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Striatal cholinergic interneurons (CINs) use acetylcholine (ACh) and glutamate (Glut) to regulate the striatal network since they express vesicular transporters for ACh (VAChT) and Glut (VGLUT3). However, whether ACh and Glut are released simultaneously and/or independently from cholinergic varicosities is an open question. The answer to that question requires the multichannel detection of vesicular transporters at the level of single synaptic vesicle (SV). Here, we used super-resolution STimulated Emission Depletion microscopy (STED) to characterize and quantify the distribution of VAChT and VGLUT3 in CINs SVs. Nearest-neighbor distances analysis between VAChT and VGLUT3- immunofluorescent spots revealed that 34% of CINs SVs contain both VAChT and VGLUT3. In addition, 40% of SVs expressed only VAChT while 26% of SVs contain only VGLUT3. These results suggest that SVs from CINs have the potential to store simultaneously or independently ACh and/or Glut. Overall, these morphological findings support the notion that CINs varicosities can signal with either ACh or Glut or both with an unexpected level of complexity. [ABSTRACT FROM AUTHOR]

The notion of resonance matrix elements is discussed. The analogue of the Lellouch-Lüscher formula for the pion photo(electro)production amplitude in the Δ(1232) resonance region is given. A prescription for the pole extraction of the ΔNγ* transition form factors is provided. Theoretical prospects on the lattice calculation of B → K*(892)l+l− form factors are briefly summarized.

Exposure to stressors has profound effects on sleep that have been linked to serotonin (5-HT) neurons of the dorsal raphe nucleus (DR). However, the DR also comprises glutamatergic neurons expressing vesicular glutamate transporter type 3 (DRVGLUT3), leading us to examine their role. Cell-type-specific tracing revealed that DRVGLUT3 neurons project to brain areas regulating arousal and stress. We found that chemogenetic activation of DRVGLUT3 neurons mimics stress-induced sleep perturbations. Furthermore, deleting VGLUT3 in the DR attenuated stress-induced sleep perturbations, especially after social defeat stress. In the DR, VGLUT3 is found in subsets of 5-HT and non-5-HT neurons. We observed that both populations are activated by acute stress, including those projecting to the ventral tegmental area. However, deleting VGLUT3 in 5-HT neurons minimally affected sleep regulation. These findings suggest that VGLUT3 expression in the DR drives stress-induced sleep perturbations, possibly involving non-5-HT DRVGLUT3 neurons. [Display omitted] • VGLUT3 expression identifies subsets of 5-HT and non-5-HT DR neurons activated by acute stress • Selective activation of VGLUT3-expressing neurons in the DR modulates wake and REMS • Reduced VGLUT3 expression in the DR attenuates stress-induced sleep perturbations • VGLUT3 expression in 5-HT neurons exerts minimal influence on sleep Psychological stressors can induce maladaptive disruptions in sleep patterns. Henderson et al. show that neurons expressing VGLUT3 in the dorsal raphe nucleus (DR) are involved in the stress response and contribute to the circuit mechanisms mediating the effects of acute stress on sleep. [ABSTRACT FROM AUTHOR]

