P142	EDHF is a potent inhibitor of platelet adhesion in vivo.
1S.Lehrer, 1N.Hellwig, 1M.Buerkle, 2H-Y.Sohn, 2F.Krötz, 1U.Pohl
1Institute of Physiology, München, DE; 2Division of Cardiology, Med. Clinic, München, DE.

Platelets express Kca channels and we have shown before that a CYP450 dependent supernatant (EDHF) from stimulated cultured endothelial hybridoma cells increased platelet membrane potential in vitro. Here we tested whether there was an EDHF related effect on platelet activity in vivo. Dorsal skinfold chambers were implanted on Golden Syrian hamsters. Platelet-rich plasma (PRP) from healthy volunteers who had not taken medication for 10 days was prepared and the platelets were labelled with calcein-AM. 2x10e8 calcein-labelled and resuspended platelets (400,000/µl) were injected via a jugular vein catheter. Platelet velocities and adhesion in the vessels of the dorsal skin muscle were monitored by intravital fluorescence microscopy. Platelet rolling was assessed according to the velocity distribution patterns of all analyzed platelets (2000-3000 per experiment). Firm platelet adhesion was classified as number of platelets per vessel area (two-dimensional reconstruction) that did not move for at least 30s. Inhibitors of NOS (L-NA, 100µM) or of CYP450 2C9 (sulfaphenazole (SULF) 100µM) were superfused (for 30 min) in MOPS-buffer locally over the skinfold tissue. Under control conditions, the labelled platelets showed a normal distribution of velocities with no detectable adhesion in all observed arterioles of the skinfold chamber. Superfusion with the inhibitor of eNOS, L-NA, induced a significant leftward shift of the velocity distribution indicating enhanced platelet rolling and, in addition,firm adhesion of a platelet fraction of about 12 %, which was not present in control animals. Similar results were obtained, when the CYP 450 inhibitor SULF was used instead of L-NA. The effects of SULF could be reversed by superfusion of the vessels by 11,12-EETs (1µM) which also induced a significant vasodilatation. Similar effects as with SULF were also observed after superfusion of the tissue with the inhbitor of KCa channels, charybdotoxin (5µM) The present study demonstrates a clear effect of the CYP inhibitor SULF and of a blockade of KCa channels on the platelet adhesion in the microcirculation in vivo. The data suggest that a continuous release of EDHF occurs in arterioles that is sufficient to control the activity of blood platelets as assessed by their rolling and adhesion.

Copyright © 2005 S. Karger AG, Basel. Any further use of this abstract requires written permission from the publisher.