The SV2A ligand levetiracetam inhibits presynaptic Ca2+ channels via an intracellular pathway

Levetiracetam (LEV) is a prominent antiepileptic drug (AED) which binds to neuronal synaptic vesicle glycoprotein 2A (SV2A) protein and has reported effects on ion channels, but retains a poorly-defined mechanism of action. Here, we investigate inhibition of voltage-dependent Ca2+ (CaV) channels as a potential mechanism by which LEV imparts effects on neuronal activity. We used electrophysiological methods to investigate the effects of LEV on cholinergic synaptic transmission and CaV channel activity in superior cervical ganglion neurons (SCGNs). In parallel, we investigated effects of the LEV ‘inactive’ R-enantiomer, UCB L060. Thus, LEV, but not UCB L060 (each 100 μM), inhibited synaptic transmission between SCGNs in long-term culture in a time-dependent manner, significantly reducing excitatory postsynaptic potentials (EPSP) following ≥30 min application. In isolated SCGNs, LEV pretreatment (≥1 h), but not acute (5 min) application, significantly inhibited whole-cell IBa amplitude. In current clamp recordings, LEV reduced the amplitude of the afterhyperpolarizing potential (AHP) in a Ca2+-dependent manner, but also increased action potential (AP) latency in a Ca2+-independent manner, suggesting further mechanisms associated with reduced excitability. Intracellular LEV application (4-5 min) caused a rapid inhibition of IBa amplitude to an extent comparable to that seen following extracellular LEV pretreatment ( ≥ 1 h). Neither pretreatment nor intracellular application of UCB L060 produced any inhibitory effects on IBa amplitude. These results identify a stereospecific intracellular pathway by which LEV inhibits presynaptic CaV channels; resultant reductions in neuronal excitability are proposed to contribute to the anticonvulsant effects of LEV.

Synaptic vesicle glycoprotein 2A modulates vesicular release and calcium channel function at peripheral sympathetic synapses

Synaptic vesicle glycoprotein (SV)2A is a transmembrane protein found in secretory vesicles and is critical for Ca2+-dependent exocytosis in central neurons, although its mechanism of action remains uncertain. Previous studies have proposed, variously, a role of SV2 in the maintenance and formation of the readily releasable pool (RRP) or in the regulation of Ca2+ responsiveness of primed vesicles. Such previous studies have typically used genetic approaches to ablate SV2 levels; here, we used a strategy involving small interference RNA (siRNA) injection to knockdown solely presynaptic SV2A levels in rat superior cervical ganglion (SCG) neuron synapses. Moreover, we investigated the effects of SV2A knockdown on voltage-dependent Ca2+ channel (VDCC) function in SCG neurons. Thus, we extended the studies of SV2A mechanisms by investigating the effects on vesicular transmitter release and VDCC function in peripheral sympathetic neurons. We first demonstrated an siRNA-mediated SV2A knockdown. We showed that this SV2A knockdown markedly affected presynaptic function, causing an attenuated RRP size, increased paired-pulse depression and delayed RRP recovery after stimulus-dependent depletion. We further demonstrated that the SV2A–siRNA-mediated effects on vesicular release were accompanied by a reduction in VDCC current density in isolated SCG neurons. Together, our data showed that SV2A is required for correct transmitter release at sympathetic neurons. Mechanistically, we demonstrated that presynaptic SV2A: (i) acted to direct normal synaptic transmission by maintaining RRP size, (ii) had a facilitatory role in recovery from synaptic depression, and that (iii) SV2A deficits were associated with aberrant Ca2+ current density, which may contribute to the secretory phenotype in sympathetic peripheral neurons.

Highly efficient neural differentiation of human somatic stem cells, isolated by minimally invasive periodontal surgery

Neural stem cells (NSCs) are potential sources for cell therapy of neurodegenerative diseases and for drug screening. Despite their potential benefits, ethical and practical considerations limit the application of NSCs derived from human embryonic stem cells (ES) or adult brain tissue. Thus, alternative sources are required to satisfy the criteria of ready accessibility, rapid expansion in chemically defined media and reliable induction to a neuronal fate. We isolated somatic stem cells from the human periodontium that were collected during minimally invasive periodontal access flap surgery as part of guided tissue regeneration therapy. These cells could be propagated as neurospheres in serum-free medium, which underscores their cranial neural crest cell origin. Culture in the presence of epidermal growth factor (EGF) and fibroblast growth factor-2 (FGF-2) under serum-free conditions resulted in large numbers of nestin-positive/Sox-2-positive NSCs. These periodontium-derived (pd) NSCs are highly proliferative and migrate in response to chemokines that have been described as inducing NSC migration. We used immunocytochemical techniques and RT-PCR analysis to assess neural differentiation after treatment of the expanded cells with a novel induction medium. Adherence to substrate, growth factor deprivation, and retinoic acid treatment led to the acquisition of neuronal morphology and stable expression of markers of neuronal differentiation by more than 90% of the cells. Thus, our novel method might provide nearly limitless numbers of neuronal precursors from a readily accessible autologous adult human source, which could be used as a platform for further experimental studies and has potential therapeutic implications.

