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Glycogen Accumulation in the Rat Uterus Containing an Intrauterine Contraceptive Device
Glycogen Accumulation in the Rat Uterus Containing an Intrauterine Contraceptive .Device EARL PARR, B.A. *
...
..
l ,"
toward an understanding of the phenomenon of intrauterine contraception occurred when Carleton and Phelps studied the effects of placing various inert objects (silver, gold, nickel, and rubber rings; rubber strips; silkworm gut; silk thread; and celluloid thread) into one horn of the bicornuate uterus of the rabbit. The uterine horn lacking the contraceptive device became normally pregnant, while pregnancy in the IUCD-containing horn was suppressed. The extent of suppression was related to the size of the device. In fact, if only a portion of a uterine horn contained an IUCD, then pregnancy could occur in the device-free region. In confirming and extending the findings of Carleton and Phelps, Adams and Eckstein reported that ovulation, fertilization, early cleavage, and tubal transport of ova appeared to be essentially normal. Mortality of ova occurred only in the uterus, both before and after implantation. Doyle and Margolis have reported that the presence of an IUCD in the rat had no effect on fertilization and tubal transport of ova, but that once the blastocysts entered the uterus, they could no longer be recovered. With the type of IueD used by these investigators, no pregnancy occurred in any part of the device-containing uterus, while the control horn became normally pregnant. It has been observed, however, that when a smaller (a few millimeters) IUCD is placed in one end of a rat uterine horn, a pregnancy may occur at the opposite end, especially if the IUCD has been in place for only about a week or lessY This observation establishes that an IUCD can cause qualitatively similar effects in rats and rabbits. Greenwald has tried to determine the fate of blastocysts that disappear from an IUCD-containing rat uterus. He reported the finding of what he THE FIRST MAJOR ADVANCE
From the Laboratory of the Population Council, Rockefeller University, New York, N. Y. *Graduate fellow of the Rockefeller University.
797
798
PARR
FERTILITY
& STERILITY
considered to be fragments of dead and degenerating blastocysts in the cervix. The studies cited above indicate the following: (1) The IUeD exerts a local effect in the uterus, more localized in the rabbit than in the rat. (2) The effects of the IueD on pregnancy become apparent only at about the time the blastocysts are entering the uterus in these species. Secretory activity in the mammalian uterus begins just prior to the arrival of blastocysts and is a local phenomenon. The two conclusions reached above, therefore, seem to apply to secretory activity as well as the IueD. In the rat, another secretory stage appears to occur at the height of estrus, but we are presently concerned with the stage that begins a few days after mating. This secretory activity is quite likely to have a function in providing nourishment for the unattached blastocysts lying free in the uterine lumen. Evidence comes from three sources: (1) glycogen content of uterine epithelial cells and their transition from cuboidal to columnar;8, 1~ (2) metabolic requirements for blastocysts cultured in vitro;3, 4, 12, 15 and ( 3) studies of cases of sterility that are well correlated to a low endometrial glycogen content. 14 It was of interest, therefore, to examine the secretory activity of normal and IUeD-containing rat uteri just prior to nidation. The height of the epithelial cells and the presence of glycogen in the endometrium were chosen as indicators of secretory activity. The histologic studies presented in this paper show that a normal columnar secretory epithelium does not develop in the rat uterus containing an IUeD, and a rather large quantity of glycogen, as compared to that in the normal uterus, accumulates in the epithelial cells and near the epithelial basement membrane. Further studies were directed at discovering when these changes first appear, how long they persist, and which ovarian hormones, if any, are involved. MATERIALS AND METHODS
Female rats of the Holtzman strain were used. IUeD's consisted of 3-0 surgical silk thread inserted with an atraumatic needle into one horn. One end of the thread was a knot drawn up against the serosal surface, and about 1 cm. of thread passed along the lumen. The other end was brought outside the uterus, where about 1 cm. was left hanging. Roughly 200 IUeD's have now been inserted this way without a case of spontaneous loss. The devices were in situ at least 2 weeks before the animals were used. When the animals were mated, the dav sperm were found in the vagina was designated as Day 0, or Lo. In all experiments that required the assay of glycogen, uterine tissue was fixed in absolute ethanol satu-
..