Several subtypes of modulatory neurons co‐express vesicular glutamate transporters (VGLUTs) in addition to their cognate vesicular transporters. These neurons are believed to establish new forms of neuronal communication. The atypical VGLUT3 is of particular interest since in the striatum this subtype is found in tonically active cholinergic interneurons (TANs) and in a subset of 5‐HT fibers. The striatum plays a major role in psychomotor effects induced by amphetamine. Whether and how VGLUT3‐operated glutamate/ACh or glutamate/5HT co‐transmissions modulates psychostimulants‐induced maladaptive behaviors is still unknown. Here, we investigate the involvement of VGLUT3 and glutamate co‐transmission in amphetamine‐induced psychomotor effects and stereotypies. Taking advantage of constitutive and cell‐type specific VGLUT3‐deficient mouse lines, we tackled the hypothesis that VGLUT3 could gate psychomotor effects (locomotor activity and stereotypies) induced by acute or chronic administration of amphetamine. Interestingly, VGLUT3‐null mice demonstrated blunted amphetamine‐induced stereotypies as well as reduced striatal ∆FosB expression. VGLUT3‐positive varicosities within the striatum arise in part from 5HT neurons. We tested the involvement of VGLUT3 deletion in serotoninergic neurons in amphetamine‐induced stereotypies. Mice lacking VGLUT3 specifically in 5HT fibers showed no alteration to amphetamine sensitivity. In contrast, specific deletion of VGLUT3 in cholinergic neurons partially phenocopied the effects observed in the constitutive knock‐out mice. Our results show that constitutive deletion of VGLUT3 modulates acute and chronic locomotor effects induced by amphetamine. They point to the fact that the expression of VGLUT3 in multiple brain areas is pivotal in gating amphetamine‐induced psychomotor adaptations. Open Science Badges: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/. VGLUT3 is detected in neurons that use other neurotransmitters than glutamate, a neurochemical feature known as vesicular synergy. Here, we investigate the involvement of VGLUT3 in amphetamine‐induced psychomotor effects. We found that deletion of VGLUT3 promoted hyperlocomotor responses whereas blunted stereotypies were observed following amphetamine administration. A reduced expression of ∆FosB in the striatum mirrored such phenotype. In the striatum, VGLUT3 is expressed in cholinergic neurons and 5HT fibers. Cell‐type specific deletion of VGLUT3 in ACh and 5HT neurons revealed a complex dynamic regulation of amphetamine‐related effects. Our study points VGLUT3 as a critical modulator of psychostimulants‐induced adaptations. Open Science: This manuscript was awarded with the Open Materials Badge For more information see: https://cos.io/our-services/open-science-badges/ [ABSTRACT FROM AUTHOR]

Hippocampal interneurons release the inhibitory transmitter GABA to regulate excitation, rhythm generation and synaptic plasticity. A subpopulation of GABAergic basket cells co-expresses the GABA/glycine vesicular transporters (VIAAT) and the atypical type III vesicular glutamate transporter (VGLUT3); therefore, these cells have the ability to signal with both GABA and glutamate. GABAergic transmission by basket cells has been extensively characterized but nothing is known about the functional implications of VGLUT3-dependent glutamate released by these cells. Here, using VGLUT3-null mice we observed that the loss of VGLUT3 results in a metaplastic shift in synaptic plasticity at Shaeffer's collaterals - CA1 synapses and an altered theta oscillation. These changes were paralleled by the loss of a VGLUT3-dependent inhibition of GABAergic current in CA1 pyramidal layer. Therefore presynaptic type III metabotropic could be activated by glutamate released from VGLUT3-positive interneurons. This putative presynaptic heterologous feedback mechanism inhibits local GABAergic tone and regulates the hippocampal neuronal network. [ABSTRACT FROM AUTHOR]

Two near infra-red (NIR) fluorescent probes HupNIR1 and HupNIR2 based on the huprine scaffold and cyanine 5.0 dye have been synthesised and evaluated in situ for the detection of acetylcholinesterases in different tissues. As anticipated by the initial properties of huprine, both probes displayed a high affinity and selectivity for AChE toward BChE, with IC50 values in the nanomolar range and without any non-specific binding in the tissues. HupNIR2 appears the best probe for AChE with a great selectivity and sensitivity for AChE even in the brain region displaying a low AChE concentration as striatum. Moreover, the binding of HupNIR2 is affected when AChE is inhibited with toxic molecules such as organophosphates. This work provides a new tool to visualize active AChE in biological applications. [ABSTRACT FROM AUTHOR]

Within the hippocampus, the major somatostatin (SRIF) receptor subtype, the sst2A receptor, is localized at postsynaptic sites of the principal neurons where it modulates neuronal activity. Following agonist exposure, this receptor rapidly internalizes and recycles slowly through the trans-Golgi network. In epilepsy, a high and chronic release of somatostatin occurs, which provokes, in both rat and human tissue, a decrease in the density of this inhibitory receptor at the cell surface. The insulin-regulated aminopeptidase (IRAP) is involved in vesicular trafficking and shares common regional distribution with the sst2A receptor. In addition, IRAP ligands display anticonvulsive properties. We therefore sought to assess by in vitro and in vivo experiments in hippocampal rat tissue whether IRAP ligands could regulate the trafficking of the sst2A receptor and, consequently, modulate limbic seizures. Using pharmacological and cell biological approaches, we demonstrate that IRAP ligands accelerate the recycling of the sst2A receptor that has internalized in neurons in vitro or in vivo. Most importantly, because IRAP ligands increase the density of this inhibitory receptor at the plasma membrane, they also potentiate the neuropeptide SRIF inhibitory effects on seizure activity. Our results further demonstrate that IRAP is a therapeutic target for the treatment of limbic seizures and possibly for other neurological conditions in which downregulation of G-protein-coupled receptors occurs. [ABSTRACT FROM AUTHOR]

