HORMONAL EFFECTS OF AN ANTIESTROGEN, TAMOXIFEN, IN NORMAL AND OLIGOSPERMIC MEN*

Transcription

1 FERTILITY AND STERILITY Copyright ~ 1978 The American Fertility Society Vol. 29, No.3, March 1978 PrintRd in U.s.A. HORMONAL EFFECTS OF AN ANTIESTROGEN, TAMOXIFEN, IN NORMAL AND OLIGOSPERMIC MEN* ALEX VERMEULENt FRANK COMHAIRE Section of Endocrinology and Metabolic Diseases, Department of Internal Medicine, Akademisch Ziekenhuis, University of Ghent, Ghent, Belgium The administration oftamoxifen, 20 mg/day for 10 days, to normal males produced a moderate increase in luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone, and estradiol levels, comparable to the effect of 150 mg of clomiphene citrate (Clomid). However, whereas Clomid produced a decrease in the LH response to LH-releasing hormone (LHRH), no such effect was seen after the administration of tamoxifen. In fact, prolonged treatment (6 weeks) with tamoxifen significantly increased the LH response to LHRH. Treatment of patients with "idiopathic" oligospermia for 6 to 9 months resulted in a significant increase in gonadotropin, testosterone, and estradiol levels. A significant increase in sperm density was observed only in subjects with oligospermia below 20 x 10 6 /ml and normal basal FSH levels. When basal FSH levels were increased or oligospermia was moderate (>20 x 10 6 /ml), no effect on sperm density was seen. As sperm density increased, FSH levels decreased, suggesting an inhibin effect. Sperm motility was not improved by tamoxifen treatment. In five boys with delayed puberty, tamoxifen treatment appeared to activate the pituitary-gonadal axis and pubertal development. It is generally accepted that the feedback of androgens at the hypothalamic level is, at least partially, mediated via their transformation to estradiol, which binds to hypothalamic receptors. Antiestrogens, such as clomiphene citrate (Clomid), tamoxifen (Nolvadex), or nafoxidin, are presumed to displace estrogens from their receptors and in this way to interfere with the normal feedback of sex steroids, resulting in increased secretion of gonadotropin-releasing factor and subsequently in increased gonadotropin release. Hence antiestrogens lead to endogenous stimulation of the gonads by both luteinizing hormone (LH) and follicle-stimulating hormone (FSH). Received August 23, 1977; accepted October 25, *Supported in part by Grant from the Fonds voor Wetenschappelijk Geneeskundig Omderzoek (FWGO), Brussels. treprint requests: Prof. Dr. A. Vermeulen, Section of Endocrinology and Metabolic Diseases, Department of Internal Medicine, Academic Hospital, University of Ghent, De Pintelaan 135, B 9000 Ghent, Belgium. Clomiphene citrate has been used for inducing ovulation in women with functional anovulatory amenorrhea, whereas in males it has been used in the treatment of idiopathic oligospermia. Results in the latter depend upon the dose used: low doses result in an increase in sperm concentration, and higher doses (200 to 400 mg/day) result in a decrease or even in azoospermia. 1, 2 Similarly, cisclomiphene, when given in low doses, results in a positive spermatogenic response,3 with variable effects at higher dosage. 4 Another possible use of anti estrogens is the induction of puberty in delayed adolescence. In comparison with clomiphene, tamoxifen has a much weaker intrinsic estrogenic activity, which in males might have a deleterious effect; therefore, we decided to study the hormonal effects of tamoxifen in both normal and oligospermic men. A preliminary study of the influence of tamoxifen in physiologically delayed puberty in males was also conducted. 320