VOL.
17, No.6, 1966
GLYCOGEN IN RAT UTERUS
799
rated with picric acid. Sections of the tissue were stained with the PAS technic given by Barka and Anderson, and the presence of glycogen was confirmed with the diastase digestion recommended by them. Histochemical technics were applied to IUCD-carrying females in three conditions: 1. Sacrificed at 12-hr. intervals from 9 P.M. of Ll to 9 A.M. of L6 2. Sacrificed in estrus and diestrus; on days L 10 ) L H ;, and L 20 of pregnancy; and on the first 3 days postpaTtum 3. Ovariectomized on Lo and treated subsequently with: (a) 5 mg. progesterone S.c. daily and 100 /-tg. estradiol-17.B S.c. on L 3 ; (b) 5 mg. progesterone S.C. daily; (c) 100 /-tg. estradiol-17.B S.c. on L;,; or (d) no hormone therapy. All animals in this group were sacrificed on L 4 • RESULTS
,
..
... ...
..
..
Except for the ovariectomized rats, the appearance of all tissue sections from all IUCD-containing horns from 9 P.M. of L3 through 9 A.M. of L6 was essentially the same. One typical section is shown in Fig. 1 and is characterized by the accumulation of glycogen in and near the epithelium and the failure of the epithelial cells to become columnar. A section of tissue from the appropriate control horn (Fig. 2) shows a secretory epithelium and almost no glycogen. Before 9 P.M. of La' however, the differences between device-containing and control tissues were less pronounced. Unfortunately, the histochemical preparations do not provide quantitative information on the amount of glycogen present, so the transition from smaller to larger deposits must remain somewhat vague. What does appear obvious from these preparations, however, is that the amount of glycogen present in device-containing tissues from 9 P.M. of Ll is decidedly less than the amount which is found at 9 P.M. of La and later. Control uterine horns never accumulated any glycogen in the epithelial region at any time prior to implantation, and they were not examined after L 6 • The accumulation of glycogen in devicecontaining tissues is therefore always relative to no accumulation in the control horn. Table 1 gives the number of animals examined at each stage and includes the results obtained from the animals in later stages of pregnancy and in the estrus cycle. The small amount of glycogen that was typical of Ll was present in the animals in diestrus, but appeared in only 1 of the 4 animals in estrus. The larger deposit of glycogen, typical of L., was maintained throughout pregnancy and was still present in the device-
800
PARR
FERTILITY
& STERILITY
containing horn 2 days after the control horn had delivered its pups. Moreover, in estrus, horns containing IUeD's had columnar epithelium identical to that in control horns at the same stage. This finding was in contrast to the situation in L4 of pregnancy where only control tissues had columnar epithelium; device-containing tissues had an epithelium that might be described as pseudostratified. In many of the observations discussed above, tissue was taken from 2 regions of the IUeD-containing horn: the region in contact with the IueD and a region remote from the device. The appearance of sections from both kinds of tissue was the same, except that some epithelium of the
Fig. 1. rUCD-contain· ing horn at 9 A.M. of L 4 , showing accumulation of glycogen as black clumps in epithelial cells and in stromal cells bordering closely on epithelial basement membrane. Epithelial cells lack columnar formation and nuclei are not migrating. (X 520)
Fig. 2. Control horn at 9 A.M. of L 4 , showing columnar epithelial cells with nuclei migrating toward uterine lumen. No glycogen can be seen. (X 520)
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.
.
VOL.
17, No.6, 1966
801
GLYCOGEN IN RAT UTERUS
TABLE 1. Glycogen Accumulation and Retention in IUCD-containing Rat Uteri
•
Time sacri jiced
No. of animals
Glycogen*
7
+ +
Diestrus
{~
Estrus
5
9 P.M. Ll 9 A.M. L2 to 9 A.M. L3 9 P.M. L3 to postpartum 2
28 29
0
+ ++ +++ +++. ++ for
*The accumulation of glycogen in and around the epithelium is scaled from 0 to The value 0 is used when no glycogen can be detected; is for small amounts; intermediate values; and for the largest accumulations.
