Mechanism of trypsin‐induced endothelium‐dependent vasorelaxation in the porcine coronary artery

Abstract

1 To investigate the mechanism underlying the trypsin-induced endothelium-dependent relaxation, cytosolic Ca 2+ concentration ([Ca 2+ ] i ) and tension development of smooth muscle were simultaneously monitored in the porcine coronary artery, and [Ca 2+ ] i of in situ endothelial cells were monitored in the porcine aortic valvular strips, using fura-2¯uorometry. 2 During the contraction induced by 30 nM U46619, a thromboxane A 2 analogue, 100 nM trypsin induced a rapid transient signi®cant decrease in both [Ca 2+ ] i (from 67.9+5.1 to 15.7+4.4%) and tension (from 97.5+9.2 to 16.8+3.5%) of smooth muscle only in the presence of endothelium (100% level was assigned to the level obtained with the 118 mM K + -induced contraction). [Ca 2+ ] i and the tension thus returned to the levels prior to the application of trypsin by 5 and 10 min, respectively. 3 The initial phase of this relaxation was partly inhibited by 100 mM N o -nitro-L-arginine (L-NOARG), and was completely inhibited by L-NOARG plus 40 mM K + or L-NOARG plus 100 nM charybdotoxin and 100 nM apamin, while the late phase of the relaxation was inhibited by L-NOARG alone. 4 Trypsin induced a transient [Ca 2+ ] i elevation in the endothelial cells mainly due to the Ca 2+ release from the intracellular stores, at the concentrations (1 ± 100 nM) similar to those required to induce relaxation. 5 In conclusion, trypsin induced an elevation in [Ca 2+ ] i mainly due to Ca 2+ release in endothelial cells, and thereby caused endothelium-dependent relaxation. The early phase of relaxation was due to nitric oxide and hyperpolarizing factors, while the late phase was mainly due to nitric oxide in the porcine coronary artery.

Redundant signaling mechanisms contribute to the vasodilatory response of the afferent arteriole to proteinase-activated receptor-2

“…These characteristics are similar to responses attributed to EDHF in other preparations (21,29,33,41). There is limited information on the nature of the EDHF involved in the responses of other vessel types to PAR-2 agonists.…”

“…In a number of vascular preparations, PAR-2-induced vasodilation is eliminated by removal of the endothelium (21,29,30,33,39). Unfortunately, we cannot remove the endothelium in our preparation without affecting vascular reactivity.…”

“…There is limited information on the nature of the EDHF involved in the responses of other vessel types to PAR-2 agonists. Nakayama et al (33) found that in the porcine coronary artery, the EDHF-like response to trypsin was fully abolished by the combined treatment with charybdotoxin plus apamin. Similarly, Kawabata et al (21) found the combination of charybdotoxin plus apamin to abolish the NO-independent dilator effects of SLIGRL-NH 2 on gastric mucosal blood flow in the rat.…”

Role of Protease-activated Receptors in the Vascular System

“…PAR-2 couples to Gαq/11 and Gαi/o (19), and thus activates phospholipase Cβ in both pertussis toxin-sensitive and insensitive manners. We observed that trypsin induced a transient [Ca 2+ ]i elevation mainly due to Ca 2+ release in the vascular endothelial cells (73). PAR-2 was also reported to activate mitogen-activated protein kinase (74)(75)(76), protein-tyrosine phosphatase SHP2 (77) and tyrosine kinase pathways (78), which could contribute to PAR-2-mediated cytokine production (74), mitogenic signaling (77) and mucin secretion in the rat sublingual gland (78).…”

The Roles of Proteinase-Activated Receptors in the Vascular Physiology and Pathophysiology

“…6,7,18,53,62,63 The NO production by endothelial NO synthase (eNOS) is primarily dependent on the Ca 2ϩ signal. 64 In line with this, thrombininduced NO production is reported to be sensitive to calmodulin inhibitors.…”