ARNOLD JOHN GRIFF CHARLES Beth.esda,
Plasma follicle-stimulating hormone (FSH) and Euteinizing hormone (LH) levels in samples taken during clomiphene-induced ovulatory cycles in 7 oligo-ovulatory or anovulatory infertile women were determined by radioimmunoassay. These results were compared with results of similar studies made during spontaneous ovulatory cycles in 21 normal volunteers. In the clomiphene-induced ovulatory cycles FSH and LH values were significantly above pretreatment levels after 2 to 3 days of therapy, but subsequently declined prior to a preovulatory peak occurring I3 days after beginning of therapy. Following the preovulatory peak during the luteal phase values declined to the lowest levels observed at any time during the treatment cycle. These changes in plasma FSH and LN concentrations during clomiphene-induced cycles were similar to those seen in normal spontaneous cycles. The early increase in FSH and LH in the induced cycle suggests that the initial site of action of clomiphene ir on the hypothalamic-pituitary portion of the hypothalamic-pituitary-ovarian axis. It also suggests that treatment with clomiphene may provide a functional test of this axis.
W x H A v E previously reported changes in plasma luteinking hormone (LH) levels1 and in preliminary form on plasma folliclestimulating hormone (FSH) levels2 during ciomiphene-induced ovulatory cycles and postulated an initial site of action of this From the Reproductive Research Branch, National Institute of Child Health and Human Development and Surgery and Endocrinology Branches, National Cancer Institute, National Institutes of Health. *Supplied by J. E. Johnson, M.D., William S. Merrill Company. ++*Present address: Department of Obstetrics and Gynecology, University of Miami School of Medicine, Miami, -Florida.
drug on the hypotha!amic-pituita~~~ axis. The development of a radioimmunoassay for follicle-stimulating hormone3 has made it possible to determine plasma FSH concentrations in aliquots of the same samples used previously for plasma LH determination. In this report changes in plasma FSH and LH occurring during clomiphene-induced ovuiatory cycles are compared to changes in plasma gonadotropins observed during spontaneous ovulatory cycles in normal subjects as reported by Cargille, ROSS, and Uoshimi.*
Clomiphene citrate (100 to 200 mg.; was given daily for 5 days to 7 infertile women.
Plasma FSH and LH during
Diagnostic groups included one patient, K. N., with secondary anovulation with lactation; one patient, P. L., with secondary anovulation without lactation; one patient, M. S., with oligo-ovulation with lactation, 3 patients, L. G., L. M., and D. P., with oligo-ovulation without lacmtion, and one patient, J. E., with a sclerocystic ovarian syndrome. Plasma from daily heparinized venous blood samples were frozen until FSH determinations were made according to the radioimmunoassay method of Cargille ;and associates.3 Basal body temperatures wlere recorded daily. Endometrial biopsies were obtained both before, during, and after cl.omiphene therapy. Diagnosis of ovulation was made only when a shift to a progestational endometrium was seen. Secre-
C. Hem. by Culdo 6x
id mg /Day L--l
I 25 Jan
tory endometrium was dated according to the criteria of Noyes, Hertig, and Rock.5 In selected cases urinary pregnanediol excretion and histologic examination of tissue obtained by ovarian biopsy under culdoscopic examinationG pro,vided additional information. Cycle days were numbered consecutively following day 1, the day on which clomiphene therapy was initiated. FSH” and LH7 determinations were made on serial plasma samples from 21 normal women between 18 and 27 years of age. Ovulatory status was indicated by measurements of basal body temperature and determination of both urinary pregnanediol excretion and plasma progesterone levek8
Fig. 1. Plasma FSH during a typical clomiphene-induced IRP 2 per milliliter of plasma. Pregnanedid is in mg. secretory endometrial biopsies are depicted, respectively, of the secretory endometrium.
Mar ovulatory cycle. FSH is in m1.U. per 24 hour urine. Proliferative and by P and S; numbers indicate dating
32.0 6.9 ..-
165 1 ‘1
consecutively. of plasma.
3 3 7 3 4 5
143 146 106 261 130 132
_____-_ subsequent days numbered SRP 2 IIMG per milliliter
value on day peak to menses.
