Search Results (37)

Calcium (Ca²⁺)signaling dysfunction is a central contributor to neuronal hyperexcitability and seizure propagation in epilepsy, yet the intracellular mechanisms underlying the actions of valproic acid (VPA) remain incompletely understood. In this study, we investigated whether VPA modulates Ca²⁺ homeostasis at the level of the endoplasmic reticulum (ER) and how this action influences cytosolic Ca²⁺ dynamics associated with epileptiform activity. ER Ca²⁺ levels were directly measured using ER-targeted aequorin in HeLa and PC12 cells, while cytosolic Ca²⁺ signals were monitored by fura-2 fluorescence imaging in bovine chromaffin cells exposed to veratridine, a model of sustained sodium channel activation and Ca²⁺ oscillations. VPA induced a concentration-dependent release of Ca²⁺ from the ER, with an IC₅₀ of approximately 17 µM. This effect was preserved in permeabilized cells and exhibited activation kinetics comparable to those elicited by inositol 1,4,5-trisphosphate (InsP₃). Pharmacological inhibition of InsP₃ receptors (InsP₃Rs), but not ryanodine receptors or SERCA, abolished VPA-induced ER Ca²⁺ release, supporting a selective InsP₃R-mediated mechanism. Functionally, VPA suppressed the repetitive cytosolic Ca²⁺ oscillations induced by veratridine, while simultaneously producing a sustained elevation of cytosolic Ca²⁺ originating from ER stores and facilitating depolarization-evoked catecholamine secretion. Together, these results support the conclusion that VPA induces InsP₃R-mediated Ca²⁺ mobilization from the endoplasmic reticulum and identify ER Ca²⁺ release as a previously unrecognized intracellular mechanism contributing to its modulatory effects on Ca²⁺ signaling and excitability in epilepsy. Full article

Store-operated Ca²⁺ entry (SOCE) is a key regulator of cytosolic Ca²⁺ (Ca²⁺_cyt). Presynaptic SOCE can be activated by ligands like α-latrotoxin, which acts through the presynaptic G-protein-coupled receptor latrophilin-1 (LPHN1), inducing Ca²⁺ influx and neurotransmitter release. To understand how SOCE-associated proteins contribute to LPHN1 signaling in neurons, we used mouse neuroblastoma NB2a cells as a genetically tractable neuronal model. The cells were stably transfected with exogenous LPHN1 or its non-signaling mutant and stimulated with the non-pore-forming α-latrotoxin mutant LTX^N4C, a known trigger of neurotransmitter release. LPHN1 expression increased the proportion of neuron-like cells and upregulated the voltage-gated Ca²⁺ channels Ca_v1.2 and Ca_v2.1. LPHN1 stimulation by LTX^N4C induced a small Ca²⁺ release sensitive to thapsigargin, and a strong, gradual influx of Ca²⁺, which was insensitive to thapsigargin. Single-cell imaging revealed that this influx consisted of desynchronized high-amplitude Ca²⁺ oscillations in individual cells. This response was reduced by Orai2 knockdown and completely blocked by the Ca_v2.1/2.2 inhibitor ω-conotoxin MVIIC. We conclude that LPHN1 activation by LTX^N4C primes Ca²⁺ stores and induces the opening of Ca_v2.1/2.2 channels. These channels mediate an initial Ca²⁺ influx that triggers Ca²⁺-induced Ca²⁺ release and SOCE. This mechanism, elucidated in model cells, can explain how LTX^N4C stimulates neurotransmitter release. Full article

Synchronized oscillatory fluctuations in intracellular calcium concentration across extended neuronal networks represent a functional indicator of connectivity and signal coordination. In this study, a model of human immature neurons (differentiated from LUHMES precursors) has been used to establish a robust protocol for generating reproducible intracellular Ca²⁺ oscillations in both two-dimensional monolayers and three-dimensional spheroids. Oscillatory activity was induced by defined ionic conditions in combination with potassium channel blockade. It was characterized by stable frequencies of approximately 0.2 Hz and high synchronization indices across millimeter-scale cultures. These properties were consistently reproduced in independent experiments and across laboratories. Single-cell imaging confirmed that oscillations were coordinated throughout large cell populations. Pharmacological interventions demonstrated that neither excitatory nor inhibitory chemical synaptic transmission influenced oscillatory dynamics. Gap junction blockers completely disrupted synchronization, while leaving individual cell activity unaffected. Functional dye-transfer assays provided additional evidence for electrical coupling. This was further supported by connexin-43 expression profiles and immunostaining. Collectively, these findings indicate that synchronized Ca²⁺ oscillations in LUHMES cultures are mediated by gap junctional communication rather than by conventional synaptic mechanisms. This system offers a practical platform for studying fundamental principles of network coordination and for evaluating pharmacological or toxicological modulators of intercellular coupling. Moreover, it may provide a relevant human-based model to explore aspects of neuronal maturation and to assess compounds with potential neurodevelopmental toxicity. Full article

