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Cell Division Cycles and Circadian Clocks ^1,2,3

Phase-Response Curves for Light Perturbations in Synchronous Cultures of Euglena

Department of Anatomical Sciences, School of Medicine, State University of New York, Stony Brook, New York 11794, Health Sciences Center, State University of New York, Stony Brook, New York 11794

The cell division rhythm in Euglena gracilis Klebs (Z strain) freeruns with a circadian period (30.2 ± 1.8 hours for 156 monitored oscillations) in aerated, magnetically stirred, 8-liter, axenic batch cultures grown photoautotrophically at 25°C in LD: 3,3, (7,500 lux, cool-white fluorescent) 6-hour light cycles from the moment of inoculation. Cell number was measured at 2-hour intervals with an automatic fraction collector and Coulter Electronic Particle Counter. At different circadian times throughout the 30-hour division cycle, 3-hour light perturbations were imposed on free-running cell populations by giving light during one of the intervals when dark would have fallen in the LD: 3,3 regimen. Using the onset of division as the phase reference point, the net steady-state phase advance or delay (±) of the rhythm was determined after transients, if any, had subsided (usually in one or two days) relative to an unperturbed control culture. Both + and – were found, with maximum values of approximately ±11 to 12 hours being obtained at circadian time (CT) 20 to 22 (the `breakpoint'); little, if any phase shift occurred if the light signal was given between CT 6 and CT 12. The phase-resetting curve obtained by plotting new phase (') versus old phase () was of the type 0 (`strong') variety. Light perturbations, no matter when imposed, engendered new phases which mapped to a relatively restricted portion (CT 6 to CT 13) of the circadian cycle.

These data provide the first detailed phase-response curve for a circadian mitotic clock. The findings, therefore, not only further support the hypothesis that a circadian oscillator (perhaps exhibiting limit cycle behavior) can modulate cell division in eukaryotic cells, but also provide a useful basis for the dissection of the nature and extent of the coupling between cell division and circadian cycles.

¹ Dedicated to the memory of William S. Hillman, critical scientist, colleague, and good friend.

² Supported in part by National Science Foundation Grant PCM76-10273 and United States Public Health Service Biomedical Research Support Grant 2S07RR07067-16 to L. N. E.

³ Some of these results were reported at the XV International Congress of the International Society for Chronobiology, September 13-16, 1981, Minneapolis.

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