Developmental refinement of the active zone nanotopography and axon wiring at the somatosensory thalamus.

Functional refinement of neural circuits is a crucial developmental process in the brain. However, how synaptic maturation and axon wiring proceed cooperatively to establish reliable signal transmission is unclear. Here, we combined nanotopography of release machinery at the active zone (AZ), nanobiophysics of neurotransmitter release, and single-neuron reconstruction of axon arbors of lemniscal fibers (LFs) in the developing mouse somatosensory thalamus. With development, the cluster of Cav2.1 enlarges and translocates closer to vesicle release sites inside the bouton, and LFs drastically shrink their arbors and form larger boutons on the perisomatic region of target neurons. Experimentally constrained simulations show that the nanotopography of mature synapses enables not only rapid vesicular release but also reliable transmission following repetitive firing. Sensory deprivation impairs the developmental shift of molecular nanotopography and axon wiring. Thus, we uncovered the cooperative nanotopographical and morphological mechanisms underlying the developmental establishment of reliable synaptic transmission.

Diagnostic Imaging Performance of Dual-Energy Computed Tomography Compared with Conventional Computed Tomography and Magnetic Resonance Imaging for Uterine Cervical Cancer.

Objective This article evaluates the ability of low-energy (40 keV) virtual monoenergetic images (VMIs) in the local diagnosis of cervical cancer compared with that of conventional computed tomography (C-CT) and magnetic resonance imaging (MRI), using clinicopathologic staging as a reference. Methods This prospective study included 33 patients with pathologically confirmed cervical cancer who underwent dual-energy CT and MRI between 2021 and 2022. The contrast-to-noise ratio (CNR) of the tumor-to-myometrium was compared between C-CT and VMI. Additionally, sensitivity, specificity, and area under the receiver operating characteristic curve (AUC) for each local diagnostic parameter were compared between C-CT, VMI, and MRI. Interradiologist agreement was also assessed. Results The mean CNR was significantly higher on VMI ( p = 0.002). No significant difference in AUC was found between C-CT and VMI for all local diagnostic parameters, and the specificity of VMI was often significantly less than that of MRI. For parametrial invasion, mean sensitivity, specificity, and AUC for C-CT, VMI, and MRI were 0.81, 0.99, 0.93; 0.64, 0.35, 0.79; and 0.73, 0.67, 0.86, respectively, and MRI had significantly higher specificity and AUC than that of VMI ( p = 0.013 and 0.008, respectively). Interradiologist agreement was higher for VMI than C-CT and for MRI than VMI. Conclusion The CNR of VMI was significantly higher than C-CT and interradiologist agreement was better than with C-CT; however, the overall diagnostic performance of VMI did not significantly differ from C-CT and was inferior to MRI. VMI was characterized by low specificity, which should be understood and used for reading.

Comparison of preoperative diagnostic performance between dual-energy CT, conventional CT, and MRI in endometrial cancer.

Purpose: To compare the diagnostic performance of virtual monoenergetic imaging (VMI), computed tomography (CT), and magnetic resonance imaging (MRI) in patients with endometrial cancer (EC). Material and methods: This retrospective study analysed 45 EC patients (mean age: 62 years, range: 44-84 years) undergoing contrast-enhanced CT with dual-energy CT (DECT) and MRI between September 2021 and October 2022. Dual-energy CT generated conventional CT (C-CT) and 40 keV VMI. Quantitative analysis compared contrast-to-noise ratio (CNR) of tumour to myometrium between C-CT and VMI. Qualitative assessment by 5 radiologists compared C-CT, VMI, and MRI for myometrial invasion (MI), cervical invasion, and lymph node metastasis. Sensitivity, specificity, accuracy, and area under the receiver operating characteristic curve (AUC) were calculated and compared for each diagnostic parameter. Results: Virtual monoenergetic imaging showed significantly higher CNR than C-CT (p < 0.001) and a higher sensitivity for MI than C-CT (p = 0.027) and MRI (p = 0.011) but lower specificity than MRI (p = 0.018). C-CT had a higher sensitivity and AUC for cervical invasion than MRI (p = 0.018 and 0.004, respectively). Conclusions: The study found no significant superiority of MRI over CT across all diagnostic parameters. VMI demonstrated heightened sensitivity for MI, and C-CT showed greater sensitivity and AUC for cervical invasion than MRI. This suggests that combining VMI with C-CT holds promise as a comprehensive preoperative staging tool for EC when MRI cannot be performed.

