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The aim of this study is to evaluate the possibility of achieving more effective and prolonged sedation in pigs by the oral administration of increased doses of azaperone and to evaluate its safety. This was performed through a prospective randomised and double blinded study. A total of 32 weaned piglets were divided into 4 groups (8 in each group). Group A was given 1 ml of saline orally and served as the control group. Group B received azaperone orally at a dose of 4 mg/kg b.w. Group C received azaperone orally at a dose of 8 mg/kg b.w. Group D was given azaperone orally at a dose of 12 mg/kg b.w. The response to the defined stimulus, movement level, degree of salivation, body temperature, respiratory frequency, blood plasma azaperone concentration and biochemical variables were included in the trial. We found that by increasing the dose of the orally administered azaperone, the onset of the sedation is faster, the end of the sedation starts later and the sedation time is longer. However, the use of higher doses of oral azaperone is not suitable for piglets because the doses negatively affect the respiratory rate, body temperature, some biochemical parameters and cause the immobility of the piglets.

N-methyl-D-aspartate receptors (NMDARs) play a significant role in developing several central nervous system (CNS) disorders. Currently, memantine, used for treating Alzheimer's disease, and ketamine, known for its anesthetic and antidepressant properties, are two clinically used NMDAR open-channel blockers. However, despite extensive research into NMDAR modulators, many have shown either harmful side effects or inadequate effectiveness. For instance, dizocilpine (MK-801) is recognized for its powerful psychomimetic effects due to its high-affinity and nearly irreversible inhibition of the GluN1/GluN2 NMDAR subtypes. Unlike ketamine, memantine and MK-801 also act through a unique, low-affinity "membrane-to-channel inhibition" (MCI). We aimed to develop an open-channel blocker based on MK-801 with distinct inhibitory characteristics from memantine and MK-801. Our novel compound, K2060, demonstrated effective voltage-dependent inhibition in the micromolar range at key NMDAR subtypes, GluN1/GluN2A and GluN1/GluN2B, even in the presence of Mg 2+ . K2060 showed reversible inhibitory dynamics and a partially trapping open-channel blocking mechanism with a significantly stronger MCI than memantine. Using hippocampal slices, 30 µM K2060 inhibited excitatory postsynaptic currents in CA1 hippocampal neurons by ∼51 %, outperforming 30 µM memantine (∼21 % inhibition). K2060 exhibited No Observed Adverse Effect Level (NOAEL) of 15 mg/kg upon intraperitoneal administration in mice. Administering K2060 at a 10 mg/kg dosage resulted in brain concentrations of approximately 2 µM, with peak concentrations (Tmax) achieved within 15 minutes. Finally, applying K2060 with trimedoxime and atropine in mice exposed to tabun improved treatment outcomes. These results underscore K2060's potential as a therapeutic agent for CNS disorders linked to NMDAR dysfunction., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in the paper "Potent and reversible open-channel blocker of NMDA receptor derived from dizocilpine with enhanced membrane-to-channel inhibition" by Misiachna et al., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Tacrine (THA), a long withdrawn drug, is still a popular scaffold used in medicinal chemistry, mainly for its good reactivity and multi-targeted effect. However, THA-associated hepatotoxicity is still an issue and must be considered in drug discovery based on the THA scaffold. Following our previously identified hit compound 7-phenoxytacrine (7-PhO-THA), we systematically explored the chemical space with 30 novel derivatives, with a focus on low hepatotoxicity, anticholinesterase action, and antagonism at the GluN1/GluN2B subtype of the NMDA receptor. Applying the down-selection process based on in vitro and in vivo pharmacokinetic data, two candidates, I-52 and II-52, selective GluN1/GluN2B inhibitors thanks to the interaction with the ifenprodil-binding site, have entered in vivo pharmacodynamic studies. Finally, compound I-52, showing only minor affinity to AChE, was identified as a lead candidate with favorable behavioral and neuroprotective effects using open-field and prepulse inhibition tests, along with scopolamine-based behavioral and NMDA-induced hippocampal lesion models. Our data show that compound I-52 exhibits low toxicity often associated with NMDA receptor ligands, and low hepatotoxicity, often related to THA-based compounds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