Two aspects of isospin breaking in the decay $$K^\pm \rightarrow \pi ^0 \pi ^0 e^\pm \mathop {\nu _e}\limits ^{_{(-)}}$$ are studied and discussed. The first addresses the possible influence of the phenomenological description of the unitarity cusp on the extraction of the normalization of the form factor from data. Using the scalar form factor of the pion as a theoretical laboratory, we find that this determination is robust under variations of the phenomenological parameterizations of the form factor. The second aspect concerns the issue of radiative corrections. We compute the radiative corrections to the total decay rate for $$K^\pm \rightarrow \pi ^0 \pi ^0 e^\pm \mathop {\nu _e}\limits ^{_{(-)}}$$ in a setting that allows comparison with the way radiative corrections were handled in the channel $$K^\pm \rightarrow \pi ^+ \pi ^- e^\pm \mathop {\nu _e}\limits ^{_{(-)}}$$ . We find that, once radiative corrections are included, the normalizations of the form factor as determined experimentally from data in the two decay channels come to a better agreement. The remaining discrepancy can easily be accounted for by other isospin-breaking corrections, mainly those due to the difference between the masses of the up and down quarks. [ABSTRACT FROM AUTHOR]

Terminal Schwann cells (TSCs) are key components of the mammalian neuromuscular junction (NMJ). How the TSCs sense the synaptic activity in physiological conditions remains unclear. We have taken advantage of the distinct localization of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) at the NMJ to bring out the function of different ACh receptors (AChRs). AChE is clustered by the collagen Q in the synaptic cleft and prevents the repetitive activation of muscle nicotinic AChRs. We found that BChE is anchored at the TSC by a proline-rich membrane anchor, the small transmembrane protein anchor of brain AChE. When BChE was specifically inhibited, ACh release was significant depressed through the activation of α7 nAChRs localized on the TSC and activated by the spillover of ACh. When both AChE and BChE were inhibited, the spillover increased and induced a dramatic reduction of ACh release that compromised the muscle twitch triggered by the nerve stimulation, α7 nAChRs at the TSC may act as a sensor for spillover of ACh adjusted by BChE and may represent an extrasynaptic sensor for homeostasis at the NMJ. In myasthenic rats, selective inhibition of AChE is more effective in rescuing muscle function than the simultaneous inhibition of AChE and BChE because the concomitant inhibition of BChE counteracts the positive action of AChE inhibition. These results show that inhibition of BChE should be avoided during the treatment of myasthenia and the pharmacological reversal of residual curarization after anesthesia. [ABSTRACT FROM AUTHOR]

For decades, neuroscientists have used enriched preparations of synaptic particles called synaptosomes to study synapse function. However, the interpretation of corresponding data is problematic as synaptosome preparations contain multiple types of synapses and non-synaptic neuronal and glial contaminants. We established a novel Fluorescence Activated Synaptosome Sorting ( FASS) method that substantially improves conventional synaptosome enrichment protocols and enables high-resolution biochemical analyses of specific synapse subpopulations. Employing knock-in mice with fluorescent glutamatergic synapses, we show that FASS isolates intact ultrapure synaptosomes composed of a resealed presynaptic terminal and a postsynaptic density as assessed by light and electron microscopy. FASS synaptosomes contain bona fide glutamatergic synapse proteins but are almost devoid of other synapse types and extrasynaptic or glial contaminants. We identified 163 enriched proteins in FASS samples, of which FXYD6 and Tpd52 were validated as new synaptic proteins. FASS purification thus enables high-resolution biochemical analyses of specific synapse subpopulations in health and disease. [ABSTRACT FROM AUTHOR]

