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Effects of a new indole derivative on guinea pig reproduction
CONTRACEPTION
EFFECTSOF A NEW INDOLEDERIVATIVE ON
GUINEAPIG REPRODUCTION J, Bergman l, M. Hammarstrdm2, P. Eneroth 3 , T&tam G1 1 Department of Organic Chemistry, Royal Institute of Technology, S- 100 44 Stockholm, Sweden 2 Department of Physiology I, Karolinska Institutet, S- 104 0 1 Sockholm, Sweden 3 Research and Development Laboratory, Department of Obstetrics and Gynecology, Karolinska sjukhuset, S- 104 0 I Stockholm, Sweden ABSTRACT The effect of a new indole derivative, ( 1,I ,3-trimethyl-3-(3’-indolyi)1.2,3,4-tetra-hydrocyclopentIbl-indole) denoted 1 related to indomethacin and yeuhchukene 2 were tested on guinea pig reproduction. In mating experiments ten females were given compound L p.o. from cycle day 14 and onwards throughout pregnancy. Two groups of ten females each were used as controls. Conception occurred significantly (~~0.001) faster in the treatment group. The litter size was significantly (~(0.00 1) smaller after treatment with the indole derivative. Dams and pups did not seem to have been adversely affected by the treatment as judged by their exterior, behavior and survival. The results are suggested to be due to a reduction in the number of ova available for conception (litter size) in combination with improved conditions for ovum implantation and/or a facilitated sperm migration in the female genital tract. These effects are discussed in relation to known effects of arachidonic acid metabolites on reproduction.
Reprint requests: Dr. Margareta Hammarstrom Department of Physiology I, Karolinska Institute& S- 104 0 1 Stockholm, Sweden Submitted Accepted
for publication for publication
APRIL 1989 VOL. 39 NO. 4
June 22, 1988 January 4, 1989
419
CONTRACEPTION
INTRODUCTION Arachidonic acid (AA) metabolites are believed to play an important role in female reproduction. Thus, the prostaglandins (P&I are known to affect the contractile response in the uterus, fallopian tube and ovary ( 12). Exogenous PGs are also known to induce luteolysis in the guinea pig, i.e. serum progesterone levels fall rapidly after PGF2o injection (i.p.1in the dioestrus phase (3,4). Prostaglandin F20 has been postulated to be the local luteolytic hormone from the guinea pig uterus, since hysterectomy results in ovulation and persistent corpus luteum(S,6,7). PGF2o induced luteolysis in the hamster but left the subsequent ovulation process intact (8). Furthermore, gonadotropins in combination with PGEl or PGE2 induced superovulation in the monkey (9). Indomethacin inhibits LH or HMWHCGinduced ovulations in several species, e.g. the monkey, rabbit and rat ( 10,I 1,121,delays spontaneous ovulation in the Rhesus monkey ( 12). It has been suggested that prostaglandins participate in the rupture of the follicle since indomethacin causes retention of ova in the rabbit ( 13) and blocks ovulation at the ovarian level in the rat ( 14). In the latter species indomethacin did not affect ovum maturation but inhibited follicular rupture (151. PGF2, and PGEl receptors have been demonstrated in the bovine ovary follicular wall ( 161 and FGF2o has been found to increase ovarian smooth muscle contractility in the rabbit whereas PGE2had the opposite effects ( 17). In the guinea pig PGF2o-receptor activities in the luteal tissue, ovarian stromal, interstitial and follicular tissue have been reported (18). Furthermore an increased number of atretic follicles in this species was observed by Tam et al. (18) after PGF2o administration. Indole derivatives (e.g. Indomethacin) have in many reports been shown to be pharmacologically active on both the ovary, fallopian tube and uterus of several species (see below). Most of the effects have been assumed to be due to inhibition of prostaglandin synthesis since all reproductive organs are rich in prostaglandins ( 19). Indole derivatives interact with the arachidonic acid cascade, an effect which may be attributed to the interaction of indole compounds with various activated oxygen species (20,21,22). Furthermore, it has been reported that the naturally occurring indole derivative yuehchukene (2) might have antiimplantation effects (23) in rats. Because of long-standing interest in indole derivatives (24,251, we decided to further explore the effects of 420
APRIL 1999 VOL. 39 NO. 4
CONTRACEPTION
this class of substances on reproduction. The guinea pig was used because of the resemblance of cycle characteristics between this animal and man. The guinea pig has a fairly long estrous cycle (16 days) with spontaneous ovulation and a pregnancy duration of 65 days (26). MATERIALANDMETHOD!?
