Invasion of host cells by Toxoplasma gondii is accompanied by secretion of parasite proteins that occurs coincident with increases in intracellular calcium. The source of calcium mobilized by the parasite and the signals that promote calcium increase remain largely undefined. We demonstrate here that intracellular stores of calcium in the parasite were both necessary and sufficient to support microneme secretion, motility and invasion of host cells. In contrast, host cell calcium was largely unaltered during parasite entry and not essential for this process. During parasite motility, cytosolic calcium levels underwent dramatic and rapid fluxes as imaged using the calcium indicator fluo-4 and time-lapse microscopy. Surprisingly, intracellular calcium in the parasite cytosol was rapidly quenched during the initial stages of host cell invasion, suggesting that while it is needed to initiate motility, it is not required to complete entry. These studies indicate that intracellular calcium stores govern secretion and motility by T. gondii and that the essential role of calcium in these events explains its requirement for cell entry.