Glutamatergic Synaptic Transmission Research Articles

Astrocytic ALKBH5 in stress response contributes to depressive-like behaviors in mice

Epigenetic mechanisms bridge genetic and environmental factors that contribute to the pathogenesis of major depression disorder (MDD). However, the cellular specificity and sensitivity of environmental stress on brain epitranscriptomics and its impact on depression remain unclear. Here, we found that ALKBH5, an RNA demethylase of N6-methyladenosine (m6A), was increased in MDD patients’ blood and depression models. ALKBH5 in astrocytes was more sensitive to stress than that in neurons and endothelial cells. Selective deletion of ALKBH5 in astrocytes, but not in neurons and endothelial cells, produced antidepressant-like behaviors. Astrocytic ALKBH5 in the mPFC regulated depression-related behaviors bidirectionally. Meanwhile, ALKBH5 modulated glutamate transporter-1 (GLT-1) m6A modification and increased the expression of GLT-1 in astrocytes. ALKBH5 astrocyte-specific knockout preserved stress-induced disruption of glutamatergic synaptic transmission, neuronal atrophy and defective Ca2+ activity. Moreover, enhanced m6A modification with S-adenosylmethionine (SAMe) produced antidepressant-like effects. Our findings indicate that astrocytic epitranscriptomics contribute to depressive-like behaviors and that astrocytic ALKBH5 may be a therapeutic target for depression.

Human iPSC-derived neurons with reliable synapses and large presynaptic action potentials

Understanding the function of the human brain requires determining basic properties of synaptic transmission in human neurons. One of the most fundamental parameters controlling neurotransmitter release is the presynaptic action potential, but its amplitude and duration remain controversial. Presynaptic action potentials have so far been measured with high temporal resolution only in a limited number of vertebrate but not in human neurons. To uncover properties of human presynaptic action potentials, we exploited recently developed tools to generate human glutamatergic neurons by transient expression of Neurogenin 2 (Ngn2) in pluripotent stem cells. During maturation for 3 to 9 weeks of culturing in different established media, the proportion of cells with multiple axon initial segments decreased, while the amount of axonal tau protein and neuronal excitability increased. Super-resolution microscopy revealed the alignment of the pre- and postsynaptic proteins, Bassoon and Homer. Synaptic transmission was surprisingly reliable at frequencies of 20, 50, and 100 Hz. The synchronicity of synaptic transmission during high-frequency transmission increased during 9 weeks of neuronal maturation. To analyze the mechanisms of synchronous high-frequency glutamate release, we developed direct presynaptic patch-clamp recordings from human neurons. The presynaptic action potentials had large overshoots to ∼25 mV and short durations of ∼0.5 ms. Our findings show that Ngn2-induced neurons represent an elegant model system allowing for functional, structural, and molecular analyses of glutamatergic synaptic transmission with high spatio-temporal resolution in human neurons. Furthermore, our data predict that glutamatergic transmission is mediated by large and rapid presynaptic action potentials in the human brain.Significance statementPresynaptic physiology remains poorly understood despite its relevance to neurological and psychiatric diseases. Studying presynaptic functions in human iPSC-derived neurons offers the important advantage of characterizing molecular mechanisms of neurotransmitter release in neurons derived from diseased patients. As a first step towards this goal, we established direct presynaptic whole-cell patch-clamp recordings from human glutamatergic neurons induced by transient Neurogenin 2 overexpression. We furthermore analyzed the structure of the synapses with super-resolution light microscopy and the synaptic short-term plasticity during high-frequency transmission. Our findings show that synchronous high-frequency transmission is mediated by rapid and large presynaptic action potentials in human neurons, similar to small conventional nerve terminals of rodent neurons.

Thermoregulatory pathway underlying the pyrogenic effects of prostaglandin E2 in the lateral parabrachial nucleus of male rats.