+
+++
tissue in contact with the device had often been torn away. This observation is pertinent to the fact that both regions are sterile. All the 19 ovariectomized animals that received hormone therapy had the maximum, or 3 plus, accumulation of glycogen on L 4 • The 7 that received no hormone therapy had the small, or 1 plus, accumulation that was characteristic of diestrus. Apparently either estrogen or progesterone can stimulate the deposition of glycogen in the epithelial region in these conditions. DISCUSSION
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.. ...
In the endometrium of the rat up to L6 of pregnancy, glycogen never accumulates except in the presence of an IUeD. This absence of any histochemically demonstrable glycogen in the endometrium of control uteri is consistent with the biochemical finding of Kostyo that the glycogen concentration of the pregnant rat uterus remains at the diestrus level through Ls' This is in contrast to primate uteri, where, during the normal secretory stage, glycogen does accumulate in the endometrium. l3 What effect an IUCD might have on endometrial glycogen in primates is still unexplored . To the extent that differences in glycogen accumulation can be visually estimated from stained sections, the quantity of glycogen in the epithelial region of IUCD-containing uteri increases during the interval between Ll and L 4 • During this period, the development of a clear morphologic difference between the epitheliums of device-containing and control uteri suggests that control epithelium is engaged in secretion and device-containing epithelium accumulates glycogen because the latter is not being released into the lumen. This possibility can only be offered as a reason-
802
PARR
FERTILITY
&
STERILITY
able hypothesis, however, since fundamental information about the secretory process in the normal uterus is lacking. Apart from the suggested hypothesis, the accumulation of glycogen by 9 A.M. of L4 is a clear and convenient marker for the activity of the device, which can be used in experimental designs where it is not possible to assay the activity of the device by its effect on pregnancy. The abnormality in uterine glycogen metabolism is observed before blastocysts enter the uterus, and hence is compatible with the fact that blastocysts die a few hours after their arrival. There remains, however, a most perplexing question. If glycogen has accumulated because of some disturbance of the normal secretory process, why are small amounts present during diestrus? To confirm the histochemical findings presented here, it would be desirable to extract and measure the glycogen biochemically. The biochemical measurement is greatly complicated, however, by the finding that both wet and dry weights of the IUCD-containing uterus are increased over control values. 10 The increase in dry weight implies an increase in tissue, quite possibly in the number of cells. Finding a component of the devicecontaining tissue that remains at the control levels is thus a problem. A further difficulty is the measurement of glycogen which is localized in the epithelial region, since the normal myometrial glycogen in a whole-uterine extract may swamp the epithelial deposit. SUMMARY
Female rats having an IUeD in one uterine horn were sacrificed in estrus and diestrus, and on various days of pregnancy. Uterine tissue was examined histologically with the PAS technic. Control sections showed normal epithelium and contained very little glycogen. Sections from IUCD-containing uteri in estrus and diestrus also showed normal epithelium, but a small deposit of glycogen had accumulated in the epithelium and near the epithelial basement membrane of the uteri in diestrus. The same description applies to animals sacrificed at 9 P.M. of L l • Between 9 P.M. of Ll and 9 A.M. of L 4 , the accumulation of glycogen gradually increased. By 9 A.M. of L4 the glycogen accumulation became a conspicuous feature of the epithelium in IUCD-containing uteri. In addition, this epithelium was pseudostratified, while control epithelium at this stage was columnar. The glycogen deposit was maintained through the remainder of pregnancy and was still present 2 days after pups had been delivered from the control horn. On the basis of these observations, it is suggested that secretion may be
VOL. 17, No.6, 1966
GLYCOGEN IN RAT UTERUS
803
defective in IUCD-containing horns of the rat. The consequences of abnormal uterine secretion would be consistent with both the timing and the local action of the IUCD in the rat. The Rockefeller University New York, N. Y.10021
r
REFERENCES
..
