OBJECTIVE: Sphingosylphosphorylcholine (SPC) is an important lipid mediator that has been implicated in vascular disease. As it has not been studied in the pulmonary circulation, we examined its mechanisms of action in rat small intrapulmonary arteries (IPA). METHODS: IPA were mounted on a myograph for recording tension and intracellular Ca2+ concentration ([Ca2+]i). Ca2+ sensitisation was examined in alpha-toxin permeabilized IPA, and by Western blot analysis of MYPT1 phosphorylation. RESULTS: SPC induced a slow but powerful vasoconstriction in IPA associated with an elevation in [Ca2+]i, with an EC50 for vasoconstriction of 12+/-2 microM. Removal of extracellular Ca2+ increased the EC50 to 76+/-33 microM (p<0.01) and abolished the rise in [Ca2+]i. Endothelial denudation or inhibition of NO synthase with L-NAME enhanced vasoconstriction. Treatment with pertussis toxin or the PLC inhibitor U731223 had no effect on SPC-induced vasoconstriction. The Rho kinase inhibitor Y27632 reduced SPC-induced vasoconstriction by approximately 70% and abolished both SPC-induced Ca2+ sensitisation in permeabilized IPA and the associated increase in MYPT1 phosphorylation; Ca2+ sensitisation was substantially inhibited by GDPbetaS. La3+ and 2-APB, at concentrations previously shown to block capacitative Ca2+ entry in IPA, suppressed SPC-induced vasoconstriction to the same extent as removal of extracellular Ca2+; residual tension was abolished by Y27632. Diltiazem was relatively ineffective. 2-APB also abolished the SPC-induced rise in [Ca2+]i. However, treatment with thapsigargin to empty intracellular stores had no effect on the elevation of [Ca2+]i induced by SPC. CONCLUSION: We present evidence that SPC is a powerful vasoconstrictor of IPA and the novel finding that SPC-induced vasoconstriction in IPA is dependent on activation of a Ca2+ entry pathway with a similar sensitivity to La3+ and 2-APB as capacitative Ca2+ entry, although its activation is not dependent on emptying of PLC/IP3 or thapsigargin-sensitive intracellular stores.
OBJECTIVE:Sphingosylphosphorylcholine (SPC) is an important lipid mediator that has been implicated in vascular disease. As it has not been studied in the pulmonary circulation, we examined its mechanisms of action in rat small intrapulmonary arteries (IPA). METHODS:IPA were mounted on a myograph for recording tension and intracellular Ca2+ concentration ([Ca2+]i). Ca2+ sensitisation was examined in alpha-toxin permeabilized IPA, and by Western blot analysis of MYPT1 phosphorylation. RESULTS:SPC induced a slow but powerful vasoconstriction in IPA associated with an elevation in [Ca2+]i, with an EC50 for vasoconstriction of 12+/-2 microM. Removal of extracellular Ca2+ increased the EC50 to 76+/-33 microM (p<0.01) and abolished the rise in [Ca2+]i. Endothelial denudation or inhibition of NO synthase with L-NAME enhanced vasoconstriction. Treatment with pertussis toxin or the PLC inhibitor U731223 had no effect on SPC-induced vasoconstriction. The Rho kinase inhibitor Y27632 reduced SPC-induced vasoconstriction by approximately 70% and abolished both SPC-induced Ca2+ sensitisation in permeabilized IPA and the associated increase in MYPT1 phosphorylation; Ca2+ sensitisation was substantially inhibited by GDPbetaS. La3+ and 2-APB, at concentrations previously shown to block capacitative Ca2+ entry in IPA, suppressed SPC-induced vasoconstriction to the same extent as removal of extracellular Ca2+; residual tension was abolished by Y27632. Diltiazem was relatively ineffective. 2-APB also abolished the SPC-induced rise in [Ca2+]i. However, treatment with thapsigargin to empty intracellular stores had no effect on the elevation of [Ca2+]i induced by SPC. CONCLUSION: We present evidence that SPC is a powerful vasoconstrictor of IPA and the novel finding that SPC-induced vasoconstriction in IPA is dependent on activation of a Ca2+ entry pathway with a similar sensitivity to La3+ and 2-APB as capacitative Ca2+ entry, although its activation is not dependent on emptying of PLC/IP3 or thapsigargin-sensitive intracellular stores.
Authors: Jiwang Chen; Haiyang Tang; Justin R Sysol; Liliana Moreno-Vinasco; Krystyna M Shioura; Tianji Chen; Irina Gorshkova; Lichun Wang; Long Shuang Huang; Peter V Usatyuk; Saad Sammani; Guofei Zhou; J Usha Raj; Joe G N Garcia; Evgeny Berdyshev; Jason X-J Yuan; Viswanathan Natarajan; Roberto F Machado Journal: Am J Respir Crit Care Med Date: 2014-11-01 Impact factor: 21.405
Authors: Vladimir A Snetkov; Gregory A Knock; Lynne Baxter; Gavin D Thomas; Jeremy P T Ward; Philip I Aaronson Journal: J Physiol Date: 2005-12-15 Impact factor: 5.182