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Molecular and Cellular Neurosciences Mar 2024The EphB receptor tyrosine kinase family participates in intricate signaling pathways that orchestrate neural networks, guide neuronal axon development, and modulate...
BACKGROUND
The EphB receptor tyrosine kinase family participates in intricate signaling pathways that orchestrate neural networks, guide neuronal axon development, and modulate synaptic plasticity through interactions with surface-bound ephrinB ligands. Additionally, Kalirin, a Rho guanine nucleotide exchange factor, is notably expressed in the postsynaptic membrane of excitatory neurons and plays a role in synaptic morphogenesis. This study postulates that Kalirin may act as a downstream effector of EphB3 in epilepsy. This investigation focuses on understanding the link between EphB3 and epilepsy.
MATERIALS AND METHODS
Chronic seizure models using LiCl-pilocarpine (LiCl/Pilo) and pentylenetetrazol were developed in rats. Neuronal excitability was gauged through whole-cell patch clamp recordings on rat hippocampal slices. Real-time PCR determined Kalirin's mRNA expression, and Western blotting was employed to quantify EphB3 and Kalirin protein levels. Moreover, dendritic spine density in epileptic rats was evaluated using Golgi staining.
RESULTS
Modulation of EphB3 functionality influenced acute seizure severity, latency duration, and frequency of spontaneous recurrent seizures. Golgi staining disclosed an EphB3-driven alteration in dendritic spine density within the hippocampus of epileptic rats, underscoring its pivotal role in the reconfiguration of hippocampal neural circuits. Furthermore, our data propose Kalirin as a prospective downstream mediator of the EphB3 receptor.
CONCLUSIONS
Our findings elucidate that EphB3 impacts the action potential dynamics in isolated rat hippocampal slices and alters dendritic spine density in the inner molecular layer of epileptic rat hippocampi, likely through Kalirin-mediated pathways. This hints at EphB3's significant role in shaping excitatory circuit loops and recurrent seizure activity via Kalirin.
Topics: Rats; Animals; Rats, Sprague-Dawley; Prospective Studies; Neurons; Epilepsy; Seizures
PubMed: 38143048
DOI: 10.1016/j.mcn.2023.103915 -
Cell & Bioscience Jan 2024Rasal1 is a Ras GTPase-activating protein which contains C2 domains necessary for dynamic membrane association following intracellular calcium elevation. Membrane-bound...
BACKGROUND
Rasal1 is a Ras GTPase-activating protein which contains C2 domains necessary for dynamic membrane association following intracellular calcium elevation. Membrane-bound Rasal1 inactivates Ras signaling through its RasGAP activity, and through such mechanisms has been implicated in regulating various cellular functions in the context of tumors. Although highly expressed in the brain, the contribution of Rasal1 to neuronal development and function has yet to be explored.
RESULTS
We examined the contributions of Rasal1 to neuronal development in primary culture of hippocampal neurons through modulation of Rasal1 expression using molecular tools. Fixed and live cell imaging demonstrate diffuse expression of Rasal1 throughout the cell soma, dendrites and axon which localizes to the neuronal plasma membrane in response to intracellular calcium fluctuation. Pull-down and co-immunoprecipitation demonstrate direct interaction of Rasal1 with PKC, tubulin, and CaMKII. Consequently, Rasal1 is found to stabilize microtubules, through post-translational modification of tubulin, and accordingly inhibit dendritic outgrowth and branching. Through imaging, molecular, and electrophysiological techniques Rasal1 is shown to promote NMDA-mediated synaptic activity and CaMKII phosphorylation.
CONCLUSIONS
Rasal1 functions in two separate roles in neuronal development; calcium regulated neurite outgrowth and the promotion of NMDA receptor-mediated postsynaptic events which may be mediated both by interaction with direct binding partners or calcium-dependent regulation of down-stream pathways. Importantly, the outlined molecular mechanisms of Rasal1 may contribute notably to normal neuronal development and synapse formation.
PubMed: 38246997
DOI: 10.1186/s13578-024-01193-w -
Biochemical and Biophysical Research... Jul 2024Alzheimer's disease is characterized by abnormal β-amyloid and tau accumulation, mitochondrial dysfunction, oxidative stress, and synaptic dysfunction. Here, we aimed...