2 Vol. 29, No.3 HORMONAL EFFECTS OF TAMOXIFEN IN NORMAL AND OLIGOSPERMIC MEN 321 Patients EXPERIMENTAL PROCEDURE Hormonal effects were studied in several groups of subjects. Short-Term Study. In a short-term study, the hormonal effects of tamoxifen, 10 mg twice daily for 10 days, in a group of normal adult male volunteers (n = 6) were compared with the hormonal effects of clomiphene citrate (Clomid), 50 mg three times daily for 10 days, administered to a comparable group of subjects. LH, FSH, testosterone (T), and estradiol (E2) levels were measured daily; moreover, the gonadotropin response to LHreleasing hormone (LHRH) before and at the end of treatment was evaluated. In the volunteers receiving tamoxifen, treatment was continued for 6 weeks and hormonal effects as well as the LHRH response were reevaluated at the end of treatment. Long-Term Study. The long-term hormonal effects of tamoxifen were studied in a group of I patients 22 to 35 years old (n = 21; Table 1) with so-called "idiopathic" oligospermia. Most of these men had previously been treated with either androgens or gonadotropins (human chorionic gonadotropin) without success, but none had received any hormonal treatment for the previous 6 months. Seven men had undergone ligation of the vena spermatica for varicocele at least 1 year previously. In none of the patients had sperm quality improved during the last 9 months, and repeated analysis had consistently revealed sperm concentrations below 40 x 10 6 /ml (in 15 of21 men the sperm concentration was below 20 x 10 6 /mi). None had testicular atrophy or a history of testicular inflammatory disease. The hormonal response was evaluated after 1, 3, 6, and 9 months, and in some men (n = 6) after 1 year of treatment. Moreover, on each occasion a specimen of semen, freshly obtained by masturbation after 48 hours of sexual abstinence, was obtained and analyzed by one of us (F. C.). Motility was graded 3+ for rapidly progressive spermatozoa and 2+ for sluggishly progressive spermatozoa. Detailed data on the influence of tamoxifen on the ejaculate have been reported previously.s Delayed Puberty. Five boys with idiopathic delayed puberty (pubertal rating of Tanner 6 : P2) were given 10 mg of tamoxifen twice daily for 3 months. Hormonal and clinical responses were evaluated after 6 weeks and 3 months of treatment, respectively; in two subjects treatment was continued for 6 months. Parents were informed about TABLE 1. Diagnosis, Sperm Concentration, Motility, Testosterone, LH, and FSH in Oligospermic Men before Treatment with Tamoxifen, 20 mg Daily Patient Age Diag- Sperm Sperm motility nosis" ~~:~fo~ T LH FSH x 10"Imi % % ngldl nglml nglml F.S.C L.L_ C_ S_ _2 2.9 V.d.V.A_ _ _2 V_M_ 25 OV V.H. 30 OV 4_ _3 D.W. 28 OV 5_ I. A. 32 OV D.B. P V.P. 29 OV D.RA. 28 OV _9 1.9 D_L.L L.R D.R 32 OV V.C C.O D.W.F D.M.S D.C.E C. J ,8 4.3 V.RA aio, Idiopathic oligospermia; OV, operated varicocele. the experimental character of the treatment and gave their consent. Methods Plasma testosterone and estradiol levels were measured by radioimmunoassay as previously described LH and FSH levels were measured by a double-antibody method using the commercial CEA-IRE-SORIN (Belgium) kits; results are expressed in nanograms per milliliter of Medical Research Council standards 68/40 and 68/39, respectively. Statistical analysis was performed by using either Student's paired t-test or Wilcoxon's signed rank test where indicated. RESULTS The administration of tamoxifen, 20 mg/day for 10 days, to normal males (n = 6, ages 33 to 70 years) resulted in a statistically significant increase (p < 0.01) in LH levels after 3 days of treatment (Fig. 1), a plateau at 150% ± 20% (mean ± standard deviation) of basal levels being reached after 6 days of treatment. FSH levels increased more progressively to a level of 155% ± 15% of basal levels. Parallel with the LH levels, mean T levels increased to a plateau of 142% ± 11% of basal values, whereas E2 levels increased to a plateau at 151% ± 20% of basal values.

