Activated somatostatin interneurons orchestrate memory microcircuits.

Despite recent advancements in identifying engram cells, our understanding of their regulatory and functional mechanisms remains in its infancy. To provide mechanistic insight into engram cell functioning, we introduced a novel local microcircuit labeling technique that enables the labeling of intraregional synaptic connections. Utilizing this approach, we discovered a unique population of somatostatin (SOM) interneurons in the mouse basolateral amygdala (BLA). These neurons are activated during fear memory formation and exhibit a preference for forming synapses with excitatory engram neurons. Post-activation, these SOM neurons displayed varying excitability based on fear memory retrieval. Furthermore, when we modulated these SOM neurons chemogenetically, we observed changes in the expression of fear-related behaviors, both in a fear-associated context and in a novel setting. Our findings suggest that these activated SOM interneurons play a pivotal role in modulating engram cell activity. They influence the expression of fear-related behaviors through a mechanism that is dependent on memory cues.

Reducing the Volume of Upper Eyelids in East Asians Increases Vertical Palpebral Height.

BACKGROUND: Correcting puffy eyelids is important for improving the first impression. The puffiness is most predictable corrected by tissue resection and fat excision. Fold asymmetry, overcorrection, and recurrence can sometimes occur after levator aponeurosis manipulation. The objective of this study was to introduce a method of volume-controlled blepharoptosis correction (VC) without levator manipulation. METHODS: The medical records of patients who had undergone upper blepharoplasty between 2017 and 2022 were retrospectively reviewed. Questionnaires, digital photographs, and charts were used to evaluate the surgical outcomes and complications. The degree of levator function was graded as poor, fair, good, or very good. Levator function must be above good (>8 mm) to employ the VC method. Poor and fair grades of levator function were excluded because they require levator aponeurosis manipulation. The margin to reflex distance (MRD) 1 was assessed preoperatively, 2 weeks postoperatively, and at follow-up visits. RESULTS: Postoperative satisfaction was 4.3 ± 0.8 with no postoperative discomfort (0%), and the duration of swelling was 10.1 ± 2.0 days. Regarding other complications, no fold asymmetry (0%) was observed, although hematoma formation was observed in 1 (2.9%) patient in the VC group. Significant differences were observed in the changes in palpebral fissure height over time (p < 0.001). CONCLUSIONS: VC can effectively correct puffy eyelids and create natural-looking, beautiful, and thin eyelids. Thus, VC is associated with higher patient satisfaction and surgical longevity without serious complications. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Caregiver burden according to ageing and type of care activity in caregivers of individuals with spinal cord injury.

OBJECTIVE: To describe caregiver burden according to the caregivers' general characteristics, especially with ageing, and type of care activities provided by caregivers of individuals with spinal cord injury. DESIGN: A cross-sectional study was conducted utilizing a structured questionnaire that included general characteristics, health conditions, and caregiver burden. SETTING: A single center study in Seoul, Korea. SUBJECTS: Participants were recruited from 87 individuals with spinal cord injuries and 87 caregivers. METHODS: The Caregiver Burden Inventory was used to assess caregiver burden. RESULTS: Caregiver burden was significantly different by age (p = 0.001), relationship (p = 0.025), sleep hours (p = <0.001), underlying disease (p = 0.018), pain (p = <0.001), and daily activities of individuals with spinal cord injury (p = 0.001). Caregiver's age (B = 0.339, p = 0.049), sleep duration (B = -2.896, p = 0.012) and pain (B = 2.558, p < 0.001) predicted caregiver burden. Toileting assistance was the most challenging and time-consuming for caregivers, while patient transfer was associated with the greatest concerns for body injury. CONCLUSION: Caregiver education should be targeted according to caregiver's age and type of assistance. Social policies need to be developed to distribute devices and care-robots to reduce caregiver burden and thereby assist caregivers.

Hippocampal engram networks for fear memory recruit new synapses and modify pre-existing synapses in vivo.

