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Synaptopodin, an actin-associated protein found in a subset of dendritic spines in telencephalic neurons, has been described to influence both functional and morphological plasticity under various plasticity paradigms. Synaptopodin is necessary and sufficient for the formation of the spine apparatus, stacks of smooth endoplasmic reticulum cisternae. The spine apparatus is a calcium store that locally regulates calcium dynamics in response to different patterns of activity and is also thought to be a site for local protein synthesis. Synaptopodin is present in ~30% of telencephalic large dendritic spines in vivo and in vitro highlighting the heterogeneous microanatomy and molecular architecture of dendritic spines, an important but not well understood aspect of neuroplasticity. In recent years, it has become increasingly clear that synaptopodin is a formidable regulator of multiple mechanisms essential for learning and memory. In fact, synaptopodin appears to be the decisive factor that determines whether plasticity can occur, acting as a key regulator for synaptic changes. In this review, we summarize the current understanding of synaptopodin’s role in various forms of Hebbian synaptic plasticity.

Information storage and transfer in the brain require a high computational power. Neuronal network display various local or global mechanisms to allow information storage and transfer in the brain. From synaptic to intrinsic plasticity, the rules of input–output function modulation have been well characterized in neurons. In the past years, astrocytes have been suggested to increase the computational power of the brain and we are only just starting to uncover their role in information processing. Astrocytes maintain a close bidirectional communication with neurons to modify neuronal network excitability, transmission, axonal conduction, and plasticity through various mechanisms including the release of gliotransmitters or local ion homeostasis. Astrocytes have been significantly studied in the context of long-term or short-term synaptic plasticity, but this is not the only mechanism involved in memory formation. Plasticity of intrinsic neuronal excitability also participates in memory storage through regulation of voltage-gated ion channels or axonal morphological changes. Yet, the contribution of astrocytes to these other forms of non-synaptic plasticity remains to be investigated. In this review, we summarized the recent advances on the role of astrocytes in different forms of plasticity and discuss new directions and ideas to be explored regarding astrocytes-neuronal communication and regulation of plasticity.

Abstract Synaptopodin (SP), an actin-associated protein found in telencephalic neurons, affects activity-dependant synaptic plasticity and dynamic changes of dendritic spines. While being required for long-term depression (LTD) mediated by metabotropic glutamate receptor (mGluR-LTD), little is known about its role in other forms of LTD induced by low frequency stimulation (LFS-LTD) or spike-timing dependent plasticity (STDP). Using electrophysiology in ex vivo hippocampal slices from SP-deficient mice (SPKO), we show that absence of SP is associated with a deficit of LTD at Sc-CA1 synapses induced by LFS-LTD and STDP. As LTD is known to require AMPA- receptors internalization and IP3-receptors calcium signaling, we tested by western blotting and immunochemistry if there were changes in their expression which we found to be reduced. While we were not able to induce LTD, long-term potentiation (LTP), albeit diminished in SPKO, can be recovered by using a stronger stimulation protocol. In SPKO we found no differences in NMDAR, which are the primary site of calcium signalling to induce LTP. Our study shows, for the first time, the key role of the requirement of SP to allow induction of activity-dependant LTD at Sc-CA1 synapses.

Spike-timing-dependent plasticity (STDP) is considered as a primary mechanism underlying formation of new memories during learning. Despite the growing interest in activity-dependent plasticity, it is still unclear whether synaptic plasticity rules inferred from in vitro experiments are correct in physiological conditions. The abnormally high calcium concentration used in in vitro studies of STDP suggests that in vivo plasticity rules may differ significantly from in vitro experiments, especially since STDP depends strongly on calcium for induction. We therefore studied here the influence of extracellular calcium on synaptic plasticity. Using a combination of experimental (patch-clamp recording and Ca2+ imaging at CA3-CA1 synapses) and theoretical approaches, we show here that the classic STDP rule in which pairs of single pre- and postsynaptic action potentials induce synaptic modifications is not valid in the physiological Ca2+ range. Rather, we found that these pairs of single stimuli are unable to induce any synaptic modification in 1.3 and 1.5 mM calcium and lead to depression in 1.8 mM. Plasticity can only be recovered when bursts of postsynaptic spikes are used, or when neurons fire at sufficiently high frequency. In conclusion, the STDP rule is profoundly altered in physiological Ca2+, but specific activity regimes restore a classical STDP profile.