It has been shown that prostaglandin (PG) E2 synthesized in the lateral parabrachial nucleus (LPBN) is involved in lipopolysaccharide-induced fever. But the neural mechanisms of how intra-LPBN PGE2 induces fever remain unclear. In this study, we investigated whether the LPBN-preoptic area (POA) pathway, the thermoafferent pathway for feed-forward thermoregulatory responses, mediates fever induced by intra-LPBN PGE2 in male rats. The core temperature (Tcore) was monitored using a temperature radiotelemetry transponder implanted in rat abdomen. We showed that microinjection of PGE2 (0.28 nmol) into the LPBN significantly enhanced the density of c-Fos-positive neurons in the median preoptic area (MnPO). The chemical lesioning of MnPO with ibotenate or selective genetic lesioning or inhibition of the LPBN-MnPO pathway significantly attenuated fever induced by intra-LPBN injection of PGE2. We demonstrated that EP3 receptor was a pivotal receptor for PGE2-induced fever, since microinjection of EP3 receptor agonist sulprostone (0.2 nmol) or EP3 receptor antagonist L-798106 (2 nmol) into the LPBN mimicked or weakened the pyrogenic action of LPBN PGE2, respectively, but this was not the case for EP4 and EP1 receptors. Whole-cell recording from acute LPBN slices revealed that the majority of MnPO-projecting neurons originating from the external lateral (el) and dorsal (d) LPBN were excited and inhibited, respectively, by PGE2 perfusion, initiating heat-gain and heat-loss mechanisms. The amplitude but not the frequency of spontaneous and miniature glutamatergic excitatory postsynaptic currents (sEPSCs and mEPSCs) in MnPO-projecting LPBel neurons increased after perfusion with PGE2; whereas the frequency and amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs) and the A-type potassium (IA) current density did not change. In MnPO-projecting LPBd neurons, neither sEPSCs nor sIPSCs responded to PGE2; however, the IA current density was significantly increased by PGE2 perfusion. These electrophysiological responses and the thermoeffector reactions to intra-LPBN PGE2 injection, including increased brown adipose tissue thermogenesis, shivering, and decreased heat dissipation, were all abolished by L-798106, and mimicked by sulprostone. These results suggest that the pyrogenic effects of intra-LPBN PGE2 are mediated by both the inhibition of the LPBd-POA pathway through the EP3 receptor-mediated activation of IA currents and the activation of the LPBel-POA pathway through the selective enhancement of glutamatergic synaptic transmission via EP3 receptors.

Prenatal exposure to valproic acid reduces synaptic δ-catenin levels and disrupts ultrasonic vocalization in neonates

Valproic acid (VPA) is an effective and commonly prescribed drug for epilepsy and bipolar disorder. However, children born from mothers treated with VPA during pregnancy exhibit an increased incidence of autism spectrum disorder (ASD). Although VPA may impair brain development at the cellular level, the mechanism of VPA-induced ASD has not been completely addressed. A previous study has found that VPA treatment strongly reduces δ-catenin mRNA levels in cultured human neurons. δ-catenin is important for the control of glutamatergic synapses and is strongly associated with ASD. VPA inhibits dendritic morphogenesis in developing neurons, an effect that is also found in neurons lacking δ-catenin expression. We thus hypothesize that prenatal exposure to VPA significantly reduces δ-catenin levels in the brain, which impairs glutamatergic synapses to cause ASD. Here, we found that prenatal exposure to VPA markedly reduced δ-catenin levels in the brain of mouse pups. VPA treatment also impaired dendritic branching in developing mouse cortical neurons, which was partially reversed by elevating δ-catenin expression. Prenatal VPA exposure significantly reduced synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor levels and postsynaptic density 95 (PSD95) in the brain of mouse pups, indicating dysfunctions in glutamatergic synaptic transmission. VPA exposure also significantly altered ultrasonic vocalization (USV) in newly born pups when they were isolated from their nest. Moreover, VPA-exposed pups show impaired hypothalamic response to isolation, which is required to produce animals’ USVs following isolation from the nest. Therefore, these results suggest that VPA-induced ASD pathology can be mediated by the loss of δ-catenin functions.

