Ovarian Ornithine Decarboxylase Research Articles

Can peri-ovulatory putrescine supplementation improve egg quality in older infertile women?

The aging-related decline in fertility is an increasingly pressing medical and economic issue in modern society where women are delaying family building. Increasingly sophisticated, costly, and often increasingly invasive, assisted reproductive clinical protocols and laboratory technologies (ART) have helped many older women achieve their reproductive goals. Current ART procedures have not been able to address the fundamental problem of oocyte aging, the increased rate of egg aneuploidy, and the decline of developmental potential of the eggs. Oocyte maturation, which is triggered by luteinizing hormone (LH) in vivo or by injection of human chorionic gonadotropin (hCG) in an in vitro fertilization (IVF) clinic, is the critical stage at which the majority of egg aneuploidies arise and when much of an egg's developmental potential is established. Our proposed strategy focuses on improving egg quality in older women by restoring a robust oocyte maturation process. We have identified putrescine deficiency as one of the causes of poor egg quality in an aged mouse model. Putrescine is a biogenic polyamine naturally produced in peri-ovulatory ovaries. Peri-ovulatory putrescine supplementation has reduced egg aneuploidy, improved embryo quality, and reduced miscarriage rates in aged mice. In this paper, we review the literature on putrescine, its occurrence and physiology in living organisms, and its unique role in oocyte maturation. Preliminary human data demonstrates that there is a maternal aging-related deficiency in ovarian ornithine decarboxylase (ODC), the enzyme responsible for putrescine production. We argue that peri-ovulatory putrescine supplementation holds great promise as a natural and effective therapy for infertility in women of advanced maternal age, applicable in natural conception and in combination with current ART therapies.

Peri-ovulatory putrescine supplementation reduces embryo resorption in older mice.

Does peri-ovulatory putrescine supplementation of older mice improve oocyte quality and reduce the incidence of embryo resorption? Peri-ovulatory putrescine supplementation in older mice improved oocyte quality, as indicated by increased blastocyst cell numbers and reduced the incidence of embryo resorption. Rodents exhibit a transient rise of ornithine decarboxylase (ODC) and putrescine in the ovaries during ovulation. Older mice exhibit reduced ovarian ODC activity during ovulation. Supplementation of in vitro maturation medium with putrescine reduces oocyte aneuploidy rates of older mice. The rationale was to correct ovarian putrescine deficiency in older mice by peri-ovulatory putrescine supplementation in drinking water and to observe the reproductive consequences of this intervention. This project was conducted between 2010 and 2014. Older mice (9-11 months of age) were given regular drinking water (control) or drinking water with 1% putrescine dihydrochloride (62 mM) for 2-4 days before mating. Plugged mice were then withdrawn from putrescine supplementation. Blastocysts were retrieved on 3.5 days post coitum (dpc) for the determination of cell numbers. For resorption analyses, mice were killed on 9.5 dp or 12.5 dpc, and implantation sites were dissected to determine the embryo status. For birth studies, mice were examined every morning between 16.5 and 23.5 dpc. Births were recorded as live or stillbirth. We demonstrated that deficiency of ovarian putrescine in older mice can be restored by peri-ovulatory putrescine supplementation in drinking water. Putrescine supplementation in older mice increased blastocyst cell numbers (from 40 to 54; P < 0.0001, t-test), reduced embryo resorption rates (from 41.1 to 15.4% in old C57BL/6 mice, P < 0.0001, Fisher's exact test; from 14.2 to 6.4% in old CF1 mice, P = 0.004, Fisher's exact test), and doubled the number of live born pups. Furthermore, exogenous putrescine exhibited rapid absorption and excretion, and showed no toxicity to mothers or fetuses. The mechanism of putrescine action in oocytes and/or ovaries remains unclear. Peri-ovulatory putrescine deficiency in older mice appears to adversely impact on oocyte maturation resulting in poor quality embryos (as assessed by blastocyst cell numbers) and early embryo death. This study demonstrates a natural and simple remedy to improve oocyte quality in older women. This study was supported by the NSERC, the March of Dimes Foundation, and the National Natural Science Foundation of China. The authors declare no competing interest.

