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The cytology of the irradiated ovary of Drosophila melanogaster
Experimental
Cell Research
THE
13, 545-552
CYTOLOGY
545
(1957)
OF THE IRRADIATED
OF DROSOPHILA
OVARY
MELANOGASTERI
R. C. KING Northwestern
University,
Evanston, ill., U.S.A.
Received May 7, 1957
RECENTstudies
[2, 31 have been published which describe the cytology of the normal ovary of the adult fruit fly, Drosophila melanogasfer and radiationinduced ovarian tumors in this insect. It is the purpose of this paper to describe other radiation-induced cytological changes in the ovary. MATERIAL
AND
METHODS
Freshly eclosed female flies belonging to the Oregon-R, wild type strain were and they were irradiated with 4000 r of CobalV” gamma rays (N 100 r/second), then placed with untreated males on medium for varying time intervals (generally 12 days). They were killed subsequently, and Feulgen whole mounts were made of their ovaries. Microscopic observations were made in normal bright field or phase contrast. In cases where the situation warranted, tracings were made of abnormal chambers using a Seibert Promi projection microscope, or photomicrographs were taken using Adox KB14 film in a Leica with a Micro-Ibso attachment. Approximately 500 ovaries were examined in both the control and the treated series. RESULTS
AND
CONCLUSIONS
Early effects--A cytological examination was made of the ovaries of flies killed 1, 2, and 6 days after irradiation. The ovaries from the l- and ‘L-day series show-ed large numbers of Feulgen-positive granules distributed homogeneously throughout the cytoplasm of all cells (cf. Fig. 1). The majority of these granules have disappeared by the Gth day. Lute efiects.-All the following information applies to the ovaries of flies killed 12 days after treatment. PII~‘7osis.-Normall~ every orariole of the 12-day-old Drosophilrr melanogder contains oocytes in stages 1, 2, 3, 6, 7 and 14. The majority of orarioles also contain stage 4 oocytcs, and roughly half of the orarioles also contain stage 8 oocytes. One stage 3 oocyte is found in every 5th ovariolc. Oocytrs 1 Research supported by the U.S. Atomic Energy Commission (contract No. AT (I 1.1).8’3, project
12) and the Graduate
School of Northwestern
University. Experimenlul
Cdl
Research
13
R. C. King
546
in stages 9 and 10 are found rarely (1 per ovary), and stages 11, 12 and 13 are absent. About 15 per cent of stage 8 chambers are pycnotic. In the irradiated ovary pycnotic chambers belonging to stages 1, 2, 3, 4, 6, 7, 9 and 10 have been observed in addition to stage 8. Degenerating nurse cell nuclei shrink and become more densely stained as a consequence. The cell walls undergo cytolysis, and the Feulgen-positive blobs representing the nurse nuclei
Fig. 1.-A camera lucida drawing of the Feulgen positive granules found distributed through the cytoplasm of a stage 4 chamber one day after irradiation. Fig. 2.-An ovariole observed 12 days after irradiation. Nurse nuclei have ruptured from the posterior-most chamber (stage 8) and have passed into the anterior portion of the ovariole. C, chamber; G, germarium; E.S., epithelial sheath; N.N., nurse nucleus.
may rupture into the oocyte in the case of stage 8, 9, 10 or 11 chambers. Alternatively, the nurse nuclei may rupture the anterior follicular epithelium of the chamber and pass into the ovariole, where they become wedged between other chambers or the germarium and the epithelial sheath (cf. Fig. 2). Subsequent to the cytolysis of several chambers the ovariole itself may begin to degenerate. Such ovarioles are seen quite commonly ( N 1 per ovary), and they generally contain debris consisting of Feulgen positive blobs of various sizes which represent nurse and follicle cell nuclei (cf. Fig. 3). Growth.-One of the commonest effects of irradiation is a general slowing down of oogenesis which in turn leads to an abnormal stage distribution. For example, the frequency with which chambers in stages 10, 11 and 12 occur is markedly increased (cf. Table I). Modified striation represents are rarer type of abnormality. Normally adjacent chambers in any ovariole are never in the same stage. However, in irradiated ovarioles chamber sequences such as the following have been observed: 1, 3, 3, 3, 3, 10; 1, 3, 3, 3, 3, 8; 1, 2, 4, 5, 5, 5, 5, 5; and 2, 3, 4, 5, 6, 6, 7, 8, 12. One ovary was found which the egg-laying apparatus contained over fifty stage 14 oocytes. Apparently failed to function normally in this case. Cell division.-A second common effect of irradiation is inhibition of cell division. Anaphase and metaphase chromosome figures are rarely found in Experimental
Cell Research 13
Irradiated fly ovary
0, Fig. 3.--A
TABLE
!