386 Seizure Presentation in IDH WT Glioma is Related to Upregulated Synaptic Signaling and Ionic Transport Genes

INTRODUCTION: IDH wildtype glioblastoma is the most common malignant brain tumor in adults and is universally lethal. Seizures often occur as a presenting sign of tumor. Recent studies have implicated tumor-neuron communication facilitating synaptic remodeling and peritumoral circuit hyperactivity influencing tumor growth. METHODS: 63 tumor samples were analyzed for clinical characteristics. Bulk RNA sequencing data was used to identify differentially expressed genes (DEGs), gene sets enrichment analysis (GSEA), and gene ontologies (GO). RESULTS: 24 IDH WT GBM patients presented with seizures compared to 39 patients presenting with other symptoms. There were no significant differences between age at presentation, sex, tumor lobe, tumor location, or grade of tumor. GSEA of gene ontologies revealed upregulated genes in the seizure cohort were largely related to synaptic signaling and ionic transport (a., p-val < 0.05). Alternatively downregulated gene sets were related to extracellular matrix organization, and inflammatory and immune response (b., p-val < 0.05). Focusing on significantly upregulated DEGs (padj < 0.05) biological processes related to glutamatergic synaptic transmission and cellular components related to synaptic structures were upregulated. Alternatively, significantly downregulated DEGs (padj < 0.05) were related to biological processes of immunity and extracellular matrix organization and cellular components related to the extracellular matrix. Upregulated genes include IGFN1 and THBS4 both involved in synapse assembly, and SLC17A3, involved in presynaptic glutamate sorting (LogFC > 1, padj < 0.05). The downregulated genes include numerous collagen related genes and immune related genes and KCNJ15 an inward rectifying potassium channel and GABRR1 a GABA-rho subunit. CONCLUSIONS: Using bulk RNA sequencing in a clinically similar set of IDH WT tumors, we found gene sets and gene ontologies related to synaptic signaling and glutamatergic signaling upregulated in patients with seizures while genes related to both extracellular matrix and immune inflammation were downregulated.

Repeated Morphine Exposure Alters Temporoamonic-CA1 Synaptic Plasticity in Male Rat Hippocampus

In this study, the electrophysiological and biochemical consequences of repeated exposure to morphine in male rats on glutamatergic synaptic transmission, synaptic plasticity, the expression of GABA receptors and glutamate receptors at the temporoammonic-CA1 synapse along the longitudinal axis of the hippocampus (dorsal, intermediate, ventral, DH, IH, VH, respectively) were investigated. Slice electrophysiological methods, qRT-PCR, and western blotting techniques were used to characterize synaptic plasticity properties. We showed that repeated morphine exposure (RME) reduced excitatory synaptic transmission and ability for long-term potentiation (LTP) in the VH as well as eliminated the dorsoventral difference in paired-pulse responses. A decreased expression of NR2B subunit in the VH and an increased expression GABAA receptor of α1 and α5 subunits in the DH were observed following RME. Furthermore, RME did not affect the expression of NR2A, AMPA receptor subunits, and γ2GABAA and GABAB receptors in either segment of the hippocampus. In sum, the impact of morphine may differ depending on the region of the hippocampus studied. A distinct change in the short- and long-term synaptic plasticity along the hippocampus long axis due to repeated morphine exposure, partially mediated by a change in the expression profile of glutamatergic receptor subunits. These findings can be useful in further understanding the cellular mechanism underlying deficits in information storage and, more generally, cognitive processes resulting from chronic opioid abuse.

Toluene alters the intrinsic excitability and excitatory synaptic transmission of basolateral amygdala neurons.

Inhalant abuse is an important health issue especially among children and adolescents who often encounter these agents in the home. Research into the neurobiological targets of inhalants has lagged behind that of other drugs such as alcohol and psychostimulants. However, studies from our lab and others have begun to reveal how inhalants such as the organic solvent toluene affect neurons in key addiction related areas of the brain including the ventral tegmental area, nucleus accumbens and medial prefrontal cortex. In the present study, we extend these findings and examine the effect of toluene on electrophysiological responses of pyramidal neurons in the basolateral amygdala BLA, a region important for generating emotional and reward based information needed to guide future behavior. Whole-cell patch-clamp electrophysiology recordings of BLA pyramidal neurons in rat brain slices were used to assess toluene effects on intrinsic excitability and excitatory glutamatergic synaptic transmission. Acute application of 3 mM but not 0.3 mM toluene produced a small but significant (~20%) increase in current-evoked action potential (AP) firing that reversed following washout of the toluene containing solution. The change in firing during exposure to 3 mM toluene was accompanied by selective changes in AP parameters including reduced latency to first spike, increased AP rise time and decay and a reduction in the fast after-hyperpolization. To examine whether toluene also affects excitatory synaptic signaling, we expressed channelrhodopsin-2 in medial prefrontal cortex neurons and elicited synaptic currents in BLA neurons via light pulses. Toluene (3 mM) reduced light-evoked AMPA-mediated synaptic currents while a lower concentration (0.3 mM) had no effect. The toluene-induced reduction in AMPA-mediated BLA synaptic currents was prevented by the cannabinoid receptor-1 antagonist AM281. These findings are the first to demonstrate effects of acute toluene on BLA pyramidal neurons and add to existing findings showing that abused inhalants such as toluene have significant effects on neurons in brain regions involved in natural and drug induced reward.