Abstract Mammary tumors in dogs hold great potential as naturally occurring breast cancer models in translational oncology, as they share the same environmental risk factors, key histological features, hormone receptor expression patterns, prognostic factors, and genetic characteristics as their human counterparts. We aimed to develop in vitro tools that allow functional analysis of canine mammary tumors (CMT), as we have a poor understanding of the underlying biology that drives the growth of these heterogeneous tumors. We established the long-term culture of 24 organoid lines from 16 dogs, including organoids derived from normal mammary epithelium or benign lesions. CMT organoids recapitulated key morphological and immunohistological features of the primary tissue from which they were derived, including hormone receptor status. Furthermore, genetic characteristics (driver gene mutations, DNA copy number variations, and single-nucleotide variants) were conserved within tumor-organoid pairs. We show how CMT organoids are a suitable model for in vitro drug assays and can be used to investigate whether specific mutations predict therapy outcomes. Specifically, certain CMT subtypes, such as PIK3CA mutated, estrogen receptor-positive simple carcinomas, can be valuable in setting up a preclinical model highly relevant to human breast cancer research. In addition, we could genetically modify the CMT organoids and use them to perform pooled CRISPR/Cas9 screening, where library representation was accurately maintained. In summary, we present a robust 3D in vitro preclinical model that can be used in translational research, where organoids from normal, benign as well as malignant mammary tissues can be propagated from the same animal to study tumorigenesis.

Synaptopodin, an actin-associated protein found in a subset of dendritic spines in telencephalic neurons, has been described to influence both functional and morphological plasticity under various plasticity paradigms. Synaptopodin is necessary and sufficient for the formation of the spine apparatus, stacks of smooth endoplasmic reticulum cisternae. The spine apparatus is a calcium store that locally regulates calcium dynamics in response to different patterns of activity and is also thought to be a site for local protein synthesis. Synaptopodin is present in ~30% of telencephalic large dendritic spines in vivo and in vitro highlighting the heterogeneous microanatomy and molecular architecture of dendritic spines, an important but not well understood aspect of neuroplasticity. In recent years, it has become increasingly clear that synaptopodin is a formidable regulator of multiple mechanisms essential for learning and memory. In fact, synaptopodin appears to be the decisive factor that determines whether plasticity can occur, acting as a key regulator for synaptic changes. In this review, we summarize the current understanding of synaptopodin's role in various forms of Hebbian synaptic plasticity. [ABSTRACT FROM AUTHOR]

Information storage and transfer in the brain require a high computational power. Neuronal network display various local or global mechanisms to allow information storage and transfer in the brain. From synaptic to intrinsic plasticity, the rules of input-output function modulation have been well characterized in neurons. In the past years, astrocytes have been suggested to increase the computational power of the brain and we are only just starting to uncover their role in information processing. Astrocytes maintain a close bidirectional communication with neurons to modify neuronal network excitability, transmission, axonal conduction, and plasticity through various mechanisms including the release of gliotransmitters or local ion homeostasis. Astrocytes have been significantly studied in the context of long-term or short-term synaptic plasticity, but this is not the only mechanism involved in memory formation. Plasticity of intrinsic neuronal excitability also participates in memory storage through regulation of voltage-gated ion channels or axonal morphological changes. Yet, the contribution of astrocytes to these other forms of non-synaptic plasticity remains to be investigated. In this review, we summarized the recent advances on the role of astrocytes in different forms of plasticity and discuss new directions and ideas to be explored regarding astrocytes-neuronal communication and regulation of plasticity. [ABSTRACT FROM AUTHOR]

