Short-term facilitation and depression in the cerebellum: some observations on wild-type and mutant rodents deficient in the extracellular matrix molecule tenascin C

Short-term plasticity was studied on synapses to Purkinje cells (PC): paired-pulse facilitation in parallel fibers (PF) and paired-pulse depression in climbing fibers (CF). Both phenomena relate to synaptic strength. These forms of short-term plasticity were tested on cerebellar slices in rat by early postnatal synchronous stimulation of olivary neurons (i.e., CFs) with harmaline and by inhibition of a metabotropic glutamate receptor (mGluR) as well as in mice that were deficient in the extracellular matrix glycoprotein tenascin-C. Harmaline stimulation delayed the developmental competition between CF inputs and maintained multiple innervation. Paired-pulse depression of the CF-PC synapse after harmaline treatment was more expressed. However, paired-pulse facilitation in PF-PC synapses remained unchanged. Electrophysiological responses of postsynaptic mGluR1 in CF-PC synapses could be obtained only with AMPA receptors blocked and glutamate uptake impaired. The mGluR1-specific antagonist CPCCOEt suppressed the CF-mGluR EPSC in some PCs and potentiated it in other PCs. CF paired-pulse depression was not changed with CPCCOEt, thus excluding a presynaptic effect. The postsynaptic effect was underlined by CPCCOEt-induced rise in amplitude of EPSC and by a prolongation of its decay time. Tenascins are extracellular matrix glycoproteins that may restrict the regenerative capacity of the nervous tissue. Testing short-term presynaptic plasticity in tenascin-C-deficient mice showed that CF paired-pulse depression was less expressed while PF paired-pulse facilitation was augmented except in a group of cells where there was even depression. The results underline differences in forms of short-term plasticity with regard to susceptibility to diverse modulatory factors

Immunoglobulins G from sera of amyotrophic lateral sclerosis patients induce oxidative stress and upregulation of antioxidative system in BV-2 microglial cell line

© 2017 Milošević, Milicević, Božić, Lavrnja, Stevanović, Bijelić, Dubaić, Živković, Stević, Giniatullin and Andjus. Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder with a very fast progression, no diagnostic tool for the presymptomatic phase, and still no effective treatment of the disease. Although ALS affects motor neurons, the overall pathophysiological condition points out to the non-cell autonomous mechanisms, where astrocytes and microglia play crucial roles in the disease progression. We have already shown that IgG from sera of ALS patients (ALS IgG) induce calcium transients and an increase in the mobility of acidic vesicles in cultured rat astrocytes. Having in mind the role of microglia in neurodegeneration, and a well-documented fact that oxidative stress is one of the many components contributing to the disease, we decided to examine the effect of ALS IgG on activation, oxidative stress and antioxidative system of BV-2 microglia, and to evaluate their acute effect on cytosolic peroxide, pH, and on reactive oxygen species (ROS) generation. All tested ALS IgGs (compared to control IgG) induced oxidative stress (rise in nitric oxide and the index of lipid peroxidation) followed by release of TNF-α and higher antioxidative defense (elevation of Mn- and CuZn-superoxide dismutase, catalase, and glutathione reductase with a decrease of glutathione peroxidase and glutathione) after 24 h treatment. Both ALS IgG and control IgG showed same localization on the membrane of BV-2 cells following 24 h treatment. Cytosolic peroxide and pH alteration were evaluated with fluorescent probes HyPer and SypHer, respectively, having in mind that HyPer also reacts to pH changes. Out of 11 tested IgGs from ALS patients, 4 induced slow exponential rise of HyPer signal, with maximal normalized fluorescence in the range 0.2-0.5, also inducing similar increase of SypHer intensity, but of a lower amplitude. None of the control IgGs induced changes with neither of the indicators. Acute ROS generation was detected in one out of three tested ALS samples with carboxy-H2DCFDA. The observed phenomena demonstrate the potential role of inflammatory humoral factors, IgGs, as potential triggers of the activation in microglia, known to occur in later stages of ALS. Therefore, revealing the ALS IgG signaling cascade in microglial cells could offer a valuable molecular biomarker and/or a potential therapeutic target

Polyacetylenes from Sardinian Oenanthe fistulosa: A Molecular Clue to risus sardonicus

An investigation of Oenanthe fistulosa from Sardinia afforded oenanthotoxin (1a) and dihydrooenanthotoxin (1b) from the roots and the diacetylenic epoxydiol 2 from the seeds. The absolute configuration of 1a and 1b was established as R by the modified Mosher's method, and the structure of 2 by chemical correlation with (+)-(3R,8S)-falcarindiol. Oenanthotoxin (1a) and dihydrooenanthotoxin (1b) were found to potently block GABAergic responses, providing a molecular rationale for the symptoms of poisoning from water-dropwort (Oenanthe crocata) and related plants. These observations bear relevance for a series of historical and ethnopharmacological observations on the identification of the Sardonic herb and the molecular details of the facial muscular contraction caused by its ingestion (risus sardonicus)

Pin1-dependent signaling negatively affects GABAergic transmission by modulating neuroligin2/gephyrin interaction