!
!
photomicrograph
I. Morpholoyicul
differences
I
yb
of a degenerating
between
control
ovariole.
and irradiated
Control Germaria Oogonial metaphases and anaphases Av. developing eggs/ovariole Av. total ovarioles/female Av. total developing eggs/female Stage 10 oocytes/female 11 12 Frequency of ovarioles developing eggs x=6 7 8 Total ovarioles
with
large, sausage-shaped frequent 6.97 ?. 0.62 (n = 30)’ 25.06 & 1.18 (n = 17)b 175 2.66L0.74 (n=lg) 0 0.44iO.20 (n= 18)
ovcrries. Treated
small, blunt absent 6.45 ?I 0.75 (II 22.90 Il.02 (n 148 17.7 * 1.03 (n 1.50 i- 0.32 (II 5.90 t 0.70 (n
= 6Qa = 20)” = 20) = 20) = 20)
z 0.10 0.53 0.07 30
0.59 0.37 0.04 68
F These means are not significantly different, P = 0.7. These means are highly significantly different, P = 0.009. Experimenfal
Cell Research 13
R. C. King
548
irradiated germaria. Many of the Feulgen-positive blobs observed in the l- and 2-day ovaries probably represent chromosome fragments. The number of developing eggs in treated ovaries is 85 per cent of the control. This is due to there being fewer ovarioles and fewer oocytes per ovariole (cf. Table I). Although radiation generally inhibits cell division, in specific rare instances it accelerates it. One example of this acceleration is the production of ovarian tumors [a]. A second is the production of chambers which contain cells which have undergone a supernumerary division. The two such eggs observed contained 23 nurse cells and the oocyte contained 2 nuclei (cf. Figs. 4, 5). Excess numbers of nurse cells (21-29) have been found in stage 7, 8, 9 and 10 chambers. These chambers all contained a single oocyte, ho\vever, and it is impossible to decide Tvhether they resulted from fusions of two chambers or from a supernumerary division. The reverse situation, namely chambers with less than the normal cell number, have also been observed. In these cases the situation is always an extreme one. That is the chambers contain a few (l-4) very large nurse cells. Cell migration.--Egg chambers pass down the ovariole as they develop. Irradiation appears to partially inhibit this process \\-ith the result that has proadjacent chambers often fuse together. In this respect irradiation duced a phenocopy of the gene comb-gap [ 11. The following classes of fusions have been observed: 213, 314, 415, 718 and lOjl4. Fusions occur in the control series, but at a rate at least 50 times lower. Fusion almost always leads to a reversal of polarity of the anterior oocyte. Normally the most posterior of the 16 cells in a chamber differentiates into the oocyte. When two chambers fuse the cells of the posterior chamber behave normally. The oocyte of the anterior chamber, holyever, moves anteriorly and eventually becomes the most anterior cell of the chamber. The result is a chamber containing two groups of nurse cells lvhich differ in size and stage and two oocytes-one at each pole (cf. Fig. 6). Often as the nurse cells grow the anterior oocyte will be crushed. The resulting chamber at a later stage may appear perfectly normal except for the abnormally high number of nurses. The chamber in Fig. 7 is probably the result of such a sequence of events. Fig. 8 shows an interesting, but extremely rare abnormality observed in irradiated material. Subsequent to its determination the oocyte seems to have migrated to the center of the chamber. In spite of this, hojvcver, yolk is being laid down at the posterior pole as \\-ould normally occur in stage 8. This bizarre chamber demonstrates that the physical presence of the oocyte in the posterior pole of the stage 8 chamber is not required for the transfer of yolk to this region. Experimental
Cell Research 13
Irrarliafed
D
549
fly ovary
SD,< FE
Y c.
