Hippocampal Synaptic Transmission Research Articles

Individual NMDA receptor GluN2 subunit signaling domains differentially regulate the postnatal maturation of hippocampal excitatory synaptic transmission and plasticity but not dendritic morphology.

N-methyl-d-aspartate receptors (NMDARs) at hippocampal excitatory synapses undergo a late postnatal shift in subunit composition, from an initial prevalence of GluN2B subunit incorporation to a later predominance of GluN2A. This GluN2B to GluN2A shift alters NMDAR calcium conductance dynamics and intracellular molecular signaling that are individually regulated by distinct GluN2 signaling domains and temporally align with developmental alterations in dendritic and synaptic plasticity. However, the impacts of individual GluN2B to GluN2A signaling domains on neuronal development remain unknown. Ionotropic and intracellular signaling domains of GluN2 subunits were separated by creating chimeric GluN2 subunits that were expressed in two transgenic mouse lines. Western blot and immunoprecipitation revealed that roughly one third of native synaptic NMDARs were replaced by transformed NMDARs without altering total synaptic NMDAR content. Schaffer collateral synaptic strength was transiently increased in acutely prepared hippocampal slices at just over 3 weeks of age in animals overexpressing the GluN2B carboxy terminus. Long-term potentiation (LTP) induction following lower frequency stimulation was regulated by GluN2 ionotropic signaling domains in an age-dependent manner and LTP maintenance was enhanced by overexpression of the GluN2B CTD in mature animals. After higher frequency stimulation, the induction and maintenance of LTP were increased in young adult animals overexpressing the GluN2B ionotropic signaling domains but reduced in juveniles just over 3 weeks of age. Confocal imaging of green fluorescent protein (GFP)- labeled CA1 pyramidal neurons revealed no alterations in dendritic morphology or spine density in mice expressing chimeric GluN2 subunits. These results illustrate how individual GluN2 subunit signaling domains do or do not control physiological and morphological development of hippocampal excitatory neurons and better clarify the neurobiological factors that govern hippocampal maturation. SIGNIFICANCE STATEMENT: A developmental reduction in the magnitude of hippocampal long-term synaptic potentiation (LTP) and a concomitant improvement in spatial maze performance coincide with greater incorporation of GluN2A subunits into synaptic NMDARs. Corroborating our prior discovery that overexpression of GluN2A-type ionotropic signaling domains enables context-based navigation in immature mice, GluN2A-type ionotropic signaling domain overexpression reduces LTP induction threshold and magnitude in immature mice. Also, we previously found that GluN2B carboxy terminal domain (CTD) overexpression enhances long-term spatial memory in mature mice and now report that the GluN2B CTD is associated with greater amplitude of LTP after induction in mature mice. Thus, the late postnatal maturation of context encoding likely relies on a shift toward GluN2A-type ionotropic signaling and a reduction in the threshold to induce LTP while memory consolidation and LTP maintenance are regulated by GluN2B subunit CTD signaling.

The gut metabolite indole-3-propionic acid activates ERK1 to restore social function and hippocampal inhibitory synaptic transmission in a 16p11.2 microdeletion mouse model

BackgroundMicrodeletion of the human chromosomal region 16p11.2 (16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document}) is a prevalent genetic factor associated with autism spectrum disorder (ASD) and other neurodevelopmental disorders. However its pathogenic mechanism remains unclear, and effective treatments for 16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document} syndrome are lacking. Emerging evidence suggests that the gut microbiota and its metabolites are inextricably linked to host behavior through the gut-brain axis and are therefore implicated in ASD development. Despite this, the functional roles of microbial metabolites in the context of 16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document} are yet to be elucidated. This study aims to investigate the therapeutic potential of indole-3-propionic acid (IPA), a gut microbiota metabolite, in addressing behavioral and neural deficits associated with 16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document}, as well as the underlying molecular mechanisms.ResultsMice with the 16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document} showed dysbiosis of the gut microbiota and a significant decrease in IPA levels in feces and blood circulation. Further, these mice exhibited significant social and cognitive memory impairments, along with hyperactivation of hippocampal dentate gyrus neurons and reduced inhibitory synaptic transmission in this region. However, oral administration of IPA effectively mitigated the histological and electrophysiological alterations, thereby ameliorating the social and cognitive deficits of the mice. Remarkably, IPA treatment significantly increased the phosphorylation level of ERK1, a protein encoded by the Mapk3 gene in the 16p11.2 region, without affecting the transcription and translation of the Mapk3 gene.ConclusionsOur study reveals that 16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document} leads to a decline in gut metabolite IPA levels; however, IPA supplementation notably reverses the behavioral and neural phenotypes of 16p11.2+/-\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$${}^{+/-}$$\\end{document} mice. These findings provide new insights into the critical role of gut microbial metabolites in ASD pathogenesis and present a promising treatment strategy for social and cognitive memory deficit disorders, such as 16p11.2 microdeletion syndrome.92Bbfs5rK2Gd_SbL4an2CwVideo