Astrocytic calcium signaling is a central mechanism of neuron-glia communication that operates across multiple spatial and temporal scales. Traditionally, research has focused on intracellular Ca²⁺ oscillations that regulate gliotransmitter release, ion homeostasis, and metabolic support. Recent evidence, however, reveals that extracellular calcium ([Ca²⁺]o) is not a passive reservoir but a dynamic signaling mediator capable of influencing neuronal excitability within milliseconds. Through mechanisms such as calcium-sensing receptor (CaSR) activation, ion channel modulation, surface charge effects, and ephaptic coupling, astrocytes emerge as active partners in both slow and rapid modes of communication. This dual perspective reshapes our understanding of brain physiology and disease. Disrupted Ca²⁺ signaling contributes to network instability in epilepsy, synaptic dysfunction in Alzheimer’s and Parkinson’s disease, and impaired maturation in neurodevelopmental disorders. Methodological advances, including Ca²⁺-selective microelectrodes, genetically encoded extracellular indicators, and computational modeling, are beginning to uncover the richness of extracellular Ca²⁺ dynamics, though challenges remain in achieving sufficient spatial and temporal resolution. By integrating classical intracellular pathways with emerging insights into extracellular signaling, this review highlights astrocytes as central architects of the ionic landscape. Recognizing calcium as both an intracellular messenger and an extracellular signaling mediator provides a unifying framework for neuron–glia interactions and opens new avenues for therapeutic intervention. Full article

Brain pathophysiology in Down syndrome (DS), the most common genetic cause of intellectual disability, has traditionally been considered a consequence of neuronal dysfunction. However, although it is well documented that astrocytes play a critical role in brain homeostasis, synaptic regulation, and neuronal support, and their malfunction has been associated with the onset and progression of different neurological disorders, only a few studies have addressed whether astrocyte dysfunction can contribute to the DS pathophysiology. Astrocytes are increased in number and size, and show increased levels of expression of astroglial markers like S100β and GFAP. In this study, we detected a region-specific increase in astrocyte population in CA1 and, to a lesser extent, in the dentate gyrus. Single-nucleus transcriptomic profiling identified markers associated with reactive astroglia, synaptic transmission, and neuroinflammation in trisomic astrocytes. Functional analysis revealed abnormal Ca²⁺ oscillations in trisomic astrocytes and impaired astrocyte-to-neuron communication in CA1, the most affected subregion, leading to astrocyte-mediated excitatory synaptic depression. Our findings demonstrate that astrocytes play an active and critical role in the pathophysiology of DS, not only as reactive responders to neuronal injury but as key contributors to the disease process itself. This astrocytic dysfunction presents a region-specific distribution within the hippocampus, suggesting localized vulnerability and complex glial involvement in DS-related neuropathology. Full article

Epilepsy is a complex neurological disorder characterized by abnormal neural synchronization and interactions between local foci and global brain networks during seizures. Understanding seizure mechanisms across multiple scales is essential for advancing our understanding of epileptic network dynamics and guiding personalized treatment strategies. However, neural recording technologies are limited by insufficient spatial resolution, signal fidelity, and the inability to simultaneously capture network- and cellular-level dynamics. To address these limitations, we developed a high-density, flexible deep-brain probe with excellent mechanical compliance and wideband recording capabilities, enabling high-fidelity recordings of high-frequency oscillations (HFOs, 80–500 Hz) and action potentials (APs). Using a pentylenetetrazol (PTZ)-induced epilepsy model, we identified distinct spatiotemporal dynamics of HFOs and APs across epileptic stages, indicating that CA3 plays a key role in seizure onset, while CA1 is crucial for propagation. AP-HFO coupling analysis further uncovered neuronal heterogeneity, offering insights into the diverse roles of neurons in epileptic networks. This study highlights the potential of a flexible deep-brain probe for advancing epilepsy research and guiding personalized therapeutic interventions. Full article