3 322 CLOMID 3x 5Omg/d n-11 VERMEULEN AND COMHAffiE March 1978 TAMOXIFEN 2x10mg/d n-7 FIG. 1. Influence of Clomid, 150 mg/day, and tamoxifen, 20 mg/day, on hormonal parameters in normal males. As compared with the hormonal effects of Clomid, 50 mg three times daily for 10 days (Fig. 1), the response was slightly weaker; however, with both drugs the range of increase in hormonal levels was rather wide. The results of the LHRH test (100 JLg intramuscularly), performed before and after treatment with either Clomid or tamoxifen for 10 days, revealed some differences (Fig. 2). After Clomid treatment, LHRH stimulation resulted in LH values constantly lower than before treatment, whereas FSH levels were constantly higher; however, the increase of FSH above basal levels (sum total of levels at 20, 40, 60, and 120 minutes less 4 times the basal level = ~.:lfsh) was not significantly higher than that observed before treatment. After 10 days oftreatment with tamoxifen, levels of LH and FSH after LHRH stimulation were constantly higher than before treatment. The increase above basal levels as well as the ratio (R) of ~.:llh to ~.:lfsh, however, did not show any significant difference from the corresponding values before treatment. Prolonged treatment with tamoxifen, 20 mg/day for 6 weeks, in four normal males resulted in a slightly greater increase in LH (172P1o ± 10%) and FSH levels (162P1o ± 12P1o) than after short-term treatment; similarly, the increase in plasma T (83% ± 36%) and E2 (104% ± 29%) levels was greater than that after 10 days of treatment. LHRH stimulation finally resulted in higher absolute LH and FSH levels and a higher increase above basal levels than after 10 days of treatment; moreover, ~.:llh increased more than ~.:lfsh, with a ~.:llh:~.:lfsh ratio of 9.5 ± 3.4 versus 6.1 ± 1.1 after 10 days of treatment. In patients with oligospermia, treatment with tamoxifen, 20 mg/day for 1 month, resulted in an increase of the mean FSH level by 44% ± 24% (not significant) (Fig. 3), of the mean LH level by 96% ± 44% (p < 0.05), and of the mean testosterone level by ± 100% (p < 0.001). In view of the duration of spermatogenesis (74 ± 4 days) and the tubular transit time,9. 10 the influence of treatment on sperm quality was not evaluated after 1 month. After 3 months of treatment, the mean FSH level was significantly increased (80% ± 21% (P < 0.01, paired t-test), whereas the increase in the mean LH level (42P1o ± 24%) was not statistically significant. T levels and E2 levels, on the other hand, remained increased by ±100% (P < 0.001). Upon continuation of tamoxifen treatment, FSH levels tended to decrease (P < 0.05), but they remained significantly increased (6 months, P < 0.01; 9 months, P < 0.05) as compared with basal levels. LH levels and T levels, however, remained at a plateau; by 6 months and 9 months the increase in LH levels was still significant (P < and P < 0.05, respectively [Wilcoxon's signed rank test]). As far as the influence of tamoxifen treatment on sperm quality is concerned, after 3 months of treatment, neither sperm concentration nor sperm motility showed any statistically significant improvement (paired analysis) when all cases (n = 21) were considered together. However, when only subjects with a sperm density below 20 x 10 6 / ml and normal basal FSH levels were considered (n = 12), a borderline significant increase (P < 0.05, paired t-test) in sperm concentration from a mean of 7.7 ± 5.9 x 10 6 /ml to 16.9 ± 15.2 (SD) x 106/ml was observed. After 6 months of treatment, the mean sperm concentration had increased to 29.2 ± 26.7 (SD) x 10 6 /ml (P < 0.02); again, the effect on sperm concentration was more pronounced (mean, 39.9 ± 24.8 x 10 6 /ml; P < 0.01) when only subjects with normal basal FSH levels and a basal sperm concentration below 20 x'10 6 / ml were considered; moreover, the increase after 3 to 6 months of treatment was statistically

