We studied the role of cytoplasmic free Ca2+ concentration ([Ca2+]i) in cytokinesis of zoosporangia of the water mold Phytophthora cinnamomi. In these cells cytokinesis is separated from nuclear division and can be triggered at precisely determined times by cold shock. Changes in [Ca2+]i were monitored by ratiometric fluorescence imaging of pressure microinjected Fura-2 dextran. Two increases in [Ca2+]i always occurred in sporangia that underwent cytokinesis in response to cold shock. Within the first minute of cold shock, [Ca2+]i rose rapidly and transiently to levels 25 to 131% higher than the resting level of 104 +/- 54 nM. By 10 min, [Ca2+]i had decreased and was near the initial resting level. The second increase in [Ca2+]i was gradual and prolonged, accompanying cell division. Near completion of cytokinesis, [Ca2+]i had risen to 231 +/- 165 nM. The initial brief rise in [Ca2+]i was absent in sporangia that did not undergo cleavage. Microinjection of the Ca2+ buffer 5,5'-dibromo-BAPTA before cold shock, blocked cytokinesis suggesting that the transient rise in [Ca2+]i may be necessary for induction. The subsequent gradual increase in [Ca2+]i may not be critical because microinjection of 5,5'-dibromo-BAPTA during cleavage plane development did not always perturb division.