Isospin breaking in the K form factors induced by the difference between charged and neutral pion masses is studied. Starting from suitably subtracted dispersion representations, the form factors are constructed in an iterative way up to two loops in the low-energy expansion by implementing analyticity, crossing, and unitarity due to two-meson intermediate states. Analytical expressions for the phases of the two-loop form factors of the K→ π π e ν channel are given, allowing one to connect the difference of form-factor phase shifts measured experimentally (out of the isospin limit) and the difference of S- and P-wave ππ phase shifts studied theoretically (in the isospin limit). The isospin-breaking correction consists of the sum of a universal part, involving only ππ rescattering, and a process-dependent contribution, involving the form factors in the coupled channels. The dependence on the two S-wave scattering lengths $a_{0}^{0}$ and $a_{0}^{2}$ in the isospin limit is worked out in a general way, in contrast to previous analyses based on one-loop chiral perturbation theory. The latter is used only to assess the subtraction constants involved in the dispersive approach. The two-loop universal and process-dependent contributions are estimated and cancel partially to yield an isospin-breaking correction close to the one-loop case. The recent results on the phases of K→ π π e ν form factors obtained by the NA48/2 collaboration at the CERN SPS are reanalysed including this isospin-breaking correction to extract values for the scattering lengths $a_{0}^{0}$ and $a_{0}^{2}$, as well as for low-energy constants and order parameters of two-flavour χPT. [ABSTRACT FROM AUTHOR]

CLIPR-59 is a new member of the cytoplasmic linker proteins (CLIP) family mainly localized to the trans-Golgi network. We show here that Clipr-59 expression in mice is restricted to specific pools of neurons, in particular motoneurons (MNs), and progressively increases from embryonic day 12.5 (E12.5) until the first postnatal days. We generated a Clipr-59 knockout mouse model that presents perinatal lethality due to respiratory defects. Physiological experiments revealed that this altered innervation prevents the normal nerveelicited contraction of the mutant diaphragm that is reduced both in amplitude and fatigue-resistance at E18.5, despite unaffected functional muscular contractility. Innervation of the mutant diaphragm is not altered until E15.5, but is then partially lost in the most distal parts of the muscle. Ultrastructural observations of neuromuscular junctions (NMJs) in the distal region of the diaphragm reveal a normal organization, but a lower density of nerve terminals capped by terminal Schwann cells in E18.5 mutant when compared with control embryos. Similar defects in NMJ stability, with a hierarchy of severity along the caudo-rostral axis, are also observed in other muscles innervated by facial and spinal MNs in Clipr-59 mutant mice. Clipr-59 deficiency therefore affects axon maintenance but not axon guidance toward muscle targets. Thus, CLIPR-59 is involved in the stabilization of specific motor axons at the NMJ during mouse late embryogenesis and its role is crucial for mouse perinatal development. [ABSTRACT FROM AUTHOR]

Background: We wished to develop alternate production strategies to generate antibodies against traditionally problematic antigens. As a model we chose butyrylcholinesterase (BChE), involved in termination of cholinergic signaling, and widely considered as a poor immunogen. Methodology/Principal Findings: Jettisoning traditional laborious in silico searching methods to define putative epitopes, we simply immunized available BChE knock-out mice with full-length recombinant BChE protein (having been produced for crystallographic analysis). Immunization with BChE, in practically any form (recombinant human or mouse BChE, BChE purified from human serum, native or denatured), resulted in strong immune responses. Native BChE produced antibodies that favored ELISA and immunostaining detection. Denatured and reduced BChE were more selective for antibodies specific in Western blots. Two especially sensitive monoclonal antibodies were found capable of detecting 0.25 ng of BChE within one min by ELISA. One is specific for human BChE; the other cross-reacts with mouse and rat BChE. Immunization of wild-type mice served as negative controls. Conclusions/Significance: We examined a simple, fast, and highly efficient strategy to produce antibodies by mining two expanding databases: namely those of knock-out mice and 3D crystallographic protein-structure analysis. We conclude that the immunization of knock-out mice should be a strategy of choice for antibody production. [ABSTRACT FROM AUTHOR]

Acetylcholinesterase (AChE) accumulates on axonal varicosities and is primarily found as tetramers associated with a proline-rich membrane anchor (PRiMA). PRiMA is a small transmembrane protein that efficiently transforms secreted AChE to an enzyme anchored on the outer cell surface. Surprisingly, in the striatum of the PRiMA knock-out mouse, despite a normal level of AChEmRNA,we find only 2-3% of wild type AChE activity, with the residual AChE localized in the endoplasmic reticulum, demonstrating that PRiMA in vivo is necessary for intracellular processing of AChE in neurons. Moreover, deletion of the retention signal of the AChE catalytic subunit in mice, which is the domain of interaction with PRiMA, does not restore AChE activity in the striatum, establishing thatPRiMAis necessary to target and/or to stabilize nascent AChE in neurons. These unexpected findings open new avenues to modulating AChE activity and its distribution in CNS disorders. [ABSTRACT FROM AUTHOR]