Unlocking cognition with pulsatile GnRH in Down Syndrome: implications for Alzheimer’s disease

AbstractBackgroundThere is currently no viable treatment for the cognitive and olfactory deficits of Trisomy 21 and Alzheimer’s disease.MethodPhysiological characterization and epigenetic, cellular, chemogenetic, and pharmacological interventions in male and female Ts65Dn mice (P9‐P360), a mouse model of Down syndrome, and 12‐month‐old THY::TAU22 male mice, a mouse model of Alzheimer’s disease, and their wild‐type littermates. Seven men with Down syndrome (20‐50 years of age) were enrolled in an open‐label pilot study to assess the effect of 6‐month pulsatile GnRH therapy on cognitive and olfactory function.ResultWe show in Ts65Dn mice that olfactory and cognitive neurological symptoms are closely related to a post‐pubertal decrease in hypothalamic and extra‐hypothalamic expression of a master molecule controlling reproduction ‐ gonadotropin‐releasing hormone (GnRH) ‐ and appear to be linked to an imbalance in a microRNA‐gene network known to regulate the maturation of GnRH neurons as well as to an alteration in hippocampal synaptic transmission. Restoring physiological GnRH levels abolishes olfactory and cognitive deficits in Ts65Dn and THY::TAU22 mice, while pulsatile GnRH treatment improves cognition and brain connectivity in adult T21 patients.ConclusionWe have discovered that GnRH plays a crucial role in olfaction and cognition, and pulsatile GnRH treatment holds promise for improving cognitive deficits in patients with Down syndrome. Such treatment could potentially also be beneficial for mobilizing cognitive reserve in pathological brain aging such as that found in Alzheimer’s disease.

The therapeutic potential of ellagic acid loaded in nanocarriers for the treatment of the Alzheimer’s disease

AbstractBackgroundEllagic acid (EA) has been shown to have protective effects on Alzheimer’s disease, but it shows poor pharmacokinetic profile. A valid approach is delivering the drug in non‐ionic surfactant vesicles (NBVs) through the nose to brain (N2B) route coupled to focused ultrasound (FUS) improving brain targeting. Several niosomal formulations composed by specific surfactants (potentially enhance neuroprotective effect) have been characterized. The contrast agent was loaded inside the nanocarriers which was subsequently characterized using FUS in vitro technique. The EA will be loaded inside the vesicles and the neuroprotective effect of NBVs and EA‐NBVs will be explored using electrophysiological recordings (field and whole cell patch clamp) of hippocampal synaptic transmission and plasticity in ex‐vivo Alzheimer model.MethodEmpty and loaded NBVs were prepared and deeply characterized. In vitro FUS analysis were performed to evaluate the ability of FUS to induce a stable and prolonged release of probe‐drug loaded. The electrophysiological experiments were conducted using C57BL6/J mice of both sexes aged between 30 and 40 days. We performed electrophysiological recordings in area CA1‐CA3. Paired pulse facilitation was assessed at inter‐stimulus intervals ranging from 50 to 500 ms. An additional recording of baseline was evaluated before inducing Long‐Term Potentiation (LTP) by theta‐burst stimulation. Responses (fEPSP) were recorded for 1 hour after tetanization. In vitro Alzheimer model was obtained applying Aβ1‐42 oligomers (200 nM). As a control, we tested the effect of Aβ1‐42 with a scrambled sequence applied in the same condition of Aβ1‐42. Empty and loaded NBVs were used in presence and not of Aβ1‐42 and of Aβ1‐42 scrambled.ResultPreliminary results of empty and loaded NBVs present an optimal range of size for N2B delivery and in vitro FUS release shows a stable and controlled release profile of loaded drug. Slices incubated in presence of Aβ1‐42 showed an impairment of LTP, while this effect didn’t occur after incubation of slices with Aβ1‐42 scrambled. Preliminary results show that NBVs formulations didn’t compromise LTP suggesting the possibility of using them as drug carriers in Alzheimer’s disease.ConclusionA better brain delivery of EA could be a valid approach in pathologies characterized by synaptic dysfunction.