Deficiency of ovarian ornithine decarboxylase contributes to aging‐related egg aneuploidy in mice

It has been known for more than four decades that during mammalian estrous cycles, luteinizing hormone stimulates a transitory rise in the ovaries of ornithine decarboxylase (ODC) activity and its enzymatic product putrescine, concurrent with oocyte maturation in vivo. Inhibition of this transitory ODC/putrescine rise, however, does not appear to affect oocyte maturation or ovulation. Using several mouse models and combining in vitro and in vivo approaches, we demonstrated that deficiency of ODC during oocyte maturation is correlated with increased levels of egg aneuploidies. These results suggest that the transitory ovarian ODC rise in late proestrus is important for ensuring proper chromosome segregation during oocyte maturation. Older mice (8 months of age) exhibited about 1/3 that of young mice in LH-stimulated ovarian ODC activity and a corresponding increase in egg aneuploidies. Moreover, a combination of putrescine supplementation in mouse drinking water leading up to oocyte retrieval and in oocyte maturation medium reduced egg aneuploidies of the older mice from 12.7% to 5.3%. Therefore, ovarian ODC deficiency might be an important etiology of maternal aging-related aneuploidies, and peri-ovulatory putrescine supplementation might reduce the risk of aneuploid conceptions in older women.

Influence of Ovarian Ornithine Decarboxylase in Folliculogenesis and Luteinization

LH plays a relevant role in folliculogenesis, ovulation, and luteinization. Although ornithine decarboxylase (ODC), a key enzyme in polyamine biosynthesis, is a target of LH in the ovary, the functional significance of ODC induction has remained elusive. Our study reveals that the blockade of the induction of ovarian ODC by means of the specific inhibitor alpha-difluoromethylornithine (DFMO) affects folliculogenesis and luteinization. In immature female mice, DFMO was found to inhibit ovarian growth, the formation of Graafian follicles, and the secretion of progesterone and estradiol. In adult cycling females, the administration of DFMO on the evening/night of proestrus markedly decreased plasma progesterone levels at diestrus, which was associated to the decrease in the expression of steroidogenic factor 1, cytochrome cholesterol side chain cleavage enzyme, and steroidogenic acute regulatory protein in the ovary and to a reduced vascularization of the corpora lutea. These effects were not reverted by the administration of gonadotropins or prolactin. ODC immunoreactivity was also stimulated by LH in theca and granulosa cells of antral follicles but not in preantral follicles. Overall, these experiments demonstrate that elevated ODC values found in the ovary of immature and adult mice play a relevant function in ovarian physiology and that ODC/polyamines must be considered as important mediators of some of the effects of LH on follicular development and luteinization.

The preovulatory rise of ovarian ornithine decarboxylase is required for progesterone secretion by the corpus luteum

Ovarian progesterone secretion during the diestrus stage of the estrous cycle is produced by luteal cells derived from granulosa and thecal cells after the differentiation process that follows ovulation. Our results show that blockade of the preovulatory rise of ovarian ornithine decarboxylase (ODC), a key enzyme in polyamine biosynthesis, by treatment with the specific inhibitor α-difluoromethylornithine (DFMO) leads to a significant decrease in the ovarian progesterone content and a dramatic fall in the plasma levels of this hormone during the following diestrus. The same inhibition was produced in spite of the fact that both luteinizing and follicle stimulating hormones were given concomitantly with DFMO. On the other hand, the acute rise in the plasma progesterone levels observed after administration of human chorionic gonadotropin to mice at different periods of the estrous cycle was not affected by DFMO administration. Our results indicate that although elevated levels of ODC are not required for acute ovarian steroidogenesis, the preovulatory peak of ovarian ODC activity observed in the evening of proestrus may be critical for the establishment of a constitutive steroidogenic pathway and progesterone secretion by the corpus luteum during the diestrus stage of the murine estrous cycle.

