Modulation of synaptic transmission using muscarinic and GABA-A receptor ligands in neuronal cultures using a high capacity assay

Poster Neuroscience 2017

Date: Nov 11, 2017
Presentation Time: 1:00 pm – 2:00 pm
Presentation Number: 039.09
Poster Board Number: D58

Abstract
Modulators of synaptic transmission, acting at various molecular targets, have great potential for development as novel therapeutic agents in an extensive range of CNS diseases, and the search for novel specific pharmacological agents is of widespread interest. However, traditional high throughput screening approaches primarily rely on heterologous expression systems which lack the ability to replicate synaptic complexity. Thus there is a need to develop screening systems that can evaluate compounds on measures of synaptic transmission.

Here, in a 96-well format, we used Cellaxess Elektra® electric field stimulation (EFS) to excite cultured mouse cortical neurones loaded with the calcium fluorophore Calcium 5 in an imaging based microplate reader. By simultaneously electrically stimulating a discrete subset of the neurones in each well we recorded transient, synaptically mediated calcium elevations in neurones away from the stimulating field. These responses were mediated by AMPA and NMDA receptors, as evidenced by concentration dependent block by NBQX and MK801, respectively. We have studied the effects of compounds acting at either muscarinic receptors or GABA-A receptors as examples of agents that can modulate synaptic transmission via G-protein coupled receptors (GPCRs) or ligand-gated ion channels (LGICs).

Addition of the broad-spectrum muscarinic agonist carbachol to the cultures resulted in a concentration-related reduction of synaptic transmission, with an EC50 of 2.2 µM – a value that is similar to the EC50 observed for the reduction of synaptic transmission in the CA1 area of the mouse hippocampal slice. The effect of carbachol could be reduced by the selective M1 receptor antagonist VU0255035. In separate experiments, addition of the GABA-A agonist muscimol or the GABA-A modulator allopregnanolone resulted in concentration-related reductions of synaptic transmission, with EC50 of 0.5 µM and 0.1 µM respectively. These results show that it is possible to evaluate the modulation of synaptic transmission in cultured neurones using a 96-well plate format and that this produces results comparable to those obtained in electrophysiological studies. Furthermore, this approach can be successfully used in studies of agents acting at GPCRs and LGICs.

Authors: J. SVENSSON DALÉN¹, C. LINDWALL-BLOM¹, Å. JÄGERVALL¹, M. KARLSSON¹, P. KARILA¹, S. A. NEALE², T. E. SALT²;
¹Cellectricon AB, Mölndal, Sweden; ²Neurexpert Ltd, London, United Kingdom

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