Interaction of adult human neural crest-derived stem cells with a nanoporous titanium surface is sufficient to induce their osteogenic differentiation

Osteogenic differentiation of various adult stem cell populations such as neural crest-derived stem cells is of great interest in the context of bone regeneration. Ideally, exogenous differentiation should mimic an endogenous differentiation process, which is partly mediated by topological cues. To elucidate the osteoinductive potential of porous substrates with different pore diameters (30 nm, 100 nm), human neural crest-derived stem cells isolated from the inferior nasal turbinate were cultivated on the surface of nanoporous titanium covered membranes without additional chemical or biological osteoinductive cues. As controls, flat titanium without any topological features and osteogenic medium was used. Cultivation of human neural crest-derived stem cells on 30 nm pores resulted in osteogenic differentiation as demonstrated by alkaline phosphatase activity after seven days as well as by calcium deposition after 3 weeks of cultivation. In contrast, cultivation on flat titanium and on membranes equipped with 100 nm pores was not sufficient to induce osteogenic differentiation. Moreover, we demonstrate an increase of osteogenic transcripts including Osterix, Osteocalcin and up-regulation of Integrin β1 and α2 in the 30 nm pore approach only. Thus, transplantation of stem cells pre-cultivated on nanostructured implants might improve the clinical outcome by support of the graft adherence and acceleration of the regeneration process.

Christian Gottlob Heyne and the changing fortunes of the commentary in the age of Altertumswissenschaft

This chapter discusses the pedagogic and scholarly priorities that informed Heyne’s commentaries on Tibullus (1755), Virgil (1767-75) and Homer (1802), as well as their initial critical reception. Like those of his teachers, Gesner and Ernesti, Heyne’s works eschew detailed textual scholarship in favour of aesthetic and historicizing appreciation of literary works as wholes. Their formal innovations – most notably the relegation of advanced philological discussions to endnotes and the inclusion of excursuses on significant historical and cultural questions – are an attempt to tailor a traditional format to the demands of an Enlightened age and the cultural-historical interests of the new Altertumswissenschaft. The chapter discusses their contrasting critical receptions in order to raise questions about the viability of Heyne’s endeavours to make a traditional medium fit new concerns.

Winckelmann in the perspective of Altertumswissenschaft: Christian Gottlob Heyne and Friedrich August Wolf

Winckelmann's writings hold an interest for modern classical studies which is not restricted to the subfield of classical archaeology. Considered in terms of methodology, his writings dramatise problems and questions which attend any attempt to provide a comprehensive account of ancient culture and society. The Geschichte der Kunst des Alterthums provided an influential model of what such a reconstruction might look like in a period when classical philology was undergoing a significant reconfiguration as „Alterthums-Wissenschaft" at the hands of scholars such as Christian Gottlob Heyne and Friedrich August Wolf. Investigation of their critical responses to Winckelmann’s works aims to contribute to understanding both of the early reception of his works and of questions which are still relevant today. Im Rahmen der modernen Altertumswissenschaften kommt den Werken Winckelmanns eine Bedeutung zu, die nicht auf den Bereich der Klassischen Archäologie beschränkt ist. Methodologisch betrachtet, dramatisieren seine Schriften Probleme und Fragen, die jedem Versuch einer umfassenden, erklärenden Rekonstruktion der antiken Kultur und Gesellschaft zugrunde liegen. Die Geschichte der Kunst des Alterthums hat ein einflussreiches Modell dafür geliefert, was eine solche Rekonstruktion in einer Zeit leisten konnte, in der die klassische Philologie einer erheblichen Umstrukturierung als „Alterthums-Wissenschaft“ durch Gelehrte wie Christian Gottlob Heyne und Friedrich August Wolf unterzogen wurde. Die vorliegende Untersuchung ihrer kritischen Reaktionen auf Winckelmanns Schriften soll dazu beitragen, sowohl die frühe Rezeption seines Werkes als auch Fragestellungen, die heute noch aktuell sind, besser zu verstehen.

Students’ perception of privacy risks in using social networking sites for learning: a study of Uganda Christian University

Although social networking sites (SNSs) present a great deal of opportunities to support learning, the privacy risk is perceived by learners as a friction point that affects their full use for learning. Privacy risks in SNSs can be divided into risks that are posed by the SNS provider itself and risks that result from user’s social interactions. Using an online survey questionnaire, this study explored the students’ perception of the benefits in using social networking sites for learning purposes and their perceived privacy risks. A sample of 214 students from Uganda Christian University in Africa was studied. The results show that although 88 % of participants indicated the usefulness of SNSs for learning, they are also aware of the risks associated with these sites. Most of the participants are concerned with privacy risks such as identity theft, cyber bullying, and impersonation that might influence their online learning participation in SNSs.