Number of rat ova implanting after sub-sterilizing X-irradiation of one or both ovaries.код для вставкиСкачать
Number of R a t Ova Implanting after Sub-sterilizing X-irradiation of One or Both Ovaries' ROBERT H. KREHBIEL AND JAMES C. PLAGGE Department of Anatomy, University of Illinois, College of Medicine, Chicago, Illinois Irradiation with €4001 of x-ray was applied to the exteriorized left ovary, to both ovaries or to the left ovary after unilateral (right) castration of 60 day old rats. Treated females were placed with the male at 60, 70, 80 or 90 days of age. Females were explored at eight or nine days after sperm were found in the morning smears to determine the number of implantations. The number of matings was reduced during the period of normal vaginal cycles. Matings did occur after the unset of continuous vaginal cornification. Animals with an irradiated left and a normal right ovary maintained an average number of implantations through pregnancies one to seven. The normal ovary comsensated for the loss of production by the irradiated ovary. Bilateral irradiation caused a decrease in implantations in the second and third pregnancies. In individual cases, however, the normal complement for a single ovary occurred as late as 201 days after irradiation. The irradiated ovary in the absence of the other ovary compensated by producing more implantations; its period of production was limited to 31 days following irradiation. ABSTRACT Large doses of x-ray to the rat ovary cause changes in the estrous cycle, and in time, produce an infertile animal. Mandl ('59a) noted occasional mating by the rat after a sterilizing dose of x-ray. These matings did not produce pregnancy although pseudopregnancy did occur. Ingram ('59), using smaller doses of x-ray, obtained pregnancies and concluded that the length of the fertile period thereafter is inversely associated with the dosage. He also suggested that the decrease in fertility was due to a reduction of available ova. Similar findings have been reported by others (Drips and Ford, '32; Rugh and Wolff, '57 and Nuzhdin et al., '55). All of these observations were made using animals in which the entire ovarian tissue was exposed to irradiation. Pencharz and Long ('32) irradiated only one ovary in the rat and found that subsequent pregnancies were produced and maintained by the untreated ovary. Krehbiel and Plagge ('62), on the other hand, observed implantations on the irradiated side after unilateral exposure to x-ray. Therefore, in the present investigation, both bilateral and unilateral irradiation of the ovary with a sub-sterilizing dose (800 r ) were undertaken in an attempt to clarify the effects on fertility. MATERIAL AND METHODS With the animal in a protective lead shield, the ovaries were exteriorized and irradiated with 800 r at 60 days of age in the manner described previously (Plagge and Krehbiel, '62). The left ovary of 22 rats and both ovaries of 22 rats were irradiated. The third group of 12 rats had the left ovary irradiated and the right ovary removed. Vaginal smears were obtained beginning two weeks prior to irradiation and were continued through the period of observation. Some females were placed with the male the evening of the day of irradition. Others were placed with the male for the first time 10, 20 or 30 days later. The number of implantations was determined by exploring the female eight or nine days after sperm were found in the morning smear. If pregant, the animal was isolated until she delivered. The young were examined superficially for abnormalities and sacrificed. In some cases the female was returned to the male cage during the postpartum period. Cycles and mating were observed up to 300 days following irradiation. 3 Aided by grants from National Institutes of Health, United States Public Health Service and the University of Illinois Research Board, The Graduate College. 257 258 ROBERT H . KREHBIEL AND JAMES C . PLAGGE OBSERVATIONS AND DISCUSSION Vaginal cycles in all animals continued more or less regularly for two months following irradiation. During this time, in animals exposed to the male, there were periods of vaginal cornification in which no sperm were found. During the early post-irradiation period (1-60 days) sperm were seen 59 times in 127 observations of vaginal cornification in females with an irradiated and an untreated ovary. Comparable results were obtained when both ovaries were irradiated (41 in 11 1 smears). In unilaterally castrated and irradiated females, sperm were found in 27 of 60 cycles. These ratios may indicate a lack of coordination of vaginal and ovarian activity, as reported by Mandl ('59b), or perhaps, a failure of the ovaries to produce a normal psychic estrus consistently. Matings occurred during the period of regular cycling and also after the onset of continuous vaginal cornification. No definite pattern was seen in the freqeuncy of the later matings. In a few cases, the females accepted the male for periods of three or four days. There also was a suggestion that these animals were mating at cyclic intervals (fig. 1; no. 1,034 and 1,048). The untreated right ovary in unilaterally