As basic units of neural networks, ensembles of synapses underlie cognitive functions such as learning and memory. These synaptic engrams show elevated synaptic density among engram cells following contextual fear memory formation. Subsequent analysis of the CA3-CA1 engram synapse revealed larger spine sizes, as the synaptic connectivity correlated with the memory strength. Here, we elucidate the synapse dynamics between CA3 and CA1 by tracking identical synapses at multiple time points by adapting two-photon microscopy and dual-eGRASP technique in vivo. After memory formation, synaptic connections between engram populations are enhanced in conjunction with synaptogenesis within the hippocampal network. However, extinction learning specifically correlated with the disappearance of CA3 engram to CA1 engram (E-E) synapses. We observed "newly formed" synapses near pre-existing synapses, which clustered CA3-CA1 engram synapses after fear memory formation. Overall, we conclude that dynamics at CA3 to CA1 E-E synapses are key sites for modification during fear memory states.

Synaptic ensembles between raphe and D1R-containing accumbens shell neurons underlie postisolation sociability in males.

Social animals expend considerable energy to maintain social bonds throughout their life. Male and female mice show sexually dimorphic behaviors, yet the underlying neural mechanisms of sociability and their dysregulation during social disconnection remain unknown. Dopaminergic neurons in dorsal raphe nucleus (DRNTH) is known to contribute to a loneliness-like state and modulate sociability. We identified that activated subpopulations in DRNTH and nucleus accumbens shell (NAcsh) during 24 hours of social isolation underlie the increase in isolation-induced sociability in male but not in female mice. This effect was reversed by chemogenetically and optogenetically inhibiting the DRNTH-NAcsh circuit. Moreover, synaptic connectivity among the activated neuronal ensembles in this circuit was increased, primarily in D1 receptor-expressing neurons in NAcsh. The increase in synaptic density functionally correlated with elevated dopamine release into NAcsh. Overall, specific synaptic ensembles in DRNTH-NAcsh mediate sex differences in isolation-induced sociability, indicating that sex-dependent circuit dynamics underlie the expression of sexually dimorphic behaviors.

Interrogating structural plasticity among synaptic engrams.

Our daily experiences and learnings are stored in the form of memories. These experiences trigger synaptic plasticity and persistent structural and functional changes in neuronal synapses. Recently, cellular studies of memory storage and engrams have emerged over the last decade. Engram cells reflect interconnected neurons via modified synapses. However, we were unable to observe the structural changes arising from synaptic plasticity in the past, because it was not possible to distinguish the synapses between engram cells. To overcome this barrier, dual-eGRASP (enhanced green fluorescent protein reconstitution across synaptic partners) technology can label specific synapses among multiple synaptic ensembles. Selective labeling of engram synapses elucidated their role by observing the structural changes in synapses according to the memory state. Dual-eGRASP extends cellular level engram studies to introduce the era of synaptic level studies. Here, we review this concept and possible applications of the dual-eGRASP, including recent studies that provided visual evidence of structural plasticity at the engram synapse.

The essence of the engram: Cellular or synaptic?

Memory is composed of various phases including cellular consolidation, systems consolidation, reconsolidation, and extinction. In the last few years it has been shown that simple association memories can be encoded by a subset of the neuronal population called engram cells. Activity of these cells is necessary and sufficient for the recall of association memory. However, it is unclear which molecular mechanisms allow cellular engrams to encode the diverse phases of memory. Further research is needed to examine the possibility that it is the synapses between engram cells (the synaptic engram) that constitute the memory. In this review we summarize recent findings on cellular engrams with a focus on different phases of memory, and discuss the distinct molecular mechanism required for cellular and synaptic engrams.

Synaptic correlates of associative fear memory in the lateral amygdala.