4 Vol. 29, No.3 HORMONAL EFFECTS OF TAMOXIFEN IN NORMAL AND OLIGOSPERMIC MEN 323 LHRH1001lll 1M Basal AfterlOd ~.~_... Basal After 1OdTamox. 15Omgld. ~L\.LH : 17.5±l.2 9.4±2.l 16.7±l ±l ±4.:2 ~L\.FSH: O'..o'.W 12c 20' 40' 50' 12cl:zO' 40' 60' FIG. 2. Gonadotropin response to LHRH before and after 10 days and 6 weeks of treatment with clomiphene citrate, 150 mg/day, and tamoxifen, 20 mg/day. Means ± standard deviation and ranges are indicated. R = ratio of ~ ~LH to ~ ~FSH. significant (P < 0,025). No further increase was observed after 9 months of treatment, On the other hand, no statistically significant effect on sperm motility was observed during the entire period of treatment (Fig. 3). In a small number of patients (n = 6) in whom the effect of 9 months of treatment with tamoxifen on sperm concentration was considered insufficient, treatment was continued for 1 year; all of these men had a basal sperm concentration below 20 x 106/ml (mean, 7,9 ± 4.3 [SD]). Although after 1 year of treatment the mean sperm concentration had increased to 17.6 ± 15.7 (SD) x 106/ml (P < 0,025, Wilcoxon's signed rank test), no significant improvement with regard to the results obtained after 9 months was observed. Sperm motility remained seriously impaired (mean, 14% 3+ and 18% 2+) and neither mean FSH levels nor LH levels were increased above basal levels, although T levels were still significantly increased <P < 0,01) (mean, 165% ± 11% of basal levels). The last group of subjects studied consisted of five boys with idiopathic delayed puberty in pubertal stage 2 according to the definition of Tanner.6 Tamoxifen, 20 mg/day, resulted in a clear-cut activation of the pituitary-gonadal axis as evidenced by the increase in gonadotropin and testosterone levels determined after 1 and 3 months of treatment, respectively; moreover, a rapid progression of pubertal development was observed (Table 2). DISCUSSION Both in normal subjects and in patients with idiopathic oligospermia, the administration of 20 mg/day of tamoxifen resulted in activation of the hypothalamic-pituitary-gonadal axis with a moderate but statistically significant increase in plasma LH, FSH, T, and E2 levels. This finding contradicts the results reported by Willis et al.,11 who did not observe any stimulation of LH and FSH levels after the administration of tamoxifen to normal males at a dose of 10 mg/day for 7 days. In the normal subjects the degree of activation of the hypothalamic-pituitary-gonadal axis obtained by tamoxifen, 20 mg/day, was slightly,

5 324 VERMEULEN AND COMHAIRE March 1978 TAMOXIFEN 20mgld.., -~:.=--,... <:0.001 <:0.01 <:0.05 o <:0.05(W) <:0.02(W) * > 15 L._ ~ ~ "5 5 2+n.S. +n.s. E O'~ ~ FIG. 3. Evolution of hormonal parameters, sperm concentration, and sperm motility under chronic treatment with tamoxifen, 20 mg/day, in oligospermic men. n.s., Not significant. W, Wilcoxon's signed rank test. although not significantly, less than that obtained with Clomid, 150 mg/day. However, the increase in LH levels observed after Clomid treatment was somewhat lower than that reported by Santen et a1.,12 who administered only 100 mg of Clomid, but rather similar to the increase observed by Bardin et a1. 