Direct and indirect pathways for heterosynaptic interaction underlying developmental synapse elimination in the mouse cerebellum.

Developmental synapse elimination is crucial for shaping mature neural circuits. In the neonatal mouse cerebellum, Purkinje cells (PCs) receive excitatory synaptic inputs from multiple climbing fibers (CFs) and synapses from all but one CF are eliminated by around postnatal day 20. Heterosynaptic interaction between CFs and parallel fibers (PFs), the axons of cerebellar granule cells (GCs) forming excitatory synapses onto PCs and molecular layer interneurons (MLIs), is crucial for CF synapse elimination. However, mechanisms for this heterosynaptic interaction are largely unknown. Here we show that deletion of AMPA-type glutamate receptor functions in GCs impairs CF synapse elimination mediated by metabotropic glutamate receptor 1 (mGlu1) signaling in PCs. Furthermore, CF synapse elimination is impaired by deleting NMDA-type glutamate receptors from MLIs. We propose that PF activity is crucial for CF synapse elimination by directly activating mGlu1 in PCs and indirectly enhancing the inhibition of PCs through activating NMDA receptors in MLIs.

Developmental trajectories of GABAergic cortical interneurons are sequentially modulated by dynamic FoxG1 expression levels.

GABAergic inhibitory interneurons, originating from the embryonic ventral forebrain territories, traverse a convoluted migratory path to reach the neocortex. These interneuron precursors undergo sequential phases of tangential and radial migration before settling into specific laminae during differentiation. Here, we show that the developmental trajectory of FoxG1 expression is dynamically controlled in these interneuron precursors at critical junctures of migration. By utilizing mouse genetic strategies, we elucidate the pivotal role of precise changes in FoxG1 expression levels during interneuron specification and migration. Our findings underscore the gene dosage-dependent function of FoxG1, aligning with clinical observations of FOXG1 haploinsufficiency and duplication in syndromic forms of autism spectrum disorders. In conclusion, our results reveal the finely tuned developmental clock governing cortical interneuron development, driven by temporal dynamics and the dose-dependent actions of FoxG1.

Role of the medial agranular cortex in unilateral spatial neglect.

Unilateral spatial neglect (USN) results from impaired attentional networks and can affect various sensory modalities, such as visual and somatosensory. The rodent medial agranular cortex (AGm), located in the medial part of the forebrain from rostral to caudal direction, is considered a region associated with spatial attention. The AGm selectively receives multisensory input with the rostral AGm receiving somatosensory input and caudal part receiving visual input. Our previous study showed slower recovery from neglect with anterior AGm lesion using the somatosensory neglect assessment. Conversely, the functional differences in spatial attention across the entire AGm locations (anterior, intermediate, and posterior parts) are unknown. Here, we investigated the relationship between the severity of neglect and various locations across the entire AGm in a mouse stroke model using a newly developed program-based analysis method that does not require human intervention. Among various positions of the lesions, the recovery from USN during recovery periods (postoperative day; POD 10-18) tended to be slower in cases with more rostral lesions in the AGm (r = - 0.302; p = 0.028). Moreover, the total number of arm entries and maximum moving speed did not significantly differ between before and after AGm infarction. According to these results, the anterior lesions may slowly recover from USN-like behavior, and there may be a weak association between the AGm infarct site and recovery rate. In addition, all unilateral focal infarctions in the AGm induced USN-like behavior without motor deficits.

DNA hypomethylator phenotype reprograms glutamatergic network in receptor tyrosine kinase gene-mutated glioblastoma.

