A clickable oxysterol photolabel retains NMDA receptor activity and accumulates in neurons

Oxysterol analogs that modulate NMDA receptor function are candidates for therapeutic development to treat neuropsychiatric disorders. However, the cellular actions of these compounds are still unclear. For instance, how these compounds are compartmentalized or trafficked in neurons is unknown. In this study, we utilized a chemical biology approach combining photolabeling and click chemistry. We introduce a biologically active oxysterol analog that contains: (1) a diazirine group, allowing for the permanent labeling of cellular targets, and (2) an alkyne group, allowing for subsequent in situ visualization using Cu2+ catalyzed cycloaddition of an azide-conjugated fluorophore. The physiological properties of this analog at NMDA receptors resemble those of other oxysterols, including occlusion with other oxysterol-like compounds. Fluorescent imaging reveals that the analog accumulates diffusely in the cytoplasm of neurons through an energy-independent mechanism. Overall, this work introduces a novel chemical biology approach to investigate oxysterol actions and introduces a tool useful for further cell biological and biochemical studies of oxysterols.</p

Effect of Central Mass Concentration on the Formation of Nuclear Spirals in Barred Galaxies

We have performed smoothed particle hydrodynamics (SPH) simulations to study the response of the central kiloparsec region of a gaseous disk to the imposition of nonaxisymmetric bar potentials. The model galaxies are composed of the three axisymmetric components (halo, disk, and bulge) and a non-axisymmetric bar. These components are assumed to be invariant in time in the frame corotating with the bar. The potential of spherical $\gamma$-models of Dehnen is adopted for the bulge component whose density varies as $r^{-\gamma}$ near the center and $r^{-4}$ at larger radiiand hence, possesses a central density core for $\gamma = 0$ and cusps for $\gamma > 0$. Since the central mass concentration of the model galaxies increases with the cusp parameter $\gamma$, we have examined here the effect of the central mass concentration by varying the cusp parameter $\gamma$ on the mechanism responsible for the formation of the symmetric two-armed nuclear spirals in barred galaxies. Our simulations show that the symmetric two-armed nuclear spirals are formed by hydrodynamic spiral shocks driven by the gravitational torque of the bar for the models with $\gamma = 0$ and 0.5. On the other hand, the symmetric two-armed nuclear spirals in the models with $\gamma=1$ and 1.5 are explained by gas density waves. Thus, we conclude that the mechanism responsible for the formation of the symmetric two-armed nuclear spirals in barred galaxies changes from the hydrodynamic shocks to the gas density waves when the central mass concentration increases from $\gamma = 0$ to 1.5.Comment: 29 pages, 5 figures (Color Figures 3-5), Accepted for Publication in Astrophysical Journal (ApJ

Allopregnanolone effects on inhibition in hippocampal parvalbumin interneurons

Allopregnanolone (AlloP) is a neurosteroid that potentiates ionotropic GABAergic (GAB

Positive allosteric modulation as a potential therapeutic strategy in anti-NMDA receptor encephalitis

N-methyl-d-aspartate receptors (NMDARs) are ionotropic glutamate receptors important for synaptic plasticity, memory, and neuropsychiatric health. NMDAR hypofunction contributes to multiple disorders, including anti-NMDAR encephalitis (NMDARE), an autoimmune disease of the CNS associated with GluN1 antibody-mediated NMDAR internalization. Here we characterize the functional/pharmacological consequences of exposure to CSF from female human NMDARE patients on NMDAR function, and we characterize the effects of intervention with recently described positive allosteric modulators (PAMs) of NMDARs. Incubation (48 h) of rat hippocampal neurons of both sexes in confirmed NMDARE patient CSF, but not control CSF, attenuated NMDA-induced current. Residual NMDAR function was characterized by lack of change in channel open probability, indiscriminate loss of synaptic and extrasynaptic NMDARs, and indiscriminate loss of GluN2B-containing and GluN2B-lacking NMDARs. NMDARs tagged with N-terminal pHluorin fluorescence demonstrated loss of surface receptors. Thus, function of residual NMDARs following CSF exposure was indistinguishable from baseline, and deficits appear wholly accounted for by receptor loss. Coapplication of CSF and PAMs of NMDARs (SGE-301 or SGE-550, oxysterol-mimetic) for 24 h restored NMDAR function following 24 h incubation in patient CSF. Curiously, restoration of NMDAR function was observed despite washout of PAMs before electrophysiological recordings. Subsequent experiments suggested that residual allosteric potentiation of NMDAR function explained the persistent rescue. Further studies of the pathogenesis of NMDARE and intervention with PAMs may inform new treatments for NMDARE and other disorders associated with NMDAR hypofunction.SIGNIFICANCE STATEMENTAnti-N-methyl-d-aspartate receptor encephalitis (NMDARE) is increasingly recognized as an important cause of sudden-onset psychosis and other neuropsychiatric symptoms. Current treatment leaves unmet medical need. Here we demonstrate cellular evidence that newly identified positive allosteric modulators of NMDAR function may be a viable therapeutic strategy.</jats:p