Alzheimer's disease is characterized by abnormal β-amyloid and tau accumulation, mitochondrial dysfunction, oxidative stress, and synaptic dysfunction. Here, we aimed to assess the mechanisms and signalling pathways in the neuroprotective effect of gastrodin, a phenolic glycoside, on murine neuroblastoma N2a cells expressing human Swedish mutant APP (N2a/APP). We found that gastrodin increased the levels of presynaptic-SNAP, synaptophysin, and postsynaptic-PSD95 and reduced phospho-tau Ser396, APP and Aβ levels in N2a/APP cells. Gastrodin treatment reduced reactive oxygen species generation, lipid peroxidation, mitochondrial fragmentation and DNA oxidation; restored mitochondrial membrane potential and intracellular ATP production. Upregulated phospho-GSK-3β and reduced phospho-ERK and phospho-JNK were involved in the protective effect of gastrodin. In conclusion, we demonstrated the neuroprotective effect of gastrodin in the N2a/APP cell line by ameliorating the impairment on synaptic and mitochondrial function, reducing tau phosphorylation, Aβ levels as well as reactive oxygen species generation. These results provide new mechanistic insights into the potential effect of gastrodin in the treatment of Alzheimer's disease.
Topics: Glucosides; Benzyl Alcohols; Oxidative Stress; Animals; Mice; Mitochondria; Neuroprotective Agents; Cell Line, Tumor; Reactive Oxygen Species; Synapses; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Humans; Alzheimer Disease; tau Proteins; Membrane Potential, Mitochondrial; Peptide Fragments
PubMed: 38761634
DOI: 10.1016/j.bbrc.2024.150127 -
Molecular Medicine Reports Nov 2023Blume has been widely used to treat various central and peripheral nerve diseases, and Para‑hydroxybenzaldehyde (PHBA) is one of the indicated components suggested to...
Blume has been widely used to treat various central and peripheral nerve diseases, and Para‑hydroxybenzaldehyde (PHBA) is one of the indicated components suggested to provide a neuroprotective effect. In our previous, it was shown that PHBA protected mitochondria against cerebral ischemia‑reperfusion (I/R) injury in rats. In the present study, how PHBA regulated the metabolic mechanism in blood following cerebral I/R was assessed to identify an effective therapeutic target for the prevention and treatment of ischemic stroke (IS). First, a rat model of cerebral ischemia‑reperfusion injury was established via middle cerebral artery occlusion/reperfusion (MCAO/R). The therapeutic effect of PHBA on brain I/R was evaluated by assessing the neurological function score, triphenyl tetrazolium chloride, hematoxylin and eosin, and Nissl staining. Next, a non‑targeted metabolomic based on high‑performance liquid chromatography quadrupole time‑of‑flight mass spectrometry was established to identify differential metabolites. Finally, a targeted metabolic spectrum was analyzed and the potential therapeutic targets were verified by Western blotting. The results showed that the neurological function score, cerebral infarction area, hippocampal morphology, and the number of neurons in the PHBA group were significantly improved compared with the model group. Metabonomic analysis showed that 13 different metabolites were identified between the model and PHBA group, which may be involved in the 'tricarboxylic acid cycle', 'glutathione metabolism', and 'mutual transformation of pentose and glucuronates', amongst others. Among these, the levels of the most significant differential metabolite, dGMP, decreased significantly following PHBA treatment. Western blotting was used to verify the expression of membrane‑associated guanosine kinase PSD‑95 and the subunit of glutamate AMPA receptor GluA1, which significantly increased after PHBA treatment. In addition, it was also found that PHBA increased the expression of the light chain‑3 protein and autophagy effector protein 1, whilst the expression of sequestosome‑1 decreased, indicating that PHBA promoted autophagy. Similarly, in TUNEL staining and detection of apoptosis‑related proteins, it was found that MCAO/R upregulated the expression of Bax and cleaved‑caspase‑3 whilst downregulating the expression of Bcl‑2 and increasing the apoptosis of hippocampal neurons; PHBA reversed this situation. These results suggest that cerebral I/R causes postsynaptic dysfunction by disrupting the interaction between PSD‑95 and AMPARs, and the inhibition of the autophagy system eventually leads to the apoptosis of hippocampal neurons.
Topics: Rats; Animals; Ischemic Attack, Transient; Brain Ischemia; Benzaldehydes; Apoptosis; Infarction, Middle Cerebral Artery; Apoptosis Regulatory Proteins; Reperfusion Injury; Neuroprotective Agents
PubMed: 37800608
DOI: 10.3892/mmr.2023.13111 -
Patterns (New York, N.Y.) Oct 2023Mapping functional connectivity between neurons is an essential step toward probing the neural computations mediating behavior. Accurately determining synaptic...