Regulation of ovarian ornithine decarboxylase activity and its mRNA by gonadotropins in the immature rat

Previous studies have demonstrated that gonadotropin increases ornithine decarboxylase (ODC) activity in the rat ovary. The increase of ODC activity following gonadotropin stimulation is localized primarily to proliferating granulosa cells. The present studies were undertaken to characterize the mechanism of this hormone-induced increase of ODC. A single intraperitoneal injection of PMSG resulted in a 15-fold increase in ODC activity ( p < 0.0001). Activity was maximal 1 day following PMSG administration, and thereafter declined with time. PMSG also induced an increase in ODC mRNA levels ( p < 0.0001). However, ODC mRNA increased only 2-fold, as compared with a 15-fold increase in ODC activity. Actin mRNA levels were unaffected by PMSG. These results demonstrate that PMSG-induced increase in ODC activity cannot be fully explained by an increase in ODC mRNA, thus indicating that part of the regulation of ODC by PMSG is occurring at the translational and/or posttranslational level. Such regulation is likely a necessary dimension of the fine control of the levels/activity of this important enzyme which has a very short half-life.

Modulation of ovarian ornithine decarboxylase activity during follicular differentiation/maturation

The role of gonadotropins and estrogen on the regulation of ovarian ornithine decarboxylase was studied during follicular differentiation/maturation. In intact immature rats follicular differentiation/maturation was initiated with sequential administration of estrogen and follicle stimulating hormone. Ornithine decarboxylase activity in response to human chorionic gonadotropin was markedly enhanced (2-fold) in rats with preovulatory antral follicles when compared to rats with non-ovulatory follicles. This increase could be attributed to the alteration in the turnover of the enzyme. Following follicle maturation the half life of the human chorionic gonadotropin stimulated ornithine decarboxylase was increased from 18 to 62 min. This increase in half life was associated with differentition of follicles. In the estrogen treated group (which does not induce follicular differentiation), the half life of the enzyme remained unaltered. The regulation of ornithine decarboxylase through the formation of protein inhibitor antizyme induced by diamino hexane, was unaltered during follicular differentiation.

Tissue specific compartmental analysis of gonadotropin stimulation of ovarian ornithine decarboxylase

Luteinizing hormone is known to stimulate the enzyme ornithine decarboxylase in the ovary. Highly purified human follicle stimulating hormone that is devoid of significant biologically active luteinizing hormone can also induce ornithine decarboxylase activity in intact immature rats with a time course of induction similar to that reported for luteinizing hormone. A maximum of 8–10-fold stimulation above controls was observed 4 h following intravenous administration of human follicle stimulating hormone. This stimulation followed a strict dose response relationship. Ovine luteinizing hormone and human chorionic gonadotropin always induced more ovarian ornithine decarboxylase activity than that achieved by maximally effective doses of follicle stimulating hormone. This could not be attributed solely to the ability of specific cell population to respond to the respective gonadotropins. Although granulosa cells contained little receptor for luteinizing hormone/human chorionic gonadotropin and the residual tissue contained little receptor for follicle stimulating hormone, each tissue responded to these gonadotropins in a manner suggestive of the mediation by one or more diffusable factors. A relationship between gonadotropin induced 3’5’-cyclic adenosine monophosphate (cyclic adenosine monophosphate) concentration and ornithine decarboxylase activity suggests that the mediation of gonadotropin stimulated ovarian ornithine decarboxylase is not solely through cyclic adenosine monophosphate, indicating the presence of other factors in the induction of gonadotropin increased ornithine decarboxylase activity.

Regulation of ovarian ornithine decarboxylase by human chorionic gonadotrophin.

Treatment of 29-day-old female Sprague-Dawley rats with human chorionic gonadotrophin (hCG) produced a large and rapid increase in the activity of ornithine decarboxylase. Measurements that use a specific radioimmunoassay showed that the increased activity could be accounted for by a parallel change in the amount of ornithine decarboxylase protein. The increased protein content was caused by an increased rate of synthesis, since the half-life of ornithine decarboxylase was not changed by the hormone treatment. The content of mRNA for ornithine decarboxylase was determined by hybridization with a cDNA probe, and it was found that the increased amount of protein was correlated with a change in the amount of mRNA. These results indicate that treatment with hCG induces ornithine decarboxylase in the rat ovary by increasing the production or the stability of the mRNA for this enzyme. The increased amount of ornithine decarboxylase led to an increase in putrescine in the ovary but did not increase the content of the polyamines spermidine and spermine. These findings show that, despite its rapid and large scale induction, ornithine decarboxylase is not the rate-limiting step that determines the content of these polyamines in this tissue. They also suggest that putrescine itself may play an important role in the ovary.