0L
.
,
.
9
20!,
Fig. &-An outline drawing of an early stage 9 chamber (la-day series). It contains 23 nurse cells and 2 oocytes. F.E., follicular epithelium; O.N., oocyte nucleus; P, plasmosome; B.C., border cells; I<, karyosome. Fig. 5.-,4n outline drawing of a late stage 10 chamber (12-day series). It contains 23 nurse cells and 2 oocytes. Fig. 6.-.Xn outline drawing of a composite chamber (la-day series) originating from the fusion of a stage 5 and a stage 6 chamber. It contains 37 nurse cells and 2 oocytes. Fig. 7.-An outline drawing of a late stage 10 chamber (12-day series) which contains 29 nurse cells and 1 ooryte. Fig. 8.--A stage 8 chamber (la-day series) containing a misplaced oocyte and a tiny nurse nucleus (N.N.). I’.& yolky cytoplasm. Fig. 9.-(A) A stage 8 chamber (12-day series) with nurse nuclei arrested iti differentiation at stage 4. The oocyte nucleus appears to have degenerated and only a series of vacuoles remain, (J3) a normal stage 4 chamber. 0, oocyte.
Differentiation.-Rare abnormalities have been observed which are interpreted as disturbances in differentiation. The first class of abnormality represents a homogeneous disturbance in that all nurse cells are similarly effected. N’hat we find are chambers which can be classified as stage 8 on Experimental
Cell Research 13
R. C. King
550
the basis of the overall size, the follicular morphology, the size of the oocyte and that of the nurse nuclei. The nurse nuclei, however, are morphologically identical to stage 3 or 4. Thus nuclear differentiation has been arrested at stage 3 or 4 for the nurse cells, but this has not prevented further growth of these nuclei, nor has it prevented nurse cell cytoplasmic growth and yolk synthesis or the differentiation of the follicular epithelium. Fig. 9 illustrates a situation observed twice where the differentiation of nurse cells was arrested at stage 4. One case was observed where they stopped at stage 3. The second class of abnormality represents a non-homogeneous disturbance in that not all the nurse cells of a chamber are affected similarly. The chamber represented in Fig. 8 shows an example of this phenomenon where the development of one of the nurse nuclei has been arrested. The reverse situation has also been observed. The case in point was a stage 3 chamber in which one of the 15 nurse nuclei had differentiated into a nucleus morphologically similar to a stage 6 nucleus but similar in size to a stage 3. A similar situation was observed once in a germarium. A stage 5 nurse nucleus was observed lying among oogonial cells. Origin of border cells.-A large number of irradiated stage 9 and 10 chambers were observed in the hope of observing the migration of the border cells (cf. [3], p. 144). Various stages in the migration of these cells were observed (cf. Fig. 4, 5, 7). Apparently the cells originate early in stage 9 at the anterior pole of the egg and migrate posteriorly through the center of the nurse chamber until they reach the surface of the oocyte. Late in stage 10 the cells migrate back the way they came. After they have migrated about 100 micra into the nurse chamber they degenerate. Whether this degeneration also occurs in non-irradiated chambers is not known. The chromosomes of the nurse cells.-We suggested in an earlier paper [3] that the chromosome number in the nurse cells remains constant and that as the chromosomes elongate they do not decrease in thickness because they simultaneously undergo a lateral replication and therefore become polytene. We observed in one stage 8 chamber a nurse nucleus which showed exceptional chromosomal detail (cf. Fig. 10). Normally the nurse nucleus at
0
I
I
I I
Experimental
I ‘?