Interleukin-15 alters hippocampal synaptic transmission and impairs episodic memory formation in mice

Cytokines are potent immunomodulators exerting pleiotropic effects in the central nervous system (CNS). They influence neuronal functions and circuit activities with effects on memory processes and behaviors. Here, we unravel a neuromodulatory activity of interleukin-15 (IL-15) in mouse brain. Acute exposure of hippocampal slices to IL-15 enhances gamma-aminobutyricacid (GABA) release and reduces glutamatergic currents, while chronic treatment with IL-15 increases the frequency of hippocampal miniature inhibitory synaptic transmission and impairs memory formation in the novel object recognition (NOR) test. Moreover, we describe that serotonin is involved in mediating the hippocampal effects of IL-15, because a selective 5-HT3A receptor antagonist prevents the effects on inhibitory neurotransmission and ameliorates mice performance in the NOR test. These findings provide new insights into the modulatory activities of cytokines in the CNS, with implications on behavior.

Mitragynine inhibits hippocampus neuroplasticity and its molecular mechanism

BackgroundMitragynine (MIT), the primary indole alkaloid of kratom (Mitragyna speciosa), has been associated with addictive and cognitive decline potentials. In acute studies, MIT decreases spatial memory and inhibits hippocampal synaptic transmission in long-term potentiation (LTP). This study investigated the impacts of 14-day MIT treatment on hippocampus synaptic transmission and its possible underlying mechanisms.MethodsUnder urethane anesthesia, field excitatory post-synaptic potentials (fEPSP) of the hippocampal CA1 region were recorded in the Sprague Dawley (SD) rats that received MIT (1, 5, and 10 mg/kg), morphine (MOR) 5 mg/kg, or vehicle (ip). The effects of the treatments on basal synaptic transmission, paired-pulse facilitation (PPF), and LTP were assessed in the CA1 region. Analysis of the brain's protein expression linked to neuroplasticity was then performed using a western blot.ResultsThe baseline synaptic transmission's amplitude was drastically decreased by MIT at 5 and 10 mg/kg doses, although the PPF ratio before TBS remained unchanged, the PPF ratio after TBS was significantly reduced by MIT (10 mg/kg). Strong and persistent inhibition of LTP was generated in the CA1 region by MIT (5 and 10 mg/kg) doses; this effect was not seen in MIT (1 mg/kg) treated rats. In contrast to MIT (1 mg/kg), MIT (5 and 10 mg/kg) significantly raised the extracellular glutamate levels. After exposure to MIT, GluR-1 receptor expression remained unaltered. However, NMDAε2 receptor expression was markedly downregulated. The expression of pCaMKII, pERK, pCREB, BDNF, synaptophysin, PSD-95, Delta fosB, and CDK-5 was significantly downregulated in response to MIT (5 and 10 mg/kg) exposure, while MOR (5 mg/kg) significantly raised synaptophysin and Delta fosB expression.ConclusionFindings from this work reveal that a smaller dose of MIT (1 mg/kg) poses no risk to hippocampal synaptic transmission. Alteration in neuroplasticity-associated proteins may be a molecular mechanism for MIT (5 and 10 mg/kg)-induced LTP disruption and cognitive impairments. Data from this work posit that MIT acted differently from MOR on neuroplasticity and its underlying mechanisms.Graphical abstract

Optoα1AR activation in astrocytes modulates basal hippocampal synaptic excitation and inhibition in a stimulation-specific manner.

Astrocytes play active roles at synapses and can monitor, respond, and adapt to local synaptic activity. While there is abundant evidence that astrocytes modulate excitatory transmission in the hippocampus, evidence for astrocytic modulation of hippocampal synaptic inhibition remains more limited. Furthermore, to better investigate roles for astrocytes in modulating synaptic transmission, more tools that can selectively activate native G protein signaling pathways in astrocytes with both spatial and temporal precision are needed. Here, we utilized AAV8-GFAP-Optoα1AR-eYFP (Optoα1AR), a viral vector that enables activation of Gq signaling in astrocytes via light-sensitive α1-adrenergic receptors. To determine if stimulating astrocytic Optoα1AR modulates hippocampal synaptic transmission, recordings were made in CA1 pyramidal cells with surrounding astrocytes expressing Optoα1AR, channelrhodopsin (ChR2), or GFP. Both high-frequency (20 Hz, 45-ms light pulses, 5 mW, 5 min) and low-frequency (0.5 Hz, 1-s pulses at increasing 1, 5, and 10 mW intensities, 90 s per intensity) blue light stimulation were tested. 20 Hz Optoα1AR stimulation increased both inhibitory and excitatory postsynaptic current (IPSC and EPSC) frequency, and the effect on miniature IPSCs (mIPSCs) was largely reversible within 20 min. However, low-frequency stimulation of Optoα1AR did not modulate either IPSCs or EPSCs, suggesting that astrocytic Gq -dependent modulation of basal synaptic transmission in the hippocampus is stimulation-dependent. By contrast, low-frequency stimulation of astrocytic ChR2 was effective in increasing both synaptic excitation and inhibition. Together, these data demonstrate that Optoα1AR activation in astrocytes changes basal GABAergic and glutamatergic transmission, but only following high-frequency stimulation, highlighting the importance of temporal dynamics when using optical tools to manipulate astrocyte function.