NMDA receptors (NMDARs) are ionotropic glutamate receptors that play a key role in excitatory neurotransmission. The number and subtype of surface NMDARs are regulated at several levels, including their externalization, internalization, and lateral diffusion between the synaptic and extrasynaptic regions. Here, we used novel anti-GFP (green fluorescent protein) nanobodies conjugated to either the smallest commercially available quantum dot 525 (QD525) or the several nanometer larger (and thus brighter) QD605 (referred to as nanoGFP-QD525 and nanoGFP-QD605, respectively). Targeting the yellow fluorescent protein-tagged GluN1 subunit in rat hippocampal neurons, we compared these two probes to a previously established larger probe, a rabbit anti-GFP IgG together with a secondary IgG conjugated to QD605 (referred to as antiGFP-QD605). The nanoGFP-based probes allowed faster lateral diffusion of the NMDARs, with several-fold increased median values of the diffusion coefficient ( D ). Using thresholded tdTomato-Homer1c signals to mark synaptic regions, we found that the nanoprobe-based D values sharply increased at distances over 100 nm from the synaptic edge, while D values for antiGFP-QD605 probe remained unchanged up to a 400 nm distance. Using the nanoGFP-QD605 probe in hippocampal neurons expressing the GFP-GluN2A, GFP-GluN2B, or GFP-GluN3A subunits, we detected subunit-dependent differences in the synaptic localization of NMDARs, D value, synaptic residence time, and synaptic-extrasynaptic exchange rate. Finally, we confirmed the applicability of the nanoGFP-QD605 probe to study differences in the distribution of synaptic NMDARs by comparing to data obtained with nanoGFPs conjugated to organic fluorophores, using universal point accumulation imaging in nanoscale topography and direct stochastic optical reconstruction microscopy. SIGNIFICANCE STATEMENT Our study systematically compared the localization and mobility of surface NMDARs containing GFP-GluN2A, GFP-GluN2B, or GFP-GluN3A subunits expressed in rodent hippocampal neurons, using anti-green fluorescent protein (GFP) nanobodies conjugated to the quantum dot 605 (nanoGFP-QD605), as well as nanoGFP probes conjugated with small organic fluorophores. Our comprehensive analysis showed that the method used to delineate the synaptic region plays an important role in the study of synaptic and extrasynaptic pools of NMDARs. In addition, we showed that the nanoGFP-QD605 probe has optimal parameters for studying the mobility of NMDARs because of its high localization accuracy comparable to direct stochastic optical reconstruction microscopy and longer scan time compared with universal point accumulation imaging in nanoscale topography. The developed approaches are readily applicable to the study of any GFP-labeled membrane receptors expressed in mammalian neurons., (Copyright © 2023 Kortus et al.)

N-methyl-D-aspartate receptors (NMDARs) play an essential role in excitatory neurotransmission in the mammalian brain, and their physiological importance is underscored by the large number of pathogenic mutations that have been identified in the receptor's GluN subunits and associated with a wide range of diseases and disorders. Here, we characterized the functional and pharmacological effects of the pathogenic N650K variant in the GluN1 subunit, which is associated with developmental delay and seizures. Our microscopy experiments showed that when expressed in HEK293 cells (from ATCC®), the GluN1-N650K subunit increases the surface expression of both GluN1/GluN2A and GluN1/GluN2B receptors, but not GluN1/GluN3A receptors, consistent with increased surface expression of the GluN1-N650K subunit expressed in hippocampal neurons (from embryonic day 18 of Wistar rats of both sexes). Using electrophysiology, we found that the GluN1-N650K variant increases the potency of GluN1/GluN2A receptors to both glutamate and glycine but decreases the receptor's conductance and open probability. In addition, the GluN1-N650K subunit does not form functional GluN1/GluN2B receptors but does form fully functional GluN1/GluN3A receptors. Moreover, in the presence of extracellular Mg 2+ , GluN1-N650K/GluN2A receptors have a similar and increased response to ketamine and memantine, respectively, while the effect of both drugs had markedly slower onset and offset compared to wild-type GluN1/GluN2A receptors. Finally, we found that expressing the GluN1-N650K subunit in hippocampal neurons reduces excitotoxicity, and memantine shows promising neuroprotective effects in neurons expressing either wild-type GluN1 or the GluN1-N650K subunit. This study provides the functional and pharmacological characterization of NMDARs containing the GluN1-N650K variant., Competing Interests: Declaration of competing interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The experimental data were fitted with Equation (1) or Equation (2); the EC(50) values (in μM), Hill coefficients (h), and numbers of cells analyzed (n) are shown; the cells were recorded at a membrane potential of −60 mV; p < 0.05 for all mutant NMDARs vs. the corresponding wild-type NMDAR (Student's t-test); the data for the wild-type receptors were published previously (Kolcheva et al., 2021)., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The aim of this study is to evaluate the possibility of achieving more effective and prolonged sedation in pigs by the oral administration of increased doses of azaperone and to evaluate its safety. This was performed through a prospective randomised and double blinded study. A total of 32 weaned piglets were divided into 4 groups (8 in each group). Group A was given 1 ml of saline orally and served as the control group. Group B received azaperone orally at a dose of 4 mg/kg b.w. Group C received azaperone orally at a dose of 8 mg/kg b.w. Group D was given azaperone orally at a dose of 12 mg/kg b.w. The response to the defined stimulus, movement level, degree of salivation, body temperature, respiratory frequency, blood plasma azaperone concentration and biochemical variables were included in the trial. We found that by increasing the dose of the orally administered azaperone, the onset of the sedation is faster, the end of the sedation starts later and the sedation time is longer. However, the use of higher doses of oral azaperone is not suitable for piglets because the doses negatively affect the respiratory rate, body temperature, some biochemical parameters and cause the immobility of the piglets., Competing Interests: The authors declare no conflict of interest., (Copyright: © 2022 Svoboda et al.)

