 10 GHz exceeding 29 octaves, the RIN is controlled to below -160 dB/Hz, approaching the shot-noise limit (SNL) level. To the best of our knowledge, this is the lowest RIN result of SFFL within such an extensive frequency range, and this is the highest output power of the near-SNL super-wideband SFFL. Furthermore, a linewidth of less than 0.8 kHz, a long-term stable polarization extinction ratio of 20 dB, and an optical signal-to-noise ratio of over 60 dB are obtained simultaneously. This start-of-the-art SFFL has provided a systematic solution for high-power and low-noise light sources, which is competitive for sophisticated applications, such as free-space laser communication, space-based gravitational wave detection, and super-long-distance space coherent velocity measurement and ranging.

Frequency-stabilized improvement of saturated absorption spectroscopy based on laser linewidth-control strategy.

Single-frequency fiber lasers (SFFLs), 1083 nm, have been extensively applied in 4He optical pumping magnetometers (OPMs) for magnetic field detection. However, the sensitivity and accuracy of OPMs are constrained by the frequency stability of SFFLs. Focusing on this concern, the frequency-stabilized performance of the 1083 nm SFFLs is successfully improved by externally tailoring the laser linewidth to match the spectral width of the error signal in saturated absorption spectroscopy. Thereinto, a high-intensity error signal of saturated absorption is generated as a large number of 4He atoms with a wide range of velocities interacting with the 1083 nm laser. Consequently, the root mean square value of the fluctuating frequency after locking is effectively decreased from 24.6 to 13.6 kHz, which achieves a performance improvement of 44.7%. Such a strategy can provide a technical underpinning for effectuating an absolute frequency stabilization with higher precision based on atomic and molecular absorption spectroscopy techniques.

An ACC-VTA-ACC positive-feedback loop mediates the persistence of neuropathic pain and emotional consequences.

The central mechanisms underlying pain chronicity remain elusive. Here, we identify a reciprocal neuronal circuit in mice between the anterior cingulate cortex (ACC) and the ventral tegmental area (VTA) that mediates mutual exacerbation between hyperalgesia and allodynia and their emotional consequences and, thereby, the chronicity of neuropathic pain. ACC glutamatergic neurons (ACCGlu) projecting to the VTA indirectly inhibit dopaminergic neurons (VTADA) by activating local GABAergic interneurons (VTAGABA), and this effect is reinforced after nerve injury. VTADA neurons in turn project to the ACC and synapse to the initial ACCGlu neurons to convey feedback information from emotional changes. Thus, an ACCGlu-VTAGABA-VTADA-ACCGlu positive-feedback loop mediates the progression to and maintenance of persistent pain and comorbid anxiodepressive-like behavior. Disruption of this feedback loop relieves hyperalgesia and anxiodepressive-like behavior in a mouse model of neuropathic pain, both acutely and in the long term.

Survival outcomes of 2018 FIGO stage IIIC versus stages IIIA and IIIB in cervical cancer: A systematic review with meta-analysis.