Changes in ornithine decarboxylase activity in ovarian follicles of the laying hen (Gallus domesticus) during the ovulatory cycle.

Fowl ovarian ornithine decarboxylase activity was measured at 20, 10 and 3 h before an expected ovulation in granulosa and thecal tissues from follicles at various stages of development. An increase in the enzyme activity between 10 and 3 h before an expected ovulation was assumed to be caused by preovulatory increase in plasma LH concentration. The activity in granulosa tissue increased with increasing size of the follicle. In the largest (F1) follicle there was an 11-fold increase in granulosa ornithine decarboxylase specific activity between 10 and 3 h before ovulation. In the third (F3) and fifth (F5) largest follicles there was a 1.9- and 2-fold increase respectively. The enzyme activity in thecal tissue from the follicular hierarchy decreased with increasing size of the follicle and the F3 thecal preparation was the only tissue to respond to the preovulatory LH surge. In contrast, ornithine decarboxylase activity in thecal tissue from small (less than 5 mm) non-atretic follicles increased by two- to threefold after the preovulatory LH surge. The activity in atretic follicles of the same size was low and remained unchanged throughout the ovulatory cycle.

Ibuprofen modulation of human chorionic gonadotropin-induced ornithine decarboxylase activity and ovulation in the rabbit ovary

The activity of ornithine decarboxylase, the rate-limiting enzyme in polyamine synthesis, increases as granulosa cells proliferate in developing follicles. Both luteinizing hormone and prostaglandins stimulate ornithine decarboxylase activity. Here, we sought to determine the relative contributions of both trophic stimuli to ornithine decarboxylase activity in the preovulatory rabbit ovary. Baseline ovarian ornithine decarboxylase activity, determined by measuring the release of CO2 from (1-14C)-ornithine, was 13.4 ± 1.27 (mean ± SD) pmol of carbon dioxide per hour per milligram of protein. Treatment with ibuprofen, a prostaglandin synthetase inhibitor, led to a significant (p < 0.05) decrease in the baseline ovarian ornithine decarboxylase activity (4.7 ± 0.29 pmol of carbon dioxide per hour per milligram of protein). Administration of human chorionic gonadotropin (hCG) (50 IU/kg intramuscularly) to adult rabbits (2.5 to 3.5 kg) elicited a 1,200% elevation of ovarian ornithine decarboxylase activity 5 hours after injection; there was a return to baseline by 11 hours after injection. Stimulation with human chorionic gonadotropin led to ovulation in 22.2%, 25%, and 60% of rabbits at 7, 9, and 11 hours after treatment, respectively. Single-dose ibuprofen treatment (5 mg/kg intramuscularly), 7 hours after human chorionic gonadotropin administration inhibited ovulation and elevated ovarian ornithine decarboxylase activity. These results indicate that ibuprofen effectively inhibits ovulation in hCG-stimulated rabbit ovaries in the presence of a significant (p < 0.001) elevation in ovarian ornithine decarboxylase activity. Thus, different intracellular mechanisms are involved in the prostaglandin modulation of basal and hCG-stimulated cells during the course of preovulatory follicle maturation.

Enhancement of ovulation in the rat by DL-α-difluoromethylornithine, an irreversible inhibitor of ornithine decarboxylase

Spontaneously cycling Sprague-Dawley Rats were treated with the irreversible ornithine decarboxylase (ODC) inhibitor DL-α-difluoromethylornithine (DFMO) on the day of proestrus. This treatment completely suppressed both the rise in ovarian ODC and the associated increase in putrescine concentration and increased the number of eggs released the next day by 91%. At the next oestrus (5 days after DFMO) there was an increase of 53% but at the third oestrus after DFMO (9 days) there was no significant change in the number of eggs released. A model is proposed in which ornithine decarboxylase forms part of a local inhibitory feed-back system controlling the number of follicles which develop to maturity and rupture during each cycle.