We investigated the cellular and neurophysiological mechanisms underlying the pro-cognitive effects of 5-HT4R activation in hippocampal–prefrontal pathways. Our findings show that, in addition to pyramidal neurons, 30–60% of parvalbumin+ interneurons in the CA1, CA3, and dentate gyrus (DG) of the hippocampus and the anterior cingulate (ACC), prelimbic (PL), and infralimbic (IL) regions of the prefrontal cortex co-express 5-HT4Rs. Additionally, 15% of somatostatin+ interneurons in CA1 and CA3 express 5-HT4Rs. Partial 5-HT4R agonist RS-67333 (1 mg/kg, i.p.) exerted anxiolytic effects and ameliorated short-term (3-min) and long-term (24-h) memory deficits in a mouse model of schizophrenia-like cognitive impairment induced by sub-chronic phencyclidine (sPCP) but did not enhance memory in healthy mice. At the neurophysiological level, RS-67333 normalized sPCP-induced disruptions in hippocampal–prefrontal neural dynamics while having no effect in healthy animals. Specifically, sPCP increased delta oscillations in CA1 and PL, leading to aberrant delta–high-frequency coupling in CA1 and delta–high-gamma coupling in PL. RS-67333 administration attenuated this abnormal delta synchronization without altering phase coherence or signal directionality within the circuit. Collectively, these results highlight the therapeutic potential of 5-HT4R activation in pyramidal, parvalbumin+, and somatostatin+ neurons of hippocampal–prefrontal pathways for mitigation of cognitive and negative symptoms associated with schizophrenia. Full article

Growth-Associated Protein-43 (GAP-43) is a calmodulin-binding protein, originally found in neurons, that in skeletal muscle regulates the handling of intracellular Ca²⁺ dynamics. According to its role in Ca²⁺ regulation, myotubes from GAP-43 knockout (GAP-43^−/−) mice display alterations in spontaneous Ca²⁺ oscillations and increased Ca²⁺ release. The emerging hypothesis is that GAP-43 regulates CaM interactions with RyR and DHPR Ca²⁺ channels. The loss of GAP-43 promotes cardiac hypertrophy in newborn GAP-43^−/− mice, extending the physiological role of GAP-43 in cardiac muscle. We investigated the role of GAP-43 in cardiomyocytes derived from the hearts of GAP-43^−/− mice, evaluating intracellular Ca²⁺ variations and the correlation with the levels of reactive oxygen species (ROS), considering their importance in cardiovascular physiology. In GAP-43^−/− cardiomyocytes, we found the increased expression of markers of cardiac hypertrophy, Ca²⁺ alterations, and high mitochondria ROS levels (O₂^•−) together with increased oxidized functional proteins. Treatment with a CaM inhibitor (W7) restored Ca²⁺ and ROS alterations, possibly due to high mitochondrial Ca²⁺ entry by a mitochondrial Ca²⁺ uniporter. Indeed, Ru360 was able to abolish O₂^•− mitochondrial production. Our results suggest that GAP-43 has a key role in the regulation of Ca²⁺ and ROS homeostasis, alterations to which could trigger heart disease. Full article

Tefluthrin (Tef) is categorized as a type-I pyrethroid insecticide, telmisartan (Tel) functions as an angiotensin II receptor blocker, and KB-R7943 has been identified as an inhibitor of the Na⁺-Ca²⁺ exchange process. However, the influence of these compounds on the amplitude and gating properties of voltage-gated Na⁺ current (I_Na) in neurons associated with pain signaling remains unclear. In cultured dorsal root ganglion (DRG) neurons, whole-cell current recordings revealed that Tef or Tel increased the peak amplitude of I_Na, concomitant with an elevation in the time constant of I_Na inactivation, particularly in the slow component. Conversely, exposure to KB-R7943 resulted in a depression in I_Na, coupled with a decrease in the slow component of the inactivation time constant of I_Na. Theoretical simulations and bifurcation analyses were performed on a modeled interneuron in the spinal dorsal horn. The occurrence of I_Na inactivation accentuated the subthreshold oscillations (SO) in the membrane potential. With an increase in applied current, SO became more pronounced, accompanied by the emergence of high-frequency spiking (HS) with a frequency of approximately 150 Hz. Moreover, an elevation in I_Na conductance further intensified both SO and HF. Consequently, through experimental and in silico studies, this work reflects that Tef, Tel, or KB-R7943 significantly impacts the magnitude and gating properties of I_Na in neurons associated with pain signaling. The alterations in I_Na magnitude and gating in these neurons suggest a close relationship with pain transmission. Full article