Selective inhibition of cannabinoid CB1 receptor-evoked signalling by the interacting protein GAP43

Cannabinoids exert pleiotropic effects on the brain by engaging the cannabinoid CB1 receptor (CB1R), a presynaptic metabotropic receptor that regulates key neuronal functions in a highly context-dependent manner. We have previously shown that CB1R interacts with growth-associated protein of 43 kDa (GAP43) and that this interaction inhibits CB1R function on hippocampal excitatory synaptic transmission, thereby impairing the therapeutic effect of cannabinoids on epileptic seizures in vivo. However, the underlying molecular features of this interaction remain unexplored. Here, we conducted mechanistic experiments on HEK293T cells co-expressing CB1R and GAP43 and show that GAP43 modulates CB1R signalling in a strikingly selective manner. Specifically, GAP43 did not affect the archetypical agonist-evoked (i) CB1R/Gi/o protein-coupled signalling pathways, such as cAMP/PKA and ERK, or (ii) CB1R internalization and intracellular trafficking. In contrast, GAP43 blocked an alternative agonist-evoked CB1R-mediated activation of the cytoskeleton-associated ROCK signalling pathway, which relied on the GAP43-mediated impairment of CB1R/Gq/11 protein coupling. GAP43 also abrogated CB1R-mediated ROCK activation in mouse hippocampal neurons, and this process led in turn to a blockade of cannabinoid-evoked neurite collapse. An NMR-based characterization of the CB1R-GAP43 interaction supported that GAP43 binds directly and specifically through multiple amino acid stretches to the C-terminal domain of the receptor. Taken together, our findings unveil a CB1R-Gq/11-ROCK signalling axis that is selectively impaired by GAP43 and may ultimately control neurite outgrowth.

Gestational dexamethasone exposure impacts hippocampal excitatory synaptic transmission and learning and memory function with transgenerational effects.

The formation of learning and memory is regulated by synaptic plasticity in hippocampal neurons. Here we explored how gestational exposure to dexamethasone, a synthetic glucocorticoid commonly used in clinical practice, has lasting effects on offspring's learning and memory. Adult offspring rats of prenatal dexamethasone exposure (PDE) displayed significant impairments in novelty recognition and spatial learning memory, with some phenotypes maintained transgenerationally. PDE impaired synaptic transmission of hippocampal excitatory neurons in offspring of F1 to F3 generations, and abnormalities of neurotransmitters and receptors would impair synaptic plasticity and lead to impaired learning and memory, but these changes failed to carry over to offspring of F5 and F7 generations. Mechanistically, altered hippocampal miR-133a-3p-SIRT1-CDK5-NR2B signaling axis in PDE multigeneration caused inhibition of excitatory synaptic transmission, which might be related to oocyte-specific high expression and transmission of miR-133a-3p. Together, PDE affects hippocampal excitatory synaptic transmission, with lasting consequences across generations, and CDK5 in offspring's peripheral blood might be used as an early-warning marker for fetal-originated learning and memory impairment.

A centronuclear myopathy-causing mutation in dynamin-2 disrupts neuronal morphology and excitatory synaptic transmission in a murine model of the disease.

