Histamine-stimulated cytosolic free Ca2+ ([Ca2+]i) oscillations in human umbilical vein endothelial cells (HUVECs) comprise repetitive spikes generated by pulsatile release from stores. We have investigated the roles of the store Ca2+-ATPases in regulating both the upstroke and downstroke of a Ca2+ spike. 2. The sarco-endoplasmic reticulum Ca2+-ATPase (SERCA) inhibitor cyclopiazonic acid (CPA) dramatically affected oscillations whereas inhibition of the plasma membrane Ca2+-ATPase (PMCA) with La3+ had little effect. This and other evidence suggested that the downstroke of a spike is predominantly mediated by SERCA. 3. Artificial [Ca2+]i spiking generated by repetitive pulsatile application of 0.3 microM histamine in Ca2+-free medium did not cause net loss of Ca2+ from the cell whereas repetitive pulsatile application of 1 and 10 microM histamine did, with the higher concentration being more effective. We conclude that there is an inverse relationship between stimulus intensity and relative SERCA activity. 4. For a Ca2+ transient, the initiation of release was suppressed by SERCA during either the lag phase or the interspike period (ISP) since: (i) the ISP was shortened by low CPA concentrations, (ii) higher concentrations of CPA stimulated an explosive Ca2+ release when applied during the ISP but not when applied in the absence of agonist, and (iii) CPA synchronized the initial Ca2+ response to a low histamine dose (even recruiting silent, histamine-unresponsive cells). 5. Two aspects of the regenerative upstroke of a spike were differently affected by SERCA inhibition: Ca2+ wave velocity was entirely unaffected by CPA whereas the local rate of rise was increased. 6. The [Ca2+]i at which a Ca2+ spike terminated depended on SERCA since CPA dose dependently enhanced the peak [Ca2+]i. 7. We conclude that SERCA plays a powerful and dynamic role in regulating [Ca2+]i oscillations in HUVECs. SERCA differentially modulates the phases of Ca2+ release in addition to bringing about the falling phase of a Ca2+ spike.
