Previous experiments from our lab have shown that the endothelium-mediated dilatation of resistance arteries to endothelial stimulators was significantly reduced after inhibition of vascular gap junctional communication. We therefore hypothesised that endothelial gap junctions might play a significant role in the net increase of calcium following endothelial stimulation, which ultimately controls the production of endothelial autacoids such as nitric oxide and prostacyclin. Experiments were performed in human umbilical vein endothelial cells (HUVEC) stimulated with histamine (HIS, 5 μM). Studies were performed in n monolayers of HUVEC (2nd passage); values are presented as means ± SEM. Statistical comparisons were performed using t tests. All comparisons reported here were significant (p<0.05). HUVECs were loaded with the calcium sensitive dye Fura 2- AM and the calcium rise in response to histamine was analysed with a camera-based system at intervals of 200 ms. HIS induced an initial Ca2+i rise in only 5±0.2% of all cells (n=13 cultures), which were randomly dispersed in the monolayers. A Ca2+i rise was finally observed in 98±1% of all cells (n=10) albeit with a delay of up to 3 s. Inhibition of GJ coupling (50μM Meclofenamic acid + 1mM heptanol) did not affect the Ca2+i increase in the initially responding cells (in 83±1% identical with controls) whereas the total number of cells responding to histamine was reduced to 38±5% (n= 6). Similar results (reduction of the slope of the calcium rise in the monolayer from 131 ± to 65±18, n=6) were obtained when the cells were exposed for 3 min to the supernatant of polymorphonuclear neutrophils being stimulated with fMLP (fMLP had no effect on HUVEC GJ coupling, data not shown). In pilot immunohistochemical experiments (n=2) we found that only very few cells in a monolayer were positive for the H1 receptor known to be responsible for HIS-mediated effects in HUVEC. The Ca2+i increase induced by histamine seems to be only partly mediated by direct receptor-mediated events occuring in a limited number of 'pacemaker cells'. In the majority of cells the calcium rise seems to be due to spread of a signal (probably calcium or IP3) via gap junctions to the neighbouring receptor-deficient cells. Thus, control of GJ coupling may be a novel pathway to regulate the EC responsiveness to autacoid-releasing receptor-mediated stimuli.