Dynamin-2 (DNM2) is a large GTPase, a member of the dynamin superfamily that regulates membrane remodelling and cytoskeleton dynamics. Mutations in DNM2 cause autosomal dominant centronuclear myopathy (CNM), a congenital neuromuscular disorder characterized by progressive weakness and atrophy of the skeletal muscles. Cognitive defects have been reported in some DNM2-linked CNM patients suggesting that these mutations can also affect the central nervous system (CNS). Here we studied how a DNM2 CNM-causing mutation influences the CNS function. Heterozygous mice harbouring the p.R465W mutation in Dnm2 (HTZ), the most common causing autosomal dominant CNM, were used as the disease model. We evaluated dendritic arborization and spine density in hippocampal cultured neurons, analysed excitatory synaptic transmission by electrophysiological field recordings in hippocampal slices, and evaluated cognitive function by performing behavioural tests. HTZ hippocampal neurons exhibited reduced dendritic arborization and lower spine density than WT neurons, which was reversed by transfecting an interference RNA against the Dnm2 mutant allele. Additionally, HTZ mice showed defective hippocampal excitatory synaptic transmission and reduced recognition memory compared to the WT condition. Our findings suggest that the Dnm2 p.R465W mutation perturbs the synaptic and cognitive function in a CNM mouse model and support the idea that Dnm2 plays a key role in regulating neuronal morphology and excitatory synaptic transmission in the hippocampus.

Schedule-induced alcohol intake during adolescence sex dependently impairs hippocampal synaptic plasticity and spatial memory

In a previous study, we demonstrated that intermittent ethanol administration in male adolescent animals impaired hippocampus-dependent spatial memory, particularly under conditions of excessive ethanol administration. In this current study, we subjected adolescent male and female Wistar rats an alcohol schedule-induced drinking (SID) procedure to obtain an elevated rate of alcohol self-administration and assessed their hippocampus-dependent spatial memory. We also studied hippocampal synaptic transmission and plasticity, as well as the expression levels of several genes involved in these mechanisms. Both male and female rats exhibited similar drinking patterns throughout the sessions of the SID protocol reaching similar blood alcohol levels in all the groups. However, only male rats that consumed alcohol showed spatial memory deficits which correlated with inhibition of hippocampal synaptic plasticity as long-term potentiation. In contrast, alcohol did not modify hippocampal gene expression of AMPA and NMDA glutamate receptor subunits, although there are differences in the expression levels of several genes relevant to synaptic plasticity mechanisms underlying learning and memory processes, related to alcohol consumption as Ephb2, sex differences as Pi3k or the interaction of both factors such as Pten.In conclusion, elevated alcohol intake during adolescence seems to have a negative impact on spatial memory and hippocampal synaptic plasticity in a sex dependent manner, even both sexes exhibit similar blood alcohol concentrations and drinking patterns.

Adenosine A2A Receptors Shut Down Adenosine A1 Receptor-Mediated Presynaptic Inhibition to Promote Implementation of Hippocampal Long-Term Potentiation.

Adenosine operates a modulation system fine-tuning the efficiency of synaptic transmission and plasticity through A1 and A2A receptors (A1R, A2AR), respectively. Supramaximal activation of A1R can block hippocampal synaptic transmission, and the tonic engagement of A1R-mediated inhibition is increased with increased frequency of nerve stimulation. This is compatible with an activity-dependent increase in extracellular adenosine in hippocampal excitatory synapses, which can reach levels sufficient to block synaptic transmission. We now report that A2AR activation decreases A1R-medated inhibition of synaptic transmission, with particular relevance during high-frequency-induced long-term potentiation (LTP). Thus, whereas the A1R antagonist DPCPX (50 nM) was devoid of effects on LTP magnitude, the addition of an A2AR antagonist SCH58261 (50 nM) allowed a facilitatory effect of DPCPX on LTP to be revealed. Additionally, the activation of A2AR with CGS21680 (30 nM) decreased the potency of the A1R agonist CPA (6-60 nM) to inhibit hippocampal synaptic transmission in a manner prevented by SCH58261. These observations show that A2AR play a key role in dampening A1R during high-frequency induction of hippocampal LTP. This provides a new framework for understanding how the powerful adenosine A1R-mediated inhibition of excitatory transmission can be controlled to allow the implementation of hippocampal LTP.

Pre- and postsynaptic modulation of hippocampal inhibitory synaptic transmission by pregnenolone sulphate