3 3 3 3
day of clomiphene administration; subsequent days numbered consecutively. .~Prctxcatment values expressed in m1.U. IRP 2 HMG per milliliter of plasma, $Per cent of pretreatment value on day 1.
Mean 5.3%. ___~~_--~~~~~-..
lPer cent of pretreatment ub3ys from preovulatory
administration; expressed in m1.U.
day of clomiphenc
11.5 17.5 16.5 11.5 16.5
G. L. M. N. P.
L. P. L. K. D.
96 114 158 205 100 100
143 146 la5 261 130 144
10 12 12 15 10 13
61 34 79 57 55 64
12 14 13 17 12 15
36 57 200 122 106 192
14 5 12 6 lR 1;
70 48 41 43
82 ?F e ‘. Q “Lti
$i. VI “?g n
Volume 102 Number
Mean values for plasma FSH during clomiphene-induced ovulatory cycles in all 7 patients studied are shown in Table I. Plasma FSH values obtained on day 3 or 4, at the time of the clomiphene peak, at the follicular nadir, and during the luteal phase were all significantly different from day 1 plasma FSH levels (p < 0.05). Two patients had unusual luteal phases. One subject with secondary amenorrhea with lactation, K. N., had a short luteal phase of 6 days. The remaining subject with secondary amenorrhea, R. L., had a Weal phase which was not associated with a basal body temperature elevation and lasted only 5 days. However, ovulation in this woman was confirmed at culdoscopy by visualization
The pattern of response for FSH in a clomiphene induced ovulatory cycle in a typical subject is shown in Fig. 1. The response of this subject was representative of responses setan in 5 patients having luteal phases which were associated with clearly elevated basal body temperatures and which lasted 12 to 15 days. The FSH pattern was typified by an early clomiphene rise which was evident by the second day following initiation of therapy and reached a maximum value on day 5, the clomiphene peak. Following this peak there was a decline to a follicular nadir and a subsequent preovulatory peak followed by very low luteal phase levels.
Size Endometriol 8x
3X2 5X5 P
t C. Hem. by Culdo 8x
a oz -
2 ? z
Clomid 200mg /Day I I I I 5 Aug
Fig. 2. Plasma FSH during a clomiphene-induced ovulatory phase. Symbols are as in Fig. 1. Ovarian size is in centimeters.
and bropsy of a corpus hemorrhagicurn
peak was greater than. Zhe ciomiphene peak in all patients studied. Changes in plasma FSW and LH during a spontaneous cycle in a typical normal woman are shown graphically in Fig. 4 and mean changesin gonadotro,pinsin 21 normal women are tabulated in Table III. Complete results of serial FSH and LH determinations jn these 21 normal women are reported by Cargille, Ross, and Yoshimi.* Day I plasma FSH values in the &go-ovulatory and anovulatory patients were not significantly different from the mean early follicular values in normal women. The 10 day interval between the FSH clomiphene peak and the preovulatory peak corresponded to the 10 day interval between the foliicular and midcycle peaks in spontaneous cycles. In both clomi;he preovulatory
Plasma iH d Plasma FSH
Feb Mar Fig- 3. Plasma FSH and LH during a typicak clomiphene-induced ovulatory are as in Fig. 1. FSH and LH are in m1.U. IRP 2 per milliliter of plasma. Jan
Plasma FSH and LH during
phene-induced and spontaneous ovulatory cycles the midcycle preovulatory FSH and LH peaks usually coincided and then both gonadotropins declined to low luteal levels. In addition to similarities, differences were observed in patterns of FSH and LH in the clomiphene-induced ovulatory cycles and the spontaneous ovulatory cycles. Plasma LH values on d.ay 1 in the oligo-ovulatory and anovulatory patients were significantly high-
s. D - NORMAL
er than normal follicular levels in 4 of 7 patients. In the clomiphene cycle there was a marked early follicular LH peak whereas in the normal ovulatory cycle, although follicular levels were higher than luteal levels there was no true peak. In the clomiphene-induced ovulatory cycle the FSH follicular nadir was significantly lower than in the normal spontaneous cycles. In the clomiphene-induced ovulatory cycle the FSH
Plasma Progesferone /mpg/m/J
0.65 I I
I ’ , ’ I ’ ’ I ’ I ’ I 1 I
d 1 \
Fig. 4. Plasma in mI.TJ. IRP
FSH and LH 2 per milliliter
FSH and LH during
1 follicular ,
13.6 -,no_.I 20.12 1.4s
Follicular nadir (% changeI) 72 2 6.6
cycle. FSH and LH are milliliter of plasma.