•
•
•
1. ADAMS, C. E., and ECKSTEIN, P. The effect of intrauterine foreign bodies on pregnancy in the rabbit. Fertil Steril16:508, 1965. 2. BARKA, T., and ANDERSON, P. J. Histochemistry: Theory, Practice, Bibliography. Hoeber, New York, 1963. 3. BRINSTER, R. L. Studies on the development of mouse embryos in vitro. II. The effect of energy source. I Exp Zool158:59, 1965. 4. BRINSTER, R. L. Studies on the development of mouse embryos in vitro. TIl. The effect of fixed-nitrogen source. I Exp Zool158:69, 1965 . 5. CARLETON, H. M., and PHELPS, H. J. Experimental observations on the Grafenberg ring contraceptive methods. I Obstet Gynaec Brit Emp 40:81, 1933. 6. DOYLE, L. L., and MARGOLIS, A. J. Intra-uterine foreign body: effect on pregnancy in the rat. Science 139:833, 1963. 7. GREENWALD, G. S. Interruption of pregnancy in the rat by a uterine suture. I Reprod Fertil 9:9, 1965. 8. HAM, A. W. Histology (ed. 5). Lippincott, Philadelphia, 1965. 9. KosTYo, J. L. A study of the glycogen levels of the rat uterus and certain skeletal muscles during pregnancy. Endocrinology 60:33,1957. 10. PARR, E. L., and SEGAL, S. J. The effect of an intrauterine contraceptive device on the weight of the rat uterus. Fertil Steril17:648, 1966. 11. PARR, E. L. Unpublished data. 12. PINCUS, G., and PuRsHOTTAM, N. In vitro cultivation of mammalian eggs. Anat Rec 140:51, 1961. 13. REYNOLDS, S. R. M. Physiology of the Uterus (ed. 2). Hafner, New York, 1965, p.422. 14. RUBULIS, A., JACOBS, R. D., and HUGHES, E. C. Glycogen synthetase in mammalian uterus. Biochim Biophys Acta 99:584, 1965. 15. WmTTEN, W. K. Culture of tubal ova. Nature 179:1081, 1957.
...
..
l ,"
toward an understanding of the phenomenon of intrauterine contraception occurred when Carleton and Phelps studied the effects of placing various inert objects (silver, gold, nickel, and rubber rings; rubber strips; silkworm gut; silk thread; and celluloid thread) into one horn of the bicornuate uterus of the rabbit. The uterine horn lacking the contraceptive device became normally pregnant, while pregnancy in the IUCD-containing horn was suppressed. The extent of suppression was related to the size of the device. In fact, if only a portion of a uterine horn contained an IUCD, then pregnancy could occur in the device-free region. In confirming and extending the findings of Carleton and Phelps, Adams and Eckstein reported that ovulation, fertilization, early cleavage, and tubal transport of ova appeared to be essentially normal. Mortality of ova occurred only in the uterus, both before and after implantation. Doyle and Margolis have reported that the presence of an IUCD in the rat had no effect on fertilization and tubal transport of ova, but that once the blastocysts entered the uterus, they could no longer be recovered. With the type of IueD used by these investigators, no pregnancy occurred in any part of the device-containing uterus, while the control horn became normally pregnant. It has been observed, however, that when a smaller (a few millimeters) IUCD is placed in one end of a rat uterine horn, a pregnancy may occur at the opposite end, especially if the IUCD has been in place for only about a week or lessY This observation establishes that an IUCD can cause qualitatively similar effects in rats and rabbits. Greenwald has tried to determine the fate of blastocysts that disappear from an IUCD-containing rat uterus. He reported the finding of what he THE FIRST MAJOR ADVANCE
From the Laboratory of the Population Council, Rockefeller University, New York, N. Y. *Graduate fellow of the Rockefeller University.