Neurosteroids are important endogenous modulators of GABAA receptor-mediated neurotransmission within the CNS and play a vital role in maintaining normal healthy brain function. Research has mainly focussed on neurosteroids such as allopregnanolone and tetrahydro-deoxycorticosterone (THDOC) which are allosteric potentiators of GABAA receptors, whilst the sulphated steroids, including pregnenolone sulphate (PS), which inhibit GABAA receptor function, have been relatively neglected. Importantly, a full description of PS effects on inhibitory synaptic transmission, at concentrations that are expected to inhibit postsynaptic GABAA receptors, is lacking. Here, we address this deficit by recording inhibitory postsynaptic currents (IPSCs) from rat hippocampal neurons both in culture and in acute brain slices and explore the impact of PS at micromolar concentrations. We reveal that PS inhibits postsynaptic GABAA receptors, evident from reductions in IPSC amplitude and decay time. Concurrently, PS also causes an increase in synaptic GABA release which we discover is due to the activation of presynaptic TRPM3 receptors located close to presynaptic GABA release sites. Pharmacological blockade of TRPM3 receptors uncovers a PS-evoked reduction in IPSC frequency. This second presynaptic effect is caused by PS activation of inwardly-rectifying Kir2.3 channels on interneurons, which act to depress synaptic GABA release. Overall, we provide a comprehensive characterisation of pre- and postsynaptic modulation by PS of inhibitory synaptic transmission onto hippocampal neurons which elucidates the diverse mechanisms by which this understudied neurosteroid can modulate brain function.

Ketamine preservative benzethonium chloride potentiates hippocampal synaptic transmission and binds neurotransmitter receptors and transporters

Benzethonium chloride (BZT) is an excipient used in numerous products including (R,S)-ketamine (ketamine) drug formulations for human and veterinary use. Emerging evidence indicates BZT is pharmacologically active. BZT may therefore contribute to some of the clinical or preclinical effects observed with ketamine. In the present study, we evaluated: (i) the affinity of BZT for neurotransmitter receptors and transporters, (ii) the effects of BZT on hippocampal synaptic transmission in vitro, and (iii) plasma and brain concentrations of BZT following its intraperitoneal administration to male CD1 mice. Radioligand binding assays determined the affinity of BZT for neurotransmitter targets. Effects of BZT on field excitatory postsynaptic potentials (fEPSPs) were established via electrophysiological recordings from slices collected from male C57BL/6J mice. The binding assays revealed that BZT binds to numerous receptors (e.g., σ2 Ki = 7 nM) and transporters (e.g., dopamine transporter Ki = 545 nM). Bath application of BZT potentiated hippocampal fEPSPs in mouse hippocampal slices with an EC50 of 2.03 nM. Following intraperitoneal administration, BZT was detected in the plasma, but not in the brain of mice. These data highlight that studies measuring peripheral endpoints or directly exposing systems, in vitro, intracerebroventricularly, or intracortically, to BZT-containing formulations should account for the direct effects of BZT. Our findings also suggest that earlier data attributing pharmacological effects to ketamine may be confounded by BZT and that additional investigation into the functional impact of BZT is warranted.This article is part of the Special Issue on ‘Ketamine and its Metabolites’.

Role of the activated microglia in hippocampal synaptic plasticity alteration in in vitro inflammatory models

AbstractBackgroundMicroglia cells are primary players in neuroinflammation, which plays a key role in the pathogenesis of Alzheimer's disease (AD). Under pathological inflammatory conditions, microglia activation leads to synaptic alterations thereby influencing the progression of AD. Increased activation of microglial colony stimulating factor 1 receptor (CSF1R) can lead to pathological microglia activation in experimental models of AD (Olmos‐Alonso et al., 2016). Using electrophysiological and immunohistochemical analysis, we investigated hippocampal synaptic transmission and plasticity in experimental models of neuroinflammation. We then tested whether pharmacological manipulation of microglia activation could restore the observed neuroplasticity alterations.MethodC57BL6/J mices of both sexes aged between 30‐ and 40‐days were used. Employing electrophysiological approaches (field and whole cell patch clamp), we recorded in area CA1 in response to stimulation of Schaffer collaterals. We evaluated Long‐Term Potentiation (LTP) induced by theta‐burst stimulation measuring field Excitatory Post‐Synaptic Potentials (fEPSPs) 1 hour after tetanization. We also investigated paired‐pulse ratio (PPR) and spontaneous EPSCs (sEPSCs). Two in vitro inflammatory models were obtained applying Lipopolysaccharide (LPS) (10 μg/ml) or Aβ 1‐42 oligomers (200 nM). As a control, we tested the effect of scrambled Aβ 1‐42. To inhibit the activation of microglia we used Pexidartinib (PLX‐3397) (10μM) and Minocycline (500 nM). For immunohistochemical experiments free‐floating sections were incubated with primary antibodies overnight at 4°C followed by Alexa‐conjugated secondary antibody and DAPI for 2 hours at room temperature in the dark. VGLUT1, Iba1 and CD68 were used as synaptic marker in a triple staining to study pathological pruning and its reversal by PLX‐3397.ResultBoth inflammatory models induced LTP impairment, that was restored by application of a no‐specific blocker, Minocycline, or a specific inhibitor of activated microglia, PLX‐3397.Using whole cell patch clamp recordings, we observed that Aβ produces a reduction in glutamatergic transmission, evaluated as amplitude or frequency of sEPSCs and PPR, that is prevented by PLX‐3397. The data were confirmed using imaging studies, by which count of CA1 microglia and spine remodeling were evaluated.ConclusionOur results demonstrate a functional role of activated microglia in synaptic plasticity alteration that occurs in neuroinflammation suggesting CSF1R as new pharmacological target for AD treatment.

