MAP-tau thr212 is dephosphorylated after agonist treatment in GSK3β^+/− platelets and in human platelets treated with GSK3 inhibitors. (A) Washed human platelets (4 × 10⁷) were incubated for the indicated times with no agonist (Untx) or SFLLRN (5 μM), lysed, and immunoblotted with antibody to phospho-GSK3β ser9, phospho-tau-thr231, or actin (the actin immunoblot shown is a reprobe of the immunoblot of phospho-GKS3β). Similar results have been obtained in 3 separate experiments. (B) Washed human platelets (4 × 10⁷) were incubated for 2 hours in Tyrode buffer (−) or 10 μM SB216763 (+), then treated with the indicated concentration of human TRAP SFLLRN (0, 1, or 5 μM) for 3 minutes. Platelets were lysed and immunoblotted with antibody to phospho-MAPtau-thr 231. The blot was stripped and reprobed with an antibody to total MAP-tau. Similar results have been obtained in 4 experiments. (C) Washed platelets (4 × 10⁷) from WT mice were incubated with varying concentrations of TRAP AYPGKF, lysed and immunoblottted with a phosphospecific antibody to MAP-tau thr 231 or GSK3β-ser9 (top blots). WT mouse platelets were incubated for the indicated times with 1 mM AYPFGF, lysed, and immunoblotted as above. The results shown are representative of 2 independent experiments each. (D) Washed platelets (4 × 10⁷) from WT or GSK3β^+/− mice were incubated with or without 1 mM AYPGKF, lysed, and immunoblottted with a phosphospecific antibody to MAP-tau thr 231. Blots were reprobed with an antibody to actin. A representative blot of 3 experiments is shown.