Role of adenosine receptors in synapse stabilization – ADONIS

This project takes advantage of the complementary skills of the different scientific leaders of the project. We have designed a multidisciplinary approach, spanning different levels of analysis (from molecule dynamics to synaptic currents), using cellular and molecular neuroanatomy, electrophysiology, and state-of-the-art techniques such as super-resolution Stochastic Optical Reconstruction Microscopy (STORM) microscopy, Quantum Dot-based Single Particle Tracking, optogenetics, two photon imaging and live video-microscopy of synaptic components.

These last 18 months, we have accumulated a number of evidences in favor of our hypothesis that the adenosine signaling through A2A receptors stabilize nascent synapses during the development of the central nervous system.
We have shown that the acute blockade of A2A receptors with a selective antagonist reduces the GABAergic functional synaptic transmission in hippocampal neurons. This reduction in synapse efficacy involves a rapid escape of GABAA receptors from synapses due to the disappearance of the main scaffolding molecule gephyrin. This destabilization of the postsynaptic element may lead to the detachment of the presynaptic element and to synapse collapse. The underlying mechanism if this destabilization will involve gephyrin dephosphorylation following a lack of activation of A2A receptor/adenyl cyclase and AMPc signaling pathway. The rapidity of the dislocation of the synapse implies a destabilization of the nascent synapses rather than a preclusion to form new synapses. The fact that the expression of the A2A receptor parallels the period of synaptogenesis and that the antagonist acts only during this critical period is also in favor of a role of this signaling pathway during the critical period of synapse stabilization.

We now need to determine whether the activity of the postsynaptic A2A receptor is enough to stabilize the synapse and to find the trans-synaptic partners of this regulation. We will also need to identify the source of adenosine (ATO-derived or direct release of adenosine, neuronal vs glial origin). Our preliminary data suggest that ATP itself may stabilize synapses through a distinct mechanism than A2A receptors. We will therefore evaluate the contribution of these two pathways in stabilizing synapses. In order to better understand the implication of these two pathways on brain development, one need to explore the impact of these pathways on glutamatergic excitatory synapses. Most of these experiments will be carried out in vitro and ex vivo. Finally, we will explore the impact of a dysregulation of these pathways on synaptogenesis in vivo and on animal behavior

This project led so far to several participations in national (1) and international (3) meetings. A first article is being written.

Adenosine, a degradation product of ATP, is a prototypical paracrine signal. In the brain, it dually controls neurotransmitter release mainly through activation of inhibitory A1 and facilitatory A2A receptors (A2AR). The function of these receptors in the mature brain is well established, but their role in the developing brain remains to be fully established. We have recently shown that A2ARs control the migration of some types of GABA neurons at early stages of development. Such mechanism allowed to us to demonstrate the deleterious consequences of caffeine (a natural A2AR antagonist) consumption during pregnancy on the foetal brain. While performing this study, we also discovered that A2ARs control the stability of GABAergic synapses. This observation constitutes the angular stone of our project.
Indeed, we found that A2AR blockade profoundly reduced the efficacy of type A GABA receptors (GABAAR)-mediated inhibitory synaptic transmission and the stabilization of GABAAR and its main scaffolding molecule gephyrin in the postsynaptic membrane. These results led us to propose that during development postsynaptic A2ARs transiently act as sensors of presynaptic GABAergic terminal activity. The release of ATP via the presynaptic terminal and/or glial cells, its transformation into adenosine in the cleft, and the subsequent activation of postsynaptic A2AR, via its signaling pathways, would stabilize the newly formed inhibitory synapse. A lack of A2AR activation would trigger synapse removal after a critical time. This, postsynaptic A2ARs would act as sensors of presynaptic activity, informing the postsynaptic compartment to maintain the synapse. The tasks of ADONIS are to validate this hypothesis and explore the A2AR-dependent mechanisms of synapse stabilization and removal during development.
Task 1 will explore when the A2AR-dependent mechanism is operant. This task will inform us about the developmental time window during which A2ARs are important for maintaining GABAergic synapses and the critical time window during which the lack of A2AR signaling triggers synapse removal. This task will also explore which steps of synapse development (formation, stabilization and elimination of newly formed synapses) are under the control of the A2AR-dependent mechanism.
Task 2 will be devoted to the molecular mechanisms of synapse stabilization/removal. In this task we will determine the mechanisms by which A2ARs control synaptic stabilization/removal.
Task 3 will enable us to identify the source of the adenosine that activates A2ARs and will validate the hypothesis that the adenosine-A2AR signaling acts as an activity-sensor to stabilize newly formed active synapses.
To address these questions, we have designed a multidisciplinary approach, spanning different levels of analysis (from molecule dynamics to synaptic currents), using cellular and molecular neuroanatomy, electrophysiology, and state-of-the-art techniques such as super-resolution Stochastic Optical Reconstruction Microscopy (STORM) microscopy, Quantum Dot-based Single Particle Tracking, optogenetics, two photon imaging and live video-microscopy of synaptic components.
The consortium gathers three leading experts in their respective fields, including a foreign partner. ADONIS is balanced between low-risk and high-risk/high-pay off tasks.