Mapping functional connectivity between neurons is an essential step toward probing the neural computations mediating behavior. Accurately determining synaptic connectivity maps in populations of neurons is challenging in terms of yield, accuracy, and experimental time. Here, we developed a compressive sensing approach to reconstruct synaptic connectivity maps based on random two-photon cell-targeted optogenetic stimulation and membrane voltage readout of many putative postsynaptic neurons. Using a biophysical network model of interconnected populations of excitatory and inhibitory neurons, we characterized mapping recall and precision as a function of network observability, sparsity, number of neurons stimulated, off-target stimulation, synaptic reliability, propagation latency, and network topology. We found that mapping can be achieved with far fewer measurements than the standard pairwise sequential approach, with network sparsity and synaptic reliability serving as primary determinants of the performance. Our results suggest a rapid and efficient method to reconstruct functional connectivity of sparsely connected neuronal networks.
PubMed: 37876895
DOI: 10.1016/j.patter.2023.100845 -
Alzheimer's Research & Therapy Mar 2024The key to the prevention and treatment of Alzheimer's disease (AD) is to be able to predict and diagnose AD at the preclinical or early stage, but the lack of a...
Characterization of preclinical Alzheimer's disease model: spontaneous type 2 diabetic cynomolgus monkeys with systemic pro-inflammation, positive biomarkers and developing AD-like pathology.
BACKGROUND
The key to the prevention and treatment of Alzheimer's disease (AD) is to be able to predict and diagnose AD at the preclinical or early stage, but the lack of a preclinical model of AD is the critical factor that causes this problem to remain unresolved.
METHODS
We assessed 18 monkeys in vivo evaluation of pro-inflammatory cytokines and AD pathological biomarkers (n = 9 / type 2 diabetic mellitus (T2DM) group, age 20, fasting plasma glucose (FPG) ≥ 100 mg/dL, and n = 9 / negative control (NC) group, age 17, FPG < 100 mg/dL). Levels of pro-inflammatory cytokines and AD pathological biomarkers was measured by ELISA and Simoa Technology, respectively. 9 monkeys evaluated ex vivo for AD-like pathology (n = 6 / T2DM group, age 22.17, FPG ≥ 126 mg/dL, and n = 3 / NC group, age 14.67, FPG < 100 mg/dL). To evaluate the pathological features of AD in the brains of T2DM monkeys, we assessed the levels of Aβ, phospho-tau, and neuroinflammation using immunohistochemistry, which further confirmed the deposition of Aβ plaques by Bielschowsky's silver, Congo red, and Thioflavin S staining. Synaptic damage and neurodegeneration were assessed by immunofluorescence.
RESULTS
We found not only increased levels of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) in peripheral blood (PB) and brain of T2DM monkeys but also changes in PB of AD pathological biomarkers such as decreased β-amyloid (Aβ) 42 and Aβ40 levels. Most notably, we observed AD-like pathological features in the brain of T2DM monkeys, including Aβ plaque deposition, p-tau from neuropil thread to pre-neurofibrillary tangles (NFTs), and even the appearance of extracellular NFT. Microglia were activated from a resting state to an amoeboid. Astrocytes showed marked hypertrophy and an increased number of cell bodies and protrusions. Finally, we observed impairment of the postsynaptic membrane but no neurodegeneration or neuronal death.
CONCLUSIONS
Overall, T2DM monkeys showed elevated levels of peripheral and intracerebral inflammation, positive AD biomarkers in body fluids, and developing AD-like pathology in the brain, including Aβ and tau pathology, glial cell activation, and partial synaptic damage, but no neuronal degeneration or death as compared to the healthy normal group. Hereby, we consider the T2DM monkeys with elevation of the peripheral pro-inflammatory factors and positive AD biomarkers can be potentially regarded as a preclinical AD model.
Topics: Animals; Alzheimer Disease; Macaca fascicularis; Amyloid beta-Peptides; Inflammation; Brain; Biomarkers; Diabetes Mellitus, Type 2; Cytokines; tau Proteins
PubMed: 38459540
DOI: 10.1186/s13195-024-01416-9 -
BioRxiv : the Preprint Server For... Apr 2024Layer 6 corticothalamic (L6 CT) neurons provide massive input to the thalamus, and these feedback connections enable the cortex to influence its own sensory input by...
