Impacted spike frequency adaptation associated with reduction of KCNQ2/3 exacerbates seizure activity in temporal lobe epilepsy.

Numerous epilepsy-related genes have been identified in recent decades by unbiased genome-wide screens. However, the available druggable targets for temporal lobe epilepsy (TLE) remain limited. Furthermore, a substantial pool of candidate genes potentially applicable to TLE therapy awaits further validation. In this study, we reveal the significant role of KCNQ2 and KCNQ3, two M-type potassium channel genes, in the onset of seizures in TLE. Our investigation began with a quantitative analysis of two publicly available TLE patient databases to establish a correlation between seizure onset and the downregulated expression of KCNQ2/3. We then replicated these pathological changes in a pilocarpine seizure mouse model and observed a decrease in spike frequency adaptation due to the affected M-currents in dentate gyrus granule neurons. In addition, we performed a small-scale simulation of the dentate gyrus network and confirmed that the impaired spike frequency adaptation of granule cells facilitated epileptiform activity throughout the network. This, in turn, resulted in prolonged seizure duration and reduced interictal intervals. Our findings shed light on an underlying mechanism contributing to ictogenesis in the TLE hippocampus and suggest a promising target for the development of antiepileptic drugs.

Corrigendum: Immunogenomic landscape in breast cancer reveals immunotherapeutically relevant gene signatures.

[This corrects the article DOI: 10.3389/fimmu.2022.805184.].

A TP53 Related Immune Prognostic Model for the Prediction of Clinical Outcomes and Therapeutic Responses in Lung Adenocarcinoma.

TP53 is the most frequently mutated gene in lung adenocarcinoma (LUAD). The tumor immune microenvironment (TIM) is considered a vital factor that influences tumor progression and survival rate. The influence of TP53 mutation on TIM in LUAD has not been fully studied. Here we systematically investigated the relationship and potential mechanisms between TP53 mutation status and immune response in LUAD. We constructed an immune prognostic model (IPM) using immune associated genes, which were expressed differentially between the TP53 mutant and wild type LUAD patients. We discovered that TP53 mutations were significantly associated with 5 immune related biological processes. Thirty-six immune genes were expressed differentially between TP53 mutant and wild type LUAD patients. An IPM was constructed using 3 immune genes to differentiate the prognostic survival in LUAD. The high-risk LUAD group displayed significantly higher proportions of dendritic cell resting, T cell CD4 memory resting and mast cell resting, and significantly low proportions of dendritic cell activated, T cell CD4 memory activated, and mast cell activated. Moreover, IPM was found to be an independent clinical feature and can be used to predict immunotherapy responses. In summary, we constructed and validated an IPM using 3 immune related genes, which provides a better understanding of the mechanism from an immunological perspectives.

Immunogenomic Landscape in Breast Cancer Reveals Immunotherapeutically Relevant Gene Signatures.

Breast cancer is characterized by some types of heterogeneity, high aggressive behaviour, and low immunotherapeutic efficiency. Detailed immune stratification is a prerequisite for interpreting resistance to treatment and escape from immune control. Hence, the immune landscape of breast cancer needs further understanding. We systematically clustered breast cancer into six immune subtypes based on the mRNA expression patterns of immune signatures and comprehensively depicted their characteristics. The immunotherapeutic benefit score (ITBscore) was validated to be a superior predictor of the response to immunotherapy in cohorts from various datasets. Six distinct immune subtypes related to divergences in biological functions, signatures of immune or stromal cells, extent of the adaptive immune response, genomic events, and clinical prognostication were identified. These six subtypes were characterized as immunologically quiet, chemokine dominant, lymphocyte depleted, wounding dominant, innate immune dominant, and IFN-γ dominant and exhibited features of the tumor microenvironment (TME). The high ITBscore subgroup, characterized by a high proportion of M1 macrophages:M2 macrophages, an activated inflammatory response, and increased mutational burden (such as mutations in TP53, CDH1 and CENPE), indicated better immunotherapeutic benefits. A low proportion of tumor-infiltrating lymphocytes (TILs) and an inadequate response to immune treatment were associated with the low ITBscore subgroup, which was also associated with poor survival. Analyses of four cohorts treated with immune checkpoint inhibitors (ICIs) suggested that patients with a high ITBscore received significant therapeutic advantages and clinical benefits. Our work may facilitate the understanding of immune phenotypes in shaping different TME landscapes and guide precision immuno-oncology and immunotherapy strategies.

Up-regulation of microRNA-19b is associated with metastasis and predicts poor prognosis in patients with colorectal cancer.

