Thrombin‐induced reversal of astrocyte stellation is mediated by activation of protein kinase C β‐1

Exogenous or endogenous injuries of the central nervous system trigger astrogliosis characterized by proliferation of astrocytes and changes in their morphology from stellate to flat polygonal. Astrocytes in culture are very sensitive to thrombin, a serine protease, which through its proteolytically activated receptor (PAR‐1) induces proliferation and morphological changes comparable to astrogliosis. Evaluation of the thrombin signal‐transduction pathway in the reversal of astrocyte stellation might help to understand astrogliosis. For this purpose, primary cultured murine cortical astrocytes were treated with H7, a protein‐kinase inhibitor, and thrombin, which resulted in an inhibition of stellation reversal. Treatments with phorbol 12‐myristate 13‐acetate (PMA), a protein kinase C (PKC) activator, mimicked the action of thrombin. Subsequently, direct assay of astrocyte PKC activity after thrombin or PMA treatment demonstrated involvement of PKC in thrombin signaling associated with shape change. Western blotting showed that PKC isoform β‐1 was involved in this pathway, while PKC α was only weakly activated and PKC β‐2 was not activated by thrombin. PKC β‐1 translocation was also elicited by a thrombin‐receptor active peptide (SFLLRN), demonstrating the involvement of PAR‐1 in this process. PKC δ and ε were located constitutively in the membrane fraction in stellate astrocytes. Isoforms γ, η, θ, and ζ were absent from astrocytes. These results suggest that astrogliosis in vivo might be regulated by modulating the activity of thrombin, PAR‐1, or specific PKC isoforms.