Regulation of ovarian ornithine decarboxylase: Role of calcium ions in enzyme induction in isolated swine granulosa cells in vitro

When swine granulosa cells were cultured in chemically defined medium selectively deficient in Ca 2+, the dose-dependent stimulation of ornithine decarboxylase (EC 4.1.1.17) activity in response to prostaglandin E 2, l-epinephrine or the somatomedin, multiplication-stimulating activity, was attenuated markedly. Putative calcium influx blockers, verapamil and diltiazem, also inhibited hormone-stimulated enzymic activity. Similar inhibitory effects were exerted by divalent (cobalt) or trivalent (lanthanum) cations believed to compete with calcium for extracellular binding sites. The suppressive effects of extracellular calcium deprivation were time-dependent (suggesting gradual depletion of intracellular calcium stores), and could be mimicked by the intracellular antagonist of calcium action, trifluoperazine. The mechanism(s) subserving diminished hormonal induction of enzyme activity could not be accounted for by alterations in cell viability, general protein synthesis, half-life of decay of enzyme activity (measured in the presence of cycloheximide), or apparent K m of ornithine decarboxylase. Ca 2+ and/or calcium antagonists did not modify enzyme activity in cell-free preparations. These observations implicate Ca 2+ in the hormonal induction of a discrete cytosolic enzyme in isolated intact ovarian cells.

Multihormonal regulation of ornithine decarboxylase activity in highly differentiated porcine granulosa cells in vitro.

Purified luteinizing hormone, but not follicle-stimulating hormone, elicited time- and dose-dependent stimulation of the cytosolic enzyme, ornithine decarboxylase, in highly differentiated, porcine granulosa cells maintained in vitro in chemically defined medium. Enzymatic induction was susceptible to inhibitors of protein and RNA synthesis, and was suppressed by selective direct and indirect inhibitors of ornithine decarboxylase. Physiologic concentrations of prostaglandin E2 and L-epinephrine also enhanced enzymatic activity in a dose-dependent and saturable manner. Systematic comparison of the hormonal induction of ornithine decarboxylase in highly differentiated versus poorly differentiated granulosa cells revealed distinctive patterns of enzymic responsivity in relation to the degree of cytodifferentiation attained in vivo. This in vitro model is likely to permit further detailed examination of the molecular mechanisms subserving the hormonal control of ovarian ornithine decarboxylase activity in spontaneously differentiated granulosa cells maintained under chemically defined conditions in vitro.

Changes in ovarian luteinizing hormone and follicle-stimulating hormone receptor content and in gonadotropin-induced ornithine decarboxylase activity during prepubertal and pubertal development of the female rat.