Recent evidence has demonstrated that microRNA-19b (miR-19b) is elevated and functions as a prognosis predictor in hepatocellular carcinoma and melanoma. However, its expression and clinical significance in colorectal cancer (CRC) remain unclear. The study aimed to identify the correlation between miR-19b expression and the clinicopathological features and prognosis of patients with CRC. In this study, we found that the levels of miR-19b were significantly up-regulated in CRC tissues and cell lines compared with matched adjacent non-cancerous tissues and human colon mucosal epithelial cell lines, and its expression was also increased in patients with lymph node metastasis compared with those patients with no lymph node metastasis. Meanwhile, the patients with distal metastasis have a higher miR-19b expression than those patients with no distal metastasis. The high expression of miR-19b in patients with CRC was associated with lymph node metastasis and distant metastasis. miR-19b expression was an independent prognostic indicator for overall survival of CRC patients. Moreover, patients with a high miR-19b expression have shorter overall survival times than those patients with a low miR-19b expression. In addition, an in vitro functional assay showed that miR-19b knockdown restrained the migration and invasion of HCT116 and SW480 cells. In summary, the study provides the first convincing statistical and experimental evidence that the up-regulation of miR-19b is associated with metastasis and predicts unfavorable prognosis in patients with CRC, suggesting that miR-19b may serve as a novel and promising prognostic biomarker in CRC.

Piriform cortical glutamatergic and GABAergic neurons express coordinated plasticity for whisker-induced odor recall.

Neural plasticity occurs in learning and memory. Coordinated plasticity at glutamatergic and GABAergic neurons during memory formation remains elusive, which we investigate in a mouse model of associative learning by cellular imaging and electrophysiology. Paired odor and whisker stimulations lead to whisker-induced olfaction response. In mice that express this cross-modal memory, the neurons in the piriform cortex are recruited to encode newly acquired whisker signal alongside innate odor signal, and their response patterns to these associated signals are different. There are emerged synaptic innervations from barrel cortical neurons to piriform cortical neurons from these mice. These results indicate the recruitment of associative memory cells in the piriform cortex after associative memory. In terms of the structural and functional plasticity at these associative memory cells in the piriform cortex, glutamatergic neurons and synapses are upregulated, GABAergic neurons and synapses are downregulated as well as their mutual innervations are refined in the coordinated manner. Therefore, the associated activations of sensory cortices triggered by their input signals induce the formation of their mutual synapse innervations, the recruitment of associative memory cells and the coordinated plasticity between the GABAergic and glutamatergic neurons, which work for associative memory cells to encode cross-modal associated signals in their integration, associative storage and distinguishable retrieval.

Functional compatibility between Purkinje cell axon branches and their target neurons in the cerebellum.

A neuron sprouts an axon, and its branches to innervate many target neurons that are divergent in their functions. In order to efficiently regulate the diversified cells, the axon branches should differentiate functionally to be compatible with their target neurons, i.e., a function compatibility between presynaptic and postsynaptic partners. We have examined this hypothesis by using electrophysiological method in the cerebellum, in which the main axon of Purkinje cell projected to deep nucleus cells and the recurrent axons innervated the adjacent Purkinje cells. The fidelity of spike propagation is superior in the recurrent branches than the main axon. The capabilities of encoding spikes and processing GABAergic inputs are advanced in Purkinje cells versus deep nucleus cells. The functional differences among Purkinje's axonal branches and their postsynaptic neurons are preset by the variable dynamics of their voltage-gated sodium channels. In addition, activity strengths between presynaptic and postsynaptic partners are proportionally correlated, i.e., active axonal branches innervate active target neurons, or vice versa. The physiological impact of the functional compatibility is to make the neurons in their circuits to be activated appropriately. In conclusion, each cerebellar Purkinje cell sprouts the differentiated axon branches to be compatible with the diversified target cells in their functions, in order to construct the homeostatic and efficient units for their coordinated activity in neural circuits.

Dual face of axonal inhibitory inputs in the modulation of neuronal excitability in cortical pyramidal neurons.

Limited by the tiny structure of axons, the effects of these axonal hyperpolarizing inputs on neuronal activity have not been directly elucidated. Here, we imitated these processes by simultaneously recording the activities of the somas and proximal axons of cortical pyramidal neurons. We found that spikes and subthreshold potentials propagate between somas and axons with high fidelity. Furthermore, inhibitory inputs on axons have opposite effects on neuronal activity according to their temporal integration with upstream signals. Concurrent with somatic depolarization, inhibitory inputs on axons decrease neuronal excitability and impede spike generation. In addition, following action potentials, inhibitory inputs on an axon increase neuronal spike capacity and improve spike precision. These results indicate that inhibitory inputs on proximal axons have dual regulatory functions in neuronal activity (suppression or facilitation) according to neuronal network patterns.