One of the major breakthroughs of neurobiology was the identification of distinct ranges of oscillatory activity in the neuronal network that were found to be responsible for specific biological functions, both physiological and pathological in nature. Astrocytes, physically coupled by gap junctions and possessing the ability to simultaneously modulate the functions of a large number of surrounding synapses, are perfectly positioned to introduce synchronised oscillatory activity into the neural network. However, astrocytic somatic calcium signalling has not been investigated to date in the frequency ranges of common neuronal oscillations, since astrocytes are generally considered to be slow responders in terms of Ca²⁺ signalling. Using high-frequency two-photon imaging, we reveal fast Ca²⁺ oscillations in the soma of astrocytes in the delta (0.5–4 Hz) and theta (4–8 Hz) frequency bands in vivo in the rat cortex under ketamine–xylazine anaesthesia, which is known to induce permanent slow-wave sleep. The high-frequency astrocytic Ca²⁺ signals were not observed under fentanyl anaesthesia, excluding the possibility that the signals were introduced by motion artefacts. We also demonstrate that these fast astrocytic Ca²⁺ signals, previously considered to be exclusive to neurons, are present in a large number of astrocytes and are phase synchronised at the astrocytic network level. We foresee that the disclosure of these high-frequency astrocytic signals may help with understanding the appearance of synchronised oscillatory signals and may open up new avenues of treatment for neurological conditions characterised by altered neuronal oscillations. Full article

Gi-coupled receptors, particularly cannabinoid receptors (CBRs), are considered perspective targets for treating brain pathologies, including epilepsy. However, the precise mechanism of the anticonvulsant effect of the CBR agonists remains unknown. We have found that WIN 55,212-2 (a CBR agonist) suppresses the synchronous oscillations of the intracellular concentration of Ca²⁺ ions (epileptiform activity) induced in the neurons of rat hippocampal neuron-glial cultures by bicuculline or NH₄Cl. As we have demonstrated, the WIN 55,212-2 effect is mediated by CB₁R receptors. The agonist suppresses Ca²⁺ inflow mediated by the voltage-gated calcium channels but does not alter the inflow mediated by NMDA, AMPA, and kainate receptors. We have also found that phospholipase C (PLC), protein kinase C (PKC), and G-protein-coupled inwardly rectifying K⁺ channels (GIRK channels) are involved in the molecular mechanism underlying the inhibitory action of CB₁R activation against epileptiform activity. Thus, our results demonstrate that the antiepileptic action of CB₁R agonists is mediated by different intracellular signaling cascades, including non-canonical PLC/PKC-associated pathways. Full article

The core pathological event in Parkinson’s disease (PD) is the specific dying of dopamine (DA) neurons of the substantia nigra pars compacta (SNc). The reasons why SNc DA neurons are especially vulnerable and why idiopathic PD has only been found in humans are still puzzling. The two main underlying factors of SNc DA neuron vulnerability appear related to high DA production, namely (i) the toxic effects of cytoplasmic DA metabolism and (ii) continuous cytosolic Ca²⁺ oscillations in the absence of the Ca²⁺-buffer protein calbindin. Both factors cause oxidative stress by producing highly reactive quinones and increasing intra-mitochondrial Ca²⁺ concentrations, respectively. High DA expression in human SNc DA neuron cell bodies is suggested by the abundant presence of the DA-derived pigment neuromelanin, which is not found in such abundance in other species and has been associated with toxicity at higher levels. The oxidative stress created by their DA production system, despite the fact that the SN does not use unusually high amounts of energy, explains why SNc DA neurons are sensitive to various genetic and environmental factors that create mitochondrial damage and thereby promote PD. Aging increases multiple risk factors for PD, and, to a large extent, PD is accelerated aging. To prevent PD neurodegeneration, possible approaches that are discussed here are (1) reducing cytoplasmic DA accumulation, (2) blocking cytoplasmic Ca²⁺ oscillations, and (3) providing bioenergetic support. Full article