The tetrahydrocyclopent-b-indole( which can be considered as a simplified analogue of yuehchukene (21,was obtained by a simple condensation (induced by trifluoroacetic acid), of indole with acetone (Fig.1 & 2). The structure of 1 has also been rigorously proven by an independent synthesis (32). Under different conditions (33,34,35) several other condensation products have been obtained. Animals Thirty sexually mature mottled or albino virgin guinea pigs (English shorthair guinea pigs) 400-450 g, were used. . . Estrouscycle deWnlnatm The animals were evaluated daily concerning the status of the vaginal closure membrane. Only animals showing regular cycles were chosen. Vaginal smears were taken when the membrane was open. After at least two normal cycles, the animals entered the study. Matinn exoeriments Thirty female animals were divided into three groups of 10 each. One group of 10 females were used for administration of compound 1. The female animals were given 10 mg/kg/day (p.01from day 14 of the estrous cycle. Drug administration was performed daily and continued throughout pregnancy. Four to five females were caged with one male. The males (n-6) were shifted from one female group to the next after two weeks. One group of twenty animals was used as control with respect to conception, pregnancy rate and litter size. The females were checked daily concerning the status of the vaginal closure membrane. Pregnancy was calculated from the second day of vaginal opening (27) and the animals were examined by palpation early in pregnancy.
Substance 1 was dissolved in sesame oil (1 ml) and administered orally with a 1 ml syringe once daily in the morning at a dosage of 10 mg/kg from cycle day 14 until parturition. Animals in control group were similarily given 1 ml sesame oil daily in the morning. APRIL 1999 VOL. 39 NO. 4
421
CONTRACEPTION
-1 Eis_L Tetrahydrocyclopent-b-indole
in this study tested for effects on
guinea pig reproduction.
CH3 I
2 = EgJ. Yuehchukene, considered rats and hamsters.
422
to have antiimplantation effects on
APRIL 1999 VOL. 39 NO. 4
CONTRACEPTION
cal evalug&g X2-analysis or Fischers exact test and Students t-test were used as described (28). RESULTS In the treatment group all animals ( 1O/ 10) became pregnant, while in the control group only 14/20 became pregnant. Most pregnancies in the treatment group occurred during the first cycle (8/ 10) while in the control group often several cycles passed until pregnancy took place (Table I).
Table I. Time of conception in guinea pigs with and without oral treatment ( 10 mg/kg/day) with an indole derivative (compound J_)
Cycle 1
Treatment group 8/l 0 (n-10)
Conceotion rate Cycle 2-4 No. of animals Significance not pregnant after 4 cycles 212
0X X
Control group (n-10)
2/20
14/18
63
Group differences were tested by X2-analysis; xxx denotes p
Apparently, the drug facilitated conception (p
APRIL 1999 VOL. 39 NO. 4
423
CONTRACEPTION
Table II. Effects of oral treatment (10 mg/kg/dayl with an indole derivative (compound 1) on litter size in guinea pigs
Treated animals Control Total number of pups/ pregnant animals
28/10
Mean litter size
2.8
Median ”
”
-xxx
55114
7
3.9
-xxY4 3
Significance of group differences was analyzed by Students t-test. xxx means p
The number of dead pups/litters in the treatment group were smaller than in the control group and there were no toxic effect on the pups or on the dams (Table III).