Cell-Type-Specific Effects of Somatostatin on Synaptic Transmission in the Anterior Cingulate Cortex.

Inhibitory modulation of glutamatergic information processing is a prerequisite for proper network function. Among the many groups of interneurons (INs), somatostatin-expressing interneurons (SOM-INs) play an important role in the maintenance of physiological brain activity. We have previously shown that somatostatin (SOM) causes a reduction in pyramidal cell (PC) excitability. However, the mechanisms of action of the peptide on cortical synaptic circuits are still unclear. To understand the effects of the neuropeptide SOM on cortical synaptic circuits, we performed a detailed side-by-side comparison of its postsynaptic effects on PCs, SOM-INs, and layer 1 interneurons (L1-INs) in the anterior cingulate cortex of male and female mice and found that SOM produced pronounced postsynaptic effects in PCs while having little to no effect on either IN type. This comparison allowed us to link the observed postsynaptic effects to SOM-induced modulations of glutamatergic and GABAergic synaptic transmission and to trace the impact of the neuropeptide on the neuronal circuitry between these three cell types. We show here that SOM depresses glutamatergic synaptic transmission via a presynaptic mechanism while exerting a differential impact on GABAA receptor- and GABAB receptor-mediated transmission at the pre- and postsynaptic level resulting in a shift of inhibition in L2/3 PCs from L1-INs to SOM-INs. In summary, this study unravels a novel aspect by which SOM modulates synaptic signaling between PCs, L1-INs, and SOM-INs.

Cacao Procyanidins-Induced Lifespan Extension in Caenorhabditis elegans in a Nervous System and CaMKII-Dependent Manner.

Procyanidins are gaining attention due to their potential health benefits. We found that cacao liquor procyanidin (CLPr) from Theobroma cacao seeds increased the lifespan of Caenorhabditis elegans, a representative model organism for aging studies. The genetic dependence of the lifespan-extending effect of CLPr was consistent with that of blueberry procyanidin, which is dependent on unc-43, osr-1, sek-1, and mev-1, but not on daf-16, sir-2.1, or skn-1. The lifespan-extending effect of CLPr was inhibited by neuron-specific RNA interference (RNAi) targeting unc-43 and pmk-1, and in worms with loss-of-function mutations in the odr-3, odr-1, or tax-4 genes, which are essential in sensory neurons, including AWC neurons. It was also inhibited in worms in which AWC neurons or AIB interneurons had been eliminated, and in worms with loss-of-function mutations in eat-4 or glr-1, which are responsible for glutamatergic synaptic transmission. These results suggest that the lifespan-extending effect of CLPr is dependent on the nervous system. In addition, it also requires unc-43 and pmk-1 expression in nonneuronal cells, as demonstrated by the experiments with RNAi in wild-type worms, the neuronal cells of which are not affected by systemic RNAi. The osr-1 gene is expressed in hypodermal and intestinal cells and regulates the response to osmotic stress along with unc-43/calcium/calmodulin-dependent protein kinase II and the p38 mitogen-activated protein kinase pathway. Consistent with this, CLPr improved osmotic stress tolerance in an unc-43- and pmk-1-dependent manner, and it was also dependent on AWC neurons. The lifespan-extending and osmotic-tolerance-improving activities were attributed to procyanidins with a tetrameric or higher-order oligomeric structure.

Retraction Note: Predator odor increases avoidance and glutamatergic synaptic transmission in the prelimbic cortex via corticotropin-releasing factor receptor 1 signaling.

Retraction Note: Predator odor increases avoidance and glutamatergic synaptic transmission in the prelimbic cortex via corticotropin-releasing factor receptor 1 signaling.

Insulin-like growth factor 2 (IGF-2) rescues social deficits in NLG3-/y mouse model of ASDs.