Fig. 10.-A exceptional
Cell Research 13
camera lucida drawing chromosomal detail.
of a stage 8 nurse cell nucleus which shows
Irradiated fly ovary
Fig. 11.--i\ photomicrograph of cytokinesis in a stage 9 nurse chamber. Note the cleavage furrow which is pinching the chromatin into two unequal masses. A, Bright field. B, Phase contrast.
stage 8 is completely tilled with chromosomal material and therefore looks amorphous. However, the nucleus in question was relatively gigantic and the chromosomes within were clearly visible. A given chromosome segment shows marked variations in circumference which appear to result from variations in coiling. Certain regions show what appears to be a series of bands, but it is impossible to rule out the possibility that these may represent a compact series of gyres. Also in the irradiated material an instance was observed of cytokincsis in a stage 9 nurse chamber. Fig. 11 shows the nurse cell nucleus in question as it is pinched in two by the cleavage furrojv. DISCUSSION
The limited state of our present knowledge makes it impossible to make any valid statements as to the specific mechanisms involved in producing the abnormalities observed. About all one can say is that current studies in developmental genetics have led to the concept that the developmental activities of cells (growth, differentiation, migration, and controlled division) are the result of the interactions of various portions of the genome of given cells with specific molecules arranged in specific gene controlled cytoplasmic patterns. Perhaps the abnormalities observed result from radiation-induced Experimental
Cell Research 13
552
R. C. King
inactiviation or loss of various portions of the genetic machinery essential to such developmental sequences. For such an injured cell and its progeny normal developmental behavior is impossible, and depending on the specific genes removed from action there follows a specific type of abnormal behavior. SUMMARY
A cytological study was made of the ovaries of 12-day-old Drosophila melanogaster given 4000 r Co63 gamma rays shortly after emergence. Many severely damaged oocytes underwent pycnotic degeneration. The number of developing eggs was reduced to 85 per cent the control value. Many of the surviving oocytes showed abnormalities which are ascribed to radiation induced disturbances of cell growth, division, migration and differentiation. The author is grateful for the conscientious Samuels. Thanks are also due to Dr. Malcolm facilities.
assistance performed by Mr. Jack Dole for providing the irradiation
REFERENCES 1. BEATTY,
2. KING, 3. KISG,
R. .4., Proc. Roy. Sot. Edinbrrrgh R. C., Growth 21 (1957). R. C., Run~,~sos, A. C. and SMITH,
Experimental
Cell Resenrcl~ 13
63B,
249 (1949).
R. F., Growth
20, 121 (1956).
Cell Research
THE
13, 545-552
CYTOLOGY
545
(1957)
OF THE IRRADIATED
OF DROSOPHILA
OVARY
MELANOGASTERI
R. C. KING Northwestern
University,
Evanston, ill., U.S.A.
Received May 7, 1957
RECENTstudies
[2, 31 have been published which describe the cytology of the normal ovary of the adult fruit fly, Drosophila melanogasfer and radiationinduced ovarian tumors in this insect. It is the purpose of this paper to describe other radiation-induced cytological changes in the ovary. MATERIAL
AND
METHODS
Freshly eclosed female flies belonging to the Oregon-R, wild type strain were and they were irradiated with 4000 r of CobalV” gamma rays (N 100 r/second), then placed with untreated males on medium for varying time intervals (generally 12 days). They were killed subsequently, and Feulgen whole mounts were made of their ovaries. Microscopic observations were made in normal bright field or phase contrast. In cases where the situation warranted, tracings were made of abnormal chambers using a Seibert Promi projection microscope, or photomicrographs were taken using Adox KB14 film in a Leica with a Micro-Ibso attachment. Approximately 500 ovaries were examined in both the control and the treated series. RESULTS
AND
CONCLUSIONS
Early effects--A cytological examination was made of the ovaries of flies killed 1, 2, and 6 days after irradiation. The ovaries from the l- and ‘L-day series show-ed large numbers of Feulgen-positive granules distributed homogeneously throughout the cytoplasm of all cells (cf. Fig. 1). The majority of these granules have disappeared by the Gth day. Lute efiects.-All the following information applies to the ovaries of flies killed 12 days after treatment. PII~‘7osis.-Normall~ every orariole of the 12-day-old Drosophilrr melanogder contains oocytes in stages 1, 2, 3, 6, 7 and 14. The majority of orarioles also contain stage 4 oocytcs, and roughly half of the orarioles also contain stage 8 oocytes. One stage 3 oocyte is found in every 5th ovariolc. Oocytrs 1 Research supported by the U.S. Atomic Energy Commission (contract No. AT (I 1.1).8’3, project
12) and the Graduate
School of Northwestern
University. Experimenlul
Cdl
Research
13
R. C. King
546
in stages 9 and 10 are found rarely (1 per ovary), and stages 11, 12 and 13 are absent. About 15 per cent of stage 8 chambers are pycnotic. In the irradiated ovary pycnotic chambers belonging to stages 1, 2, 3, 4, 6, 7, 9 and 10 have been observed in addition to stage 8. Degenerating nurse cell nuclei shrink and become more densely stained as a consequence. The cell walls undergo cytolysis, and the Feulgen-positive blobs representing the nurse nuclei
Fig. 1.-A camera lucida drawing of the Feulgen positive granules found distributed through the cytoplasm of a stage 4 chamber one day after irradiation. Fig. 2.-An ovariole observed 12 days after irradiation. Nurse nuclei have ruptured from the posterior-most chamber (stage 8) and have passed into the anterior portion of the ovariole. C, chamber; G, germarium; E.S., epithelial sheath; N.N., nurse nucleus.