Table III. Number of dead pups in litters from guinea pigs given oral treatment (I 0 mg/kg/day) with an indole derivative (compound 11
Treatment group (n=lOl No. of dead pups/ total no. of pups Pup deaths/litter size
4128
212; 213
Control (n=141 12155
214; 4/4; l/3 314 215
Fischer’s exact test was used for testing of significance; no significant difference between the groups was found.
424
APRIL 1989 VOL. 39 NO. 4
CONTRACEPTION
DISCUSSION The indole compound 1 structurally and functionally related to indomethacin likely caused a reduction in the number of ova available for conception as indicated by the reduced litter size. However, an effect on ova quality or tubal and/or uterine motility cannot be excluded. Resorption of ova after indomethacin has been shown in the rabbit (13) but whether this was due to a nonspecific toxic effect or to specific changes at the implantation sites is uncertain. In the guinea pig about 20 follicles (ovulation points) were reported to develop per cycle and 20 ova were found by Maia et al. (30). Furthermore, after indomethacin ( 12 mg/kg/24h on cycle day 16) treatment these investigators found that ovulation points were reduced to 13 and only 10 ova could be recovered. The ova transport was suggested to be unaffected. However, Sisk (27) considers the normal number of ovulation points to be 3.33.4 and the number of corpora lutea on the average 3.5 in the guinea pig (3 1). The indole compound l-treated animals in the present study conceived faster than the control females. The possibility that a prolonged progesterone secretion favored implantation can not be excluded since Poyser (6) reported a prolongation (3 days) of the estrous cycle after indomethacin treatment in high doses (40 mg/d) to the guinea pig. Apart from a facilitated implantation induced by progesterone, the rapid conception observed in the female guinea pig receiving compound 1 may also be explained by an improved sperm penetration and/or capacitation. Female genital tract secretion is generally considered to be at least in part regulated by prostaglandins ( 19). Our results are somewhat in contrast to the results of Kong et al. (23) who found that the indole yeuhchukene counteracted reproduction in the female rat. This difference may be due to species differences or due to the fact that the indole 1 is structurally different from yeuhchukene. There were no toxic effects noted on the dams or on the litters (with compound 1) despite continued treatment throughout pregnancy. The number of dead pups in our study did not exceed that in the control groups. The mean litter size in the indole (compound l&exposed group was 2.8. Interestingly Wright (29) has pointed out that a litter size of 2 in the guinea pig is optimal for pup survival which is in agreement with the increased mortality in our control groups. Our results thus indicate that indole 1 is comparatively nontoxic.
APRIL 1999 VOL. 39 NO. 4
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CONTRACEPTION
Whatever mechanisms that were involved, the present results indicate that our indole compounds may improve reproductive performance in the female guinea pig. ACKNOWLEDGEMENT The skilful technical assistance of Ms. Eva Andersson, Margareta TapperPersson and Ann Hagstrem is gratefully acknowledged. For typing the manuscript we are indebted to Ms. Ann Nicolausson. This project has been supported by research funds from the Karolinska Institute. REFERENCES 1. Couthino, E.M.and H.S.Maia: The contractile response of the human uterus, fallopian tubes, and ovary to prostaglandins in vivo. Fert. Ster. 1971; 22,539-543. 2.
Christensen, N.J:Studies on the bioconversion of arachidonic acid in human pregnant reproductive tissues. Thesis. Stockholm, Sweden 1984.
