Modulation of neurotransmitter release by cannabinoid receptors at individual cortical synapses

Lead Research Organisation: University College London
Department Name: Institute of Neurology

Abstract

Cannabis is a very widely used drug, which has many well-documented deleterious effects on mental and physical health. In recent years it has emerged that the active constitutent hijacks a natural form of signalling between neurons, which acts in the opposite direction to the normal flow of information in the brain: in response to activity in one neuron, a signal is sent back to synapses that normally release chemical messengers to that same neuron. The result of this is that the strength of communication is diminished. How this happens is poorly understood. We have developed the ability to examine the actions of both cannabis derivatives and naturally occurring chemicals in the brain on the behaviour of individual synapses at unprecedented resolution. We expect that the results obtained with this cutting-edge methodology will shed light both on the normal workings of the brain, and on disorders of brain excitability such as epilepsy, and ultimately reveal previously unknown targets for pharmacological intervention.

Technical Summary

Ca2+-dependent release of neurotransmitters underlies fast information flow in the brain. This release can be strongly modulated by retrograde signalling from the postsynaptic neuron to the presynaptic terminal. Although CB1 cannabinoid receptors are implicated in this modulation, its physiological and/or pathological triggers are incompletely understood, as is the molecular machinery that enables a change in Ca2+-dependent exocytosis. Our aims are, first, to establish the molecular mechanisms that underlie the actions of presynaptic cannabinoid receptors on neurotransmitter release at the level of individual cortical synapses; second, to elucidate the physiological consequences of such effects, and their spatiotemporal extent, in the synaptic circuitry; and third, to understand the relationship between these mechanisms and disorders of circuit excitability, in particular those associated with seizures. We will apply two-photon excitation microscopy combined with patch-clamp electrophysiology and optical quantal analysis in acute brain slices. The experimental protocols are in place, and we have obtained substantial pilot data, including the first evidence that endocannabinoids alter presynaptic Ca2+ signalling and probabilistic neurotransmitter release at individual hippocampal synapses. The proposed research will provide novel insights into the fundamental mechanisms of cannabinoid receptor actions in the brain at the highest resolution in live tissue available to date.

Publications

10 25 50
 
Description ANR, National Awarding Panel Member (France)
Geographic Reach Asia 
Policy Influence Type Participation in advisory committee
 
Description Chair, Physiology and Medicine Section, Academia Europaea
Geographic Reach Asia 
Policy Influence Type Implementation circular/rapid advice/letter to e.g. Ministry of Health
Impact Influenced academic science policies and award criteria in Europe.
 
Description Member Academia Europaea
Geographic Reach Asia 
Policy Influence Type Participation in advisory committee
 
Description National ANR Awarding Panel Member (France)
Geographic Reach Asia 
Policy Influence Type Participation in advisory committee
 
Description l'AERES (Research and Higher Education Evaluation Agency, France) Review Committee on Institute of Neuroscience (Bordeaux, 2009)
Geographic Reach Asia 
Policy Influence Type Influenced training of practitioners or researchers
Impact Influenced the strategy of a major biomedical Research Institution
 
Description l'AERES Review Committee on Laboratoire de Physiologie Cérébrale (Universite Paris Decartes, 2013)
Geographic Reach Asia 
Policy Influence Type Influenced training of practitioners or researchers
Impact Influenced the research strategy of a major biomedical research Institution.
 
Description ERC Advanced Grant
Amount € 2,496,000 (EUR)
Funding ID 323113 
Organisation European Research Council (ERC) 
Sector Public
Country Belgium
Start 06/2013 
End 05/2018
 
Description EU Integrated Project FP6
Amount € 495,000 (EUR)
Funding ID 512012 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 04/2005 
End 03/2009
 
Description FP7 Initial Training Network
Amount € 250,000 (EUR)
Funding ID 606950 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 10/2013 
End 09/2016
 
Description UCL Excellence Fellowship for Dr Hennberger
Amount £250,000 (GBP)
Organisation University College London 
Sector Academic/University
Country United Kingdom
Start 09/2010 
End 02/2011
 
Description Wellcome Trust Enhancement Grant
Amount £102,612 (GBP)
Funding ID 5772 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 06/2012 
End 09/2013
 
Description Wellcome Trust Principal Fellowship
Amount £2,790,000 (GBP)
Funding ID 101896/Z/13/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2013 
End 09/2018
 
