SYNAPTIC CHANGES IN GLUN2A KNOCKDOWN OF MATURE PRIMARY NEURONAL CULTURES

SALVETTI, ANNA; BAEZ, MARÍA VERÓNICA; VAZQUEZ, CECILIA; JERUSALINSKY, DIANA ALICIA; ACUTAIN, M. FLORENCIA

Mar del Plata

Congreso; Reunión Conjunta de la Sociedad Argentina de Investigación Clínica (SAIC), la Sociedad Argentina de Inmunología (SAI) y la Sociedad Argentina de Fisiología (SAFIS); 2018

NMDA Receptors (NMDAR) are glutamatergicreceptors involved in synaptic plasticity, learning and memory processes aswell as in several neuropathologies. NMDAR are composed by two GluN1 obligatorysubunits and two regulatory subunits: GluN2 (A-D) or GluN3 (A-B). In memoryrelated brain structures, like the hippocampus, GluN2A and GluN2B are the mostexpressed regulatory subunits. Transcription and translation of both subunits aretightly regulated; while GluN2B expression is characteristic of immaturesynapses, GluN2A is present in mature and stable synapses. However, changes inGluN2A subunit expression has been shown in some synaptopathologies. In orderto better understand the role of GluN2A in synapsis, we built two AAV-eGFPvectors: one codifying a specific shRNA anti GluN2A (AAV-sh2A), and the othercarrying a shRNA scramble as control (AAV-shSc). In mature primary neuronalcultures transduced with AAV-sh2A or AAV-shSc we analyzed dendritic spines andalso the expression of two synaptic proteins: Syn-1 and PSD95. As was expected,we observed by qPCR a decrease in GluN2A mRNA only in primary cultures transducedwith AAVsh2A, without modifications in the other NMDAR subunits expression.Interestingly in those cultures where GluN2A was knockdown, we saw asignificant decrease in GluN1 protein, while GluN2B protein levels did not change.Furthermore, in those cultures we found an increase in dendritic spines number,at expenses of immature spines. In addition, the expression of Syn-1 and PSD95 wasraised up in GluN2A knockdown cultures. These results suggest that GluN2Adecreased expression seems to induce a rise in the synaptic spines, that wouldprovoke changes in neuronal architecture and the return of the system to animmature state. All these observed changes could help to explain, at least inpart, the molecular pathway of some variants of epileptic disease related toGluN2A mutations.