3. Wehrenberg, W.B.,D.J.Dierschke, and R.C.Wolf: The effect of prostaglandin F2o on ovarian blood flow, and progesterone concentration in cyclic guinea pigs. Biol. Reprod. 1979; 2 1, 187- 19 1. 4. Tso, B.C.-F.and W. H. Tam: The luteolytic effect of prostaglandin F2o on the guinea pig corpus luteum in relation to gonadotropins and stages of the oestrous cycle. Can. J. Physiol. Pharmacol. 1978; 56 828833. 5. Blatchley, P.R.and B. T. Donovan: The effect of prostaglandin F2o and prostaglandins E2 upon luteal function and ovulation in the guineapig.J. Endocr. 1972; 493-50 1. 6. Poyser, N. L: Prostaglandin F2a in the uterine luteolytic hormone in the guniea pig: The evidence reviewed. In: Advances in Prostaglandins and Tromboxane Research, ~012. (ed. B. Samuelsson and R. Paoletti). Raven Press, New York, 1976, pp. 633-640.
426
APRIL 1999 VOL. 39 NO. 4
CONTRACEPTION
7. Deansly, R. and J.S. Perry: Independent regression of normal and induced corpora lutea in hysterectomized guinea-pig. J. Reprod. Fert. 1969; 20,503-508. 8.
Labhsetwar, A. P: Luteolysis and ovulation induced by prostaglandin F2o in the hamster. Nature 1971; 230,528-529.
9. Batta, SK and B.G.Bracket: Ovulation in Rhesus monkeys by treatment with gonadotrophin and prostaglandins. Prostaglandins 1974; 6,45-54. 10. Armstrong, D.T.,D.L.Grinwich, Y.S.Moon and J. Zamecknik: Inhibition of ovulation in rabbits by intrafollicular injection of indomethacin and prostaglandin F antiserum. Life Sciences 1974: 14, 129-140. 11. Grinwhich, D.L.,T.G.Kennedy and D.T.Armstrong: Dissociation of ovulatory and steroidogenic actions of luteinizing hormone in rabbit with indomethacin, an inhibitor of prostaglandin biosynthesis. Prostaglandins 1972;1, 89- 106. 12.Wallach, E.,A.de la Cruz, A.J.Hunt, K.H.Wright, and V.C.Stevens: The effect of indeomethacin on HMG-HCG induced ovulation in the Rhesus monkey. Prostaglandins 1975; 9,645-658. 13. O’Grady,J.P., B.W.Caldewell, F.J. Auletta, and L. Speroff. The effect of an inhibitor of prostaglandin synthesis (indomethacin) on ovulation, pregnancy and pseudopregnancy in the rabbit. Prostaglandins 1972; 1, 97-106. 14. Behrman, H.R.,P. Gayle, P. Orcyk, and R.O.Greep: Effect of synthetic gonadotrophin releasing hormone (Gn-Rh) on ovulation blockade by aspirin and indomethacin. Prostaglandins 1972; 1,245-258. 1S.Tsafriri, A., H.R.Lindner, U. Zor, and S.A. Lamprecht: Psychological role of prostaglandins in the induction of ovulation. Prostaglandins 1972; 2, l-10. 16. Kimball, F.A and J.W. Lauerdale: Prostaglandin El and F2o specific binding in bovine corpora lutea: Comparison with luteolytic effects, Prostaglandins 1975; 10, 3 13-322. APRIL 1989 VOL. 39 NO. 4
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17. Virutamasen, P., K.H.Wright and E.E.Wallach: Effects of prostaglandins E2 and F2o on ovarian contractility in the rabbit. Fert. Ster 1972; 23,675682. 18Tam, W.H.,W.K.Beveridge and C.-F.Tso: Effects of gonadotropins, hypophysectomy and prostaglandin F-2o on corpora lutea in ovarian follicles, an evidence for the presence of PGF-2o receptors in the ovarian follicles and interstitium of non-pregnant guinea-pig. J. Reprod. Fert. 1982; 64,9-17. 19.Labhsetwar, AI? Prostaglandins and studies related to reproduction in laboratory animals. In: Prostaglandins and Reproduction. ted. SM. Karim). MTP Press Ltd. Edinburgh, 1975; pp.241-270. 20. Bodaness, R.S.and P.C.Chan: Reaction of indomethacin with singlet molecular oxygen. B&hem. Pharmacol. 1980; 29, 1337-1340. 2 1. Durchstein, H.-J:Reaktionen von antiphlogistisch wirkenden Indolderivaten mit Singulett-Sauerstoff, I. Mittl: Beteiligung von reaktiven Sauerstoppspezies am Mechanismus der Prostaglandinsynthese. Arch. Pharm. (Weinheim) 19853 18, 127- 134. 