Coordinated Plasticity among Glutamatergic and GABAergic Neurons and Synapses in the Barrel Cortex Is Correlated to Learning Efficiency.

Functional plasticity at cortical synapses and neurons is presumably associated with learning and memory. Additionally, coordinated refinement between glutamatergic and GABAergic neurons occurs in associative memory. If these assumptions are present, neuronal plasticity strength and learning efficiency should be correlated. We have examined whether neuronal plasticity strength and learning efficiency are quantitatively correlated in a mouse model of associative learning. Paired whisker and odor stimulations in mice induce odorant-induced whisker motions. The fully establishment of this associative memory appears fast and slow, which are termed as high learning efficiency and low learning efficiency, respectively. In the study of cellular mechanisms underlying this differential learning efficiency, we have compared the strength of neuronal plasticity in the barrel cortices that store associative signals from the mice with high vs. low learning efficiencies. Our results indicate that the levels of learning efficiency are linearly correlated with the upregulated strengths of excitatory synaptic transmission on glutamatergic neurons and their excitability, as well as the downregulated strengths of GABAergic neurons' excitability, their excitatory synaptic inputs and inhibitory synaptic outputs in layers II~III of barrel cortices. The correlations between learning efficiency in associative memory formation and coordinated plasticity at cortical glutamatergic and GABAergic neurons support the notion that the plasticity of associative memory cells is a basis for memory strength.

Associative Memory Extinction Is Accompanied by Decayed Plasticity at Motor Cortical Neurons and Persistent Plasticity at Sensory Cortical Neurons.

Associative memory is essential for cognition, in which associative memory cells and their plasticity presumably play important roles. The mechanism underlying associative memory extinction vs. maintenance remains unclear, which we have studied in a mouse model of cross-modal associative learning. Paired whisker and olfaction stimulations lead to a full establishment of odorant-induced whisker motion in training day 10, which almost disappears if paired stimulations are not given in a week, and then recovers after paired stimulation for an additional day. In mice that show associative memory, extinction and recovery, we have analyzed the dynamical plasticity of glutamatergic neurons in layers II-III of the barrel cortex and layers IV-V of the motor cortex. Compared with control mice, the rate of evoked spikes as well as the amplitude and frequency of excitatory postsynaptic currents increase, whereas the amplitude and frequency of inhibitory postsynaptic currents (IPSC) decrease at training day 10 in associative memory mice. Without paired training for a week, these plastic changes are persistent in the barrel cortex and decayed in the motor cortex. If paired training is given for an additional day to revoke associative memory, neuronal plasticity recovers in the motor cortex. Our study indicates persistent neuronal plasticity in the barrel cortex for cross-modal memory maintenance as well as the dynamical change of neuronal plasticity in the motor cortex for memory retrieval and extinction. In other words, the sensory cortices are essential for long-term memory while the behavior-related cortices with the inability of memory retrieval are correlated to memory extinction.

PKC and CaMK-II inhibitions coordinately rescue ischemia-induced GABAergic neuron dysfunction.

Cerebral ischemia leads to neuronal death for stroke, in which the imbalance between glutamatergic neurons and GABAergic neurons toward neural excitotoxicity is presumably involved. GABAergic neurons are vulnerable to pathological factors and impaired in an early stage of ischemia. The rescue of GABAergic neurons is expected to be the strategy to reserve ischemic neuronal impairment. As protein kinase C (PKC) and calmodulin-dependent protein kinase II (CaMK-II) are activated during ischemia, we have investigated whether the inhibitions of these kinases rescue the ischemic impairment of cortical GABAergic neurons. The functions of GABAergic neurons were analyzed by whole-cell recording in the cortical slices during ischemia and in presence of 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (CaMK-II inhibitor) and chelerythrine chloride (PKC inhibitor). Our results indicate that PKC inhibitor or CaMK-II inhibitor partially prevents ischemia-induced functional deficits of cortical GABAergic neurons. Moreover, the combination of PKC and CaMK-II inhibitors synergistically reverses this ischemia-induced deficit of GABAergic neurons. One of potential therapeutic strategies for ischemic stroke may be to rescue the ischemia-induced deficit of cortical GABAergic neurons by inhibiting PKC and CaMK-II.

Ischemia deteriorates spike encoding at cortical GABAergic neurons and cerebellar Purkinje cells by increasing the intracellular Ca2.