Epilepsy is a chronic condition characterized by recurrent spontaneous seizures. The interaction between astrocytes and neurons has been suggested to play a role in the abnormal neuronal activity observed in epilepsy. However, the exact way astrocytes influence neuronal activity in the epileptogenic brain remains unclear. Here, using the PTZ-induced kindling mouse model, we evaluated the interaction between astrocyte and synaptic function by measuring astrocytic Ca²⁺ activity, neuronal excitability, and the excitatory/inhibitory balance in the hippocampus. Compared to control mice, hippocampal slices from PTZ-kindled mice displayed an increase in glial fibrillary acidic protein (GFAP) levels and an abnormal pattern of intracellular Ca²⁺-oscillations, characterized by an increased frequency of prolonged spontaneous transients. PTZ-kindled hippocampal slices also showed an increase in the E/I ratio towards excitation, likely resulting from an augmented release probability of excitatory inputs without affecting inhibitory synapses. Notably, the alterations in the release probability seen in PTZ-kindled slices can be recovered by reducing astrocyte hyperactivity with the reversible toxin fluorocitrate. This suggests that astroglial hyper-reactivity enhances excitatory synaptic transmission, thereby impacting the E/I balance in the hippocampus. Altogether, our findings support the notion that abnormal astrocyte–neuron interactions are pivotal mechanisms in epileptogenesis. Full article

Abnormal depolarization of neuronal membranes called paroxysmal depolarization shift (PDS) represents a cellular correlate of interictal spikes. The mechanisms underlying the generation of PDSs or PDS clusters remain obscure. This study aimed to investigate the role of ionotropic glutamate receptors (iGluRs) in the generation of PDS and dependence of the PDS pattern on neuronal membrane potential. We have shown that significant depolarization or hyperpolarization (by more than ±50 mV) of a single neuron does not change the number of individual PDSs in the cluster, indicating the involvement of an external stimulus in PDS induction. Based on this data, we have suggested reliable protocols for stimulating single PDS or PDS clusters. Furthermore, we have found that AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptors are necessary for PDS generation since AMPAR antagonist NBQX completely suppresses bicuculline-induced paroxysmal activity. In turn, antagonists of NMDA (N-methyl-D-aspartate) and kainate receptors (D-AP5 and UBP310, respectively) caused a decrease in the amplitude of the first action potential in PDSs and in the amplitude of the oscillations of intracellular Ca²⁺ concentration occurring alongside the PDS cluster generation. The effects of the NMDAR (NMDA receptor) and KAR (kainate receptor) antagonists indicate that these receptors are involved only in the modulation of paroxysmal activity. We have also shown that agonists of some G_i-coupled receptors, such as A₁ adenosine (A₁Rs) or cannabinoid receptors (CBRs) (N₆-cyclohexyladenosine and WIN 55,212-2, respectively), completely suppressed PDS generation, while the A₁R agonist even prevented it. We hypothesized that the dynamics of extracellular glutamate concentration govern paroxysmal activity. Fine-tuning of neuronal activity via action on G_i-coupled receptors or iGluRs paves the way for the development of new approaches for epilepsy pharmacotherapy. Full article

Terahertz waves lie within the rotation and oscillation energy levels of biomolecules, and can directly couple with biomolecules to excite nonlinear resonance effects, thus causing conformational or configuration changes in biomolecules. Based on this mechanism, we investigated the effect pattern of 0.138 THz radiation on the dynamic growth of neurons and synaptic transmission efficiency, while explaining the phenomenon at a more microscopic level. We found that cumulative 0.138 THz radiation not only did not cause neuronal death, but that it promoted the dynamic growth of neuronal cytosol and protrusions. Additionally, there was a cumulative effect of terahertz radiation on the promotion of neuronal growth. Furthermore, in electrophysiological terms, 0.138 THz waves improved synaptic transmission efficiency in the hippocampal CA1 region, and this was a slow and continuous process. This is consistent with the morphological results. This phenomenon can continue for more than 10 min after terahertz radiation ends, and these phenomena were associated with an increase in dendritic spine density. In summary, our study shows that 0.138 THz waves can modulate dynamic neuronal growth and synaptic transmission. Therefore, 0.138 terahertz waves may become a novel neuromodulation technique for modulating neuron structure and function. Full article