22.Laegreid, W.W and R.G.Breeze: The effect of 3methylindole on superoxide and hydrogen peroxide production and NADPHoxidation by goat lung microsomes. Research Communication in Chemical Pathology and Pharmacology 1985; 47,387-392. 23.Kong, Y.-C.,K.F.Cheng, R.C.Cambie and P.G.Waterman: Yuehchukene, a novel indole alkaloid with anti-implantation activity. J. Chem, Sot., Chem. Commun. 1985,47-48. 24.Bergman, J. and J.-E. Backvall: Base-induced rearrangement of 3-(ahaloacyl)indoles. Tetrahedron 1975; 3 1, 2063-2073. 25.Bergman. J. and S. Bergman: Studies on rutaecarpine and related quinazolincarboline alkaloids. J Org. Chem. 1985; 50, 1246-1255. 26. Hammarstrom M: Uterine secretomotor innervation. Thesis, Acta Physiol. Stand 109. Suppl. 484, 1980.
429
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27Sisk, D.B:In: The Biology of Guinea Pig, chapter 7, Physiology ted. J.E. Wagner and PJ. Manning). Academic Press, New York, 1976, pp. 7997. 28Colquohoun D:Lectures on Biostatistics. Clarendon Press, Oxford, 1971. 29.Wright, S: The genetics of vital characters of the guniea pig. J. Cell. Camp. Physiol. 1960: 56, Suppl. 1, 123-151. 30.Maia, H. Jr., L.A. Salina, E.O.Fernandez and C.J.Paurstein: Pharmacologic modification of the time course of ovum transport in guinea pig. Fert. Ster. 1977; 28, 1361-1364. 3 1.Reed. M. and W.F.Hounslow: Induction of ovulation in the guinea-pig. J. Endocrinol. 1971; 49,203-2 11. 32Bergman J,, P-P. Norrby, U. Tilstam and L. Venemalm: Structure elucidation of some products obtained by acid catalyzed condensation of indole with acetone. To be published. 33.Noland, W.E.and M.R.Veinkiteswaren: Cyclizative condensation. J. Org. Chem. 1961; 26,42-63. 34Chatterjee, AS. Manna, J. Banerji. C. Pascard, Th. Prange. and J.N. Schoolery: Lewis acid induced electrophilic substitution in indoles with acetone. Part 2. J. Chem. Sot. 553-555, 1980. 35. Banerji, J., Chatterjee. A., Manna, S., Pascard. C..Prange. Thand J.N. Schoolery: Lewis acid induced substitution of indole. Part 3. J Chem Socl5.325. 1981.
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EFFECTSOF A NEW INDOLEDERIVATIVE ON
GUINEAPIG REPRODUCTION J, Bergman l, M. Hammarstrdm2, P. Eneroth 3 , T&tam G1 1 Department of Organic Chemistry, Royal Institute of Technology, S- 100 44 Stockholm, Sweden 2 Department of Physiology I, Karolinska Institutet, S- 104 0 1 Sockholm, Sweden 3 Research and Development Laboratory, Department of Obstetrics and Gynecology, Karolinska sjukhuset, S- 104 0 I Stockholm, Sweden ABSTRACT The effect of a new indole derivative, ( 1,I ,3-trimethyl-3-(3’-indolyi)1.2,3,4-tetra-hydrocyclopentIbl-indole) denoted 1 related to indomethacin and yeuhchukene 2 were tested on guinea pig reproduction. In mating experiments ten females were given compound L p.o. from cycle day 14 and onwards throughout pregnancy. Two groups of ten females each were used as controls. Conception occurred significantly (~~0.001) faster in the treatment group. The litter size was significantly (~(0.00 1) smaller after treatment with the indole derivative. Dams and pups did not seem to have been adversely affected by the treatment as judged by their exterior, behavior and survival. The results are suggested to be due to a reduction in the number of ova available for conception (litter size) in combination with improved conditions for ovum implantation and/or a facilitated sperm migration in the female genital tract. These effects are discussed in relation to known effects of arachidonic acid metabolites on reproduction.