N -methyl-D-aspartate receptors (NMDARs) belong to a family of ionotropic glutamate receptors that play essential roles in excitatory neurotransmission and synaptic plasticity in the mammalian central nervous system (CNS). Functional NMDARs consist of heterotetramers comprised of GluN1, GluN2A-D, and/or GluN3A-B subunits, each of which contains four membrane domains (M1 through M4), an intracellular C-terminal domain, a large extracellular N-terminal domain composed of the amino-terminal domain and the S1 segment of the ligand-binding domain (LBD), and an extracellular loop between M3 and M4, which contains the S2 segment of the LBD. Both the number and type of NMDARs expressed at the cell surface are regulated at several levels, including their translation and posttranslational maturation in the endoplasmic reticulum (ER), intracellular trafficking via the Golgi apparatus, lateral diffusion in the plasma membrane, and internalization and degradation. This review focuses on the roles played by the extracellular regions of GluN subunits in ER processing. Specifically, we discuss the presence of ER retention signals, the integrity of the LBD, and critical N -glycosylated sites and disulfide bridges within the NMDAR subunits, each of these steps must pass quality control in the ER in order to ensure that only correctly assembled NMDARs are released from the ER for subsequent processing and trafficking to the surface. Finally, we discuss the effect of pathogenic missense mutations within the extracellular domains of GluN subunits with respect to ER processing of NMDARs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Horak, Barackova, Langore, Netolicky, Rivas-Ramirez and Rehakova.)

Although numerous pathogenic mutations have been identified in various subunits of N-methyl-D-aspartate receptors (NMDARs), ionotropic glutamate receptors that are central to glutamatergic neurotransmission, the functional effects of these mutations are often unknown. Here, we combined in silico modelling with microscopy, biochemistry, and electrophysiology in cultured HEK293 cells and hippocampal neurons to examine how the pathogenic missense mutation S688Y in the GluN1 NMDAR subunit affects receptor function and trafficking. We found that the S688Y mutation significantly increases the EC 50 of both glycine and D-serine in GluN1/GluN2A and GluN1/GluN2B receptors, and significantly slows desensitisation of GluN1/GluN3A receptors. Moreover, the S688Y mutation reduces the surface expression of GluN3A-containing NMDARs in cultured hippocampal neurons, but does not affect the trafficking of GluN2-containing receptors. Finally, we found that the S688Y mutation reduces Ca 2+ influx through NMDARs and reduces NMDA-induced excitotoxicity in cultured hippocampal neurons. These findings provide key insights into the molecular mechanisms that underlie the regulation of NMDAR subtypes containing pathogenic mutations.

The aim of this study is to evaluate the results of dentigerous cysts enucleation with preservation of cyst-associated teeth as a single procedure in a group of children and preadolescents. A retrospective case series of seven patients with 15 unerupted developing teeth associated with or adjacent to dentigerous cysts was conducted. Patients were treated using the uniform surgical protocol (cystectomy, preservation of cyst-associated teeth, without use of orthodontic traction, primary wound closure). The metric analysis of the cysts and unerupted teeth on CBCT and an evaluation of the state of involved teeth after the treatment was performed. The mean age of patients was 9.0 ± 2.1 years (range 5-12 years, median = 9 years). All teeth erupted spontaneously. No serious postoperative complications or recurrences of lesions were detected. The mean follow-up was 53.1 ± 15.7 months (range 25-71 months, median = 52 months). The cystectomy of dentigerous cysts with preservation of associated teeth may be considered an alternative to marsupialization in children and preadolescents., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2020 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.)

The aim of this study was to investigate pre-injury factors, causes of dental injuries and healing complications after traumatic injuries to permanent teeth. The analysed sample comprised 889 permanent teeth of 384 patients, who were treated in the Dentistry Department in Faculty Hospital in Pilsen. Enamel-dentin fractures [233 teeth (26.2%)] and lateral luxations [207 teeth (23.3%)] were the most frequent injuries. The age of the patients at the time of injury varied between 7 and 65 years. Predominantly, children were affected [587 injured teeth (66.0%)]. The most frequent causes of injuries in patients older than 11 years were various sport activities, predominantly bicycling. Pulp necrosis was observed in 239 teeth (26.9%). It was the most frequent post-traumatic complication in all types of dental traumas. Teeth with a completed root formation demonstrated a higher prevalence of pulp necrosis than teeth with an incomplete root formation in all types of luxation injuries. External root resorption was observed in 144 teeth. The rate of inflammatory resorption differed between the various types of luxation injuries (extrusive luxation 5.6%, lateral luxation 11.6%, intrusive luxation 33.3%). Following avulsion and replantation, active inflammatory resorptions were diagnosed in 13 (26.5%) of 49 replanted teeth and ankylosis/replacement resorptions were observed in 21 (42.9%) of 49 replanted teeth. After avulsion, primarily, immature teeth were affected by these complications. Within the observation period of 5 years, 39 teeth (4.4%) had to be removed (16 teeth with root fractures, 19 avulsed and replanted teeth, 3 luxated teeth, 1 tooth with crown-root fracture)., (© 2010 John Wiley & Sons A/S.)