797
798
PARR
FERTILITY
& STERILITY
considered to be fragments of dead and degenerating blastocysts in the cervix. The studies cited above indicate the following: (1) The IUeD exerts a local effect in the uterus, more localized in the rabbit than in the rat. (2) The effects of the IueD on pregnancy become apparent only at about the time the blastocysts are entering the uterus in these species. Secretory activity in the mammalian uterus begins just prior to the arrival of blastocysts and is a local phenomenon. The two conclusions reached above, therefore, seem to apply to secretory activity as well as the IueD. In the rat, another secretory stage appears to occur at the height of estrus, but we are presently concerned with the stage that begins a few days after mating. This secretory activity is quite likely to have a function in providing nourishment for the unattached blastocysts lying free in the uterine lumen. Evidence comes from three sources: (1) glycogen content of uterine epithelial cells and their transition from cuboidal to columnar;8, 1~ (2) metabolic requirements for blastocysts cultured in vitro;3, 4, 12, 15 and ( 3) studies of cases of sterility that are well correlated to a low endometrial glycogen content. 14 It was of interest, therefore, to examine the secretory activity of normal and IUeD-containing rat uteri just prior to nidation. The height of the epithelial cells and the presence of glycogen in the endometrium were chosen as indicators of secretory activity. The histologic studies presented in this paper show that a normal columnar secretory epithelium does not develop in the rat uterus containing an IUeD, and a rather large quantity of glycogen, as compared to that in the normal uterus, accumulates in the epithelial cells and near the epithelial basement membrane. Further studies were directed at discovering when these changes first appear, how long they persist, and which ovarian hormones, if any, are involved. MATERIALS AND METHODS
Female rats of the Holtzman strain were used. IUeD's consisted of 3-0 surgical silk thread inserted with an atraumatic needle into one horn. One end of the thread was a knot drawn up against the serosal surface, and about 1 cm. of thread passed along the lumen. The other end was brought outside the uterus, where about 1 cm. was left hanging. Roughly 200 IUeD's have now been inserted this way without a case of spontaneous loss. The devices were in situ at least 2 weeks before the animals were used. When the animals were mated, the dav sperm were found in the vagina was designated as Day 0, or Lo. In all experiments that required the assay of glycogen, uterine tissue was fixed in absolute ethanol satu-
..
VOL.
17, No.6, 1966
GLYCOGEN IN RAT UTERUS
799
rated with picric acid. Sections of the tissue were stained with the PAS technic given by Barka and Anderson, and the presence of glycogen was confirmed with the diastase digestion recommended by them. Histochemical technics were applied to IUCD-carrying females in three conditions: 1. Sacrificed at 12-hr. intervals from 9 P.M. of Ll to 9 A.M. of L6 2. Sacrificed in estrus and diestrus; on days L 10 ) L H ;, and L 20 of pregnancy; and on the first 3 days postpaTtum 3. Ovariectomized on Lo and treated subsequently with: (a) 5 mg. progesterone S.c. daily and 100 /-tg. estradiol-17.B S.c. on L 3 ; (b) 5 mg. progesterone S.C. daily; (c) 100 /-tg. estradiol-17.B S.c. on L;,; or (d) no hormone therapy. All animals in this group were sacrificed on L 4 • RESULTS
,
..
... ...
..
..