may rupture into the oocyte in the case of stage 8, 9, 10 or 11 chambers. Alternatively, the nurse nuclei may rupture the anterior follicular epithelium of the chamber and pass into the ovariole, where they become wedged between other chambers or the germarium and the epithelial sheath (cf. Fig. 2). Subsequent to the cytolysis of several chambers the ovariole itself may begin to degenerate. Such ovarioles are seen quite commonly ( N 1 per ovary), and they generally contain debris consisting of Feulgen positive blobs of various sizes which represent nurse and follicle cell nuclei (cf. Fig. 3). Growth.-One of the commonest effects of irradiation is a general slowing down of oogenesis which in turn leads to an abnormal stage distribution. For example, the frequency with which chambers in stages 10, 11 and 12 occur is markedly increased (cf. Table I). Modified striation represents are rarer type of abnormality. Normally adjacent chambers in any ovariole are never in the same stage. However, in irradiated ovarioles chamber sequences such as the following have been observed: 1, 3, 3, 3, 3, 10; 1, 3, 3, 3, 3, 8; 1, 2, 4, 5, 5, 5, 5, 5; and 2, 3, 4, 5, 6, 6, 7, 8, 12. One ovary was found which the egg-laying apparatus contained over fifty stage 14 oocytes. Apparently failed to function normally in this case. Cell division.-A second common effect of irradiation is inhibition of cell division. Anaphase and metaphase chromosome figures are rarely found in Experimental
Cell Research 13
Irradiated fly ovary
0, Fig. 3.--A
TABLE
!
!
!
photomicrograph
I. Morpholoyicul
differences
I
yb
of a degenerating
between
control
ovariole.
and irradiated
Control Germaria Oogonial metaphases and anaphases Av. developing eggs/ovariole Av. total ovarioles/female Av. total developing eggs/female Stage 10 oocytes/female 11 12 Frequency of ovarioles developing eggs x=6 7 8 Total ovarioles
with
large, sausage-shaped frequent 6.97 ?. 0.62 (n = 30)’ 25.06 & 1.18 (n = 17)b 175 2.66L0.74 (n=lg) 0 0.44iO.20 (n= 18)
ovcrries. Treated
small, blunt absent 6.45 ?I 0.75 (II 22.90 Il.02 (n 148 17.7 * 1.03 (n 1.50 i- 0.32 (II 5.90 t 0.70 (n
= 6Qa = 20)” = 20) = 20) = 20)
z 0.10 0.53 0.07 30
0.59 0.37 0.04 68
F These means are not significantly different, P = 0.7. These means are highly significantly different, P = 0.009. Experimenfal
Cell Research 13
R. C. King
548
irradiated germaria. Many of the Feulgen-positive blobs observed in the l- and 2-day ovaries probably represent chromosome fragments. The number of developing eggs in treated ovaries is 85 per cent of the control. This is due to there being fewer ovarioles and fewer oocytes per ovariole (cf. Table I). Although radiation generally inhibits cell division, in specific rare instances it accelerates it. One example of this acceleration is the production of ovarian tumors [a]. A second is the production of chambers which contain cells which have undergone a supernumerary division. The two such eggs observed contained 23 nurse cells and the oocyte contained 2 nuclei (cf. Figs. 4, 5). Excess numbers of nurse cells (21-29) have been found in stage 7, 8, 9 and 10 chambers. These chambers all contained a single oocyte, ho\vever, and it is impossible to decide Tvhether they resulted from fusions of two chambers or from a supernumerary division. The reverse situation, namely chambers with less than the normal cell number, have also been observed. In these cases the situation is always an extreme one. That is the chambers contain a few (l-4) very large nurse cells. Cell migration.