Intrinsic Activity, 2013; 1(Suppl.1)

withdrawn before publication A1.3 Activation of kappa opioid receptors reduces seizure activity in a mouse model of temporal lobe epilepsy Luca Zangrandi and Christoph Schwarzer* Department of Pharmacology, Innsbruck Medical University, Austria *E-mail: [email protected] Intrinsic Activity, 2013; 1(Suppl. 1): A1.3 Background: Neuropsychiatric disorders are one of the main challenges of medicine with epilepsies representing some of the most frequent. Temporal lobe epilepsy (TLE) is the most common type of epilepsies and is often accompanied by marked neuronal degeneration. It was shown that the deletion of prodynorphin in mice and low expression in humans is associated with increased epilepsy vulnerability. Dynorphin targets opioid receptors and in particular the kappa opioid receptor (KOP). KOP receptors are located in strategically ideal places to control hippocampal excitability also in chronic TLE. The aim of this study was to investigate the potential of KOP receptor agonists as antiepileptic drugs and compare it to new generation of anti-epileptic drugs (AEDs) in a mouse model of drug-resistant TLE. Methods: Fifteen C57BL/6N male mice were injected unilaterally with saline (n = 5) or kainic acid (KA; 1 nmol in 50 nl saline; n = 10) into dorsal hippocampus. While saline-injected animals did not show any signs of EEG or histopathological alterations, KA caused acute and delayed behavioral, pathological and EEG effects. Four-channel EEG traces were recorded from ipsiand controlateral hippocampi and motorcortices applying depth and surface electrodes from freely moving mice, respectively. The KOP receptor-specific agonist U-50488H, saline, or one the new AEDs oxcarbazepine, lamotrigine and levetiracetam were applied i.p. at different doses. Number and duration of EEG seizures were automatically evaluated (Sciworks software) for 45 min preceding and following the injections. Results: Spike trains, sharp waves and paroxysmal discharges in the ipsilateral hippocampus were observed starting from about 14 days after KA injection. Prolonged EEG seizures occurred frequently after about 4 weeks. Such paroxysmal discharges were accompanied by behavioural arrest and stereotypic behaviour like head nodding. Application of KOP receptor agonists decreased EEG abnormalities caused by KA up to 2 hours in a dosedependent manner, whereas the AEDs didn’t show any significant effect. Animals treated with U-50488H were awake during the time of recordings, unlike those treated with diazepam. Discussion: Our data demonstrate an anticonvulsant action of KOP receptor agonists in the chronic phase of epilepsy, comparable to the effect of 2.5 mg/kg diazepam. Acknowledgements: This work was supported by the Austrian Science Fund FWF grant W1206-BO5. A1.4 Galanin-receptor-3-deficient male mice exhibit anxiety-like phenotype Susanne Brunner, Aitak Farzi, Felix Locker, Sabine Ebner, Barbara Holub, Andreas Lang, Johannes Mayr, Roland Lang, Peter Holzer and Barbara Kofler* Laura Bassi Centre of Expertise – THERAPEP, Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Paracelsus Medical University, Salzburg, Austria; Research Unit Translational Neurogastroenterology, Institute of Experimental and Clinical Pharmacology, Medical University of Graz, Austria; Department of Dermatology, Paracelsus Medical University, Salzburg, Austria *E-mail: [email protected] Intrinsic Activity, 2013; 1(Suppl. 1): A1.4 Background: The neuropeptide galanin (GAL) has a wide distribution in the central and peripheral nervous system and is a modulator of various physiological and pathological processes. The GAL receptors, GAL1, GAL2, and GAL3, are all G protein-coupled receptors. They show substantial differences in functional coupling and subsequent signaling activities contributing to the diversity of physiological effects of GAL. A role for GAL as modulator of mood and anxiety was suggested as GAL and its receptors are highly expressed in limbic brain structures of rodents. Injections of GAL into the ventricular system of the brain induce anxiolytic-like effects in Vogel’s punished drinking test. Furthermore, increased anxietylike behavior in the elevated plus maze (EPM) was reported for GAL1and GAL2-receptor-deficient mice. However, up to date there is sparse literature implicating GAL3 receptors in behavioral functions. Therefore, we measured behavior of GAL3-receptordeficient mice (GalR3-KO), to elucidate whether the GAL3 receptor is involved in mediating behavior-associated actions of GAL. Methods: Disruption of the GAL3 receptor locus was achieved by homologous recombination with targeting both coding exons, and mice were maintained on a C57BL/6 background. Results: The GalR3-KO mouse line exhibits normal breeding and physical development. Hematology and amino acid profile from serum and whole brain of male GalR3-KO mice were not different from wild-type (WT) littermates. In contrast to WT littermates male GalR3-KO mice exhibited an anxiety-like phenotype in the EPM, open field (OF) and light-dark (L/D) test. In the EPM, GalR3-KO mice spent significantly less time on the open arms and, conversely, significantly more time on the closed arms. In the OF test, time spent in the central area and number of visits to this area were significantly reduced in GAL3-receptor-deficient mice, whereas the number of fecal boli shed was significantly enhanced compared to WT animals. GalR3-KO mice spent significantly less time in the light compartment of the L/D test and also displayed significantly decreased locomotion in the light compartment. Furthermore, GalR3-KO mice were less socially affiliated to a stranger mouse than WT animals in the social interaction test. Discussion: Our data suggest an involvement of the GAL3 receptor on GAL-mediated effects on mood, anxiety and behaviour, making it a possible target for new treatment strategies of mood disorders. Additionally, similar anxiety-related results for GAL1 and GAL2 receptors hint at a possible heterodimerzation of GAL receptors in the context of mediating anxiety and related behaviors. A1.5 Expression and function of the skeletal muscle calcium channel splice variant CaV1.1∆E29 Nasreen Sultana, Gerald J. Obermair, Ariane Benedetti, Petronel Tuluc and Bernhard E. Flucher* Intrinsic Activity, 2013; 1(Suppl. 