OBJECTIVE: To assess the difference in survival outcomes between stage IIIC and stages IIIA and IIIB in the 2018 FIGO cervical cancer staging system. METHODS: The PubMed, EMBASE, MEDLINE and Web of Science were searched for articles published from November 1, 2018 to January 31, 2023. Articles published in English were considered. The included studies compared the survival outcomes of patients with cervical cancer in FIGO 2018 stage IIIC with those in stages IIIA and IIIB. Studies focused on rare histopathological types were excluded. The statistical analyses were performed using Stata 17 software. The endpoints were overall survival (OS) and progression-free survival (PFS). RESULTS: Ten retrospective cohort studies were eligible, involving 2113 (6.2%), 9812 (28.6%), 44 (0.1%), 10 171 (29.7%), 11 677 (34.1%) and 445 (1.3%) patients in stage IIIA, IIIB, IIIA&B, IIIC, IIIC1, and IIIC2, respectively. In the OS group, stage IIIC/C1 was significantly associated with superior survival compared with stage IIIA (hazard risk [HR] 0.62, 95% confidence interval [CI] 0.41-0.93, P = 0.022; I2 = 92.9%) and stage IIIB(A&B) (HR 0.56, 95% CI 0.44-0.71, P < 0.001; I2 = 94.0%). The FIGO 2018 stage IIIC2 was not associated with an increased mortality risk compared with stage IIIA and stage IIIB(A&B). In the PFS group, the outcome of FIGO 2018 stage IIIC/C1 was similar to stage IIIA (HR 0.66, 95% CI 0.27-1.64, P = 0.371; I2 = 65.6%), but better than stage IIIB(A&B) (HR 0.75, 95% CI 0.68-0.83, P < 0.001; I2 = 0.0%). The FIGO 2018 stage IIIC2 has similar PFS outcomes to stage IIIA and stage IIIB(A&B). CONCLUSION: Our findings demonstrate that survival outcomes of stage IIIC are no worse than those of stage IIIA and stage IIIB in the 2018 FIGO cervical cancer staging system. In cervical cancer, FIGO 2018 stage IIIC1 has significantly better OS outcomes than stage IIIA and stage IIIB.

Ultrafine electro-optical frequency comb based on cascade phase modulation with cyclic frequency shifting.

An ultrafine electro-optical frequency comb (EOFC) with plentiful comb teeth is demonstrated. Adopting a single-frequency fiber laser as a light source, cascade phase modulation based on a sinusoidal signal and a frequency-time transformation (FTT) signal is executed to generate the EOFC with high fineness. Meanwhile, a cyclic fast frequency shifting strategy is introduced to boost the number of comb teeth and the bandwidth of the EOFC. As a result, an EOFC with 12600 comb lines covering a broad bandwidth from -6.3 GHz to 6.3 GHz is established, corresponding to an ultrafine comb space of 1 MHz. Moreover, the power fluctuation of a comb tooth is less than 0.5 dBm. This state-of-the-art EOFC has significant potential in the field of precision spectroscopy.

Chinese Mothers' Intent to Disclose Their HIV Status to Their Children: The Role of Positive Outcome Expectations and Reward Responsiveness.

Objective: The current study investigated the role of positive outcome expectations and reward responsiveness in intention to disclose HIV status to children among women living with HIV in China. The moderating role of reward responsiveness was also explored. Method: A 1-year longitudinal survey was conducted. 269 women living with HIV who had at least one child aged >5 years and had not yet disclosed their HIV status to their oldest child were selected from a larger sample of women living with HIV at baseline, with a total of 261 respondents completing the follow-up survey. Results: After adjusting for significant socio-demographic and medical variables, positive outcome expectations positively predicted mothers' intention to disclose HIV, while reward responsiveness had a negative effect. A moderation effect of reward responsiveness was found, with further analysis showing that reward responsiveness has strengthened the relationship between positive outcome expectations and intention to disclose HIV. Conclusion: Findings support the relevance of positive outcome expectations and reward responsiveness to intention of disclosure among women living with HIV in China.

Cocaine induces locomotor sensitization through a dopamine-dependent VTA-mPFC-FrA cortico-cortical pathway in male mice.

As a central part of the mammalian brain, the prefrontal cortex (PFC) has been implicated in regulating cocaine-induced behaviors including compulsive seeking and reinstatement. Although dysfunction of the PFC has been reported in animal and human users with chronic cocaine abuse, less is known about how the PFC is involved in cocaine-induced behaviors. By using two-photon Ca2+ imaging to simultaneously record tens of intact individual networking neurons in the frontal association cortex (FrA) in awake male mice, here we report that a systematic acute cocaine exposure decreased the FrA neural activity in mice, while the chemogenetic intervention blocked the cocaine-induced locomotor sensitization. The hypoactivity of FrA neurons was critically dependent on both dopamine transporters and dopamine transmission in the ventromedial PFC (vmPFC). Both dopamine D1R and D2R neurons in the vmPFC projected to and innervated FrA neurons, the manipulation of which changed the cocaine-induced hypoactivity of the FrA and locomotor sensitization. Together, this work demonstrates acute cocaine-induced hypoactivity of FrA neurons in awake mice, which defines a cortico-cortical projection bridging dopamine transmission and cocaine sensitization.