Acetate supplementation restores cognitive deficits caused by ARID1A haploinsufficiency in excitatory neurons.

Mutations in AT-rich interactive domain-containing protein 1A (ARID1A) cause Coffin-Siris syndrome (CSS), a rare genetic disorder that results in mild to severe intellectual disabilities. However, thebiological role of ARID1A in the brain remains unclear. In thisstudy, we report that the haploinsufficiency of ARID1A in excitatory neurons causes cognitive impairment and defects in hippocampal synaptic transmission and dendritic morphology in mice. Similarly, human embryonic stem cell-derived excitatory neurons with deleted ARID1A exhibit fewer dendritic branches and spines, and abnormal electrophysiological activity. Importantly, supplementation of acetate, an epigenetic metabolite, can ameliorate the morphological and electrophysiological deficits observed in mice with Arid1a haploinsufficiency, as well as in ARID1A-null human excitatory neurons. Mechanistically, transcriptomic and ChIP-seq analyses demonstrate that acetate supplementation can increase the levels of H3K27 acetylation at the promoters of key regulatory genes associated with neural development and synaptic transmission. Collectively, these findings support the essential roles of ARID1A in the excitatory neurons and cognition and suggest that acetate supplementation could be a potential therapeutic intervention for CSS.

Neonatal ketamine exposure impairs infrapyramidal bundle pruning and causes lasting increase in excitatory synaptic transmission in hippocampal CA3 neurons

Preclinical models demonstrate that nearly all anesthetics cause widespread neuroapoptosis in the developing brains of infant rodents and non-human primates. Anesthesia-induced developmental apoptosis is succeeded by prolonged neuropathology in the surviving neurons and lasting cognitive impairments, suggesting that anesthetics interfere with the normal developmental trajectory of the brain. However, little is known about effects of anesthetics on stereotyped axonal pruning, an important developmental algorithm that sculpts neural circuits for proper function. Here, we proposed that neonatal ketamine exposure may interfere with stereotyped axonal pruning of the infrapyramidal bundle (IPB) of the hippocampal mossy fiber system and that impaired pruning may be associated with alterations in the synaptic transmission of CA3 neurons. To test this hypothesis, we injected postnatal day 7 (PND7) mouse pups with ketamine or vehicle over 6 h and then studied them at different developmental stages corresponding to IPB pruning (PND20–40). Immunohistochemistry with synaptoporin (a marker of mossy fibers) revealed that in juvenile mice treated with ketamine at PND7, but not in vehicle-treated controls, positive IPB fibers extended farther into the stratum pyramidale of CA3 region. Furthermore, immunofluorescent double labeling for synaptoporin and PSD-95 strongly suggested that the unpruned IPB caused by neonatal ketamine exposure makes functional synapses. Importantly, patch-clamp electrophysiology for miniature excitatory postsynaptic currents (mEPSCs) in acute brain slices ex vivo revealed increased frequency and amplitudes of mEPSCs in hippocampal CA3 neurons in ketamine-treated groups when compared to vehicle controls. We conclude that neonatal ketamine exposure interferes with normal neural circuit development and that this interference leads to lasting increase in excitatory synaptic transmission in hippocampus.

Synthetic cannabinoid JWH-073 alters both acute behavior and in vivo/vitro electrophysiological responses in mice.

