title: Mechanical lesion activates newly identified NFATc1 in primary astrocytes: implication of ATP and purinergic receptors.
abstract: Ca2+-dependent calcineurin is upregulated in reactive astrocytes in neuroinflammatory models. Therefore, the fact that the nuclear factor of activated T cells (NFAT) is activated in response to calcineurin qualifies this family of transcription factors with immune functions as candidates to mediate astrogliosis. Brain trauma induces a neuroinflammatory state in which ATP is released from astrocytes, stimulating calcium signalling. Our goal here is to characterize NFATc1 and NFATc2 in mouse primary astrocyte cultures, also exploring the implication of NFAT in astrocyte activation by mechanical lesion. Quantitative reverse transcriptase-polymerase chain reaction, Western blot analysis and immunofluorescence microscopy identified NFATc1 in astrocytes, but not NFATc2. Moreover, NFATc1 was expressed in the cytosol of resting astrocytes, whereas activation of the Ca2+-calcineurin pathway by ionomycin translocated NFATc1 to the nucleus, which is a requirement for activation. The implication of astrocytic NFAT in brain trauma was analysed using an in vitro scratch lesion model. Mechanical lesion caused a rapid NFATc1 translocation that progressed throughout the culture as a gradient and was maintained for at least 4 h. We also demonstrate that ATP, released by lesion, is a potent inducer of NFATc1 translocation and activation. Moreover, the use of P2Y receptor modulators showed that such ATP action is mediated by stimulation of several G(q)-protein-coupled P2Y purinergic receptors, among which P2Y(1) and P2Y(6) are included. In conclusion, this work provides evidence that newly identified NFATc1 is translocated in astrocytes in response to lesion following a pathway that involves ATP release and activation of metabotropic purinergic receptors.
authors.affiliation: Department of Medical Sciences, University of Castilla-La Mancha, Av. Almansa 14, Albacete 02006, Spain.
authors.forename: José M
authors.initials: JM
authors.lastname: Pérez-Ortiz
authors.forename: María C
authors.initials: MC
authors.lastname: Serrano-Pérez
authors.forename: María D
authors.initials: MD
authors.lastname: Pastor
authors.forename: Eduardo D
authors.initials: ED
authors.lastname: Martín
authors.forename: Soledad
authors.initials: S
authors.lastname: Calvo
authors.forename: Mercedes
authors.initials: M
authors.lastname: Rincón
authors.forename: Pedro
authors.initials: P
authors.lastname: Tranque
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001253:Astrocytes / Q000378:metabolism* / Q000473:pathology; D015153:Blotting, Western; D002478:Cells, Cultured; D003593:Cytoplasm / Q000737:chemistry / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D005455:Fluorescent Antibody Technique; D051379:Mice; D008810:Mice, Inbred C57BL; D050778:NFATC Transcription Factors / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D011983:Receptors, Purinergic / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
chemical_list: D050778:NFATC Transcription Factors; C494005:Nfatc1 protein, mouse; D011983:Receptors, Purinergic; D000255:Adenosine Triphosphate