Cytoskeletal regulation of the L-arginine/NO pathway in pulmonary artery endothelial cells

We investigated possible involvement of the actin cytoskeleton in the regul ation of the L-arginine/nitric oxide (NO) pathway in pulmonary artery endot helial cells (PAEC). We exposed cultured PAEC to swinholide A (Swinh), whic h severs actin microfilaments, or jasplakinolide (Jasp), which stabilizes a ctin filaments and promotes actin polymerization, or both. After treatment, the state of the actin cytoskeleton, L-arginine uptake mediated by the cat ionic amino acid transporter-1 (CAT-1), Ca2+/calmodulin-dependent (endothel ial) NO synthase (eNOS) activity and content, and NO production were examin ed. Jasp (50-100 nM, 2 h treatment) induced a reversible activation of L-[H -3] arginine uptake by PAEC, whereas Swinh (10-50 nM) decreased L-[H-3] arg inine uptake. The two drugs could abrogate the effect of each other on L-[H -3] arginine uptake. The effects of both drugs on L-[H-3] arginine transpor t were not related to changes in expression of CAT-1 transporters. Swinh (5 0 nM, 2 h) and Jasp (100 nM, 2 h) did not change eNOS activities and conten ts in PAEC. Detection of NO in PAEC by the fluorescent probe 4,5-diaminoflu orescein diacetate showed that Swinh (50 nM) decreased and Jasp (100 nM) in creased NO production by PAEC. The stimulatory effect of Jasp on NO product ion was dependent on the availability of extracellular L-arginine. Our resu lts indicate that the state of actin microfilaments in PAEC regulates L- ar ginine transport and that this regulation can affect NO production by PAEC.