Autism spectrum disorders (ASDs) comprise developmental disabilities characterized by impairments of social interaction and repetitive behavior, often associated with cognitive deficits. There is no current treatment that can ameliorate most of the ASDs symptomatology; thus, identifying novel therapies is urgently needed. Here, we used the Neuroligin 3 knockout mouse (NLG3-/y), a model that recapitulates the social deficits reported in ASDs patients, to test the effects of systemic administration of IGF-2, a polypeptide that crosses the blood-brain barrier and acts as a cognitive enhancer. We show that systemic IGF-2 treatment reverses the typical defects in social interaction and social novelty discrimination reflective of ASDs-like phenotypes. This effect was not accompanied by any change in spontaneous glutamatergic synaptic transmission in CA2 hippocampal region, a mechanism found to be crucial for social novelty discrimination. However, in both NLG3+/y and NLG3-/y mice IGF-2 increased cell excitability. Although further investigation is needed to clarify the cellular and molecular mechanisms underpinning IGF-2 effect on social behavior, our findings highlight IGF-2 as a potential pharmacological tool for the treatment of social dysfunctions associated with ASDs.

α7 nicotinic acetylcholine receptors induce long-term synaptic enhancement in the dorsal but not ventral hippocampus.

Agents that positively modulate the activity of α7nAChRs are used as cognitive enhancers and for the treatment of hippocampus-dependent functionaldecline. However, it is not known whether the expression and the effects of α7nAChRs apply to the entire longitudinal axis of the hippocampus equally. Given that cholinergic system-involving hippocampal functions are not equally distributed along the hippocampus, we comparatively examined the expression and the effects of α7nAChRs on excitatory synaptic transmission between the dorsal and the ventral hippocampal slices from adult rats. We found that α7nAChRs are equally expressed in the CA1 field of the two segments of the hippocampus. However, activation of α7nAChRs by their highly selective agonist PNU 282987 induced a gradually developing increase in field excitatory postsynaptic potential only in the dorsal hippocampus. This long-term potentiation was not reversed upon application of nonselective nicotinic receptor antagonist mecamylamine, but the induction of potentiation was prevented by prior blockade of α7nAChRs by their antagonist MG 624. In contrast to the long-term synaptic plasticity, we found that α7nAChRs did not modulate short-term synaptic plasticity in either the dorsal or the ventral hippocampus. These results may have implications for the role that α7nAChRs play in specifically modulating functions that depend on the normal function of the dorsal hippocampus. We propose that hippocampal functions that rely on a direct α7 nAChR-mediated persistent enhancement of glutamatergic synaptic transmission are preferably supported by dorsal but not ventral hippocampal synapses.

TAAR1 agonist ulotaront modulates striatal and hippocampal glutamate function in a state-dependent manner

Aberrant dopaminergic and glutamatergic function, particularly within the striatum and hippocampus, has repeatedly been associated with the pathophysiology of schizophrenia. Supported by preclinical and recent clinical data, trace amine-associated receptor 1 (TAAR1) agonism has emerged as a potential new treatment approach for schizophrenia. While current evidence implicates TAAR1-mediated regulation of dopaminergic tone as the primary circuit mechanism, little is known about the effects of TAAR1 agonists on the glutamatergic system and excitation-inhibition balance. Here we assessed the impact of ulotaront (SEP-363856), a TAAR1 agonist in Phase III clinical development for schizophrenia, on glutamate function in the mouse striatum and hippocampus. Ulotaront reduced spontaneous glutamatergic synaptic transmission and neuronal firing in striatal and hippocampal brain slices, respectively. Interestingly, ulotaront potentiated electrically-evoked excitatory synaptic transmission in both brain regions, suggesting the ability to modulate glutamatergic signaling in a state-dependent manner. Similar striatal effects were also observed with the TAAR1 agonist, RO5166017. Furthermore, we show that ulotaront regulates excitation-inhibition balance in the striatum by specifically modulating glutamatergic, but not GABAergic, spontaneous synaptic events. These findings expand the mechanistic circuit hypothesis of ulotaront and TAAR1 agonists, which may be uniquely positioned to normalize both the excessive dopaminergic tone and regulate abnormal glutamatergic function associated with schizophrenia.

Protein 4.1N Plays a Cell Type-Specific Role in Hippocampal Glutamatergic Synapse Regulation.

