P2Y4/TSP-1/TGF-β1/pSmad2/3 pathway contributes to acute generalized seizures induced by kainic acid

Abstract Epilepsy is accompanied by angiogenesis and blood-brain barrier (BBB) disruption. The transforming growth factor-β1 (TGF-β1)/phosphorylated small mothers against decapentaplegic 2 and 3 (pSmad2/3)/vascular endothelial growth factor (VEGF) pathway, activated by thrombospondin-1 (TSP-1), which is further regulated by Y type P2 purinergic receptor activity, may participate in angiogenesis. We sought to investigate the relationship between the P2R/TSP-1/TGF-β1/pSmad2/3/VEGF pathway, angiogenesis, and BBB damage in a kainic acid (KA) model of acute generalized seizure. Our results demonstrated that KA-induced seizures were accompanied by angiogenesis and BBB damage. In addition, expression of TSP-1, TGF-β1, and pSmad2/3 was increased. Rats treated with pyridoxal phosphate-6-azophenyl-2′, 4′-disulfonic acid, a broad P2 receptor antagonist, or Reactive Blue 2, a potent P2Y4 receptor antagonist, showed significant attenuation of TSP-1 expression and Smad2/3 phosphorylation levels. Furthermore, angiogenesis, BBB damage, and acute seizure severity were also reduced. The inhibition of TSP-1 expression by siRNA or TGF-β1 activation by Leu-Ser-Lys-Leu (LSKL) treatment prevented KA-induced phosphorylation of Smad2/3, angiogenesis, BBB damage, and acute seizures. Our results strongly indicate that the P2Y4/TSP-1/TGF-β1/pSmad2/3/VEGF pathway plays an essential role in seizure pathophysiology and angiogenesis. Therapeutic interventions targeting this pathway may offer new treatment options for acute seizures.