DNA methylation is crucial for chromatin structure and gene expression and its aberrancies, including the global "hypomethylator phenotype", are associated with cancer. Here we show that an underlying mechanism for this phenotype in the large proportion of the highly lethal brain tumor glioblastoma (GBM) carrying receptor tyrosine kinase gene mutations, involves the mechanistic target of rapamycin complex 2 (mTORC2), that is critical for growth factor signaling. In this scenario, mTORC2 suppresses the expression of the de novo DNA methyltransferase (DNMT3A) thereby inducing genome-wide DNA hypomethylation. Mechanistically, mTORC2 facilitates a redistribution of EZH2 histone methyltransferase into the promoter region of DNMT3A, and epigenetically represses the expression of DNA methyltransferase. Integrated analyses in both orthotopic mouse models and clinical GBM samples indicate that the DNA hypomethylator phenotype consistently reprograms a glutamate metabolism network, eventually driving GBM cell invasion and survival. These results nominate mTORC2 as a novel regulator of DNA hypomethylation in cancer and an exploitable target against cancer-promoting epigenetics.

Functional and structural synaptic remodeling mechanisms underlying somatotopic organization and reorganization in the thalamus.

The somatosensory system organizes the topographic representation of body maps, termed somatotopy, at all levels of an ascending hierarchy. Postnatal maturation of somatotopy establishes optimal somatosensation, whereas deafferentation in adults reorganizes somatotopy, which underlies pathological somatosensation, such as phantom pain and complex regional pain syndrome. Here, we focus on the mouse whisker somatosensory thalamus to study how sensory experience shapes the fine topography of afferent connectivity during the critical period and what mechanisms remodel it and drive a large-scale somatotopic reorganization after peripheral nerve injury. We will review our findings that, following peripheral nerve injury in adults, lemniscal afferent synapses onto thalamic neurons are remodeled back to immature configuration, as if the critical period reopens. The remodeling process is initiated with local activation of microglia in the brainstem somatosensory nucleus downstream to injured nerves and heterosynaptically controlled by input from GABAergic and cortical neurons to thalamic neurons. These fruits of thalamic studies complement well-studied cortical mechanisms of somatotopic organization and reorganization and unveil potential intervention points in treating pathological somatosensation.

Distinct nociception processing in the dysgranular and barrel regions of the mouse somatosensory cortex.

Nociception, a somatic discriminative aspect of pain, is, like touch, represented in the primary somatosensory cortex (S1), but the separation and interaction of the two modalities within S1 remain unclear. Here, we show spatially distinct tactile and nociceptive processing in the granular barrel field (BF) and adjacent dysgranular region (Dys) in mouse S1. Simultaneous recordings of the multiunit activity across subregions revealed that Dys neurons are more responsive to noxious input, whereas BF neurons prefer tactile input. At the single neuron level, nociceptive information is represented separately from the tactile information in Dys layer 2/3. In contrast, both modalities seem to converge on individual layer 5 neurons of each region, but to a different extent. Overall, these findings show layer-specific processing of nociceptive and tactile information between Dys and BF. We further demonstrated that Dys activity, but not BF activity, is critically involved in pain-like behavior. These findings provide new insights into the role of pain processing in S1.

Involvement of Cdk5 activating subunit p35 in synaptic plasticity in excitatory and inhibitory neurons.

Cyclin-dependent kinase 5 (Cdk5) /p35 is involved in many developmental processes of the central nervous system. Cdk5/p35 is also implicated in synaptic plasticity, learning and memory. Several lines of conditional Cdk5 knockout mice (KO) have been generated and have shown different outcomes for learning and memory. Here, we present our analysis of p35 conditional KO mice (p35cKO) in hippocampal pyramidal neurons or forebrain GABAergic neurons using electrophysiological and behavioral methods. In the fear conditioning task, CamKII-p35cKO mice showed impaired memory retention. Furthermore, NMDAR-dependent long-term depression (LTD) induction by low-frequency stimuli in hippocampal slices from CamkII-p35cKO mice was impaired compared to that in control mice. In contrast, Dlx-p35cKO mice showed no abnormalities in behavioral tasks and electrophysiological analysis in their hippocampal slices. These results indicated that Cdk5/p35 in excitatory neurons is important for the hippocampal synaptic plasticity and associative memory retention.

Dysfunction of parvalbumin-expressing cells in the thalamic reticular nucleus induces cortical spike-and-wave discharges and an unconscious state.