Reprint requests: Dr. Margareta Hammarstrom Department of Physiology I, Karolinska Institute& S- 104 0 1 Stockholm, Sweden Submitted Accepted
for publication for publication
APRIL 1989 VOL. 39 NO. 4
June 22, 1988 January 4, 1989
419
CONTRACEPTION
INTRODUCTION Arachidonic acid (AA) metabolites are believed to play an important role in female reproduction. Thus, the prostaglandins (P&I are known to affect the contractile response in the uterus, fallopian tube and ovary ( 12). Exogenous PGs are also known to induce luteolysis in the guinea pig, i.e. serum progesterone levels fall rapidly after PGF2o injection (i.p.1in the dioestrus phase (3,4). Prostaglandin F20 has been postulated to be the local luteolytic hormone from the guinea pig uterus, since hysterectomy results in ovulation and persistent corpus luteum(S,6,7). PGF2o induced luteolysis in the hamster but left the subsequent ovulation process intact (8). Furthermore, gonadotropins in combination with PGEl or PGE2 induced superovulation in the monkey (9). Indomethacin inhibits LH or HMWHCGinduced ovulations in several species, e.g. the monkey, rabbit and rat ( 10,I 1,121,delays spontaneous ovulation in the Rhesus monkey ( 12). It has been suggested that prostaglandins participate in the rupture of the follicle since indomethacin causes retention of ova in the rabbit ( 13) and blocks ovulation at the ovarian level in the rat ( 14). In the latter species indomethacin did not affect ovum maturation but inhibited follicular rupture (151. PGF2, and PGEl receptors have been demonstrated in the bovine ovary follicular wall ( 161 and FGF2o has been found to increase ovarian smooth muscle contractility in the rabbit whereas PGE2had the opposite effects ( 17). In the guinea pig PGF2o-receptor activities in the luteal tissue, ovarian stromal, interstitial and follicular tissue have been reported (18). Furthermore an increased number of atretic follicles in this species was observed by Tam et al. (18) after PGF2o administration. Indole derivatives (e.g. Indomethacin) have in many reports been shown to be pharmacologically active on both the ovary, fallopian tube and uterus of several species (see below). Most of the effects have been assumed to be due to inhibition of prostaglandin synthesis since all reproductive organs are rich in prostaglandins ( 19). Indole derivatives interact with the arachidonic acid cascade, an effect which may be attributed to the interaction of indole compounds with various activated oxygen species (20,21,22). Furthermore, it has been reported that the naturally occurring indole derivative yuehchukene (2) might have antiimplantation effects (23) in rats. Because of long-standing interest in indole derivatives (24,251, we decided to further explore the effects of 420
APRIL 1999 VOL. 39 NO. 4
CONTRACEPTION
this class of substances on reproduction. The guinea pig was used because of the resemblance of cycle characteristics between this animal and man. The guinea pig has a fairly long estrous cycle (16 days) with spontaneous ovulation and a pregnancy duration of 65 days (26). MATERIALANDMETHOD!?
