Thrombin is a pro-coagulant inflammatory agonist that can disrupt the endothelium-lumen barrier in the lung by causing contraction of endothelial cells and promote pulmonary cell proliferation. Both contraction and proliferation require increases in [Ca²⁺)_cyt. In this study, we compared the effect of thrombin on Ca²⁺ signaling in human PASMC and PAEC. Thrombin increased the [Ca²⁺)_cyt in both cell types, however the transient response was significantly higher and recovered quicker in the PASMC suggesting different mechanisms may contribute to thrombin mediated increases in [Ca²⁺)_cyt in these cell types. Depletion of intracellular stores with cyclopiazonic acid (CPA) in the absence of extracellular Ca²⁺ induced Ca²⁺ transients representative of those observed in response to thrombin in both cell types. Interestingly, CPA pretreatment significantly attenuated thrombin-induced Ca²⁺ release in PASMC; this attenuation was not apparent in PAEC, indicating that a PAEC-specific mechanism was targeted by thrombin. Treatment with a combination of CPA, caffeine and ryanodine also failed to abolish thrombin-induced Ca²⁺ transient in PAEC. Notably, thrombin-induced receptor-mediated Ca²⁺ influx was still observed in PASMC after CPA pre-treatment in the presence of extracellular Ca²⁺. Ca²⁺ oscillations were triggered by thrombin in PASMC resulting from a balance of Ca²⁺ influx and Ca²⁺ reuptake by the SR/ER. The data shows that thrombin induces increases in intracellular calcium in PASMC and PAEC with a distinct CPA, Caffeine and Ryanodine insensitive release existing only in PAEC. Furthermore, a dynamic balance between Ca²⁺ influx, intracellular Ca²⁺ release and reuptake underlie the Ca²⁺ transients evoked by thrombin in some PASMC. Understanding of such mechanisms will provide an important insight into thrombin mediated vascular injury during hypertension.