Fast phasic release properties of dopamine studied with a channel biosensor

Few other neurotransmitters are of as intense interest to neuropsychiatry and neurology as dopamine, yet existing techniques to monitor dopamine release leave an important spatiotemporal gap in our understanding. Electrochemistry and fluorescence imaging tools have been developed to fill the gap, but these methods have important limitations. We circumvent these limitations by introducing a dopamine-gated chloride channel into rat dorsal striatal medium spiny neurons, targets of strong dopamine innervation, thereby transforming dopamine from a slow transmitter into a fast transmitter and revealing new opportunities for studying moment-to-moment regulation of dopamine release. We demonstrate pharmacological and biophysical properties of the channel that make it suitable for fast, local dopamine measurements, and we demonstrate for the first time spontaneous and evoked responses to vesicular dopamine release in the dorsal striatum. Evoked dopamine currents were separated into a fast, monosynaptic component and a slower-rising and decaying disynaptic component mediated by nicotinic receptor activation. In summary, LGC-53 represents a dopamine biosensor with properties suitable for temporal separation of distinct dopamine signals in targets of dopamine innervation

Presynaptically Silent Synapses Studied with Light Microscopy

Synaptic plasticity likely underlies the nervous system's ability to learn and remember and may also represent an adaptability that prevents otherwise damaging insults from becoming neurotoxic. We have been studying a form of presynaptic plasticity that is interesting in part because it is expressed as a digital switching on and off of a presynaptic terminal s ability to release vesicles containing the neurotransmitter glutamate. Here we demonstrate a protocol for visualizing the activity status of presynaptic terminals in dissociated cell cultures prepared from the rodent hippocampus. The method relies on detecting active synapses using staining with a fixable form of the styryl dye FM1-43, commonly used to label synaptic vesicles. This staining profile is compared with immunostaining of the same terminals with an antibody directed against the vesicular glutamate transporter 1 (vGluT-1), a stain designed to label all glutamate synapses regardless of activation status. We find that depolarizing stimuli induce presynaptic silencing. The population of synapses that is silent under baseline conditions can be activated by prolonged electrical silencing or by activation of cAMP signaling pathways

Non-sedative cortical EEG signatures of allopregnanolone and functional comparators

Neurosteroids that positively modulate GAB

A Specific Role for Ca\u3csup\u3e2+\u3c/sup\u3e-Dependent Adenylyl Cyclases in Recovery from Adaptive Presynaptic Silencing

Glutamate generates fast postsynaptic depolarization throughout the CNS. The positive-feedback nature of glutamate signaling likely necessitates flexible adaptive mechanisms that help prevent runaway excitation. We have previously explored presynaptic adaptive silencing, a form of synaptic plasticity produced by ongoing neuronal activity and by strong depolarization. Unsilencing mechanisms that maintain active synapses and restore normal function after adaptation are also important, but mechanisms underlying such presynaptic reactivation remain unexplored. Here we investigate the involvement of the cAMP pathway in the basal balance between silenced and active synapses, as well as the recovery of baseline function after depolarization-induced presynaptic silencing. Activation of the cAMP pathway activates synapses that are silent at rest, and pharmacological inhibition of cAMP signaling silences basally active synapses. Adenylyl cyclase (AC) 1 and AC8, the major Ca2+-sensitive AC isoforms, are not crucial for the baseline balance between silent and active synapses. In cells from mice doubly deficient in AC1 and AC8, the baseline percentage of active synapses was only modestly reduced compared with wild-type synapses, and forskolin unsilencing was similar in the two genotypes. Nevertheless, after strong presynaptic silencing, recovery of normal function was strongly inhibited in AC1/AC8-deficient synapses. The entire recovery phenotype of the double null was reproduced in AC8-deficient but not AC1-deficient cells.Weconclude that, under normal conditions, redundant cyclase activity maintains the balance between presynaptically silent and active synapses, but AC8 plays a particularly important role in rapidly resetting the balance of active to silent synapses after adaptation to strong activity

Topic Study Group No. 1: Early Childhood Mathematics Education (Up to Age 7)

This book is open access under a CC BY 4.0 license. The book presents the Proceedings of the 13th International Congress on Mathematical Education (ICME-13) and is based on the presentations given at the 13th International Congress on Mathematical Education (ICME-13). ICME-13 took place from 24th- 31st July 2016 at the University of Hamburg in Hamburg (Germany). The congress was hosted by the Society of Didactics of Mathematics (Gesellschaft für Didaktik der Mathematik - GDM) and took place under the auspices of the International Commission on Mathematical Instruction (ICMI). ICME-13 brought together about 3.500 mathematics educators from 105 countries, additionally 250 teachers from German speaking countries met for specific activities. Directly before the congress activities were offered for 450 Early Career Researchers. The proceedings give a comprehensive overview on the current state-of-the-art of the discussions on mathematics education and display the breadth and deepness of current research on mathematical teaching-and-learning processes. The book introduces the major activities of ICME-13, namely articles from the four plenary lecturers and two plenary panels, articles from the five ICMI awardees, reports from six national presentations, three reports from the thematic afternoon devoted to specific features of ICME-13. Furthermore, the proceedings contain descriptions of the 54 Topic Study Groups, which formed the heart of the congress and reports from 29 Discussion Groups and 31 Workshops. The additional important activities of ICME-13, namely papers from the invited lecturers, will be presented in the second volume of the proceedings