Successful adaptation to the environment requires an accurate response to external threats by recalling specific memories. Memory formation and recall require engram cell activity and synaptic strengthening among activated neuronal ensembles. However, elucidation of the underlying neural substrates of associative fear memory has remained limited without a direct interrogation of extinction-induced changes of specific synapses that encode a specific auditory fear memory. Using dual-eGRASP (enhanced green fluorescent protein reconstitution across synaptic partners), we found that synapses among activated neuronal ensembles or activated synaptic ensembles showed a significantly larger spine morphology at auditory cortex (AC)-to-lateral amygdala (LA) projections after auditory fear conditioning in mice. Fear extinction reversed these enhanced synaptic ensemble spines, whereas re-conditioning with the same tone and shock restored the spine size of the synaptic ensemble. We suggest that synaptic ensembles encode and represent different fear memory states.

Conditional knock out of transcription factor CTCF in excitatory neurons induces cognitive deficiency.

CCCTC-binding factor (CTCF) is a transcription factor that is involved in organizing chromatin structure. A reduction of CTCF expression is known to develop distinct clinical features. Furthermore, conditional knock out (cKO) study revealed reactive gliosis of astrocytes and microglia followed by age-dependent cell death in the excitatory neurons of CTCF cKO mice. To assess the cognitive ability in CTCF cKO mice of over 20 weeks of age, we examined pairwise discrimination (PD), PD reversal learning (PDr), and different paired-associate learning (dPAL) tasks using a touch screen apparatus. We found cognitive impairment in dPAL touch screen tests, suggesting that prolonged Ctcf gene deficiency results in cognitive deficits.

Loss of the neuronal genome organizer and transcription factor CTCF induces neuronal death and reactive gliosis in the anterior cingulate cortex.

CCCTC-binding factor (CTCF) is a genome organizer that regulates gene expression through transcription and chromatin structure regulation. CTCF also plays an important role during the developmental and adult stages. Cell-specific CTCF deletion studies have shown that a reduction in CTCF expression leads to the development of distinct clinical features and cognitive disorders. Therefore, we knocked out Ctcf (CTCF cKO) in the excitatory neurons of the forebrain in a Camk2a-Cre mouse strain to examine the role of CTCF in cell death and gliosis in the cortex. CTCF cKO mice were viable, but they demonstrated an age-dependent increase in reactive gliosis of astrocytes and microglia in the anterior cingulate cortex (ACC) from 16 weeks of age prior to neuronal loss observed at over 20 weeks of age. Consistent with these data, qRT-PCR analysis of the CTCF cKO ACC revealed changes in the expression of inflammation-related genes (Hspa1a, Prokr2 and Itga8) linked to gliosis and neuronal death. Our results suggest that prolonged Ctcf gene deficiency in excitatory neurons results in neuronal cell death and gliosis, possibly through functional changes in inflammation-related genes.

Reconstruction of medial epicanthal fold using v-y advancement and turnover flap.

BACKGROUND: Since the 1900s, many methods have been developed to correct the epicanthal fold. Increased use of epicanthoplasty has yielded unsatisfactory results. OBJECTIVES: To describe a method of epicanthal fold reconstruction using V-Y advancement and turnover flap for clinical application. This method is simpler than conventional surgery. It is easy to perform with excellent esthetic results. METHODS: The authors performed medial epicanthal fold reconstruction in 82 patients (16 males and 66 females) between April 2014 and September 2017. All patients enrolled in this retrospective study underwent surgical procedures at the authors' institution. Interepicanthal distance was the distance between medial epicanthal folds. It was measured with a surgical ruler. RESULTS: Before surgery, mean interepicanthal distance was 35.4 mm. Using our surgical technique, successful outcome was achieved in 79 (96.3%) patients with satisfactory results. The mean distance between the medial epicanthi post-surgery was 38.6 mm, increasing the total length by 3.2 mm without showing any major postsurgical complications. DISCUSSION: Epicanthal fold reconstruction using V-Y advancement and turnover flap is a simple and effective technique that can readily improve the frontal view. It improves periorbital contouring, makes eyes look natural without fully showing the caruncle, and yields excellent esthetic results. In particular, there were no major visible scars following eversion suture.

Incisional Ptosis Correction with Hidden Double Fold in Asian Patients.