The tetrahydrocyclopent-b-indole( which can be considered as a simplified analogue of yuehchukene (21,was obtained by a simple condensation (induced by trifluoroacetic acid), of indole with acetone (Fig.1 & 2). The structure of 1 has also been rigorously proven by an independent synthesis (32). Under different conditions (33,34,35) several other condensation products have been obtained. Animals Thirty sexually mature mottled or albino virgin guinea pigs (English shorthair guinea pigs) 400-450 g, were used. . . Estrouscycle deWnlnatm The animals were evaluated daily concerning the status of the vaginal closure membrane. Only animals showing regular cycles were chosen. Vaginal smears were taken when the membrane was open. After at least two normal cycles, the animals entered the study. Matinn exoeriments Thirty female animals were divided into three groups of 10 each. One group of 10 females were used for administration of compound 1. The female animals were given 10 mg/kg/day (p.01from day 14 of the estrous cycle. Drug administration was performed daily and continued throughout pregnancy. Four to five females were caged with one male. The males (n-6) were shifted from one female group to the next after two weeks. One group of twenty animals was used as control with respect to conception, pregnancy rate and litter size. The females were checked daily concerning the status of the vaginal closure membrane. Pregnancy was calculated from the second day of vaginal opening (27) and the animals were examined by palpation early in pregnancy.
Substance 1 was dissolved in sesame oil (1 ml) and administered orally with a 1 ml syringe once daily in the morning at a dosage of 10 mg/kg from cycle day 14 until parturition. Animals in control group were similarily given 1 ml sesame oil daily in the morning. APRIL 1999 VOL. 39 NO. 4
421
CONTRACEPTION
-1 Eis_L Tetrahydrocyclopent-b-indole
in this study tested for effects on
guinea pig reproduction.
CH3 I
2 = EgJ. Yuehchukene, considered rats and hamsters.
422
to have antiimplantation effects on
APRIL 1999 VOL. 39 NO. 4
CONTRACEPTION
cal evalug&g X2-analysis or Fischers exact test and Students t-test were used as described (28). RESULTS In the treatment group all animals ( 1O/ 10) became pregnant, while in the control group only 14/20 became pregnant. Most pregnancies in the treatment group occurred during the first cycle (8/ 10) while in the control group often several cycles passed until pregnancy took place (Table I).
Table I. Time of conception in guinea pigs with and without oral treatment ( 10 mg/kg/day) with an indole derivative (compound J_)
Cycle 1
Treatment group 8/l 0 (n-10)
Conceotion rate Cycle 2-4 No. of animals Significance not pregnant after 4 cycles 212
0X X
Control group (n-10)
2/20
14/18
63
Group differences were tested by X2-analysis; xxx denotes p
Apparently, the drug facilitated conception (p
APRIL 1999 VOL. 39 NO. 4
423
CONTRACEPTION
Table II. Effects of oral treatment (10 mg/kg/dayl with an indole derivative (compound 1) on litter size in guinea pigs
Treated animals Control Total number of pups/ pregnant animals
28/10
Mean litter size
2.8
Median ”
”
-xxx
55114
7
3.9
-xxY4 3
Significance of group differences was analyzed by Students t-test. xxx means p
The number of dead pups/litters in the treatment group were smaller than in the control group and there were no toxic effect on the pups or on the dams (Table III).
Table III. Number of dead pups in litters from guinea pigs given oral treatment (I 0 mg/kg/day) with an indole derivative (compound 11
Treatment group (n=lOl No. of dead pups/ total no. of pups Pup deaths/litter size
4128
212; 213
Control (n=141 12155
214; 4/4; l/3 314 215
Fischer’s exact test was used for testing of significance; no significant difference between the groups was found.