Spike-and-wave discharges and an accompanying loss of consciousness are hallmarks of absence seizure, which is a childhood generalized epilepsy disorder. In absence seizure, dysfunction of the cortico-thalamo-cortico circuitry is thought to engage in abnormal cortical rhythms. Previous studies demonstrated that the thalamic reticular nucleus has a critical role in the formation of normal cortical rhythms; however, whether thalamic reticular nucleus dysfunction leads directly to abnormal rhythms, such as epilepsy, is largely unknown. We found that expressing the inhibitory opsin, archaerhodopsin, including in the thalamic reticular nucleus, caused abnormal cortical rhythms in Pvalb-tetracycline transactivator::tetO-ArchT (PV-ArchT) double transgenic mice. We validated the PV-ArchT line as a new mouse model of absence seizure through physiological and pharmacological analyses, as well as through examining their behavioural features. We then discovered that archaerhodopsin expression exclusively in thalamic reticular nucleus parvalbumin-positive neurons was sufficient to induce cortical spike-and-wave discharges using adeno-associated virus-mediated thalamic reticular nucleus targeting. Furthermore, we found that archaerhodopsin expression impaired rebound burst firing and T-current in thalamic reticular nucleus parvalbumin-positive cells by slice physiology. Although T-current in the thalamic reticular nucleus was impaired, the T-current blocker ethosuximide still had a therapeutic effect in PV-ArchT mice, suggesting a gain of function of T-type calcium channels in this absence seizure model. However, we did not find any over- or misexpression of T-type calcium channel genes in the thalamus or the cortex. Thus, we demonstrated that thalamic reticular nucleus dysfunction led to an absence seizure-like phenotype in mice. In a final set of experiments, we showed that the archaerhodopsin-mediated absence seizure-like phenotype disappeared after the removal of archaerhodopsin by using a time-controllable transgenic system. These data may provide a hint as to why many absence seizures naturally regress.

Facial nerve regeneration with bioabsorbable collagen conduits filled with collagen filaments: An experimental study.

INTRODUCTION: A bioabsorbable collagen conduit (Renerve™) filled with collagen filaments is currently approved as an artificial nerve conduit in Japan and is mainly used for connecting and repairing peripheral nerves after traumatic nerve injury. However, there are few reports on its applications for reconstructing and repairing the facial nerve. The present study evaluated the efficacy of the conduit on promoting nerve regeneration in a murine model with a nerve defect at the buccal branch of the facial nerve. METHODS: Under inhalational anesthesia and microscopic guidance, the buccal branch of the left facial nerve in an 8-week-old Lewis rat was exposed, and a 7 mm gap was created in the nerve. The gap was then connected with either the nerve conduits (NC group) or an autologous nerve graft (the autograft group). At 13 weeks after the procedure, we compared the histological and physiological regenerations in the both groups. RESULTS: We found compound muscle action potential amplitude is significantly larger in the autograft group (2.8 ± 1.4 mV) than in NC group (1.3 ± 0.5 mV) (p < 0.05). The number of myelinated fibers of the autograft group was higher (3634 ± 1645) than that of NC group (1112 ± 490) (p < 0.01). The fiber diameter of the autograft group (4.8 ± 1.9 µm) was larger than that of NC group (3.8 ± 1.4 µm) (p < 0.05). The myelin thickness of the autograft group was thicker than that of NC group (0.6 ± 0.3 µm vs. 0.4 ± 0.1 µm) (p < 0.05). G-ratio of the autograft group (0.74 ± 0.19) was lower than that of NC group (0.79 ± 0.10) (p < 0.05). CONCLUSION: This study demonstrated the efficacy of collagen nerve conduit for facial nerve reconstruction following nerve injury. However, the effectiveness of the conduit on the promotion of nerve regeneration was inferior to that of the autograft.

Electrophysiological and anatomical characterization of synaptic remodeling in the mouse whisker thalamus.

In the central nervous system, developmental and pathophysiologic conditions cause a large-scale reorganization of functional connectivity of neural circuits. Here, by using a mouse model for peripheral sensory nerve injury, we present a protocol for combined electrophysiological and anatomical techniques to identify neural basis of synaptic remodeling in the mouse whisker thalamus. Our protocol provides comprehensive approaches to analyze both structural and functional components of synaptic remodeling. For complete details on the use and execution of this protocol, please refer to Ueta and Miyata, (2021).