BACKGROUND: Korean males and a few females desire to have larger eyes; however, they often wish to enlarge their eyes and conceal their double eyelids. This paper attempts to describe how to make the eyes bigger and brighter without showing double-fold eyelids. METHODS: The authors performed cosmetic ptosis correction in 121 cases from April 2013 to December 2017. All patients enrolled in this retrospective study underwent surgical procedures at the author's institutions. Patients were included that had mild-to-moderate degrees of ptosis and levator function greater than 5 mm, ages greater than 16 years, and no prior ptosis surgery. RESULTS: A successful outcome was achieved with this surgical approach in 113 (93.4%) patients. Complications potentially associated with ptosis surgery were not observed. DISCUSSION: A refined method of preoperative evaluation for incisional ptosis correction to conceal a double fold with no visible signs of surgery is described. Ptosis correction without the formation of double eyelids will result in skin hooding and visible scarring, and thus, it is recommended to lower the height of the double eyelids. The lower height of double eyelids can cover the incisional scar and make it appear there are no double eyelids. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Strengthened connections between engrams encode specific memories.

In previous studies, memory storage was localized to engram cells distributed across the brain. While these studies have provided an individual cellular profile of engram cells, their synaptic connectivity, or whether they follow Hebbian mechanisms, remains uncertain. Therefore, our recent study investigated whether synapses between engram cells exhibit selectively enhanced structural and functional properties following memory formation. This was accomplished using a newly developed technique called "dual-eGRASP". We found that the number and size of spines on CA1 engram cells that receive inputs from CA3 engram cells were larger than at other synapses. We further observed that this enhanced connectivity correlated with induced memory strength. CA3 engram synapses exhibited increased release probability, while CA1 engram synapses produced enhanced postsynaptic responses. CA3 engram to CA1 engram projections showed strong occlusion of long-term potentiation. We demonstrated that the synaptic connectivity of CA3 to CA1 engram cells was strengthened following memory formation. Our results suggest that Hebbian plasticity occurs during memory formation among engram cells at the synapse level. [BMB Reports 2018; 51(8): 369-370].

Remote Memory and Cortical Synaptic Plasticity Require Neuronal CCCTC-Binding Factor (CTCF).

The molecular mechanism of long-term memory has been extensively studied in the context of the hippocampus-dependent recent memory examined within several days. However, months-old remote memory maintained in the cortex for long-term has not been investigated much at the molecular level yet. Various epigenetic mechanisms are known to be important for long-term memory, but how the 3D chromatin architecture and its regulator molecules contribute to neuronal plasticity and systems consolidation is still largely unknown. CCCTC-binding factor (CTCF) is an 11-zinc finger protein well known for its role as a genome architecture molecule. Male conditional knock-out mice in which CTCF is lost in excitatory neurons during adulthood showed normal recent memory in the contextual fear conditioning and spatial water maze tasks. However, they showed remarkable impairments in remote memory in both tasks. Underlying the remote memory-specific phenotypes, we observed that female CTCF conditional knock-out mice exhibit disrupted cortical LTP, but not hippocampal LTP. Similarly, we observed that CTCF deletion in inhibitory neurons caused partial impairment of remote memory. Through RNA sequencing, we observed that CTCF knockdown in cortical neuron culture caused altered expression of genes that are highly involved in cell adhesion, synaptic plasticity, and memory. These results suggest that remote memory storage in the cortex requires CTCF-mediated gene regulation in neurons, whereas recent memory formation in the hippocampus does not.SIGNIFICANCE STATEMENT CCCTC-binding factor (CTCF) is a well-known 3D genome architectural protein that regulates gene expression. Here, we use two different CTCF conditional knock-out mouse lines and reveal, for the first time, that CTCF is critically involved in the regulation of remote memory. We also show that CTCF is necessary for appropriate expression of genes, many of which we found to be involved in the learning- and memory-related processes. Our study provides behavioral and physiological evidence for the involvement of CTCF-mediated gene regulation in the remote long-term memory and elucidates our understanding of systems consolidation mechanisms.

Interregional synaptic maps among engram cells underlie memory formation.