ABSTRACT Wild-type canine distemper virus (CDV) is an important pathogen of dogs as well as wildlife that can infect immune and epithelial cells through two known receptors: the signaling lymphocytic activation molecule (SLAM) and nectin-4, respectively. Conversely, the ferret and egg-adapted CDV-Onderstepoort strain (CDV-OP) is employed as an effective vaccine for dogs. CDV-OP also exhibits promising oncolytic properties, such as its abilities to infect and kill multiple cancer cells in vitro. Interestingly, several cancer cells do not express SLAM or nectin-4, suggesting the presence of a yet unknown entry factor for CDV-OP. By conducting a genome-wide CRISPR/Cas9 knockout (KO) screen in CDV-OP-susceptible canine mammary carcinoma P114 cells, which neither express SLAM nor nectin-4, we identified low-density lipoprotein receptor-related protein 6 (LRP6) as a host factor that promotes CDV-OP infectivity. Whereas the genetic ablation of LRP6 rendered cells resistant to infection, ectopic expression in resistant LRP6KO cells restored susceptibility. Furthermore, multiple functional studies revealed that (i) the overexpression of LRP6 leads to increased cell-cell fusion, (ii) a soluble construct of the viral receptor-binding protein (solHOP) interacts with a soluble form of LRP6 (solLRP6), (iii) an H-OP point mutant that prevents interaction with solLRP6 abrogates cell entry in multiple cell lines once transferred into recombinant viral particles, and (iv) vesicular stomatitis virus (VSV) pseudotyped with CDV-OP envelope glycoproteins loses its infectivity in LRP6KO cells. Collectively, our study identified LRP6 as the long sought-after cell entry receptor of CDV-OP in multiple cell lines, which set the molecular bases to refine our understanding of viral-cell adaptation and to further investigate its oncolytic properties. IMPORTANCE Oncolytic viruses (OV) have gathered increasing interest in recent years as an alternative option to treat cancers. The Onderstepoort strain of canine distemper virus (CDV-OP), an enveloped RNA virus belonging to the genus Morbillivirus, is employed as a safe and efficient vaccine for dogs against distemper disease. Importantly, although CDV-OP can infect and kill multiple cancer cell lines, the basic mechanisms of entry remain to be elucidated, as most of those transformed cells do not express natural receptors (i.e., SLAM and nectin-4). In this study, using a genome-wide CRISPR/Cas9 knockout screen, we describe the discovery of LRP6 as a novel functional entry receptor for CDV-OP in various cancer cell lines and thereby uncover a basic mechanism of cell culture adaptation. Since LRP6 is upregulated in various cancer types, our data provide important insights in order to further investigate the oncolytic properties of CDV-OP.

The integrated stress response (ISR), a pivotal protein homeostasis network, plays a critical role in the formation of long-term memory (LTM). The precise mechanism by which the ISR controls LTM is not well understood. Here, we report insights into how the ISR modulates the mnemonic process by using targeted deletion of the activating transcription factor 4 (ATF4), a key downstream effector of the ISR, in various neuronal and non-neuronal cell types. We found that the removal of ATF4 from forebrain excitatory neurons (but not from inhibitory neurons, cholinergic neurons, or astrocytes) enhances LTM formation. Furthermore, the deletion of ATF4 in excitatory neurons lowers the threshold for the induction of long-term potentiation, a cellular model for LTM. Transcriptomic and proteomic analyses revealed that ATF4 deletion in excitatory neurons leads to upregulation of components of oxidative phosphorylation pathways, which are critical for ATP production. Thus, we conclude that ATF4 functions as a memory repressor selectively within excitatory neurons. [ABSTRACT FROM AUTHOR]

Since its discovery, spike timing-dependent synaptic plasticity (STDP) has been thought to be a primary mechanism underlying the brain’s ability to learn and to form new memories. However, despite the enormous interest in both the experimental and theoretical neuroscience communities in activity-dependent plasticity, it is still unclear whether plasticity rules inferred from in vitro experiments apply to in vivo conditions. Among the multiple reasons why plasticity rules in vivo might differ significantly from in vitro studies is that extracellular calcium concentration use in most studies is higher than concentrations estimated in vivo. STDP, like many forms of long-term synaptic plasticity, strongly depends on intracellular calcium influx for its induction. Here, we discuss the importance of considering physiological levels of extracellular calcium concentration to study functional plasticity.