424
APRIL 1989 VOL. 39 NO. 4
CONTRACEPTION
DISCUSSION The indole compound 1 structurally and functionally related to indomethacin likely caused a reduction in the number of ova available for conception as indicated by the reduced litter size. However, an effect on ova quality or tubal and/or uterine motility cannot be excluded. Resorption of ova after indomethacin has been shown in the rabbit (13) but whether this was due to a nonspecific toxic effect or to specific changes at the implantation sites is uncertain. In the guinea pig about 20 follicles (ovulation points) were reported to develop per cycle and 20 ova were found by Maia et al. (30). Furthermore, after indomethacin ( 12 mg/kg/24h on cycle day 16) treatment these investigators found that ovulation points were reduced to 13 and only 10 ova could be recovered. The ova transport was suggested to be unaffected. However, Sisk (27) considers the normal number of ovulation points to be 3.33.4 and the number of corpora lutea on the average 3.5 in the guinea pig (3 1). The indole compound l-treated animals in the present study conceived faster than the control females. The possibility that a prolonged progesterone secretion favored implantation can not be excluded since Poyser (6) reported a prolongation (3 days) of the estrous cycle after indomethacin treatment in high doses (40 mg/d) to the guinea pig. Apart from a facilitated implantation induced by progesterone, the rapid conception observed in the female guinea pig receiving compound 1 may also be explained by an improved sperm penetration and/or capacitation. Female genital tract secretion is generally considered to be at least in part regulated by prostaglandins ( 19). Our results are somewhat in contrast to the results of Kong et al. (23) who found that the indole yeuhchukene counteracted reproduction in the female rat. This difference may be due to species differences or due to the fact that the indole 1 is structurally different from yeuhchukene. There were no toxic effects noted on the dams or on the litters (with compound 1) despite continued treatment throughout pregnancy. The number of dead pups in our study did not exceed that in the control groups. The mean litter size in the indole (compound l&exposed group was 2.8. Interestingly Wright (29) has pointed out that a litter size of 2 in the guinea pig is optimal for pup survival which is in agreement with the increased mortality in our control groups. Our results thus indicate that indole 1 is comparatively nontoxic.
APRIL 1999 VOL. 39 NO. 4
425
CONTRACEPTION
Whatever mechanisms that were involved, the present results indicate that our indole compounds may improve reproductive performance in the female guinea pig. ACKNOWLEDGEMENT The skilful technical assistance of Ms. Eva Andersson, Margareta TapperPersson and Ann Hagstrem is gratefully acknowledged. For typing the manuscript we are indebted to Ms. Ann Nicolausson. This project has been supported by research funds from the Karolinska Institute. REFERENCES 1. Couthino, E.M.and H.S.Maia: The contractile response of the human uterus, fallopian tubes, and ovary to prostaglandins in vivo. Fert. Ster. 1971; 22,539-543. 2.
Christensen, N.J:Studies on the bioconversion of arachidonic acid in human pregnant reproductive tissues. Thesis. Stockholm, Sweden 1984.
3. Wehrenberg, W.B.,D.J.Dierschke, and R.C.Wolf: The effect of prostaglandin F2o on ovarian blood flow, and progesterone concentration in cyclic guinea pigs. Biol. Reprod. 1979; 2 1, 187- 19 1. 4. Tso, B.C.-F.and W. H. Tam: The luteolytic effect of prostaglandin F2o on the guinea pig corpus luteum in relation to gonadotropins and stages of the oestrous cycle. Can. J. Physiol. Pharmacol. 1978; 56 828833. 5. Blatchley, P.R.and B. T. Donovan: The effect of prostaglandin F2o and prostaglandins E2 upon luteal function and ovulation in the guineapig.J. Endocr. 1972; 493-50 1. 6. Poyser, N. L: Prostaglandin F2a in the uterine luteolytic hormone in the guniea pig: The evidence reviewed. In: Advances in Prostaglandins and Tromboxane Research, ~012. (ed. B. Samuelsson and R. Paoletti). Raven Press, New York, 1976, pp. 633-640.
426
APRIL 1999 VOL. 39 NO. 4
CONTRACEPTION
7. Deansly, R. and J.S. Perry: Independent regression of normal and induced corpora lutea in hysterectomized guinea-pig. J. Reprod. Fert. 1969; 20,503-508. 8.
Labhsetwar, A. P: Luteolysis and ovulation induced by prostaglandin F2o in the hamster. Nature 1971; 230,528-529.
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