Exploração do sistema de modulação operado pela adenosina para controlar o desempenho cognitivo e a doença de Alzheimer

The scientific and translational aims of the project are as follows: 1) Define the hippocampal brain circuits where A2AR overfunction is critical to control dysfunction: this will be tackled in the two mouse models of AD, combining in vivo and in vitro approaches; first, we will inject a selective A2AR antagonist [6] in different hippocampal fields of AD mouse models (both sexes) to define in which circuits A2AR overfunction critically controls memory dysfunction in different reference memory tests (MWM-Morris water maze, OD-object displacement); we will then resort to slice electrophysiology to detail where in the circuit A2AR play a key role [4,7,8,15]. 2) Define which phases of reference memory are controlled by A2AR in AD mouse models; we will resort to a photoactivable A2AR antagonist (MRS7145), which will be light activated [21] in the previously identified circuits either during acquisition or consolidation or retrieval or extinction of reference memory in a MWM test; this will be confirmed using a spatiotemporally-controlled optoA2AR stimulation in each memory phase [5,22]. 3) Define in control mice the spatiotemporal dynamics of extracellular adenosine (and ATP) levels in different cell compartments during the different phases of memory performance, combining the use of new GRAB sensors [23,24] and cell-conditional knockouts of different sources of extracellular adenosine, based on viral-mediated CRE-expression under cell-specific promoters in floxed-ENT-2, -CD73 or -VNUT mice [15,25]. 4) Define the dynamics and source of the excessive adenosine extracellular levels in the afflicted hippocampal circuits during the phase(s) of memory controlled by A2AR in AD mouse models; this will then be confirmed in slice electrophysiology experiments and will be the basis to design novel combination strategies targeting A2AR and sources of excessive formation of extracellular adenosine; additionally, we will test if particular AD-related A2AR polymorphisms [26,27] are associated with the age of onset and/or progression of different AD markers using the CUCH data bank, which will prompt A2AR polymorphic analysis as biomarker of disease progression as a new potential start-up/service to clinics. 5) Define the form of A2AR present in the circuit related with AD-associated memory deficits in AD, using first a mass-spectrometry interactomics analysis (Fig.2 of Preliminary data), followed by co-immunoprecipitations and receptor binding studies [28]; this will be carried out in control mice, AD mice and then in hippocampal tissue of different circuits dissected during autopsy from the brain of adults and aged individuals; we will then test the selectivity of novel compounds (different families and bi-functional A2AR-based compounds). The main training objectives are to form a PhD using this golden opportunity to train and hook to science both MD students and science-Master students. The collaborative and translational aims are to develop novel drug strategies (new families, bifunctional or combinations) to selectively interfere with A2AR controlling memory deficits, with a define posology schedule. All this will be done with long-term chemistry collaborators with care to secure our share of intellectual property emerging from this project. All these activities will support an active communication based on open publications and engagement with the general public to increase science culture and awareness (i.e. citizenship).