Layer 6 corticothalamic (L6 CT) neurons provide massive input to the thalamus, and these feedback connections enable the cortex to influence its own sensory input by modulating thalamic excitability. However, the functional role(s) feedback serves during sensory processing is unclear. One hypothesis is that CT feedback is under the control of extra-sensory signals originating from higher-order cortical areas, yet we know nothing about the mechanisms of such control. It is also unclear whether such regulation is specific to CT neurons with distinct thalamic connectivity. Using mice (either sex) combined with electrophysiology techniques, optogenetics, and retrograde labeling, we describe studies of vibrissal primary motor cortex (vM1) influences on different CT neurons in the vibrissal primary somatosensory cortex (vS1) with distinct intrathalamic axonal projections. We found that vM1 inputs are highly selective, evoking stronger postsynaptic responses in Dual ventral posterior medial nucleus (VPm) and posterior medial nucleus (POm) projecting CT neurons located in lower L6a than VPm-only projecting CT cells in upper L6a. A targeted analysis of the specific cells and synapses involved revealed that the greater responsiveness of Dual CT neurons was due to their distinctive intrinsic membrane properties and synaptic mechanisms. These data demonstrate that vS1 has at least two discrete L6 CT subcircuits distinguished by their thalamic projection patterns, intrinsic physiology, and functional connectivity with vM1. Our results also provide insights into how a distinct CT subcircuit may serve specialized roles specific to contextual modulation of tactile-related sensory signals in the somatosensory thalamus during active vibrissa movements.
PubMed: 38712153
DOI: 10.1101/2024.04.22.590613 -
Cell Reports Jul 2023Dorsal and ventral medial entorhinal cortex (mEC) regions have distinct neural network firing patterns to differentially support functions such as spatial memory....
Dorsal and ventral medial entorhinal cortex (mEC) regions have distinct neural network firing patterns to differentially support functions such as spatial memory. Accordingly, mEC layer II dorsal stellate neurons are less excitable than ventral neurons. This is partly because the densities of inhibitory conductances are higher in dorsal than ventral neurons. Here, we report that T-type Ca currents increase 3-fold along the dorsal-ventral axis in mEC layer II stellate neurons, with twice as much Ca3.2 mRNA in ventral mEC compared with dorsal mEC. Long depolarizing stimuli trigger T-type Ca currents, which interact with persistent Na currents to elevate the membrane voltage and spike firing in ventral, not dorsal, neurons. T-type Ca currents themselves prolong excitatory postsynaptic potentials (EPSPs) to enhance their summation and spike coupling in ventral neurons only. These findings indicate that T-type Ca currents critically influence the dorsal-ventral mEC stellate neuron excitability gradient and, thereby, mEC dorsal-ventral circuit activity.
Topics: Entorhinal Cortex; Neurons; Action Potentials
PubMed: 37368752
DOI: 10.1016/j.celrep.2023.112699 -
International Journal of Molecular... May 2024In all cell types, small EVs, very abundant extracellular vesicles, are generated and accumulated within MVB endocytic cisternae. Upon MVB fusion and exocytosis with the... (Review)
Review
In all cell types, small EVs, very abundant extracellular vesicles, are generated and accumulated within MVB endocytic cisternae. Upon MVB fusion and exocytosis with the plasma membrane, the EVs are released to the extracellular space. In the central nervous system, the release of neuronal EVs was believed to occur only from the surface of the body and dendrites. About 15 years ago, MVB cisternae and EVs were shown to exist and function at synaptic boutons, the terminals' pre- and post-synaptic structures essential for canonical neurotransmitter release. Recent studies have revealed that synaptic EVs are peculiar in many respects and heterogeneous with respect to other neuronal EVs. The distribution of synaptic EVs and the effect of their specific molecules are found at critical sites of their distribution. The role of synaptic EVs could consist of the modulation of canonical neurotransmitter release or a distinct, non-canonical form of neurotransmission. Additional roles of synaptic EVs are still not completely known. In the future, additional investigations will clarify the role of synaptic EVs in pathology, concerning, for example, circuits, trans-synaptic transmission, diagnosis and the therapy of diseases.
Topics: Humans; Extracellular Vesicles; Animals; Neurons; Synapses; Signal Transduction; Synaptic Transmission; Exocytosis; Neurotransmitter Agents; Synaptic Vesicles
PubMed: 38791143
DOI: 10.3390/ijms25105103 -
Neurophotonics Oct 2023Membrane trafficking of post-synaptic cargo is a key determinant of synaptic transmission and synaptic plasticity. We describe here the latest developments in...
Membrane trafficking of post-synaptic cargo is a key determinant of synaptic transmission and synaptic plasticity. We describe here the latest developments in visualizing individual exocytosis and endocytosis events in neurons using pH-sensitive tags. We show how these tools help decipher the spatial and temporal regulation of membrane trafficking steps during synaptic plasticity.
PubMed: 37547562
DOI: 10.1117/1.NPh.10.4.044404