Title 64-core Parallel computing unit 
Description Parallel computing 64-core PC cluster has been bult to carry out Monter Carlo simulation of molecular diffusion 
Type Of Material Technology assay or reagent 
Year Produced 2010 
Provided To Others? Yes  
Impact Several publications have used the cluster 
 
Title FLIM monitoring of Ca2+ 
Description Using FLIM to monitor intracellular Ca2+ with nanomolar sensitivity 
Type Of Material Data analysis technique 
Year Produced 2012 
Provided To Others? Yes  
Impact papers in preparation 
 
Title Single-bouton recordings and imaging 
Description A technique to carry out patch-clamp recording and and simultaneously image Ca2+ in small axonal boutons 
Type Of Material Technology assay or reagent 
Year Produced 2006 
Provided To Others? Yes  
Impact Publications and new techniques 
 
Title The neurophysiological role of cannabinoids in cannabis users 
Description The neurophysiological role of cannbinoid in cannabis users 
Type Of Material Model of mechanisms or symptoms - in vitro 
Provided To Others? No  
Impact The finding could help to understand mechsnism of cannabis abuse 
 
Title Two-photon excitation T1117 probing 
Description The use of T1117 for GPR55 detection in live cells 
Type Of Material Physiological assessment or outcome measure 
Year Produced 2011 
Provided To Others? Yes  
Impact A new method to detect a previously undetectable receptors in live cells in organised brain tissue 
 
Title point-source line-scan diffusion measurement 
Description A new point-source line-scan imaging technique has been developed to assess extracellular molecular mobility (diffusivity) in brain tisssue. 
Type Of Material Technology assay or reagent 
Provided To Others? No  
Impact This technique has allowed measurements of extracellular molecular mobility in the hippocampus providing an important alternative to the only existing technique. 
 
Title A large scale biophysical computer model of synaptic microenvironment 
Description A large scale biophysical computer model of synaptic microenvironment is designed for Monte-Carlo simulations of rapid molecular evemts inside and outside the synaptic cleft, ion 3D brain tissue. 
Type Of Material Computer model/algorithm 
Year Produced 2010 
Provided To Others? Yes  
Impact High-impact publications. 
 
Title A large-scale computer model of hippocamapal neural networks 
Description A large-scale computer model of hippocamapal neural networks is designed for realistic modelling of inter-connected active neurons in teh hippocampus 
Type Of Material Computer model/algorithm 
Year Produced 2013 
Provided To Others? Yes  
Impact High-impact publications reflecting scientific discoveries. 
 
Title Concentration mapping algorithm for time-resolved fluoresence imaging 
Description Concentration mapping algorithm for time-resolved fluorescence imaging is designed to traslate FLIM data into a two or three-dimensional colour-coded map of fluorescence lifetime (pre-calibrated for free Ca2+ concentration). 
Type Of Material Data analysis technique 
Year Produced 2012 
Provided To Others? Yes  
Impact Publications being prepared. 
 
Description Fluctuations in release probability 
Organisation University College London
Department Institute of Neurology
Country United Kingdom 
Sector Academic/University 
PI Contribution Electrophysiological data
Collaborator Contribution Simulation data
Impact artilce in Nature Neuroscience
 
Description Fluorescence anisotropy imaging 
Organisation King's College London
Country United Kingdom 
Sector Academic/University 
PI Contribution We design and run experiments in acute brain slices
Collaborator Contribution Development of a novel method of diffusion measurement on a nano-scale
Impact To date, the outcome of the aforementioned collaboration could be summarised as follows: 1. A brand new methodology of fluorescence anisotropy imaging, which enables nano-scale diffusion and miscro-viscosity measurements in live tissue has been set up at the PI's laboratory. It is currently being tested for optics optimisation and single-cell imaging in acute brain slices. 2. The paper dealing with nano-diffusion measurements in the brain extracellular space is in its final preparation stages.
Start Year 2006
 
Description GABA signalling in neurogliaform cells 
Organisation University of Oxford
Department MRC Anatomical Neuropharmacology Unit
Country United Kingdom 
Sector Academic/University 
PI Contribution Specifci research ideas, experiments and designs
Collaborator Contribution Specific experiments
Impact Two papers in the Journal of Neuroscience
Start Year 2006
 