--Egg chambers pass down the ovariole as they develop. Irradiation appears to partially inhibit this process \\-ith the result that has proadjacent chambers often fuse together. In this respect irradiation duced a phenocopy of the gene comb-gap [ 11. The following classes of fusions have been observed: 213, 314, 415, 718 and lOjl4. Fusions occur in the control series, but at a rate at least 50 times lower. Fusion almost always leads to a reversal of polarity of the anterior oocyte. Normally the most posterior of the 16 cells in a chamber differentiates into the oocyte. When two chambers fuse the cells of the posterior chamber behave normally. The oocyte of the anterior chamber, holyever, moves anteriorly and eventually becomes the most anterior cell of the chamber. The result is a chamber containing two groups of nurse cells lvhich differ in size and stage and two oocytes-one at each pole (cf. Fig. 6). Often as the nurse cells grow the anterior oocyte will be crushed. The resulting chamber at a later stage may appear perfectly normal except for the abnormally high number of nurses. The chamber in Fig. 7 is probably the result of such a sequence of events. Fig. 8 shows an interesting, but extremely rare abnormality observed in irradiated material. Subsequent to its determination the oocyte seems to have migrated to the center of the chamber. In spite of this, hojvcver, yolk is being laid down at the posterior pole as \\-ould normally occur in stage 8. This bizarre chamber demonstrates that the physical presence of the oocyte in the posterior pole of the stage 8 chamber is not required for the transfer of yolk to this region. Experimental
Cell Research 13
Irrarliafed
D
549
fly ovary
SD,< FE
Y c.
0L
.
,
.
9
20!,
Fig. &-An outline drawing of an early stage 9 chamber (la-day series). It contains 23 nurse cells and 2 oocytes. F.E., follicular epithelium; O.N., oocyte nucleus; P, plasmosome; B.C., border cells; I<, karyosome. Fig. 5.-,4n outline drawing of a late stage 10 chamber (12-day series). It contains 23 nurse cells and 2 oocytes. Fig. 6.-.Xn outline drawing of a composite chamber (la-day series) originating from the fusion of a stage 5 and a stage 6 chamber. It contains 37 nurse cells and 2 oocytes. Fig. 7.-An outline drawing of a late stage 10 chamber (12-day series) which contains 29 nurse cells and 1 ooryte. Fig. 8.--A stage 8 chamber (la-day series) containing a misplaced oocyte and a tiny nurse nucleus (N.N.). I’.& yolky cytoplasm. Fig. 9.-(A) A stage 8 chamber (12-day series) with nurse nuclei arrested iti differentiation at stage 4. The oocyte nucleus appears to have degenerated and only a series of vacuoles remain, (J3) a normal stage 4 chamber. 0, oocyte.