GABAergic neurons play a critical role in the central nervous system, such as well-organized behaviors. The ischemic cell death is presumably initiated by neuronal excitotoxicity resulted from the dysfunction of GABAergic neurons. It is not clear how ischemia influences different types of GABAergic neurons and whether intracellular Ca2+ plays a key role in the ischemic excitotoxicity. We have investigated this issue at cortical GABAergic neurons and cerebellar Purkinje cells by whole-cell recording in mouse brain slices, and the roles of intracellular Ca2+ are examined by BABTA infusion. Compare with the data from a group of control, ischemia causes by lowering purfusion rate lowers spike encoding at cortical GABAergic neurons and enhances encoding ability at cerebellar Purkinje cells. These differential effects of ischemia on spike encoding are mechanistically associated with the changes in the refractory periods and threshold potentials of sequential spikes. These ischemia-induced dysfunction of spike encoding at two types of GABAergic cells are prevented by BABTA infusion. Therefore, the ischemia destabilizes the spike encoding of GABAergic cells via raising intracellular Ca2+. Our findings indicate that ischemia preferentially causes the dysfunction of spike encoding at GABAergic neurons by the up-regulation of intracellular Ca2+ level, which leads to neuronal excitotoxicity.

Activity strengths of cortical glutamatergic and GABAergic neurons are correlated with transgenerational inheritance of learning ability.

The capabilities of learning and memory in parents are presumably transmitted to their offsprings, in which genetic codes and epigenetic regulations are thought as molecular bases. As neural plasticity occurs during memory formation as cellular mechanism, we aim to examine the correlation of activity strengths at cortical glutamatergic and GABAergic neurons to the transgenerational inheritance of learning ability. In a mouse model of associative learning, paired whisker and odor stimulations led to odorant-induced whisker motion, whose onset appeared fast (high learning efficiency, HLE) or slow (low learning efficiency, LLE). HLE male and female mice, HLE female and LLE male mice as well as HLE male and LLE female mice were cross-mated to have their first generation of offsprings, filials (F1). The onset of odorant-induced whisker motion appeared a sequence of high-to-low efficiency in three groups of F1 mice that were from HLE male and female mice, HLE female and LLE male mice as well as HLE male and LLE female mice. Activities related to glutamatergic neurons in barrel cortices appeared a sequence of high-to-low strength in these F1 mice from HLE male and female mice, HLE female and LLE male mice as well as HLE male and LLE female mice. Activities related to GABAergic neurons in barrel cortices appeared a sequence of low-to-high strength in these F1 mice from HLE male and female mice, HLE female and LLE male mice as well as HLE male and LLE female mice. Neuronal activity strength was linearly correlated to learning efficiency among three groups. Thus, the coordinated activities at glutamatergic and GABAergic neurons may constitute the cellular basis for the transgenerational inheritance of learning ability.

A portion of inhibitory neurons in human temporal lobe epilepsy are functionally upregulated: an endogenous mechanism for seizure termination.

MAIN PROBLEM: Epilepsy is one of the more common neurological disorders. The medication is often ineffective to the patients suffering from intractable temporal lobe epilepsy (TLE). As their seizures are usually self-terminated, the elucidation of the mechanism underlying endogenous seizure termination will help to find a new strategy for epilepsy treatment. We aim to examine the role of inhibitory interneurons in endogenous seizure termination in TLE patients. METHODS: Whole-cell recordings were conducted on inhibitory interneurons in seizure-onset cortices of intractable TLE patients and the temporal lobe cortices of nonseizure individuals. The intrinsic property of the inhibitory interneurons and the strength of their GABAergic synaptic outputs were measured. The quantitative data were introduced into the computer-simulated neuronal networks to figure out a role of these inhibitory units in the seizure termination. RESULTS: In addition to functional downregulation, a portion of inhibitory interneurons in seizure-onset cortices were upregulated in encoding the spikes and controlling their postsynaptic neurons. A patch-like upregulation of inhibitory neurons in the local network facilitated seizure termination. The upregulations of both inhibitory neurons and their output synapses synergistically shortened seizure duration, attenuated seizure strength, and terminated seizure propagation. CONCLUSION: Automatic seizure termination is likely due to the fact that a portion of the inhibitory neurons and synapses are upregulated in the seizure-onset cortices. This mechanism may create novel therapeutic strategies to treat intractable epilepsy, such as the simultaneous upregulation of cortical inhibitory neurons and their output synapses.

Input-dependent subcellular localization of spike initiation between soma and axon at cortical pyramidal neurons.