irradiated females usually insured pregnancy (table 1). Following each of the 50 fertile matings, implantations were found on the non-irradiated side whereas they occurred in only 26 uteri of the irradiated side. When both ovaries were irradiated there was a marked drop in fertile matings. However, many of the 98 negative matings occurred after irregular cycles or continuous cornification had started. In females which were unilaterally castrated and irradiated the infertility Fig. 1 Typical vaginal smear records of female rats receiving 800 r of x-ray to the left ovary and the right ovary removed (1,033, 1,034); to both ovaries (1,041, 1,048) and to the left ovary only (1,004). The heavy vertical line indicates the irradiation date (60 days of age); m, date female was placed in male cage; rn-, date female was removed from male cage. Sperm i n the morning smear is indicated by a small vertical line above the smear record. Number of implantation sites in the right and left uterine horns is shown by numbers separated by a dash. Individual numbers indicate the number of young found a t birth. ? I 1 - IMPLANTING OVA AFTER X-RAY was even more marked. In these females, only five of 12 animals produced implantations and no positive matings occurred after 31 days following irradiation. TABLE 1 Matings following irradiation of ovaries 800 r to No. of animals Fertile matmgs Sterile matings Both Left R. out 22 22 12 50 27 5 9 98 ovary Left 53 When only the left ovary was exposed the average number of implantations per pregnancy was rather constant (table 2). As the number of implantations on the irradiated side (left) decreased, there was a corresponding increase in the productivity of the normal ovary. There was, however, a period of three to four weeks following irradiation in which the productivity of the irradiated ovary was comparable to that of an untreated ovary. In ten pregnancies of this period the left ovary produced 54 implantations in contrast to the 63 produced by the normal ovary. The irradiated ovary in five of these animals produced as many implantations, or more, than the normal ovary. Furthermore, one animal (1,002) accepted the male two days after irradiation and two others (1,003 and 1,004) mated at three days. These three females had been in estrus two days prior to irradiation and the irradiated ovaries produced a total of 15 ova, which implanted, at the subsequent ovulation. Whatever the damage or other effects of irradiation on the maturation of the ovum are (Harvey and Chang, '62; Mandl, '59c; Spalding et al., '52; Russell and Russell, '56 and others) the irradiation of these three ovaries did not alter the estrous cycles nor the number of viable ova produced. There was a marked decrease in the number of implantations on the left side after the three to four week period. This was also observed in subsequent pregnancies. However, ova were implanted on the irradiated side as late as 213 days folfollowing irradiation. The decreased fertility of the irradiated ovary may have been due to a decrease in the number of 259 ova as suggested by Ingram ('58). On the other hand, Plagge and Krehbiel ('56) noted that the irradiated ovary did not respond as well to gonadotrophins supplied by a castrate parabiont in the presence of the normal ovary. Also, Fels ('61) noted that the damaged ovary reacted differently in the presence of a normal ovary. This suggests a threshold difference in the ovarian response to gonadotrophins after the effects of unilateral irradiation become apparent. During the early post irradiation period a normal number of implantations also occurred after bilateral irradiation. Thereafter, these ovaries had a marked decrease in productivity. There were instances, however, of a single ovary producing a normal complement of implanting ova at relatively late dates following irradiation. Examples were : four implantations on days 93 and 145 (1,041), five on day 201 ( 1,039) seven on days 75 (1,040) and 134 (1,039). In most of these instances the opposite irradiated ovary failed to produce an implantation. This may have been due to the gonadotrophic stimulus being directed to but one ovary, or a given dose of irradiation may not be uniform in its effects, with one ovary being damaged more than the other. This cannot, however, account for an animal (1,039) in which all implantations of the second pregnancy were on the right side and those of the third pregnancy on the left side. Also, the left ovary of another female ( 1,041) produced two implantating ova at 145 days after failing to contribute to earlier pregnancies. The present evidence does not provide an adequate explanation for these variations in ovarian activity. When only one ovary produced implantations after bilateral irradiation a compensatory increase in numbers was not generally observed. This was not due to the length of the post irradiation period since the average number of implantations was 4.8 in five such pregancies during the first 30 days after irradiation. The second ovary may have ovulated ova which did not implant. On the other hand, even if not productive, this ovary may have utilized its share of the gonadotrophins. The decrease in response to gonadotrophins by irradiated ovaries (Ingram and Mandl, '58) 260 ROBERT H. KREHBIEL AND JAMES C. PLAGGE TABLE 2 Implantations per pregnancy after irradiation of ovaries Pregnancy number 2 1 No. Days post r Imp. in R. L. Days post r 4 3 Imp. in R. L. Days post r Imp. in R. L. 5 Days post r Imp. R. inL. Days r post R.in L. Imp. - ~ 800 T to left ovary 1002 1003 1004' 1001 1007 1006 1009 1005 1016 1008 1012 865 866 1018 1023 1019 1025 1021 1020 894 855 2 3 3 11 12 16 18 21 21 23 27 31 34 38 47 63 72 74 84 145 171 7 9 7 6 5 4 6 6 9 4 5 9 10 5 8 8 8 9 11 10 7 153 AvJpreg. 