Many glutamatergic synapse proteins contain a 4.1N protein binding domain. However, a role for 4.1N in the regulation of glutamatergic neurotransmission has been controversial. Here, we observe significantly higher expression of protein 4.1N in granule neurons of the dentate gyrus (DG granule neurons) compared with other hippocampal regions. We discover that reducing 4.1N expression in rat DG granule neurons of either sex results in a significant reduction in glutamatergic synapse function that is caused by a decrease in the number of glutamatergic synapses. By contrast, we find reduction of 4.1N expression in hippocampal CA1 pyramidal neurons has no impact on basal glutamatergic neurotransmission. We also find 4.1N's C-terminal domain (CTD) to be nonessential to its role in the regulation of glutamatergic synapses of DG granule neurons. Instead, we show that 4.1N's four-point-one, ezrin, radixin, and moesin (FERM) domain is essential for supporting synaptic AMPA receptor (AMPAR) function in these neurons. Altogether, this work demonstrates a novel, cell type-specific role for protein 4.1N in governing glutamatergic synapse function.SIGNIFICANCE STATEMENT Glutamatergic synapses exhibit immense molecular diversity. In comparison to heavily studied Schaffer collateral, CA1 glutamatergic synapses, significantly less is known about perforant path-dentate gyrus (DG) synapses. Our data demonstrate that compromising 4.1N function in CA1 pyramidal neurons produces no alteration in basal glutamatergic synaptic transmission. However, in DG granule neurons, compromising 4.1N function leads to a significant decrease in the strength of glutamatergic neurotransmission at perforant pathway synapses. Together, our data identifies 4.1N as a cell type-specific regulator of synaptic transmission within the hippocampus and reveals a unique molecular program that governs perforant pathway synapse function.

Activation of Metabotropic Glutamate Receptor 3 Modulates Thalamo-accumbal Transmission and Rescues Schizophrenia-like Physiological and Behavioral Deficits

BackgroundPolymorphisms in the gene encoding for metabotropic glutamate receptor 3 (mGlu3) are associated with an increased likelihood of schizophrenia diagnosis and can predict improvements in negative symptoms following treatment with antipsychotics. However, the mechanisms by which mGlu3 can regulate brain circuits involved in schizophrenia pathophysiology are not clear. MethodsWe employed selective pharmacological tools and a variety of approaches including whole-cell patch-clamp electrophysiology, slice optogenetics, and fiber photometry to investigate the effects of mGlu3 activation on phencyclidine (PCP)-induced impairments in thalamo-accumbal transmission and sociability deficits. A chemogenetic approach was used to evaluate the role of thalamo-accumbal transmission in PCP-induced sociability deficits. ResultsWe first established that PCP treatment augmented excitatory transmission onto dopamine D1 receptor–expressing medium spiny neurons (D1-MSNs) in the nucleus accumbens (NAc) and induced sociability deficits. Our studies revealed a selective increase in glutamatergic synaptic transmission from thalamic afferents to D1-MSNs in the NAc shell. Chemogenetic silencing of thalamo-accumbal inputs rescued PCP-induced sociability deficits. Pharmacological activation of mGlu3 normalized PCP-induced impairments in thalamo-accumbal transmission and sociability deficits. Mechanistic studies revealed that mGlu3 activation induced robust long-term depression at synapses from the thalamic projections onto D1-MSNs in the NAc shell. ConclusionsThese data demonstrate that activation of mGlu3 decreases thalamo-accumbal transmission and thereby rescues sociability deficits in mouse modeling schizophrenia-like symptoms. These findings provide novel insights into the NAc-specific mechanisms and suggest that agents modulating glutamatergic signaling in the NAc may provide a promising approach for treating negative symptoms in schizophrenia.

APP/AICD‐dependent expression of Glypican 4 mediates accumulation of tau oligomers in astrocytes and their synaptotoxic action