The title molecular salts, C16H16NO+·PF6−, (I), and C21H21N2+·PF6−, (II), are pyridine derivatives. In compound (I), the cation comprises a methyl N-substituted pyridine ring and a methoxy-substituted benzene ring connected by a C=C double bond. The F atoms of the PF6− anion are disordered over two sets of sites with refined occupancy factors of 0.614 (7):0.386 (7). In compound (II), the cation comprises a pyridine ring attached to unsubstituted phenyl ring and a dimethylaniline ring, which are connected by a C=C double bond. The anion is PF6−. In both salts, the cation adopts an E configuration with respect to the C=C bond. The pyridine ring makes a dihedral angle of 9.86 (12)° with the methoxy-substituted benzene ring in compound (I) and 11.2 (3)° with the dimethylamine-substituted benzene ring in compound (II). In compound (I), the crystal packing is stabilized by weak C—H...F intermolecular interactions which result in R43(14) ring motifs, forming molecular sheets running parallel to (\overline{1}03). These are further stabilized by weak P—F...π interactions. In compound (II), the crystal packing is stabilized by C—H...F interactions, which result in R66(40) ring motifs, forming molecular sheets running parallel to (101) and these are further connected by π–π interactions.

The title molecular salts, (C18H23N2)2[CdI4], (I), and C18H23N2+·C7H7O4S−·H2O, (II), are stilbazole, or 4-styrylpyridine, derivatives. The cation, (E)-4-[4-(diethylamino)styryl]-1-methylpyridin-1-ium, has a methyl group attached to pyridine ring and a diethyl amine group attached to the benzene ring. The asymmetric unit of salt (I), comprises one cationic molecule and half a CdI4 dianion. The Cd atom is situated on a twofold rotation axis and has a slightly distorted tetrahedral coordination sphere. In (II), the anion consists of a 4-methoxybenzenesulfonate and it crystallizes as a monohydrate. In both salts, the cations adopt an E configuration with respect to the C=C bond and the pyridine and benzene rings are inclined to each other by 10.7 (4)° in (I) and 4.6 (2)° in (II). In the crystals of both salts, the packing is consolidated by offset π–π stacking interactions involving the pyridinium and benzene rings, with centroid–centroid distances of 3.627 (4) Å in (I) and 3.614 (3) Å in (II). In the crystal of (II), a pair of 4-methoxybenzenesulfonate anions are bridged by Owater—H...Osulfonate hydrogen bonds, forming loops with an R24(8) motif. These four-membered units are then linked to the cations by a number of C—H...O hydrogen bonds, forming slabs lying parallel to the ab plane.

Background: Paroxysmal Permeability Disorders (PPDs) are pathological conditions caused by periodic short lasting increase of endothelial permeability, in the absence of inflammatory, degenerative, ischemic vascular injury. PPDs include primary angioedema, idiopathic systemic capillary leak syndrome and some rare forms of localized retroperitoneal-mediastinal edema.Aim: to validate a microfluidic device to study endothelial permeability in flow conditions.Materials and Methods: we designed a microchannel network (the smallest channel is 30μm square section). Human Umbilical Vein Endothelial Cells (HUVECs) were cultured under constant shear stress in the networks. Endothelial permeability assessment was based on interaction of biotinylated fibronectin used as a matrix for HUVECs and FITC-conjugated avidin. The increase in endothelial permeability was identified as changes in fluorescence intensity detected by confocal fluorescent microscopy.Results: The microchannels were constantly perfused with a steady flow of culture medium, ensuring a physiologically relevant level of shear stress at the wall of ~0.2 Pa. Our preliminary results demonstrated that circulation of culture medium or plasma from healthy volunteers was associated with low fluorescence of fibronectin matrix. When bradykinin diluted in culture medium was perfused, an increase in average fluorescence was detected.Conclusion: Our microvasculature model is suitable to study endothelial functions in physiological flow conditions and in the presence of factors like bradykinin known as mediator of several PPDs. Therefore, it can be a promising tool to better understand the mechanisms underlying disorders of endothelial permeability.