13 In accordance with results published by Hashimoto et a1.14 and Dhont et a1.,15 the pituitary response to LHRH after the administration of clomiphene citrate, 150 mg/day for 10 days, was characterized by a decrease in the LH response in comparison with the pretreatment response; tamoxifen treatment for 10 days did not change the response to LHRH. Indeed, although the absolute levels of LH and FSH were higher after treatment than before, the increases in both LH (I ~LH) and FSH levels (I ~FSH) were similar. The difference in response might be attributable to the weak intrinsic estrogenic effect of Clomid, which in this study manifested itself by an increase in transcortin and testosterone/estradiolbinding globulin levels; this increase was not observed after tamoxifen treatment. The hypoth- ffi r.. "i1 },...-IC'jMCOcn :3 tr.i""; <>:i c-i ai I E-< "0.~... ~ E-< ~~g~~~ ~ ;::,...-t~c'1oot.o 1n.~ ::t 00>"<1'(00 1! ~ ~o,...i~c"i (J - "i1 ~ ~ ~ ::t ~ "<1'",ao~~ -...:l c-i,...;,...;"";c-i p.; E-< ~g~~~~ ~ ;:: M C'l1"""l 1n

6 Vol. 29, No.3 HORMONAL EFFECTS OF TAMOXIFEN IN NORMAL AND OLIGOSPERMIC MEN 325 esis put fo.rward by Hashimo.to. et al. 14 that the decreased LH respo.nse is the co.nsequence o.f increased T levels do.es no.t explain the no.rmal LH respo.nse after tamo.xifen treatment, which induced a T increase similar to. that induced by Clo.mid. Mo.reo.ver, Twas fo.und by Franchimo.nt et al. 16 no.t to. impair the respo.nse to. LHRH. Wang et al.,17 who. also. o.bserved a decreased go.nado. tro.pin respo.nse to. LHRH after clo.miphene administratio.n, interpreted this decrease as a co.nsequence o.f the increased endo.geno.us estradio.i levels. This also. seems rather unlikely, as a co.mparable increase in estradio.i levels was o.bserved after tamo.xifen administratio.n. Therefo.re, a ro.le o.fthe intrinsic estro.genic activity o.fcio.mid which is practically absent with tamo.xifen seems the mo.re pro.bable explanatio.n. It is interesting that Santen18 repo.rted a blunted LH respo.nse but a no.rmal FSH respo.nse to. LHRH in males during estradio.i infusio.n at a rate o.f 3.5 ILg/ho.Ur. Pro.Io.nged treatment with tamo.xifen fo.r 6 weeks resulted in an enhanced respo.nse o.f bo.th LH and FSH to. LHRH stimulatio.n, the difference in the respo.nse after 10 days o.f treatment being statistically significant (P < 0.01). On the basis o.fthese results sho.wing an intense activatio.n o.fthe hypo. thalamic-pituitary-go.nadal axis, we decided to. study the ho.rmo.nal respo.nse to. tamo.xifen treatment in o.ligo.spermic men and to. co.mpare the respo.nse with the changes in sperm quality. In all subjects studied (n = 21), tamo.xifen treatment resulted in a highly significant increase in T levels (P < 0.001), a significant increase in FSH levels (P < 0.01), and a bo.rderline significant increase in LH levels (P < 0.05). The effect o.n sperm quality was hetero.geneo.us, ho.wever. As expected, no. significant effect was seen in patients with increased basal FSH levels (Table 3), altho.ugh the increase in T levels was similar to. the increase in no.rmal subjects o.r in o.ligo.spermic patients with no.rmal basal FSH levels. Ho.wever, a significant impro.vement in sperm co.ncentratio.n was o.bserved within 3 mo.nths o.ftreatment o.fpatients with o.ligo.spermia o.f less than 20 x 106/ml; generally, impro.vement co.ntinued during the next 3 mo.nths, and after 6 mo.nths o.ftreatment 6 o.f7 patients with an initial sperm co.ncentration belo.w 20 x 106/ml (mean, 8.7; range, 2.3 to 14.6) and no.rmal FSH levels had a sperm co.ncentratio.n o.f mo.re than 20 x 106/ml (mean, 39.9; range, 0.4 to. 81). The effects o.n sperm mo.tility were disappo.inting, ho.wever, as no. significant impro.vement was o.bserved (Fig. 3). These results are at variance with tho.se repo.rted by Willis et ai.,l1 who. did no.t see any effect o.f tamo.xifen (10 mg/day) o.n sperm quality, but co.nfirm previo.us results o.btained in this labo.rato.ry.5 It is remarkable that FSH levels tended to. decline after pro.io.nged tamo.xifen treatment (and impro.vement o.f sperm density), and it is