Except for the ovariectomized rats, the appearance of all tissue sections from all IUCD-containing horns from 9 P.M. of L3 through 9 A.M. of L6 was essentially the same. One typical section is shown in Fig. 1 and is characterized by the accumulation of glycogen in and near the epithelium and the failure of the epithelial cells to become columnar. A section of tissue from the appropriate control horn (Fig. 2) shows a secretory epithelium and almost no glycogen. Before 9 P.M. of La' however, the differences between device-containing and control tissues were less pronounced. Unfortunately, the histochemical preparations do not provide quantitative information on the amount of glycogen present, so the transition from smaller to larger deposits must remain somewhat vague. What does appear obvious from these preparations, however, is that the amount of glycogen present in device-containing tissues from 9 P.M. of Ll is decidedly less than the amount which is found at 9 P.M. of La and later. Control uterine horns never accumulated any glycogen in the epithelial region at any time prior to implantation, and they were not examined after L 6 • The accumulation of glycogen in devicecontaining tissues is therefore always relative to no accumulation in the control horn. Table 1 gives the number of animals examined at each stage and includes the results obtained from the animals in later stages of pregnancy and in the estrus cycle. The small amount of glycogen that was typical of Ll was present in the animals in diestrus, but appeared in only 1 of the 4 animals in estrus. The larger deposit of glycogen, typical of L., was maintained throughout pregnancy and was still present in the device-
800
PARR
FERTILITY
& STERILITY
containing horn 2 days after the control horn had delivered its pups. Moreover, in estrus, horns containing IUeD's had columnar epithelium identical to that in control horns at the same stage. This finding was in contrast to the situation in L4 of pregnancy where only control tissues had columnar epithelium; device-containing tissues had an epithelium that might be described as pseudostratified. In many of the observations discussed above, tissue was taken from 2 regions of the IUeD-containing horn: the region in contact with the IueD and a region remote from the device. The appearance of sections from both kinds of tissue was the same, except that some epithelium of the
Fig. 1. rUCD-contain· ing horn at 9 A.M. of L 4 , showing accumulation of glycogen as black clumps in epithelial cells and in stromal cells bordering closely on epithelial basement membrane. Epithelial cells lack columnar formation and nuclei are not migrating. (X 520)
Fig. 2. Control horn at 9 A.M. of L 4 , showing columnar epithelial cells with nuclei migrating toward uterine lumen. No glycogen can be seen. (X 520)
..
.
.
VOL.
17, No.6, 1966
801
GLYCOGEN IN RAT UTERUS
TABLE 1. Glycogen Accumulation and Retention in IUCD-containing Rat Uteri
•
Time sacri jiced
No. of animals
Glycogen*
7
+ +
Diestrus
{~
Estrus
5
9 P.M. Ll 9 A.M. L2 to 9 A.M. L3 9 P.M. L3 to postpartum 2
28 29
0
+ ++ +++ +++. ++ for
*The accumulation of glycogen in and around the epithelium is scaled from 0 to The value 0 is used when no glycogen can be detected; is for small amounts; intermediate values; and for the largest accumulations.
+
+++
tissue in contact with the device had often been torn away. This observation is pertinent to the fact that both regions are sterile. All the 19 ovariectomized animals that received hormone therapy had the maximum, or 3 plus, accumulation of glycogen on L 4 • The 7 that received no hormone therapy had the small, or 1 plus, accumulation that was characteristic of diestrus. Apparently either estrogen or progesterone can stimulate the deposition of glycogen in the epithelial region in these conditions. DISCUSSION
.. ...
.. ...
In the endometrium of the rat up to L6 of pregnancy, glycogen never accumulates except in the presence of an IUeD. This absence of any histochemically demonstrable glycogen in the endometrium of control uteri is consistent with the biochemical finding of Kostyo that the glycogen concentration of the pregnant rat uterus remains at the diestrus level through Ls' This is in contrast to primate uteri, where, during the normal secretory stage, glycogen does accumulate in the endometrium. l3 What effect an IUCD might have on endometrial glycogen in primates is still unexplored . To the extent that differences in glycogen accumulation can be visually estimated from stained sections, the quantity of glycogen in the epithelial region of IUCD-containing uteri increases during the interval between Ll and L 4 • During this period, the development of a clear morphologic difference between the epitheliums of device-containing and control uteri suggests that control epithelium is engaged in secretion and device-containing epithelium accumulates glycogen because the latter is not being released into the lumen. This possibility can only be offered as a reason-