JWH-073 is a synthetic cannabinoid (SCB) that is illegally marketed within an "herbal blend", causing psychoactive effects more intense than those produced by Cannabis. Users report that JWH-073 causes less harmful effects than other SCBs, misrepresenting it as a "safe JWH-018 alternative", which in turn prompts its recreational use. The present study is aimed to investigate the in vivo pharmacological activity on physiological and neurobehavioral parameters in male CD-1 mice after acute 1 mg/kg JWH-073 administration. To this aim we investigate its effect on sensorimotor (visual, acoustic, and tactile), motor (spontaneous motor activity and catalepsy), and memory functions (novel object recognition; NOR) in mice coupling behavioral and EEG data. Moreover, to clarify how memory function is affected by JWH-073, we performed in vitro electrophysiological studies in hippocampal preparations using a Long-Term Potentiation (LTP) stimulation paradigm. We demonstrated that acute administration of JWH-073 transiently decreased motor activity for up to 25 min and visual sensorimotor responses for up to 105 min, with the highest effects at 25 min (~48 and ~38%, respectively), while the memory function was altered up to 24 h (~33%) in treated-mice as compared to the vehicle. EEG in the somatosensory cortex showed a maximal decrease of α (~23%) and γ (~26%) bands at 15 min, β (~26%) band at 25 min, a maximal increase of θ (~14%) band at 25 min and δ (~35%) band at 2 h, and a significant decrease of θ (~18%), α (~26%), and β (~10%) bands during 24 h. On the other hand, EEG in the hippocampus showed a significant decrease of all bands from 10 min to 2 h, with the maximal effect at 30 min for θ (~34%) and γ (~26%) bands and 2 h for α (~36%), β (~29%), and δ (~15%) bands. Notably, the δ band significant increase both at 5 min (~12%) and 24 h (~19%). Moreover, in vitro results support cognitive function impairment (~60% of decrease) by interfering with hippocampal synaptic transmission and LTP generation. Our results suggest that JWH-073 deeply alters brain electrical responsiveness with minor behavioral symptoms. Thus, it poses a subtle threat to consumers who mistakenly consider it safer than other SCBs.

The role of AMPA and NMDA receptors in mitragynine effects on hippocampal synaptic plasticity

Mitragynine, an indole alkaloid from the plant Mitragyna speciosa (Kratom), has been reported to modify hippocampal synaptic transmission. However, the role of glutamatergic neurotransmission modulating synaptic plasticity in mitragynine-induced synaptic changes is still unknown. Here, we determined the role of AMPA- and NMDA glutamate receptors in mitragynine-induced synaptic plasticity in the hippocampus. Male Sprague Dawley rats received either vehicle or mitragynine (10 mg/kg), with or without the AMPA receptor antagonist, NBQX (3 mg/kg), or the NMDA receptor antagonist, MK-801 (0.2 mg/kg). Field excitatory postsynaptic potentials (fEPSP) during baseline, paired-pulse facilitation (PPF) and long-term potentiation (LTP) were recorded in-vivo in the hippocampal CA1 area of anaesthetised rats. Basal synaptic transmission and LTP were significantly impaired after mitragynine, NBQX, and MK-801 alone, without an effect on PPF. Combined effects suggest a weak functional AMPA- as well as NMDA receptor antagonist action of mitragynine.

Cognitive and behavioral effects of the anti-epileptic drug cenobamate (YKP3089) and underlying synaptic and cellular mechanisms

Antiseizure medication is the mainstay of treatment for seizures, and adjunctive therapy is widely used to achieve adequate seizure control in patients with epilepsy who fail to respond to the first monotherapy. The newly developed antiepileptic drug cenobamate (YKP3089) as an adjunctive therapy improved seizure control in patients with uncontrolled focal seizures. Cenobamate is thought to reduce neuronal excitability through action on multiple targets, including GABA A receptors (GABAARs) and voltage-gated sodium channels. However, its effects on brain function and synaptic plasticity are unclear. Here, we explored the behavioral, synaptic, and cellular actions of cenobamate. Cenobamate influenced novel object recognition, object location memory, and Morris water maze performance in mice. Cenobamate enhanced inhibitory postsynaptic potentials by prolonging inhibitory postsynaptic current (IPSC) decay without affecting presynaptic GABA release or the peak amplitude of IPSCs. In addition, cenobamate suppressed hippocampal excitatory synaptic transmission by reducing the excitability of Schaffer collaterals and interfered with the induction of long-term potentiation. A reduction in neuronal excitability induced by cenobamate was associated with an elevation of action potential (AP) threshold, and which progressively increased in later APs during repetitive firing, indicating the activity-dependent modulation of neuronal sodium currents. Cenobamate suppressed neuronal excitability under the condition that GABAergic neurotransmission is excitatory, and administration of cenobamate rapidly enhanced the phosphorylation of eukaryotic elongation factor 2 in the hippocampus of adult and neonatal mice. Collectively, these results suggest that the combined action of cenobamate on sodium currents and GABAAR-mediated synapse responses results in reduced excitability in neurons.