Memory resides in engram cells distributed across the brain. However, the site-specific substrate within these engram cells remains theoretical, even though it is generally accepted that synaptic plasticity encodes memories. We developed the dual-eGRASP (green fluorescent protein reconstitution across synaptic partners) technique to examine synapses between engram cells to identify the specific neuronal site for memory storage. We found an increased number and size of spines on CA1 engram cells receiving input from CA3 engram cells. In contextual fear conditioning, this enhanced connectivity between engram cells encoded memory strength. CA3 engram to CA1 engram projections strongly occluded long-term potentiation. These results indicate that enhanced structural and functional connectivity between engram cells across two directly connected brain regions forms the synaptic correlate for memory formation.

Clinical benefit and residual kidney function of en bloc nephrectomy for perirenal retroperitoneal sarcoma.

AIM: The purpose of this study was to evaluate the efficacy of en bloc nephrectomy for perirenal retroperitoneal sarcoma (RPS) with respect to postoperative kidney function and oncological benefits. METHODS: We performed a comparative study of 114 patients undergoing surgery for primary RPS, classifying cases as nephrectomy (NPX, n = 65) versus no nephrectomy (no-NPX, n = 49). The Δ and % change between preoperative and postoperative estimated glomerulus filtration rate (eGFR) were analyzed to compare renal function changes after surgery. Kaplan-Meier analysis was performed to verify the incidence of local relapse between the two groups. RESULTS: During a median follow-up of 29 months, median postoperative GFR of 65 patients in the NPX group decreased to 73.5% of preoperative eGFR. Although 38 patients (58%) in the NPX group experienced a progression in chronic kidney disease stage after nephrectomy, no patients progressed to end-stage renal disease (ESRD). In French Federation of Cancer Centers Sarcoma grade 2, the NPX group had statistically significant local control benefits, compared with the no-NPX group (P = 0.048). CONCLUSIONS: Residual renal function after en bloc nephrectomy was stabilized without progression to ESRD. Moreover, en bloc nephrectomy for perirenal RPS might secure a complete resection margin for local tumor control.

Deep Learning-Based Survival Analysis Identified Associations Between Molecular Subtype and Optimal Adjuvant Treatment of Patients With Gastric Cancer.

PURPOSE: Gastric cancer (GC) is the third-leading cause of cancer-related deaths. Several pivotal clinical trials of adjuvant treatments were performed during the previous decade; however, the optimal regimen for adjuvant treatment of GC remains controversial. PATIENTS AND METHODS: We developed a novel deep learning-based survival model (survival recurrent network [SRN]) in patients with GC by including all available clinical and pathologic data and treatment regimens. This model uses time-sequential data only in the training step, and upon being trained, it receives the initial data from the first visit and then sequentially predicts the outcome at each time point until it reaches 5 years. In total, 1,190 patients from three cohorts (the Asian Cancer Research Group cohort, n = 300; the fluorouracil, leucovorin, and radiotherapy cohort, n = 432; and the Adjuvant Chemoradiation Therapy in Stomach Cancer cohort, n = 458) were included in the analysis. In addition, we added Asian Cancer Research Group molecular classifications into the prediction model. SRN simulated the sequential learning process of clinicians in the outpatient clinic using a recurrent neural network and time-sequential outcome data. RESULTS: The mean area under the receiver operating characteristics curve was 0.92 ± 0.049 at the fifth year. The SRN demonstrated that GC with a mesenchymal subtype should elicit a more risk-adapted postoperative treatment strategy as a result of its high recurrence rate. In addition, the SRN found that GCs with microsatellite instability and GCs of the papillary type exhibited significantly more favorable survival outcomes after capecitabine plus cisplatin chemotherapy alone. CONCLUSION: Our SRN predicted survival at a high rate, reaching 92% at postoperative year 5. Our findings suggest that SRN-based clinical trials or risk-adapted adjuvant trials could be considered for patients with GC to investigate more individualized adjuvant treatments after curative gastrectomy.

Clinical Application of Targeted Deep Sequencing in Solid-Cancer Patients and Utility for Biomarker-Selected Clinical Trials.