Simultaneous achievement of power boost and low-frequency intensity noise suppression in a bidirectional pumping fiber amplifier based on saturated even-distribution gain.

An optimized bidirectional pumping fiber amplifier is demonstrated to achieve low-frequency intensity noise suppression and effective power enhancement simultaneously. Based on the concept analysis of the gain saturation effect, the influence of input signal power and pump power on intensity noise suppression is investigated and optimized systematically. Further combining with the optimization of the pumping configuration to achieve the even-distribution gain, the relative intensity noise (RIN) of 1083 nm single-frequency fiber laser (SFFL) is suppressed with 9.1 dB in the frequency range below 10 kHz. Additionally, the laser power is boosted from 10.97 dBm to 25.02 dBm, and a power instability of ±0.31% is realized. This technology has contributed to simultaneously improving the power and noise performance of the 1083 nm SFFL, which can be applied to a multi-channel helium (He) optically pumping magnetometer. Furthermore, this technique has broken the mindset that power amplification of the conventional fiber amplifiers will inevitably cause the degradation of intensity noise property, and provided a valuable guidance for the development of high-performance SFFLs.

Screening potential anaphylactoid components in vinpocetine injection using a high expression Mas-related G-protein-coupled receptor X2 cell membrane chromatography.

Vinpocetine injection is often used in clinical treatment of acute cardiovascular and cerebrovascular diseases. However, it was reported that vinpocetine injection caused allergic reactions in clinical use; therefore, its safety needs urgent attention. Until now, research on its sensitization is rarely reported. Here, the components contained in three vinpocetine injections were examined. It was found that besides vinpocetine, the synthetic raw material vincamine, the excipients benzyl alcohol and ethyl p-toluenesulfonate, and the impurities A, B, C, and D, which are excipients specified in the European Pharmacopoeia, were also present in them. Then the Mas-related G-protein-coupled receptor X2 (MRGPRX2)-HEK293 cell membrane chromatography was used to investigate the affinity of them with MRGPRX2 and found that vinpocetine, vincamine, and impurities A, B, C, and D bind to MRGPRX2. Afterwards, these compounds were further used to investigate the local sensitization ability in vivo. The results showed that vinpocetine, vincamine, and impurity C could induce swelling of the paw and decrease body temperature in mice, but only impurity C could cause local skin mast cell degranulation and serum histamine release increase. In vitro, the results also indicated that impurity C could increase intracellular [Ca2+ ] in MRGPRX2-HEK293 cells, whereas vinpocetine and vincamine did not. Therefore, the impurity C was the potential anaphylactoid component in vinpocetine injection, which may be one of the reasons for the occurrence of allergic reactions in the clinical use of vinpocetine injection. This work provides evidence on the sensitization of impurity C and also contributes to promoting the clinical safety of vinpocetine injection.

Impact of cystectomy versus ablation for endometrioma on ovarian reserve: a systematic review and meta-analysis.