Abstract: We perform a matching of the two loop-chiral perturbation theory representation of the scalar Kπ form factor to a dispersive one. Knowing the value of and in the Standard Model (SM) allows to determine two LECs, the slope of the scalar form factor and the deviation of the Callan–Treiman theorem. Going beyond the SM and assuming the knowledge of the slope of the scalar form factor from experiment, the matching allows us to determine the ratio of , , a certain combination of non-standard couplings, the deviation of the Callan–Treiman theorem and the two LECs. [Copyright &y& Elsevier]

Abstract: We study the extraction of Δ-resonance parameters from lattice data for small quark masses, corresponding to the case of an unstable Δ. To this end, we calculate the spectrum of the correlator of two Δ-fields in a finite Euclidean box up-to-and-including in the small scale expansion using infrared regularization. On the basis of our numerical study, we argue that the extraction of the parameters of the Δ-resonance (in particular, of the mass and the pion–nucleon–delta coupling constant) from the measured volume dependence of the lowest energy levels should be feasible. [Copyright &y& Elsevier]

Abstract: Theoretical as well as experimental progress has been made in the last decade in describing the properties of baryons. In this review I will mostly report on the theoretical issues. Two non-perturbative methods are privileged frameworks for studying their properties in the low-energy domain: chiral perturbation theory, the effective field theory of the Standard Model at energies below 1 GeV and lattice QCD. I will mainly concentrate here on the first one but I will also discuss the complementarity of the two methods. Chiral extrapolations for lattice simulations of some nucleon properties will be investigated. I will then concentrate on processes involving at most two nucleons, describing for example pion–nucleon and pion–deuteron scattering, pion photo- and electro-production off the nucleon and the deuteron and doubly virtual Compton scattering. Three flavor calculations will also be reviewed. [Copyright &y& Elsevier]

This review gives a brief introduction to chiral perturbation theory in its various settings. We discuss some applications of recent interest, including chiral extrapolations for lattice gauge theory. [ABSTRACT FROM AUTHOR]

Abstract: We analyze the nucleon axial-vector coupling to two loops in chiral perturbation theory. We show that chiral extrapolations based on this representation require lattice data with pion masses below 300 MeV. [Copyright &y& Elsevier]

Abstract: Clean tests of a small admixture of right-handed quark currents directly coupled to the standard W are still lacking. We show that such non-standard couplings can be significantly constrained measuring the value of the scalar Kπ form factor at the Callan–Treiman point to a few percent. A realistic prospect of such a measurement in decay based on an accurate dispersive representation of the scalar form factor is presented. The inadequacy of the currently used linear parametrization is explained and illustrated using recent KTeV data. We briefly comment on the charged kaon mode. [Copyright &y& Elsevier]

In vitro studies have widely demonstrated that the abundance and availability of G-protein-coupled receptors (GPCRs) at the cell surface is regulated by the neuronal environment and is the result of complex intraneuronal trafficking. However, this regulation is still poorly understood in vivo. Modulation of receptor availability at the neuronal membrane is a key event in the regulation of neuronal functions (e.g. neurotransmitter release or neuronal excitability in physiological, pathological or therapeutic conditions). We discuss the effects of duration of receptor stimulation (acute versus chronic) on the intraneuronal trafficking of GPCRs in vivo, and we show that this trafficking might differ according to subcellular compartment (soma, dendrites or axon terminals). [Copyright &y& Elsevier]

Abstract: We calculate the nucleon and the delta mass to fourth order in a covariant formulation of the small scale expansion. We analyze lattice data from the MILC Collaboration and demonstrate that the available lattice data combined with our knowledge of the physical values for the nucleon and delta masses lead to consistent chiral extrapolation functions for both observables up to fairly large pion masses. This holds in particular for very recent data on the delta mass from the QCDSF Collaboration. The resulting pion–nucleon sigma term is . This first quantitative analysis of the quark-mass dependence of the structure of the in full QCD within chiral effective field theory suggests that (the real part of) the nucleon–delta mass-splitting in the chiral limit, , is slightly larger than at the physical point. Further analysis of simultaneous fits to nucleon and delta lattice data are needed for a precision determination of the properties of the first excited state of the nucleon. [Copyright &y& Elsevier]