Description GPR55 collaboration 
Organisation University of Aberdeen
Department School of Medicine & Dentistry Aberdeen
Country United Kingdom 
Sector Academic/University 
PI Contribution Supply of GPR55 knockout mice
Collaborator Contribution Supply of GPR55 mice
Impact We have identified the role of GPR55 in central neural circuits
Start Year 2009
 
Description LTP and D-serine release from astrocytes 
Organisation National Institute of Health and Medical Research (INSERM)
Country France 
Sector Academic/University 
PI Contribution Dr Stephane Oliet, INSERM Bordeaux, France.
Collaborator Contribution Provided electropohysiological data
Impact Christian Henneberger, Thomas Papouin,3, Stéphane H. R. Oliet, & Dmitri A. Rusakov. Nature (in press).
Start Year 2006
 
Description Light- and filed-control microcapsules 
Organisation Queen Mary University of London
Country United Kingdom 
Sector Academic/University 
PI Contribution Carrying out imaging experiments in live brain tissue
Collaborator Contribution Design, production and supply of environment-controlled micro-capsules for drug delivery
Impact To be announced
Start Year 2009
 
Description NMDA receptor function 
Organisation University College London
Department Centre for Clinical Pharmacology & Therapeutics
Country United Kingdom 
Sector Academic/University 
PI Contribution Caried out imaging experiments
Collaborator Contribution Supplied mouse model
Impact Article in EMBO Journal
Start Year 2006
 
Title A large-scale Monte-Carlo model of synaptic environment 
Description A library of programs for Monte Carlo simulations of rapid molecular events inside and outside the synaptic cleft. Written by Drs L Savtchenko and K. Zheng. 
Type Of Technology Software 
Year Produced 2007 
Impact Several high-profile publications 
 
Title A large-scale computer model environment for exploring realistic neuronal networks 
Description A large-scale computer model environment for exploring realistic neuronal networks. Allows inclusion of >100,000 individual NEURON-type cells. Enabled for parallel computing on the cloud. 
Type Of Technology Software 
Year Produced 2013 
Impact High-impact publications reflecting some fundamental scientific observations. 
 
Title Dedicate 64-core computer cluster 
Description A dedicated in-house 64(+) computer cluster (parallel computing enabled) for biophysical and neural network modelling including computational environment. Designed and built by Dr K. Zheng. 
Type Of Technology e-Business Platform 
Year Produced 2008 
Impact High-profile publications reflecting several scientific discoveries. 
 
Title Fluorecence anisotropy imaging 
Description An image analysis tool allowing to translate two FLIM signals (parallel and perpendicular) into local rotation correlation parameter across the imaged region of interest. Applied to map local instantaneous molecular diffusion. 
Type Of Technology Software 
Year Produced 2010 
Impact Book chapter. Being implemented to gauge molecular diffusivity at nanoscale. 
 
Title Submillisecond ligand application probe with multiple solution exchange 
Description Sub-millisecond ligand application probe with multiple solution exchange enables application of receptor ligands to excised membrane patches of live cells with ~0.2 ms resolution and a 10s duty cycle for up to four solution exchanges in both theta-glass channels. Deisgned and built by Dr. S. Sylantiev. 
Type Of Technology Physical Model/Kit 
Year Produced 2009 
Impact High-profile publications reflecting several scientific discoveries. A dedicated Nature Protocols methods article. 
 
Description Henry Stewart Talks 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Open access on-line lecture

This is not a serious question or a serious example.
Year(s) Of Engagement Activity 2011,2012,2013,2014
 
Description International lectures at major research conferences and university deprtments 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talks were noted internationally.

This question and example are not serious.
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,2009,2010,2011,2012,2013,2014
 
Description Media coverage 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Editorial pieces in Nature, La Monde, The Neuroscientist, Scientist, Pour la Madicine

Large audience reached, high citation of the paper
Year(s) Of Engagement Activity 2010,2011,2012
 
Description Membership of international advisory bodies and reviewing panels: 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Decisions made

Influencing science policy and funding
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,2009,2010,2011,2012,2013,2014
 
Description Peer-reviewing for major funding bodies 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact N/A

Decision made
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,2009,2010,2011,2012,2013,2014
 
Description Peer-reviewing for top-tier science journals 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? Yes
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Publications

N/A
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,2009,2010,2011,2012,2013,2014