The ultrastructure of Chinese hamster ovary cells was examined immediately after heating cells in mitosis, and the findings were compared with (a) the behavior of heated cells monitored with time lapse cinematography following heat shocks and (b) proliferative survival of individual cells followed for 7 days after heating. Treatment of dividing cells at 45.5 degrees (5 to 15 min) disassembled the spindle and disrupted both the contractile ring and the midbody-cytoplasmic bridge complex to varying degrees depending on the length of heating. The spindle did not reform upon return to 37.0 degrees. Microtubular proteins were heated in vitro to determine if their inactivation was responsible for the inability of the spindle to reform. Heat completely disassembled the intact microtubules and inactivated a proportion of the microtubular proteins in vitro; however, a fraction of the microtubular proteins from heat-disassembled microtubules still was capable of reassembly. The time lapse studies indicated that cells in division at the time of heating entered G1 without completing division. Of the resultant tetraploid cells, 88% had greater than or equal to 2 nuclei; 59% of the tetraploid cells divided 35 +/- 7 (S.D.) hr following the heat shock (control generation time, 13 +/- 2 hr), and 95% of the flattened progeny had more than one nucleus. The fate of individual cells that were in mitosis or G1 when treated at 45.5 degrees for 4.5 min was monitored for 7 days. The survival of the total heated population of cells was 19%, but the surviving cells were almost totally accounted for by the G1 cells present as contaminants in the heated population. Less than 2% (2 of 115) of the monitored cells that were heated in mitosis formed colonies. Therefore, the morphological disruption of the spindle, contractile ring, or midbody-cytoplasmic bridge complex by a heat of 45.5 degrees prevented cytokinesis, and the resultant tetraploid cells became proliferatively dead.