802
PARR
FERTILITY
&
STERILITY
able hypothesis, however, since fundamental information about the secretory process in the normal uterus is lacking. Apart from the suggested hypothesis, the accumulation of glycogen by 9 A.M. of L4 is a clear and convenient marker for the activity of the device, which can be used in experimental designs where it is not possible to assay the activity of the device by its effect on pregnancy. The abnormality in uterine glycogen metabolism is observed before blastocysts enter the uterus, and hence is compatible with the fact that blastocysts die a few hours after their arrival. There remains, however, a most perplexing question. If glycogen has accumulated because of some disturbance of the normal secretory process, why are small amounts present during diestrus? To confirm the histochemical findings presented here, it would be desirable to extract and measure the glycogen biochemically. The biochemical measurement is greatly complicated, however, by the finding that both wet and dry weights of the IUCD-containing uterus are increased over control values. 10 The increase in dry weight implies an increase in tissue, quite possibly in the number of cells. Finding a component of the devicecontaining tissue that remains at the control levels is thus a problem. A further difficulty is the measurement of glycogen which is localized in the epithelial region, since the normal myometrial glycogen in a whole-uterine extract may swamp the epithelial deposit. SUMMARY
Female rats having an IUeD in one uterine horn were sacrificed in estrus and diestrus, and on various days of pregnancy. Uterine tissue was examined histologically with the PAS technic. Control sections showed normal epithelium and contained very little glycogen. Sections from IUCD-containing uteri in estrus and diestrus also showed normal epithelium, but a small deposit of glycogen had accumulated in the epithelium and near the epithelial basement membrane of the uteri in diestrus. The same description applies to animals sacrificed at 9 P.M. of L l • Between 9 P.M. of Ll and 9 A.M. of L 4 , the accumulation of glycogen gradually increased. By 9 A.M. of L4 the glycogen accumulation became a conspicuous feature of the epithelium in IUCD-containing uteri. In addition, this epithelium was pseudostratified, while control epithelium at this stage was columnar. The glycogen deposit was maintained through the remainder of pregnancy and was still present 2 days after pups had been delivered from the control horn. On the basis of these observations, it is suggested that secretion may be
VOL. 17, No.6, 1966
GLYCOGEN IN RAT UTERUS
803
defective in IUCD-containing horns of the rat. The consequences of abnormal uterine secretion would be consistent with both the timing and the local action of the IUCD in the rat. The Rockefeller University New York, N. Y.10021
r
REFERENCES
..
•
•
•
1. ADAMS, C. E., and ECKSTEIN, P. The effect of intrauterine foreign bodies on pregnancy in the rabbit. Fertil Steril16:508, 1965. 2. BARKA, T., and ANDERSON, P. J. Histochemistry: Theory, Practice, Bibliography. Hoeber, New York, 1963. 3. BRINSTER, R. L. Studies on the development of mouse embryos in vitro. II. The effect of energy source. I Exp Zool158:59, 1965. 4. BRINSTER, R. L. Studies on the development of mouse embryos in vitro. TIl. The effect of fixed-nitrogen source. I Exp Zool158:69, 1965 . 5. CARLETON, H. M., and PHELPS, H. J. Experimental observations on the Grafenberg ring contraceptive methods. I Obstet Gynaec Brit Emp 40:81, 1933. 6. DOYLE, L. L., and MARGOLIS, A. J. Intra-uterine foreign body: effect on pregnancy in the rat. Science 139:833, 1963. 7. GREENWALD, G. S. Interruption of pregnancy in the rat by a uterine suture. I Reprod Fertil 9:9, 1965. 8. HAM, A. W. Histology (ed. 5). Lippincott, Philadelphia, 1965. 9. KosTYo, J. L. A study of the glycogen levels of the rat uterus and certain skeletal muscles during pregnancy. Endocrinology 60:33,1957. 10. PARR, E. L., and SEGAL, S. J. The effect of an intrauterine contraceptive device on the weight of the rat uterus. Fertil Steril17:648, 1966. 11. PARR, E. L. Unpublished data. 12. PINCUS, G., and PuRsHOTTAM, N. In vitro cultivation of mammalian eggs. Anat Rec 140:51, 1961. 13. REYNOLDS, S. R. M. Physiology of the Uterus (ed. 2). Hafner, New York, 1965, p.422. 14. RUBULIS, A., JACOBS, R. D., and HUGHES, E. C. Glycogen synthetase in mammalian uterus. Biochim Biophys Acta 99:584, 1965. 15. WmTTEN, W. K. Culture of tubal ova. Nature 179:1081, 1957.