Ovarian hCG and FSH receptor content was measured at different postnatal ages (days 4-32), at 4-day intervals, and expressed as counts per min 125I-labeled hormone bound/micrograms DNA. Specific FSH binding was minimal at day 4 and increased to a maximum at day 28, with the greatest rate of increase between day 4 and 16. hCG receptor content increased 6-fold between day 4 and 32, but contrasting with FSH, its greatest rate of increase occurred between day 16 and 28. The effectiveness of in vivo treatment with LH in inducing ovarian ornithine decarboxylase (ODC) activity, measured in 20,000 X g supernatants by the in vitro formation of 14CO2 from labeled ornithine, increased significantly during prepubertal development (days 21-33), paralleling the changes in hCG receptors. Ovarian hCG and FSH receptor content was also measured during the time of puberty. Specific hCG binding increased from anestrus (juvenile 32-day-old animals) to 1300 h of the first proestrus. The levels declined significantly by 1600 h of first proestrus and reached minimal values by the morning of estrus. Specific FSH binding also increased from anestrus to first proestrus, but to a much lesser degree, the levels showing only minor fluctuations during the rest of the cycle. hCG receptors measured in granulosa cells during puberty increased to an even greater extent than in the whole ovary; binding exhibited an 8-fold increase from anestrus to 1300 h of first proestrus and then declined to much lower values on estrus and the first diestrus. The capacity of LH to induce ovarian ODC activity increased markedly between anestrus and first proestrus, declining thereafter, and again paralleled the changes in LH receptor content. Evaluation of the kinetics of binding of hCG and FSH with their respective receptors in ovaries of 28-day-old rats revealed that the binding of hCG reached equilibrium within 4-8 h, whereas the binding of FSH attained equilibrium between 12-24 h. In both cases, however, the labeled hormones were clearly dissociable from their respective receptors by addition of an excess of unlabeled hormone. Moreover, although the ovarian receptor content of both hCG and FSH increased during prepubertal development, the equilibrium association constant of either hormone with its receptor remained unchanged. The results suggest that the infantile increase in FSH receptors may be induced by the high serum FSH levels present at that age and that, as shown by others in hypophysectomized rats, the subsequent increase in hCG receptors is, at least in part, an FSH-dependent phenomenon. The parallel changes in hCG receptors and LH-induced ODC activity during pre- and peripubertal development strongly suggest that an increase in LH binding capacity plays a fundamental role in the process of ovarian growth. It is also suggested that the changes in the steroidogenic ovarian response to hCG previously observed in peripubertal rats are, at least in part, determined by changes in ovarian LH receptor content.

Inhibition of rat ovarian ornithine decarboxylase by ethanol in vivo and in vitro

Intragastric administration of ethanol greatly inhibited ovarian ornithine decarboxylase ( l-ornithine carboxy-lyase EC 4.1.1.17) stimulated by human chorionic gonadotropin in vivo. The inhibition occurred only if the treatment with ethanol was started before the injection of hormone. The use of inhibitors for alcohol dehydrogenase and aldehyde dehydrogenase clearly showed that the observed inhibition was a direct effect of ethanol itself. When rat ovarian cells were incubated in vitro with human chorionic gonadotropin the activity of ornithine decarboxylase was also markedly stimulated. This stimulation could also be inhibited by ethanol. Moreover, actinomycin D and α-amanitin inhibited the stimulation of ornithine decarboxylase, showing that the enhanced activity in vitro resulted from the synthesis of new mRNA for ornithine decarboxylase. The time dependence of the inhibition caused by ethanol additionl resembled that after addition of actinomycin D. This supports the view that one site where ethanol inhibits protein synthesis is at the transcriptional level.

Ovarian function in the rat following irreversible inhibition of L-ornithine decarboxylase

The possibility that the transient rise in rat ovarian ornithine decarboxylase (ODC) activity and the associated increase in putrescine which occur under luteinizing hormone control late in proestrus have an essential role in ovarian function has been tested using DL-α-difluoromethylornithine (DFMO) an irreversible inhibitor of ODC. Treatment with DFMO, 500 mg/kg s.c. at 12:00 h on the day of proestrus and 200 mg/kg, 6, 12 and 18 h thereafter completely suppressed both the rise in ovarian ODC activity and the associated increase in putrescine concentrations. However, ovulation took place normally under these conditions and the course of the resulting pregnancies was also normal. Similarly, combined treatment with DFMO and an inhibitor of S-adenosyl-L-methionine decarboxylase, 1,1-((methylethanediylidine)-dinitrilo) bis (3-aminoguanidine), 25 mg/kg, given at 10:00 h on the morning of proestrus failed to influence either ovulation or the subsequent period of gestation. These data provide no support for a functional role of the pre-ovulatory rise of ODC in rat ovary in the major peri-ovulatory events of that particular cycle, although they do not exclude effects on systems (e.g. steroidogenesis) not directly examined with our experimental approach. On the other hand, inhibition of ODC resulted in an increase in the number of uterine implantation sites following mating at the next proestrusestrus 4 days later. These data would support the previously expressed view that ODC may be associated with the gonadotrophin-induced initiation of follicular development for ovulation in the succeeding cycle. Moreover, since inhibition of the enzyme resulted in facilitation of the process, the normal physiological function of ODC, and/or the putrescine generated through its action, would appear to be inhibitory.