Molecular profiling of actionable mutations in refractory cancer patients has the potential to enable "precision medicine," wherein individualized therapies are guided based on genomic profiling. The molecular-screening program was intended to route participants to different candidate drugs in trials based on clinical-sequencing reports. In this screening program, we used a custom target-enrichment panel consisting of cancer-related genes to interrogate single-nucleotide variants, insertions and deletions, copy number variants, and a subset of gene fusions. From August 2014 through April 2015, 654 patients consented to participate in the program at Samsung Medical Center. Of these patients, 588 passed the quality control process for the 381-gene cancer-panel test, and 418 patients were included in the final analysis as being eligible for any anticancer treatment (127 gastric cancer, 122 colorectal cancer, 62 pancreatic/biliary tract cancer, 67 sarcoma/other cancer, and 40 genitourinary cancer patients). Of the 418 patients, 55 (12%) harbored a biomarker that guided them to a biomarker-selected clinical trial, and 184 (44%) patients harbored at least one genomic alteration that was potentially targetable. This study demonstrated that the panel-based sequencing program resulted in an increased rate of trial enrollment of metastatic cancer patients into biomarker-selected clinical trials. Given the expanding list of biomarker-selected trials, the guidance percentage to matched trials is anticipated to increase. IMPLICATIONS FOR PRACTICE: This study demonstrated that the panel-based sequencing program resulted in an increased rate of trial enrollment of metastatic cancer patients into biomarker-selected clinical trials. Given the expanding list of biomarker-selected trials, the guidance percentage to matched trials is anticipated to increase.

Foreign body reaction from anti-adhesion material during follow-up of gynaecological malignancies: Mimicking local recurrence.

BACKGROUND: Gynaecological surgery techniques and instruments have advanced and there is increasing effort to reduce operative complications by preventing adhesion complications after surgery. AIMS: The purpose of this study was to evaluate a possible limitation of the Surgi-Wrap(®) anti-adhesion material during clinical follow-up of surgically managed gynaecological malignancies. METHODS: We retrospectively analysed the medical records and imaging findings of 92 patients who received the Surgi-Wrap(®) anti-adhesion material. RESULTS: Nine of the 92 patients had local recurrence based on the imaging findings. The positive imaging findings showed focal, isolated and small pelvic lesions without other distant metastasis or recurrence and normal tumour marker levels. Laparoscopic exploration and biopsy were performed in six patients and close clinical follow-up was performed for the other three patients, who had a strong diagnostic impression of a foreign body reaction mimicking a focal recurrence of the tumour. The histological findings of the six laparoscopically-explored patients revealed a foreign body reaction without malignancy in five and recurrence in one case. The rate of foreign body reaction, mimicking a local recurrence, was 5/92 (5.4% of histologically confirmed cases) and 8/92 cases had ambiguous findings between a foreign reaction and local recurrence (8.7% of clinically suspected cases). CONCLUSIONS: It is important to avoid confusion between benign and recurrent conditions during follow-up for gynaecological malignancies. We suggest avoiding use of Surgi-wrap(®) during cancer surgeries and a need for further studies on the safety of Surgi-wrap(®) in patients with cancer.

Multiple repressive mechanisms in the hippocampus during memory formation.

Memory stabilization after learning requires translational and transcriptional regulations in the brain, yet the temporal molecular changes that occur after learning have not been explored at the genomic scale. We used ribosome profiling and RNA sequencing to quantify the translational status and transcript levels in the mouse hippocampus after contextual fear conditioning. We revealed three types of repressive regulations: translational suppression of ribosomal protein-coding genes in the hippocampus, learning-induced early translational repression of specific genes, and late persistent suppression of a subset of genes via inhibition of estrogen receptor 1 (ESR1/ERα) signaling. In behavioral analyses, overexpressing Nrsn1, one of the newly identified genes undergoing rapid translational repression, or activating ESR1 in the hippocampus impaired memory formation. Collectively, this study unveils the yet-unappreciated importance of gene repression mechanisms for memory formation.