interesting to. no.te that Reyes and Faiman3 o.bserved a nadir in FSH levels at the end o.f treatment (6 mo.nths) with cisclo.miphene, 1 mg/day, whereas LH levels had declined to. co.ntro.l levels. Mo.reo.ver, these autho.rs o.bserved, as we did in o.ur subjects treated with tamo.xifen, that in different individuals the endo.crine respo.nses to. the antiestro.gens characteristically fluctuated. In the study by Reyes and Faiman,3 the FSH nadir also. co.incided with significant elevatio.ns in sperm co.ncentratio.n; this co.incidence was interpreted by Reyes and Faiman as an indicatio.n o.f the ro.le o.f inhibin as a feedback regulato.r o.f FSH secretio.n. The evo.lutio.n o.f the mean values in o.ur study Po.ints to.ward a similar inverse co.rrelatio.n, al- TABLE 3. Effects of Tamoxifen, 20 mg/day, on Sperm Quality in Oligospermic Men with Increased FSH Levels Patient Age Duration of treatment Sperm concentration Sperm motility mo x 100Iml % % nglml ngldl V. d.v. A. 33 Control mo mo mo mo Verm.M. 25 Control mo mo mo mo VanP. 29 Control mo mo mo FSH T

7 326 VERMEULEN AND COMHAIRE March 1978 TABLE 4. Effects of Tamoxifen, 20 mgiday, on Sperm Quality in Subjects with Low-Normospermia (>20 x 10"/ml) Patient Age Duration of treatment Sperm concentration x lo'/ml COS. 27 Control mo 33.0 De W.Fr. 25 Control mo mo 23.7 DeM.J. 25 Control mo 25.2 De Cl. E. 32 Control mo 26.7 C. J. 35 Control mo 40.0 V.Kerk. A. 28 Control mo mo 33.0 Sperm motility FSH % % ng/ml ng/dl T though the values observed in individual subjects were rather erratic. On the other hand, our study (Table 4) confirms the observation by Reyes and Faiman3 that subjects with low-normospermia (sperm density> 20 x l06/m!) did not appear to show a positive germinal cell response, although the hormonal response was similar to the response in more severe oligospermia. As a rule, the increase in sperm concentration in oligospermic men was greater after 6 months than after 3 months of treatment and remained stable afterward. If the duration of human spermatogenesis and tubular transit time are considered, this finding is not unexpected and stresses the necessity of continuing treatment for a sufficient period before concluding that treatment has failed. Besides inadequate doses, an insufficient duration of treatment may be responsible for contradictory results of treatment of oligospermia with antiestrogens. In boys with delayed puberty, the intrinsic estrogenic activity ofclomid precludes any activation ofthe hypothalamic-pituitary-gonadal axis, a consequence of the higher sensitivity of the gonadostat for sex hormone feedback in prepuberty19.20; indeed, Kulin et al. 19 showed that clomiphene citrate administration does not result in activation of testicular secretion until the middle of stage 3 or the beginning of stage 4 in Tanner's classification. 6 Cathro et al.,20 however, observed a stimulation of adrenal steroid secretion. Tamoxifen, which has a much weaker estrogenic activity, seemed to activate this hypothalamic-pituitary-testicular axis. Indeed, although it is impossible to eliminate a spontaneous activation of this axis in these boys in early puberty (Tanner stage 2), we have never seen a spontaneous increase in plasma T levels to normal adult values within 3 months, as was observed in this group. Moreover, gonadal growth and the development of pubic hair increased rapidly during treatment. Therefore, we think it warranted to conclude that tamoxifen activates the hypothalamic-pituitary-gonadal axis in these patients. Androgen treatment as well as chorionic gonadotropin treatment has been advocated in physiologically delayed puberty. Since androgens as well as human chorionic gonadotropin (the latter via stimulation of testosterone