AbstractBackgroundSeveral studies including ours reported the detrimental effects of extracellular tau oligomers (ex‐oTau) on glutamatergic synaptic transmission and plasticity. Astrocytes greatly internalize ex‐oTau thus contributing to their clearance. However, glial cell engulfment of oTau leads to alteration of neuro/gliotransmitter handling that negatively affects synaptic function. We previously reported that amyloid precursor protein (APP) is required for oTau internalization in astrocytes but the molecular mechanisms underlying this phenomenon has not been clearly identified yet. Here we investigated the role of glypican 4 (GPC4), a member of the large family of heparan sulfate proteoglycans that are membrane glycoproteins highly expressed in astrocytes playing a critical role in protein trafficking through the plasma membrane.MethodThe role of GPC4 in ex‐oTau accumulation in astrocytes and the resulting synaptotoxic action were studied in in vitro and ex‐vivo models obtained from wild‐type C57/bl6 and transgenic mice in which either APP was knocked‐out or it contained the non‐phosphorylatable amino acid alanine replacing threonine 688 (APPTA mice). In these experimental models we performed immunofluorescence analyses, Western blot and real‐time PCR experiments, confocal Ca2+ imaging, high‐pressure liquid chromatography, FM1‐43 imaging, electrophysiological recordings and Chromatin immunoprecipitation assays.ResultTreatment of cultured astrocytes with a specific anti‐GPC4 antibody significantly reduced oTau internalization in astrocytes and prevented oTau‐induced alterations of intracellular Ca2+ signaling and gliotransmitter release. As such, anti‐GPC4 treatment spared neurons from the astrocyte‐mediated synaptotoxic action of ex‐oTau. Specifically, after counteracting the interaction between ex‐oTau and GPC4 no significant decreases in synaptic vesicular release and synaptic protein expression were observed in neurons co‐cultured with astrocytes following exposure to ex‐oTau. The latter also failed to inhibit hippocampal LTP at CA3‐CA1 synapses. Of note, we found that the expression of GPC4 depended on APP and, in particular, on its C‐terminal domain, AICD, generated by γ‐secretase‐dependent cleavage of APP, that can bind Gpc4 promoter. Accordingly, GPC4 expression was significantly reduced in both APP‐KO ad APPTA mice, and ex‐oTau did not exert any significant detrimental action on synaptic function in APP‐KO mice.ConclusionCollectively, our data indicate that GPC4 expression is APP/AICD‐dependent, it mediates oTau accumulation in astrocytes and the resulting synaptotoxic effects.

Pathogenesis of Depression in Alzheimer's Disease.

Depression is a prevalent occurrence among Alzheimer's disease (AD) patients, yet its underlying mechanism remains unclear. Recent investigations have revealed that several pathophysiological changes associated with Alzheimer's disease can lead to mood disorders. These alterations include irregularities in monoamine neurotransmitters, disruptions in glutamatergic synaptic transmission, neuro-inflammation, dysfunction within the hypothalamic-pituitary-adrenocortical (HPA) axis, diminished levels of brain-derived neurotrophic factor (BDNF), and hippocampal atrophy. This review consolidates research findings from pertinent fields to elucidate the mechanisms underlying depression in Alzheimer's disease, aiming to provide valuable insights for the study of its mechanisms and clinical treatment.

Effects of propofol on presynaptic synapsin phosphorylation in the mouse brain in vivo

The commonly used general anesthetic propofol can enhance the γ-aminobutyric acid-mediated inhibitory synaptic transmission and depress the glutamatergic excitatory synaptic transmission to achieve general anesthesia and other outcomes. In addition to the actions at postsynaptic sites, the modulation of presynaptic activity by propofol is thought to contribute to neurophysiological effects of the anesthetic, although potential targets of propofol within presynaptic nerve terminals are incompletely studied at present. In this study, we explored the possible linkage of propofol to synapsins, a family of neuron-specific phosphoproteins which are the most abundant proteins on presynaptic vesicles, in the adult mouse brain in vivo. We found that an intraperitoneal injection of propofol at a dose that caused loss of righting reflex increased basal levels of synapsin phosphorylation at the major representative phosphorylation sites (serine 9, serine 62/67, and serine 603) in the prefrontal cortex (PFC) of male and female mice. Propofol also elevated synapsin phosphorylation at these sites in the striatum and S9 and S62/67 phosphorylation in the hippocampus, while propofol had no effect on tyrosine hydroxylase phosphorylation in striatal nerve terminals. Total synapsin protein expression in the PFC, hippocampus, and striatum was not altered by propofol. These results reveal that synapsin could be a novel substrate of propofol in the presynaptic neurotransmitter release machinery. Propofol possesses the ability to upregulate synapsin phosphorylation in broad mouse brain regions.

