Mechanism of contraction induced by bradykinin in the rabbit saphenous vein

Abstract

1. By using fura-PE3 fluorometry and receptor-coupled permeabilization by α-toxin, the mechanism of the bradykinin (BK)-induced contraction was determined in the rabbit saphenous vein (RSV). The receptor subtype responsible for the BK-induced contraction of RSV was determined by means of a pharmacological blocker study and reverse transcription polymerase chain reaction (RT-PCR). 2. In the presence of extracellular Ca2+ (1.25 mM), BK (10-11-3 x 10-7 M) induced increases in both the cytosolic Ca2+ concentration ([Ca2+](i)) and force, in a concentration-dependent manner. Both the release of Ca2+ from the store site and the influx of extracellular Ca2+ contribute to an increase in [Ca2+](i) induced by BK. 3. In the absence of extracellular Ca2+, the application of 10-7 M BK induced transient elevations of [Ca2+](i) and force, both of which thereafter declined to the levels observed before the application of BK. When extracellular Ca2+ was replenished (1.25 mM), [Ca2+](i) and force increased to form a peak, followed by a sustained elevation in the presence of BK. When an RSV strip was pretreated with 10-5 M thapsigargin for 20 min, the BK-induced transient increases in both [Ca2+](i) and force were markedly inhibited. 4. These responses induced by BK were inhibited by Hoe 140 (D-Arg-[Hyp3, Thi3, D-Tic7, Oic8] bradykinin), a highly specific bradykinin B2 receptor antagonist, in a concentration-dependent manner. In RT-PCR, B2-receptor mRNA was expressed in the smooth muscle of RSV. 5. The [Ca2+](i)-force relationships, which were determined by cumulative applications of extracellular Ca2+ (0-5 mM) during 118 mM K+-depolarization, shifted to the upper left in the presence of BK, thus indicating that BK induced a greater force than 118 mM K+-depolarization for a given level of [Ca2+](i). 6. In α-toxin-permeabilized preparations of RSV, application of 10-7 M BK after a steady state contraction had been induced by a mixture of 5 x 10-7 M Ca2+, 10-6 M GTP and 10-6 M captopril caused an additional force development at a constant [Ca2+](i). However, treatment with 1. mM guanosine-5'-O-(β-thiodiphosphate) (GDPβS) for 5 min before and during the application of BK (10-7 M), abolished this BK-induced additional contraction. 7. These results indicated that in RSV: (1.) BK elicits vasoconstriction by increasing the Ca2+ influx from the extracellular space, Ca2+ release from intracellular thapsigargin-sensitive storage sites and increasing the Ca2+ sensitivity of the contractile apparatus, (2) the BK-induced increase in Ca2+ sensitivity is mediated by G-protein, (3) the BK-induced contractions are mediated via B2-receptors and (4) the smooth muscle cells express B2-receptor mRNA.