Endothelial cells form a multifunctional signal transducing surface that regulates many fundamental processes in vascular system. A number of protein kinases have been implicated in controlling calcium signaling in a variety of cells. PKC_α and Rho-kinase have been implicated to play an important role in control of calcium entry through store operated channels in cultured endothelium [Mehta D et al, 2003; Ahmmed GU et al, 2004]. However expression and functional role of PKC and Rho-kinase in intact endothelium were not elucidated. In the present study we have used immunohistochemistry and confocal imaging to investigate role of PKC and Rho-kinase in intact endothelium of conduit artery. Rats were humanely killed under CO₂ anaesthesia; their tail artery removed from the ventral grove, cleaned of fat and loaded with Fluo-4 AM (Molecular Probes, 15um) with pluronic. Confocal imaging was done using Nipkow disc based confocal imaging system (Ultraview Perkin Elmer, UK). Minimum of 5 animals were used in each set of experiments. We have found that in endothelium of rat tail artery PKC_α was predominantly expressed. Direct activation of PKC in endothelial cells by PDBu 0.1µM caused activation of monophasic non-oscillatory calcium transient dependent exclusively on calcium entry which was inhibited by PKC inhibitor Ro-32-0432 (5µM), and store operated Ca²⁺ channel blocker SKF-96365 (50µM) but was resistant to Rho-kinase inhibitor H-1152 (0.2µM). Stimulation of intact endothelial cells by carbachol (0.1µM, 1µM, 10µM) produced translocation of PKC_α to the plasma membrane and activation of a calcium transient which consisted of two components: initial fast – dependent on Ca²⁺ release and subsequent, sustained dependent on Ca²⁺ entry. Sustained component of CCh induced Ca²⁺ transient was 63.8±3.5% of the peak taken for 100% (n=10, 5 vessels). Inhibition of PKC by Ro-32-0432 (5µM) and Rho-kinase by H-1152 (0.2µM) reduced the sustained component of CCh induced Ca²⁺ transient to 15.9±4.3 and 34.9±4.6% (n=10, 5 vessels) of the peak, respectively. Thus the data obtained suggests that PKC and Rho-kinase are both involved in control of calcium entry activated by carbachol in the endothelial cells of large conduit arteries.