Stimulation of rat ovarian ornithine decarboxylase in vitro by hCG, amino acids and bovine serum albumin

Rat ovarian ornithine decarboxylase activity could be stimulated in vitro by a variety of factors, which apparently have different modes of action. Ovarian cells prepared from pre-pubertal rats by collagenase dispersion exhibited a low but detectable ornithine decarboxylase activity after a 6-h incubation in a defined medium. The enzyme activity as markedly enhanced in vitro by hCG, which also produced increased accumulation of cyclic AMP and stimulated the secretion of progesterone. In addition to the gonadotrophin, ovarian ornithine decarboxylase activity was strikingly stimulated by some non-essential amino acids, and especially by bovine serum albumin. While markedly enhancing ornithine decarboxylase activity, none of the latter additions increased the accumulation of cyclic AMP or enhanced the secretion of progesterone. Bovine serum albumin enhanced powerfully ornithine decarboxylase activity in vitro at very small concentrations (from 0.75 muM). The half-life of the enzyme remained unchanged (26-28 min) upon stimulation indicating that the stimulation mechanism did not involve any stabilization of the enzyme.

Studies on sex organ development. Effect of hormones on ornithine decarboxylase activity in the developing chick ovary.

The response of ornithine decarboxylase activity to hormones in the embryonic left ovary was measured throughout the stages of development. During the early stage of ovarian development (9th day of incubation), the ornithine decarboxylase activity (in terms of pmol CO(2)/30min per mg of protein) was high (766); it decreased from the 10th to the 12th day (575-239), increased slightly from the 13th to the 15th day (306) and finally fell to a low value (192-20) from the 18th day of development to birth. Administration of an optimal dose of oestrogen to the 9-10-day embryo stimulated the ovarian ornithine decarboxylase activity by 48-53%. If the same dose of oestrogen was administered to the 15-18-day embryo, the ovarian enzyme activity was suppressed by 32-43%. This biphasic response to oestrogen for enzyme induction is characteristic of the developing ovary and is not observed in other genital organs of the chick. In the early developmental stage (9-10th day) testosterone has no effect on ovarian ornithine decarboxylase activity, but in the late stage testosterone inhibits the activity by 41%. Organ culture techniques have been used to test the ovarian response to lutropin (luteinizing hormone). Lutropin stimulated ornithine decarboxylase activity by approx. 99-155% in the ovary of the early embryonic stage (10-13th day), and by 175-200% in the ovary of the late embryonic stage (15-18th day). The alteration in enzyme activity in the ovary as assayed in vitro during development is not due to the effect of the size of the endogenous ornithine pool. The relationship of ornithine decarboxylase activity to the morphological and biochemical changes in the developing ovary is discussed.

Indomethacin blocks gonadotropin stimulation of ovarian ornithine decarboxylase activity by inhibiting protein synthesis

Both gonadotropins and prostaglandins stimulate the ornithine decarboxylase (ODC) activity of porcine granulosa cells in vitro (1,2). To investigate a possible intermediary role of prostaglandins in this gonadotropin action, the effects of indomethacin on gonadotropin-induced ODC activity were studied. Indomethacin had no effect at concentrations lower than 10 −5 M; at higher concentrations indomethacin exerted a dose-dependent suppression of LH-stimulated ODC activity which was essentially complete at 5 × 10 −4 M. The effects of PGE 2 and 8-Bromo-cAMP, potent stimulators of ODC, were also blocked by indomethacin (5 × 10 −4 M). This effect did not represent direct inhibition of enzyme activity, but appeared to be due to inhibition of protein synthesis by the drug. Thus, incorporation of 14C-leucine into proteins by these cells was blocked by indomethacin with a dose-response curve similar to that for ODC suppression. This distal effect of indomethacin may complicate the interpretation of some experiments if the inhibitor is assumed to act only at the prostaglandin synthetase step.