BACKGROUND: Action potentials can be initiated at various subcellular compartments, such as axonal hillock, soma and dendrite. Mechanisms and physiological impacts for this relocation remain elusive, which may rely on input signal patterns and intrinsic properties in these subcellular compartments. We examined this hypothesis at the soma and axon of cortical pyramidal neurons by analyzing their spike capability and voltage-gated sodium channel dynamics in response to different input signals. RESULTS: Electrophysiological recordings were simultaneously conducted at the somata and axons of identical pyramidal neurons in the cortical slices. The somata dominantly produced sequential spikes in response to long-time steady depolarization pulse, and the axons produced more spikes in response to fluctuated pulse. Compared with the axons, the somata possessed lower spike threshold and shorter refractory periods in response to long-time steady depolarization, and somatic voltage-gated sodium channels demonstrated less inactivation and easier reactivation in response to steady depolarization. Based on local VGSC dynamics, computational simulated spike initiation locations were consistent with those from the experiments. In terms of physiological impact, this input-dependent plasticity of spike initiation location made neuronal encoding to be efficient. CONCLUSIONS: Long-time steady depolarization primarily induces somatic spikes and short-time pulses induce axonal spikes. The input signal patterns influence spike initiations at the axon or soma of cortical pyramidal neurons through modulating local voltage-gated sodium channel dynamics.

Physiological synaptic signals initiate sequential spikes at soma of cortical pyramidal neurons.

The neurons in the brain produce sequential spikes as the digital codes whose various patterns manage well-organized cognitions and behaviors. A source for the physiologically integrated synaptic signals to initiate digital spikes remains unknown, which we studied at pyramidal neurons of cortical slices. In dual recordings from the soma vs. axon, the signals recorded in vivo induce somatic spikes with higher capacity, which is associated with lower somatic thresholds and shorter refractory periods mediated by voltage-gated sodium channels. The introduction of these parameters from the soma and axon into NEURON model simulates sequential spikes being somatic in origin. Physiological signals integrated from synaptic inputs primarily trigger the soma to encode neuronal digital spikes.

Axons amplify somatic incomplete spikes into uniform amplitudes in mouse cortical pyramidal neurons.

BACKGROUND: Action potentials are the essential unit of neuronal encoding. Somatic sequential spikes in the central nervous system appear various in amplitudes. To be effective neuronal codes, these spikes should be propagated to axonal terminals where they activate the synapses and drive postsynaptic neurons. It remains unclear whether these effective neuronal codes are based on spike timing orders and/or amplitudes. METHODOLOGY/PRINCIPAL FINDINGS: We investigated this fundamental issue by simultaneously recording the axon versus soma of identical neurons and presynaptic vs. postsynaptic neurons in the cortical slices. The axons enable somatic spikes in low amplitude be enlarged, which activate synaptic transmission in consistent patterns. This facilitation in the propagation of sequential spikes through the axons is mechanistically founded by the short refractory periods, large currents and high opening probability of axonal voltage-gated sodium channels. CONCLUSION/SIGNIFICANCE: An amplification of somatic incomplete spikes into axonal complete ones makes sequential spikes to activate consistent synaptic transmission. Therefore, neuronal encoding is likely based on spike timing order, instead of graded analogues.

Real-time neuronal homeostasis by coordinating VGSC intrinsic properties.

Homeostasis of internal environment and cellular metabolism ensures cells' functions to be stable in living organisms. Cellular homeostasis is believed to be maintained via feedback or feedforward manners. We report a novel mechanism that maintains neuronal homeostasis through coordinating the intrinsic properties of single molecules concurrently. Spike encoding and sodium channel dynamics at cortical neurons were studied by patch-clamp recording. Voltage-gated sodium channels set refractory period and threshold potential toward different directions to stabilize the energetic barrier for firing sequential action potentials. This neuronal homeostasis is not affected by intracellular Ca(2+) signals and membrane potentials. Real-time homeostasis maintains precise and reliable neuronal encoding without any destabilization.

The postnatal development of intrinsic properties and spike encoding at cortical GABAergic neurons.

GABAergic neurons play a critical role in maintaining the homeostasis of brain functions for well-organized behaviors. It is not known about the dynamical change in signal encoding at these neurons during postnatal development. We investigated this issue at GFP-labeled GABAergic neurons by whole-cell recording in cortical slices of mice. Our results show that the ability of spike encoding at GABAergic neurons is improved during postnatal development. This change is associated with the reduction of refractory periods and threshold potentials of sequential spikes, as well as the improvement of linear correlations between intrinsic properties and spike capacity. Therefore, the postnatal maturation of the spike encoding capacity at GABAergic neurons will stabilize the excitatory state of cerebral cortex.