secretion) inhibit endogenous gonadotropin release, a similar activation of pubertal development-with fewer side effects-may be obtained by tamoxifen. However, although no side effects were observed in this study and although in animal experiments no impairment of spermatogenesis has been observed even after long-term treatment,21 further studies are necessary before tamoxifen can be advocated for the treatment of delayed puberty. REFERENCES 1. Heller CG, Rowley MJ, Heller GV: Clomiphene citrate: a correlation of its effects on sperm concentration and morphology, total gonadotropin and testicular cytology in normal men. J Clin Endocrinol Metab 29:638, Paulson DF: Endocrine therapy of male infertility, with special reference to clomiphene citrate (abstrl. Fertil Steril 28:329, Reyes FI, Faiman C: Long term therapy with low-dose cisclomiphene in male infertility: effects on semen, serum FSH, LH, testosterone and estradiol, and carbohydrate tolerance. Int J FertilI9:49, Wieland RG, Ansari AH, Klein DE, Doshi NS, Hallberg MC, Chen JC: Idiopathic oligospermia: control observations and response to cisclomiphene. Fertil Steril 23:471, Comhaire F: Treatment of oligospermia with tamoxifen. Int J Fertil 21:232, 1976

8 Vol. 29, No.3 HORMONAL EFFECTS OF TAMOXIFEN IN NORMAL AND OLIGOSPERMIC MEN Tanner JM: Growth at Adolescence, Second Edition. Ox- 14. Hashimoto T, Miayi K, Matsumoto K, Izumi K, Kumaford, Blackwell Scientific Publications, 1962 hara Y: LH and FSH response to synthetic LHRH after 7. Vermeulen A: Determination of androgens in plasma. In consecutive administration of clomiphene citrate in The Endocrine Function of Human Testis, Edited by normal males. J Clin Endocrinol Metab 41:1110, 1975 VHT James, M Serio, L Martini. New York, Academic 15. Dhont M. De Gezelle H, Vandekerkhove D: Modulation of Press, 1973, p 91 pituitary responsiveness to exogenous LHRH by an oes- 8. Verdonck L, Vermeulen A: Comparison of quick methods trogenic and an antioestrogenic compound in the normal for the estimation of estradiol in plasma by radioimmuno- males. Clin Endocrinol (Old) 5:175, 1976 assay. J Steroid Biochem 5:471, Franchimont P, Chari S, Demoulin A: Hypothalamus- 9. Heller CG, Clermont Y: Kinetics of the germinal epi- pituitary-testis interaction. J Reprod Fertil 44:338, 1975 thelium in man. Recent Prog Horm Res 20:545, Wang CF, Lasley BF, Yen SSC: The role of estrogen in 10. Rowley MS, Tishima F, Heller CG: Duration of transit of the modulation of pituitary sensitivity to LRF (luteinspermatozoa through the human male ductular system. izing hormone-releasing factor) in men. J Clin Endo- Fertil Steril 21:390, 1970 crinol Metab 41:41, Willis KS, London DR, Bevis BA, Butt WR, Lynch SS, 18. San ten RJ: Independent effects of testosterone and estra- Holder G: Hormonal effects of tamoxifen in oligospermic diol on the secretion of gonadotropins in man. In the men. J Endocrinol 73:171, 1977 Testis in Normal and Infertile Men, Edited by P Troen, 12. Santen RJ, Leonard JM, Sherins RJ, Gandy HM, Paulsen HR Nankin. New York, Raven Press, 1977, p 197 CA: Short and longterm effects of clomiphene citrate on 19. Kulin HE, Grumbach MM, Kaplan SL: Changing senthe pituitary-testicular axis. J Clin Endocrinol Metab 33: sitivity of the pubertal gonadal hypothalamic feedback 970, 1971 mechanism in man. Science 166:1012, Bardin CW, Ross GT, Lipsett MB: Site of action of clomi- 20. Cathro DM, Saez JM, Bertrand J: The effect of clomiphene citrate in men: a study of the pituitary-leydig cell phene on the plasma androgens of prepubertal and axis. J Clin Endocrinol Metab 27:1558, 1967 pubertal boys. J Endocrinol 50:387, Hemworth BM: Effect of the anti-cancer drug tamoxifen on spermatogenesis. IRCS Med Sci 3:627,1975