1060 1057 1059 1063 1061 1064 1056 1058 1041 867 1042 1048 1039 1040 1044 1043 7 8 12 12 15 15 17 22 23 32 34 41 47 75 76 77 5 6 4 0 7 7 6 7 3 9 2 3 3 2 0 0 0 0 2 1 0 67 10.5 7 8 6 4 5 0 0 5 2 7 7 4 8 6 7 3 1 72 Av./preg. 9 4 0 4 3 0 5 0 6 5 5 5 1 0 0 55 7.9 38 44 8 4 11 2 81 81 12 0 72 134 123 13 0 9 2 201 154 80 8 0 46 68 70 87 80 69 78 88 84 80 94 10 6 10 14 11 5 2 130 12 0 213 72 154 4 0 138 1 0 0 0 0 2 0 0 0 0 95 119 141 92 9 0 71 123 8 0 123 11 0 10 0 201 81 7 10 14 7 11 135 11 10.4 71 6 9.6 41 800 r to both ovaries 4 2 45 1 45 0 6 96 02 56 93 4 0 4 0 117 145 1 0 42 80 134 2 1 7 0 201 0 5 38 2 10.0 23 2 8.3 1 22 10 4.6 59 3.5 800 T to left w a r y - right ovary removed 1027 1068 1029 1031 1033 Av./preg. 8 10 11 20 31 7 10 4 9 12 42 8.4 1 Had sixth pregnancy at 205 days with 11-1, and a seventh pregnancy at 236 days with 9-0 implantations (see fig. 1). IMPLANTING OVA AFTER X-RAY and the decrease in the number of eggs in the ovary (Ingram, '58) should also be considered. When both ovaries produced during this period, however, the number of implantations was normal. In contrast to the extended period of fertility seen occasionally in bilaterally irradiated females, fertility in rats with irradiated left ovary and with the right ovary removed was limited to a very short period. No pregnancies could be initiated in this group after 31 days post irradiation. The average number of implantations was 8.4. This number is significantly greater ( p < 0.02) than the number of implantations produced by the irradiated ovary in the presence of a normal or a n irradiated ovary during a comparable post irradiation period. It is evident that the irradiated ovary during this period can compensate by producing more ova if the other ovary has been removed. There is no evident explanation for the failure of these animals to implant at a later date. Resorptions were noted at the time of exploration. No attempt was made to determine the time or the cause of these. The single daily observation of the isolated female did not give an accurate count of the number of young born. There was, however, attrition of some magnitude between the implantation and birth periods. An occasional prolonged pregnancy, terminating with a difficult labor, was noted. LITERATURE CITED Drips, D. G., and F. A. Ford 1932 The study of the effects of roentgen rays on the estrual cycle and the ovaries of the white rat. Surg. Gyn. and Obst., 55: 596-606. Fels, E. 1961 La ligadura unilateral del pediculo ovarico. Rev. SOC.Argentina Biol., 37: 4447. 26 1 Harvey, E. B., and M. C. Chang 1962 Effects of x-irradation of ovarian ova of the golden hamster. Anat. Rec., 142: 239. Ingram, D. L. 1958 Fertility and oocyte numbers after x-irradiation of the ovary. J. Endocrin., 17: 61-90. Ingram, D. L., and Anita M. Mandl 1958 The hypophysial control of the x-ray sterilized ovary. Ibid., 17: 1-12. Krehbiel, R. H., and J. C. Plagge 1962 Distribution of ova in the rat uterus. Anat. Rec., 143: 239-242. Mandl, Anita M. 1959a Mating behaviour of the adult rat after x-ray sterilization. J. Endocrin., 18: 434437. 195913 The oestrous cycle of the adult rat after x-ray sterilization. Ibid., 18: 426-433. 1959c A quantitative study of the sensitivity of oocytes to x-irradiation. Proc. Roy. SOC.B, 150: 53-71. Nuzhdin, N. I., N. I. Shapiro, 0. N. Petrova and 0. N. Kitaeva 1955 Effect of ionizing radiations on the fertility of mice and the viability of their progeny. C o d . Acad. Sci., USSR on the peaceful uses of atomic energy. U. S. Atomic Energy Com. Washington 1956, 4: 21-31. Pencharz, R. I., and J. A. Long 1932 The effect on a subsequent pregancy after x-raying one ovary of a rat. Amer. J. Anat., 50: 1-8. Plagge, J. C., and R. H. Krehbiel 1956 Histology of irradiated ovaries before and after stimulation by gonadotrophins from a castrate parabiont rat. Anat. Rec., 124: 348. 1962 Vaginal cycles in the rat after xray or other damage to one or both ovaries. Ibid., 144: 287-293. Roberts, Rough and Joan Wolff 1957 Threshold x-irradiation sterilization of the ovary. Fert. Steril., 8: 428437. Russell, Liane B., and W. L. Russell 1956 The sensitivity of different stages i n oogenesis to the radiation induction of dominant lethals and other changes in the mouse. Progress i n radiobiology. Mitchell, Holmes and Smith. Edinburgh, Oliver and Boyd, 187-195. Spalding, J. F., J. M. Wellnitz and W. H. Schweitzer 1957 The effects of high dosage x-ray on the maturation of the rat ovum and their modification by gonadotrophins. Fert. Steril., 8: 80-88.