The cell adhesion molecule Neuroligin2 (NL2) is localized selectively at GABAergic synapses, where it interacts with the scaffolding protein gephyrin in the post-synaptic density. However, the role of this interaction for formation and plasticity of GABAergic synapses is unclear. Here, we demonstrate that endogenous NL2 undergoes proline-directed phosphorylation at its unique S714-P consensus site, leading to the recruitment of the peptidyl-prolyl cis-trans isomerase Pin1. This signalling cascade negatively regulates NL2' s ability to interact with gephyrin at GABAergic post-synaptic sites. As a consequence, enhanced accumulation of NL2, gephyrin and GABA A receptors was detected at GABAergic synapses in the hippocampus of Pin1-knockout mice (Pin1\ufffd/\ufffd) associated with an increase in amplitude of spontaneous GABA A -mediated post-synaptic currents. Our results suggest that Pin1-dependent signalling represents a mechanism to modulate GABAergic transmission by regulating NL2/gephyrin interaction. \ufffd 2014 Macmillan Publishers Limited. All rights reserved

What’s special about the ethical challenges of studying disorders with altered brain activity?

Where there is no viable alternative, studies of neuronal activity are conducted on animals. The use of animals, particularly for invasive studies of the brain, raises a number of ethical issues. Practical or normative ethics are enforced by legislation, in relation to the dominant welfare guidelines developed in the UK and elsewhere. Guidelines have typically been devised to cover all areas of biomedical research using animals in general, and thus lack any specific focus on neuroscience studies at the level of the ethics, although details of the specific welfare recommendations are different for invasive studies of the brain. Ethically there is no necessary distinction between neuroscience and other biomedical research in that the brain is a final common path for suffering, irrespective of whether this involves any direct experience of pain. One exception arises in the case of in vitro studies, which are normally considered as an acceptable replacement for in vivo studies. However, to the extent sentience is possible, maintaining central nervous system tissue outside the body naturally raises ethical questions. Perhaps the most intractable challenge to the ethical use of animals in order to model neuronal disorder is presented by the logical impasse in the argument that the animal is similar enough to justify the validity of the experimental model, but sufficiently different in sentience and capacity for suffering, for the necessary experimental procedures to be permissible

Immunoglobulins From Amyotrophic Lateral Sclerosis Patients Enhance The Frequency Of Glycine-mediated Spontaneous Inhibitory Postsynaptic Currents In Rat Hypoglossal Motoneurons

Abstract — Amyotrophic lateral sclerosis (ALS) is a devastating, still incurable neurological disorder affecting upper and lower motoneurons. Passive transfer of the disease occurs when immunoglobulins from ALS patients are injected into experimental animals. It is suggested that ALS IgGs cause excitotoxicity by acting on voltage-gated Ca 2+ channels. We reported previously that ALS IgGs increase spontaneous release of glutamate in hippocampal neurons. Since these cells are not normally affected in ALS, we here studied the effect of ALS IgGs on hypoglossal motoneurons in rat brain-stem slices. The frequency of spontaneous glycine-mediated inhibitory postsynaptic currents (sIPSCs) was augmented, but not that of miniature ones (mIPSCs), thus pointing to an indirect effect on release

Pharmacology of the metabotropic glutamate receptor mediated current at the climbing fiber to Purkinje cell synapse.

Different forms of synaptic plasticity in the cerebellum are mediated by metabotropic glutamate receptors (mGluRs). At parallel fiber (PF) to Purkinje cell (PC) synapses activation of mGluR gives rise to a well known slow synaptic current inhibited by antagonists of mGluR1. The distribution of mGluR types in the climbing fiber (CF) to PC synapses is not well known. However, a mGluR1alpha-mediated all-or-none postsynaptic current was also demonstrated at the CF-PC synapse (Dzubay and Otis, Neuron 36, 1159, 2002). Using whole cell patch-clamp recording from PCs in rat cerebellar slices with AMPA receptors blocked and glutamate uptake impaired we demonstrate a more complex pharmacology of a current obtained by single or train CF stimulation. The mGluR1 specific antagonist CPCCOEt in a group of cells suppressed this response while in a similar number of other cells it induced a potentiating effect. The antagonists of mGluR groups II and III (LY341495 and MSOP, respectively) predominantly suppressed the current. The ambiguous effect of CPCCOEt was checked by measuring the paired-pulse depression of the CF EPSC, which was not changed with the antagonist in normal as well as in low (0.5 mM) external Ca(2+) (used to prevent saturation of AMPARs), thus excluding a presynaptic effect. However, CPCCOEt induced a rise in the amplitude (by approximately 50%) as well as a prolongation (p<0.05) of the decay time of CF EPSCs at normal 2 mM Ca(2+), i.e. under conditions of AMPAR saturation, thus indicating an effect of postsynaptic origin. In 0.5 mM Ca(2+) the decay of CF EPSCs was longer but it was also significantly prolonged (p?0.01) by CPCCOEt. However, the CF EPSC amplitude was not significantly affected indicating an underlying Ca(2+)-dependent mechanism. Thus, the pharmacology of the PC mGluR-mediated response points to a dual postsynaptic role of mGluR1 giving rise to a slow postsynaptic current but also regulating other presumably mGluR-dependent currents via second messenger molecules and Ca(2+). The additional electrophysiological role of mGluR II & III types was also indicated. Such a complex regulatory mechanism may have an important role in the mGluR-dependent forms of homosynaptic plasticity and motor learning at the CF-PC synapse