FoxG1 regulates the formation of cortical GABAergic circuit during an early postnatal critical period resulting in autism spectrum disorder-like phenotypes.

Abnormalities in GABAergic inhibitory circuits have been implicated in the aetiology of autism spectrum disorder (ASD). ASD is caused by genetic and environmental factors. Several genes have been associated with syndromic forms of ASD, including FOXG1. However, when and how dysregulation of FOXG1 can result in defects in inhibitory circuit development and ASD-like social impairments is unclear. Here, we show that increased or decreased FoxG1 expression in both excitatory and inhibitory neurons results in ASD-related circuit and social behavior deficits in our mouse models. We observe that the second postnatal week is the critical period when regulation of FoxG1 expression is required to prevent subsequent ASD-like social impairments. Transplantation of GABAergic precursor cells prior to this critical period and reduction in GABAergic tone via Gad2 mutation ameliorates and exacerbates circuit functionality and social behavioral defects, respectively. Our results provide mechanistic insight into the developmental timing of inhibitory circuit formation underlying ASD-like phenotypes in mouse models.

Distinct functional developments of surviving and eliminated presynaptic terminals.

For neuronal circuits in the brain to mature, necessary synapses must be maintained and redundant synapses eliminated through experience-dependent mechanisms. However, the functional differentiation of these synapse types during the refinement process remains elusive. Here, we addressed this issue by distinct labeling and direct recordings of presynaptic terminals fated for survival and for elimination in the somatosensory thalamus. At surviving terminals, the number of total releasable vesicles was first enlarged, and then calcium channels and fast-releasing synaptic vesicles were tightly coupled in an experience-dependent manner. By contrast, transmitter release mechanisms did not mature at terminals fated for elimination, irrespective of sensory experience. Nonetheless, terminals fated for survival and for elimination both exhibited developmental shortening of action potential waveforms that was experience independent. Thus, we dissected experience-dependent and -independent developmental maturation processes of surviving and eliminated presynaptic terminals during neuronal circuit refinement.

Brainstem local microglia induce whisker map plasticity in the thalamus after peripheral nerve injury.

Whisker deafferentation in mice disrupts topographic connectivity from the brainstem to the thalamic ventral posteromedial nucleus (VPM), which represents whisker map, by recruiting "ectopic" axons carrying non-whisker information in VPM. However, mechanisms inducing this plasticity remain largely unknown. Here, we show the role of region-specific microglia in the brainstem principal trigeminal nucleus (Pr5), a whisker sensory-recipient region, in VPM whisker map plasticity. Systemic or local manipulation of microglial activity reveals that microglia in Pr5, but not in VPM, are necessary and sufficient for recruiting ectopic axons in VPM. Deafferentation causes membrane hyperexcitability of Pr5 neurons dependent on microglia. Inactivation of Pr5 neurons abolishes this somatotopic reorganization in VPM. Additionally, microglial depletion prevents deafferentation-induced ectopic mechanical hypersensitivity. Our results indicate that local microglia in the brainstem induce peripheral nerve injury-induced plasticity of map organization in the thalamus and suggest that microglia are potential therapeutic targets for peripheral nerve injury-induced mechanical hypersensitivity.

Macrophages are activated in the rat anterior pituitary under chronic inflammatory conditions.

In the anterior lobe of the pituitary gland (AP), non-endocrine cells regulate hormone secretion by endocrine cells. However, the functions of non-endocrine cells in the AP during chronic pain are largely unclear. Here, we show that macrophages, but not folliculostellate (FS) cells, were selectively increased in the AP in the complete Freund's adjuvant (CFA)-induced chronic inflammatory pain model in rats. In addition, IL-1ß expression was increased in the AP, and the IL-1ß-immunopositive cells were identified as macrophages. On the other hand, increased macrophage density and IL-1ß expression were not detected in a neuropathic pain model induced by partial sciatic nerve ligation (PSL). Furthermore, we found c-Fos expression specifically in the somatotrophs under the chronic inflammatory pain condition. Because IL-1ß promotes growth hormone (GH) synthesis and release, our results suggest that AP macrophage contributes to GH release through IL-1ßduring chronic inflammatory pain.　.