OBJECTIVE: To investigate whether cystectomy or ablation for endometrioma has less impact on ovarian reserve as evaluated by antral follicle count (AFC) and antimüllerian hormone (AMH) levels. DESIGN: Systematic review and meta-analysis. SETTING: Not applicable. PATIENT(S): Patients with endometriomas undergoing cystectomy or ablation. INTERVENTION(S): All prospective studies comparing cystectomy with ablation for endometrioma in the PubMed, EMBASE, MEDLINE and Web of Science until April 3, 2022 were retrieved and reviewed. Medical treatment used as adjuvant therapy for the surgery was excluded. Two authors assessed eligibility and risk of bias independently. The statistical data were pooled using the Review Manager software. MAIN OUTCOME MEASURE(S): The changes of AMH levels and AFC values in cystectomy group and ablation group, including intergroup comparisons and intragroup comparisons. RESULT(S): Four randomized clinical trials and 2 prospective cohort studies were eligible for the meta-analysis, with a total of 294 patients. In the intergroup comparisons, preoperative AFC values were similar with low heterogeneity, but postoperative AFC values were significantly lower in cystectomy than ablation (mean differences [MD], -1.33; 95% credible interval, -2.15 to -0.51; I2 = 57%). In the intragroup comparisons of AFC values, sensitivity analyses showed a significant decrease in cystectomy (MD, -1.93; 95% credible interval, -2.40 to -1.45; I2 = 0%) at 6-month follow-up, compared with no reduction in ablation. The intragroup comparisons of AMH levels supported negative effects on ovarian reserve of both cystectomy (MD, -1.26; 95% credible interval, -1.64 to -0.88; I2 = 45%) and ablation (MD, -0.70; 95% credible interval, -1.07 to -0.32; I2 = 0%). CONCLUSION(S): Both ablation and cystectomy have significantly detrimental effects on ovarian reserve as evaluated by AMH, but the ablation causes relatively less damage to ovarian reserve as appraised by AFC. CLINICAL TRIAL REGISTRATION NUMBER: CRD42020152823;PROSPERO (york.ac.uk).

Ca2+ -independent transmission at the central synapse formed between dorsal root ganglion and dorsal horn neurons.

A central principle of synaptic transmission is that action potential-induced presynaptic neurotransmitter release occurs exclusively via Ca2+ -dependent secretion (CDS). The discovery and mechanistic investigations of Ca2+ -independent but voltage-dependent secretion (CiVDS) have demonstrated that the action potential per se is sufficient to trigger neurotransmission in the somata of primary sensory and sympathetic neurons in mammals. One key question remains, however, whether CiVDS contributes to central synaptic transmission. Here, we report, in the central transmission from presynaptic (dorsal root ganglion) to postsynaptic (spinal dorsal horn) neurons in vitro, (i) excitatory postsynaptic currents (EPSCs) are mediated by glutamate transmission through both CiVDS (up to 87%) and CDS; (ii) CiVDS-mediated EPSCs are independent of extracellular and intracellular Ca2+ ; (iii) CiVDS is faster than CDS in vesicle recycling with much less short-term depression; (iv) the fusion machinery of CiVDS includes Cav2.2 (voltage sensor) and SNARE (fusion pore). Together, an essential component of activity-induced EPSCs is mediated by CiVDS in a central synapse.

Over-20-octaves-bandwidth ultralow-intensity-noise 1064-nm single-frequency fiber laser based on a comprehensive all-optical technique.

An over-20-octaves-bandwidth ultralow-intensity-noise 1064-nm single-frequency fiber laser (SFFL) is demonstrated based on a comprehensive all-optical technique. With a joint action of booster optical amplifier (BOA) and reflective Yb-doped fiber amplifier (RYDFA), two-fold optical gain saturation effects, respectively occurring in the media of semiconductor and fiber, have been synthetically leveraged. Benefiting from the gain dynamics in complementary time scales, i.e., nanosecond-order carrier lifetime in BOA and millisecond-order upper-level lifetime in RYDFA, the relative intensity noise (RIN) is reduced to -150 dB/Hz from 0.2 kHz to 350 MHz, which exceeds 20-octaves bandwidth. Remarkably, a maximum suppressing ratio of >54 dB is obtained, and the RIN in the range of 0.09-10 GHz reaches -161 dB/Hz which is only 2.3 dB above the shot-noise limit. This broad-bandwidth ultralow-intensity-noise SFFL can serve as an important building block for squeezed light generation, space laser communication, space gravitational wave detection, etc.

Cocaine increases quantal norepinephrine secretion through NET-dependent PKC activation in locus coeruleus neurons.

