Figure 4.
Contraction agonists activate LIMK2 in canine PASMCs. Canine PASMCs were incubated with thrombin (Thr), norepinephrine (NE), endothelin-1 (ET1), angiotensin II (AngII), and 5-hydroxytryptamine (5-HT) for 5 min. LIMK2 was immunoprecipitated from total protein homogenates and kinase activity was assayed by the phosphorylation of recombinant CF in the presence of [γ-32 P]ATP in vitro. Reaction protein was resolved by SDS-PAGE, gels were stained with Coomassie brilliant blue and then exposed on phosphorimaging screens. CF radioactive bands (top inset) were normalized to protein bands (lower inset) and the agonist-mediated activation of LIMK2 was expressed as percentage of the activity of untreated cell controls (Ctr, bar graph). Mean ± SD, (∗) P < .05 compared to untreated controls, n = 3.