Ipsilesional spatial bias after a focal cerebral infarction in the medial agranular cortex: A mouse model of unilateral spatial neglect.

Unilateral spatial neglect is a disorder of higher brain function that occurs after a brain injury, such as stroke, traumatic brain injury, brain tumor, and surgical procedures etc., and leads to failure to attend or respond to stimuli presented to the side contralateral to the lesioned cerebral hemisphere. Because patients with this condition often have other symptoms due to the presence of several brain lesions, it is difficult to evaluate the recovery mechanisms and effect of training on unilateral spatial neglect. In this study, a mouse model of unilateral spatial neglect was created to investigate whether the size of the lesion is related to the severity of ipsilesional spatial bias and the recovery process. Focal infarction was induced in the right medial agranular cortex (AGm) of mice via photothrombosis. After induction of cerebral infarction, ipsilesional spatial bias was evaluated for 9 consecutive days. The major findings were as follows: (1) unilateral local infarction of the AGm resulted in ipsilateral bias during internally guided decision-making; (2) the lesion size was correlated with the degree of impairment rather than slight differences in the lesion site; and (3) mice with anterior AGm lesions experienced lower recovery rates. These findings suggest that recovery from ipsilesional spatial bias requires neural plasticity within the anterior AGm. This conditional mouse model of ipsilesional spatial bias may be used to develop effective treatments for unilateral spatial neglect in humans.

Prevention of denervated muscle atrophy with accelerated nerve-regeneration by babysitter procedure in rat facial nerve paralysis model.

PURPOSE: The "babysitter" procedure is a reconstruction technique for facial nerve complete paralysis and uses the movement source from the healthy facial nerve with a cross-nerve graft. First, an end-to-side neurorrhaphy is performed between the affected facial nerve trunk and hypoglossal nerve for continuously delivering stimuli to the mimetic muscles for preventing the atrophy of mimetic muscles. Despite favorable clinical results, histological and physiological mechanisms remain unknown. This study attempted to establish a model for the "babysitter" procedure and find its efficacy in rats with facial nerve complete paralysis. MATERIALS AND METHODS: A total of 16 Lewis rats were used and divided into 2 groups; cross nerve graft (n = 8) and babysitter groups (n = 8). The facial nerve trunk was transected in both groups. Babysitter group underwent a two-stage procedure. Cross nerve graft group underwent only the transfer of nerve graft from the healthy side to affected side. The animals were assessed physiologically by compound muscle action potential (CMAP), and the regenerated nerve tissues were evaluated histopathologically at 13 weeks after surgery. RESULTS: Facial nucleus stained with retrograde tracers proved the re-innervation of affected facial muscle by the babysitter procedure. In CMAP, the amplitude of babysitter group was significantly higher than that of the cross-facial nerve graft group (p < .05). Histological examination found a significant difference in myelin g-ratio between two groups (p < .05). CONCLUSION: This study investigated the "babysitter" procedure for rat facial nerve palsy. Babysitter procedure shortened the denervation period without mimic muscle atrophy.

Tonic GABAergic Inhibition Is Essential for Nerve Injury-Induced Afferent Remodeling in the Somatosensory Thalamus and Ectopic Sensations.

Peripheral nerve injury induces functional and structural remodeling of neural circuits along the somatosensory pathways, forming the basis for somatotopic reorganization and ectopic sensations, such as referred phantom pain. However, the mechanisms underlying that remodeling remain largely unknown. Whisker sensory nerve injury drives functional remodeling in the somatosensory thalamus: the number of afferent inputs to each thalamic neuron increases from one to many. Here, we report that extrasynaptic γ-aminobutyric acid-type A receptor (GABAAR)-mediated tonic inhibition is necessary for that remodeling. Extrasynaptic GABAAR currents were potentiated rapidly after nerve injury in advance of remodeling. Pharmacological activation of the thalamic extrasynaptic GABAARs in intact mice induced similar remodeling. Notably, conditional deletion of extrasynaptic GABAARs in the thalamus rescued both the injury-induced remodeling and the ectopic mechanical hypersensitivity. Together, our results reveal a molecular basis for injury-induced remodeling of neural circuits and may provide a new pharmacological target for referred phantom sensations after peripheral nerve injury.