Modulation of neurotransmitter release by cannabinoid receptors at individual cortical synapses

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.

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.