1) published online: 1 October 2013 http://www.intrinsicactivity.org © 2013 Intrinsic Activity, ISSN 2309-8503; Austrian Pharmacological Society (APHAR) page 3 of 42 (not for citation purpose) Department of Physiology, Innsbruck Medical University, Austria; Department of Pharmacology, University of Innsbruck, Austria *E-mail: [email protected] Intrinsic Activity, 2013; 1(Suppl. 1): A1.5 Background: The CaV1.1 voltage-gated calcium channel is the voltage sensor in skeletal muscle excitation–contraction coupling. Its primary role is to activate gating of the calcium release channel (type 1 ryanodine receptor) in the sarcoplasmic reticulum. Recently, our lab discovered a CaV1.1 splice variant with increased voltage sensitivity and current amplitude. The splice variant lacking exon 29 (CaV1.1∆E29) is expressed at low levels in adult muscle, but it is the dominant isoform in embryonic muscle. A recent study showed the presence of CaV1.1∆E29 in patients with muscle weakness and the requirement of the CaV1.1 channel in neuromuscular junction development. Methods: In order to reveal the role of CaV1.1∆E29 in developing muscle and its possible involvement in muscle disease we generated a mouse model where we specifically knocked out exon 29. Quantitative real-time PCR analysis of specific muscle types at different developmental stages was performed to analyze the expression levels of both splice variants (CaV1.1 and CaV1.1∆E29). Phenotypic characterization to analyze muscle strength and motor activity was done using home-cage activity, rotarod and treadmill. In order to study the development and patterning of neuromuscular junctions (NMJs) we stained diaphragms isolated from mouse embryos (E14) with α-bungarotoxin conjugated with Alexa 488. Results: Quantitative real-time PCR analysis of specific muscle types at different developmental stages indicate the possible requirement of CaV1.1∆E29 in prenatal development of muscle as postnatal development was associated with a rapid decrease of CaV1.1∆E29 paralleled by a strong increase of CaV1.1. In contrast to a published report showing upregulation of CaV1.1∆E29 in patients with muscle weakness, exclusive expression of CaV1.1∆E29 in exon 29 knockout mice did not result in significant muscle weakness or altered motor activity examined with a battery of behavioral tests. Although published data also suggest the requirement of CaV1.1 in neuromuscular junction development, preliminary results indicate that lack of the long CaV1.1 isoform does not result in aberrations of acetylcholine receptor prepatterning in embryonic mouse diaphragm. Discussion: So far our experiments did not reveal effects of the exclusive expression of the embryonic CaV1.1∆E29 on the normal development and function of the mature muscle. Ongoing experiments on single muscle fibers are expected to expose potential effects of increased calcium influx on excitation– contraction coupling. Further analysis of older and damaged muscle will indicate an involvement of CaV1.1∆E29 in disease or reveal potential compensatory mechanisms in the knockout mice. Acknowledgements: FWF P23479, W1101. A1.6 Asymmetry in the song of crickets: preferences of females and proximate mechanism of discrimination Heiner Römer and Stefan Hirtenlehner* Institute of Zoology, Karl-Franzens University of Graz, Austria *E-mail: [email protected] Intrinsic Activity, 2013; 1(Suppl. 1): A1.6 Background: Subtle random deviations from perfect symmetry in bilateral traits are suggested to signal reduced phenotypic and genetic quality of a sender, but only little is known about the related receiver mechanisms for discriminating symmetrical from asymmetrical traits. We investigated these mechanisms in behavioural and neurophysiological experiments in the Mediterranean field cricket, Gryllus bimaculatus. A downward frequency modulation at the end of each syllable in the calling song has been suggested to indicate morphological asymmetry in sound radiating structures between left and right forewings. Methods: Phonotaxis of female crickets was quantified on a trackball system in two-choice situations. Under identical stimulus paradigms, the discharge of the pair of AN1 neurons was recorded, known for its importance in female phonotaxis. Results: Even under ideal laboratory conditions on a trackball system, female crickets only discriminated between songs of symmetrical and asymmetrical males in two-choice experiments at carrier frequencies of 4.4 kHz and large modulation depth of 600 and 800 Hz. Under these conditions they preferred the pure tone calling songs over the modulated (asymmetrical) alternative, whereas no preference was observed at carrier frequencies of 4.9 and 5.2 kHz. These preferences correlate well with responses of a pair of identified auditory interneurons (AN1). The interneuron is tuned to an average frequency of 4.9 kHz, and the roll-off towards lower and higher frequencies determines the magnitude of responses to pure tone and frequency-modulated calling songs. The difference in response magnitude between the two neurons appears to drive the decision of females towards the song alternatives. Discussion: It is unlikely that song differences based on asymmetry in the morphology of song-producing structures play an essential role under natural conditions. Acknowledgements: The research was funded by the Austrian Science Fund (FWF): P20882-B09. A1.7 Levels of α5-containing nicotinic receptors in animal models of neuropathic pain Bogdan Ianosi, Dimitris Xanthos, Johannes Beiersdorf, Ariane Thrun, Sigismund Huck and Petra Scholze* Department of Pathobiology of the Nervous System, Center for Brain Research, Medical University of Vienna, Austria; Department of Neurophysiology, Center for Brain Research, Medical University