Differentiation.-Rare abnormalities have been observed which are interpreted as disturbances in differentiation. The first class of abnormality represents a homogeneous disturbance in that all nurse cells are similarly effected. N’hat we find are chambers which can be classified as stage 8 on Experimental
Cell Research 13
R. C. King
550
the basis of the overall size, the follicular morphology, the size of the oocyte and that of the nurse nuclei. The nurse nuclei, however, are morphologically identical to stage 3 or 4. Thus nuclear differentiation has been arrested at stage 3 or 4 for the nurse cells, but this has not prevented further growth of these nuclei, nor has it prevented nurse cell cytoplasmic growth and yolk synthesis or the differentiation of the follicular epithelium. Fig. 9 illustrates a situation observed twice where the differentiation of nurse cells was arrested at stage 4. One case was observed where they stopped at stage 3. The second class of abnormality represents a non-homogeneous disturbance in that not all the nurse cells of a chamber are affected similarly. The chamber represented in Fig. 8 shows an example of this phenomenon where the development of one of the nurse nuclei has been arrested. The reverse situation has also been observed. The case in point was a stage 3 chamber in which one of the 15 nurse nuclei had differentiated into a nucleus morphologically similar to a stage 6 nucleus but similar in size to a stage 3. A similar situation was observed once in a germarium. A stage 5 nurse nucleus was observed lying among oogonial cells. Origin of border cells.-A large number of irradiated stage 9 and 10 chambers were observed in the hope of observing the migration of the border cells (cf. [3], p. 144). Various stages in the migration of these cells were observed (cf. Fig. 4, 5, 7). Apparently the cells originate early in stage 9 at the anterior pole of the egg and migrate posteriorly through the center of the nurse chamber until they reach the surface of the oocyte. Late in stage 10 the cells migrate back the way they came. After they have migrated about 100 micra into the nurse chamber they degenerate. Whether this degeneration also occurs in non-irradiated chambers is not known. The chromosomes of the nurse cells.-We suggested in an earlier paper [3] that the chromosome number in the nurse cells remains constant and that as the chromosomes elongate they do not decrease in thickness because they simultaneously undergo a lateral replication and therefore become polytene. We observed in one stage 8 chamber a nurse nucleus which showed exceptional chromosomal detail (cf. Fig. 10). Normally the nurse nucleus at
0
I
I
I I
Experimental
I ‘?
Fig. 10.-A exceptional
Cell Research 13
camera lucida drawing chromosomal detail.
of a stage 8 nurse cell nucleus which shows
Irradiated fly ovary
Fig. 11.--i\ photomicrograph of cytokinesis in a stage 9 nurse chamber. Note the cleavage furrow which is pinching the chromatin into two unequal masses. A, Bright field. B, Phase contrast.
stage 8 is completely tilled with chromosomal material and therefore looks amorphous. However, the nucleus in question was relatively gigantic and the chromosomes within were clearly visible. A given chromosome segment shows marked variations in circumference which appear to result from variations in coiling. Certain regions show what appears to be a series of bands, but it is impossible to rule out the possibility that these may represent a compact series of gyres. Also in the irradiated material an instance was observed of cytokincsis in a stage 9 nurse chamber. Fig. 11 shows the nurse cell nucleus in question as it is pinched in two by the cleavage furrojv. DISCUSSION
The limited state of our present knowledge makes it impossible to make any valid statements as to the specific mechanisms involved in producing the abnormalities observed. About all one can say is that current studies in developmental genetics have led to the concept that the developmental activities of cells (growth, differentiation, migration, and controlled division) are the result of the interactions of various portions of the genome of given cells with specific molecules arranged in specific gene controlled cytoplasmic patterns. Perhaps the abnormalities observed result from radiation-induced Experimental
Cell Research 13
552
R. C. King
inactiviation or loss of various portions of the genetic machinery essential to such developmental sequences. For such an injured cell and its progeny normal developmental behavior is impossible, and depending on the specific genes removed from action there follows a specific type of abnormal behavior. SUMMARY
A cytological study was made of the ovaries of 12-day-old Drosophila melanogaster given 4000 r Co63 gamma rays shortly after emergence. Many severely damaged oocytes underwent pycnotic degeneration. The number of developing eggs was reduced to 85 per cent the control value. Many of the surviving oocytes showed abnormalities which are ascribed to radiation induced disturbances of cell growth, division, migration and differentiation. The author is grateful for the conscientious Samuels. Thanks are also due to Dr. Malcolm facilities.
assistance performed by Mr. Jack Dole for providing the irradiation
REFERENCES 1. BEATTY,
2. KING, 3. KISG,
R. .4., Proc. Roy. Sot. Edinbrrrgh R. C., Growth 21 (1957). R. C., Run~,~sos, A. C. and SMITH,
Experimental
Cell Resenrcl~ 13
63B,
249 (1949).
R. F., Growth
20, 121 (1956).