The norepinephrine neurons in locus coeruleus (LC-NE neurons) are essential for sleep arousal, pain sensation, and cocaine addiction. According to previous studies, cocaine increases NE overflow (the profile of extracellular NE level in response to stimulation) by blocking the NE reuptake. NE overflow is determined by NE release via exocytosis and reuptake through NE transporter (NET). However, whether cocaine directly affects vesicular NE release has not been directly tested. By recording quantal NE release from LC-NE neurons, we report that cocaine directly increases the frequency of quantal NE release through regulation of NET and downstream protein kinase C (PKC) signaling, and this facilitation of NE release modulates the activity of LC-NE neurons and cocaine-induced stimulant behavior. Thus, these findings expand the repertoire of mechanisms underlying the effects of cocaine on NE (pro-release and anti-reuptake), demonstrate NET as a release enhancer in LC-NE neurons, and provide potential sites for treatment of cocaine addiction.

Maternal Methamphetamine Exposure Influences Behavioral Sensitization and Nucleus Accumbens DNA Methylation in Subsequent Generation.

The deleterious effects of methamphetamine (METH) exposure extend beyond abusers, and may potentially impact the vulnerability of their offspring in developing addictive behaviors. Epigenetic signatures have been implicated in addiction, yet the characteristics to identify prenatal METH abuse to offspring addiction risk remains elusive. Here, we used escalating doses of METH-exposed mouse model in F0 female mice before and during pregnancy to simulate the human pattern of drug abuse and generated METH-induced behavioral sensitization to investigate the addictive behavior in offspring mice. We then utilized whole genome-bisulfite sequencing (WGBS) to investigate the methylation signature of nucleus accumbens (NAc) in male METH-sensitized mice. Interestingly, male but not female offspring exhibited an enhanced response to METH-induced behavioral sensitization. Additionally, the METH-exposed group of male mice underwent a more comprehensive wave of epigenome remodeling over all genomic elements compared with unexposed groups due to drug exposure history. 104,219 DMCs (METH-SAL vs. SAL-SAL) induced by prenatal METH-exposure were positively correlated with that of postnatal METH-exposure (38,570, SAL-METH vs. SAL-SAL). Moreover, 4,983 DMCs induced by pre- and postnatal METH exposure (METH-METH vs. SAL-METH) were negatively correlated with that of postnatal METH exposure, and 371 commonly changed DMCs between the two comparison groups also showed a significantly negative correlation and 86 annotated genes functionally enriched in the pathways of neurodevelopment and addiction. Key annotated genes included Kirrel3, Lrpprc, and Peg3, implicated in neurodevelopmental processes, were down-regulated in METH-METH group mice compared with the SAL-METH group. Taken together, we render novel insights into the epigenetic correlation of drug exposure and provide evidence for epigenetic characteristics that link maternal METH exposure to the intensity of the same drug-induced behavioral sensitization in adult offspring.

Tuning the Size of Large Dense-Core Vesicles and Quantal Neurotransmitter Release via Secretogranin II Liquid-Liquid Phase Separation.

Large dense-core vesicles (LDCVs) are larger in volume than synaptic vesicles, and are filled with multiple neuropeptides, hormones, and neurotransmitters that participate in various physiological processes. However, little is known about the mechanism determining the size of LDCVs. Here, it is reported that secretogranin II (SgII), a vesicle matrix protein, contributes to LDCV size regulation through its liquid-liquid phase separation in neuroendocrine cells. First, SgII undergoes pH-dependent polymerization and the polymerized SgII forms phase droplets with Ca2+ in vitro and in vivo. Further, the Ca2+ -induced SgII droplets recruit reconstituted bio-lipids, mimicking the LDCVs biogenesis. In addition, SgII knockdown leads to significant decrease of the quantal neurotransmitter release by affecting LDCV size, which is differently rescued by SgII truncations with different degrees of phase separation. In conclusion, it is shown that SgII is a unique intravesicular matrix protein undergoing liquid-liquid phase separation, and present novel insights into how SgII determines LDCV size and the quantal neurotransmitter release.