CNSC-11. SEIZURE PRESENTATION IN IDH WT GLIOMA IS RELATED TO UPREGULATED SYNAPTIC SIGNALING AND IONIC TRANSPORT GENES

Abstract Isocitrate dehydrogenase wildtype (IDH WT) glioblastoma (GBM) is the most common malignant brain tumor in adults and is universally lethal. Seizures often occur as a presenting sign of tumor. Recent studies have implicated tumor-neuron communication facilitating synaptic remodeling and peritumoral circuit hyperactivity influencing tumor growth. We assessed the clinical and transcriptional effect of seizure presentation on GBM biology. 63 tumor samples were analyzed for clinical characteristics. Bulk RNA sequencing data was used to identify differentially expressed genes (DEGs), gene sets enrichment analysis (GSEA), and gene ontologies (GO). 24 IDH WT GBM patients presented with seizures compared to 39 patients presenting with other symptoms. There were no significant differences between age at presentation, sex, tumor lobe, tumor location, or grade of tumor. GSEA of gene ontologies revealed upregulated genes in the seizure cohort were largely related to synaptic signaling, ionic transport, and receptor signaling (p-val< 0.05). Alternatively downregulated gene sets were related to extracellular matrix organization, and inflammatory and immune response (p-val< 0.05). Focusing on significantly upregulated DEGs (padj< 0.05,) biological processes related to glutamatergic synaptic transmission and cellular components related to synaptic structures were upregulated. Alternatively, significantly downregulated DEGs (padj< 0.05) were related to biological processes of immunity and extracellular matrix organization and cellular components related to the extracellular matrix. Upregulated genes include IGFN1 and THBS4, both involved in synapse assembly, and SLC17A3, involved in presynaptic glutamate sorting (Log2FC >1,padj <0.05). The downregulated genes include numerous collagen related genes and immune related genes and KCNJ15, an inward rectifying potassium channel and GABRR1, a GABA-rho subunit. Using bulk RNA sequencing in a clinically similar set of IDH WT tumors, we found gene sets and gene ontologies related to synaptic signaling and glutamatergic signaling upregulated in patients with seizures while genes related to both extracellular matrix and immune inflammation were downregulated.

CNSC-14. T-TYPE CALCIUM CHANNELS REGULATE NEURON-GLIOBLASTOMA INTERACTIONS AND PROMOTE TUMOR GROWTH

Abstract Glioblastoma (GBM) is the most common primary malignant brain tumor with a dismal median survival of 15 months. Glioblastoma utilizes calcium signaling to regulate communication between tumor cells and the neuronal microenvironment. T-type calcium channels (TTCCs) are voltage gated calcium channels expressed in various types of neurons throughout the brain regulating glutamatergic synaptic transmission. We analyzed spatial transcriptomics of human GBM tumors and found that TTCCs are expressed in infiltrating tumor regions and enriched for neuronal development spatial patterns and exhibit neurodevelopmental signatures. Analysis of single cells transcriptomic of human GBM tumors demonstrated that TTCCs are enriched in neuronal subtypes. To complement publicly available data,we treated Glioblastoma Stem cells (GSCs) with an FDA-approved repurposed TTCC blocker, mibefradil, and found downregulation of genes associated with neuron processes such as synapse organization, postsynaptic density and glutamatergic synapses. Treatment of GSCs with mibefradil abrogated cell proliferation in the presence of glutamate. We examined the effects of neuronal Cav3.2 on GBM growth. Genetic Cav3.2 knockout (Cav3.2 KO) in the microenvironment of mice xenografted with syngeneic cell lines (GL261 and CT2A) exhibited significant tumor volume reduction as well as increased survival compared to WT mice in both cell lines. Cav3.2 KO tumors exhibited a decrease in proliferation (Ki67+) and neuronal connections (VGlut1) measured by immunofluorescence compared to WT mice. Cav3.2KO neuron-GBM co-cultures exhibited a significant decrease in proliferation compared to WT neuron-GBM co-cultures. Cav3.2KO neuron-GBM co-cultures exhibited a decrease in synaptic connections and a decrease in neuronal projections to GBM cells compared to WT neuron-GBM co-cultures. Transcriptomic analysis of tumors from Cav3.2KO tumors revealed deregulation of genes associated synaptic transmission, glutamatergic synapses and postsynaptic specialization. Altogether, our findings indicate a role of Cav3.2 in regulating tumors and neurons and promoting GBM progression.