THE CYTOLOGICAL EFFECTS OF PODOPHYLLIN AND PODOPHYLLOTOXIN ON THE FERTILIZED EGGS OF CHAETOPTERUS

Abstract

1. Fertilized eggs of the polychaete annelid, Chaetopterus pergamentaceus, were treated with various dilutions of podophyllin and podophyllotoxin, beginning five minutes after insemination. Observations were made on the living eggs and on whole-mount cytological preparations made from samples fixed at various intervals during development. Dosages ranging from 0.1 to 0.00001 mg./ml. were tested.2. Beginning as soon as 60 seconds after the beginning of treatment with moderate to high dosages (0.1 to 0.005 mg./ml.) of podophyllin, the asters of the egg maturation figure began to fade, followed by disappearance of the spindle by about three minutes after the initiation of treatment. The nine egg chromosomes remained in the ring configuration (characteristic of the first maturation division in this form) for approximately two to five minutes longer (depending on the exact dosage), after which they were gradually enclosed in a membrane-surrounded area which appears to have been derived from the spindle substance. By this time they had taken on a somewhat vesicular appearance. The sperm chromosomes were often similarly affected by the podophyllin, although the onset of these effects was usually somewhat slower than for the eggs. Both egg and sperm chromosomes remained contained in this membrane for approximately an hour, after which they were released to lie loose in the egg cytoplasm. No further development occurred at these dosages.3. After treatments with podophyllin concentrations of 0.00005 to 0.00001 mg./ml., a semblance of normal development ensued, although the resulting larvae were usually abnormal. In egg-samples fixed 24 minutes after the beginning of treatment, there was occasional evidence of reduction in size of the asters; by 47-57 minutes after the initiation of treatment, multipolar figures were observed, as well as two entirely separate complex metaphases, often in uncleaved cytoplasm. There was frequently karyokinesis without accompanying cytokinesis. When cleavage continued to occur, it was often retarded and/or abnormal.4. Shape changes ("pear" and polar lobe stages) characteristic of the normal development of Chaetopterus eggs were observed, even after treatment with high dosages which destroyed the achromatic figure. Pseudo-polar bodies, usually without chromatin, were also noted, as well as giant polar bodies.5. The effects of podophyllotoxin were qualitatively much like those of podophyllin, but the dosage required to produce them was very much lower (0.001 mg./ml. for podophyllotoxin vs. 0.01 mg./ml. for podophyllin to completely block division, for example).6. Even short durations of exposure to either podophyllin or podophyllotoxin (10-20 minutes, as compared with the 60-minute duration routinely used in the other experiments) were sufficient to induce marked abnormality with moderate dosages.7. The effects of mitomycin C, N-dichloroacetyl DL serine (sodium) and quercetin were also tested. None of these agents was effective in producing cytological abnormality in Chaetopterus eggs, at the concentrations tested.