sarly rise 1 (74 Ghanget)
*Obtained from Cargille and Ross. fm1.U. IRP 2 HMG per milliliter plasma +_ standard error. $Per cent of first half follicular values k standard error. $LH
17 21 OF CYCLE
during a typical spontaneous ovulatory of plasma. Progesterone is in mpg
Midcycle peak (% change$) 144+ 351k
Luteal phase (% changef) 54 It 3.0 68 Lb 4.0
clomiphene peak was usua!ly greater than the midcycle peak whereas in a spontaneous cycle the FSH midcycle peak usually exceded the maximum value of the follicular phase.
In all patients reported upon here, ovulation was always associated “with an early rise in both FSH and LH followed by preovulatory peaks of both hormones. However, occurrence of the early clomiphene peak did not insure ovulation, since we have observed among patients who did not ovulate following clomiphene an early rise in both FSH and LH without a second preovulatory peak. Among such patients, the administration of HCG a.t appropriate intervals following clomiphene as proposed by KistneP may increase the number of ovulatory responses. The similarities between the early follicular phase FSH levels in the clomipheneinduced and spontaneous ovulatory cycles suggest that the clomiphene FSH peak is of more functional significance than the clomiphene LH peak which has no counterpart in the normal ovulatory cycle. The greater
prominence of the FSH clomiph.ene peak Eompared to the early follicular rise in spantaneous cycles may account for the mild ovarian hyperstimulation frequently seen in the clomiphene-induced cycles. In addition, the correspondingly Iower FSH follicular nadir in the induced cycle may be due to increased ovarian steroid hormone production. On the basis of studies of plasma LH levels during clomiphene therapy in these women, we suggested that the initial action of clomiphene was on the hypothalamic pituitary axis.l The early rise in plasma FSH noted is consistent with this hypothesis, since these changes in plasma gonadotropins precede all previously reported changes in vivo estrogen metabolism.10T I1 These early increases in FSH and LH also precede a.ll previously reported changes in gonadotropin excretion.12j I3 In addition, the early rise in plasma gonadotropins following domiphene suggested that clomiphene may provide a test of the functional competence of the hypothalamic pituitary gonadal axis in women. The technical assistance of Mr. Mathews is gratefully acknowledged.
Jacobson, A., Marshall, J. R., and Ross, G. T.: AM. J. OBST. & GYNEC. 101: 1025, 1968. A., Marshall, J. R., Ross, G. T., 2. Jacobson,. and Carzille. C. M.: Lancet 1: 1371. 1968. 3. C&gilIe,“C. &., Ross, G. T., Howland; L. A., and Rayfard, P. L.: Clin. Res. 14: 33, 1968. 4. Cargiile, C. M., Ross, G. T., and Yoshimi, T.: In preparation. 5. Noyes, R. W., Hertig, A. T., and Rock, J.: Pertil. & Steril. 1: 3, 1950. 6. Marshall, J. R., and Hammond, C. B.: AM. J. OBST. & GYNEC. 96: 1022, 1966. 7. Odell, W. D., Ross, 6. T., and Rayford, P. L.: J. Clin. Invest. 46: 248, 1967,
Yoshlmi, T., and Lips&, ha. B.: Steroids 11: 527, 1968. 9. Kistner, R. W.: Ferril. & Steril. 17: 569, 1966. PO. Barlow, J. J., and Logan, C. M.: Ax J. OBST. & GYNEC. 98: 687, 1967. 11. Smith, 0. W.: Aivr. J. OBST. & GYNK. 94: 440, 1966. 12. Dickey, R. P., Vorys, N., Stevens, V, C.: Besch, P. K., Hamwi, G. J., and Xlery, J. C.: Fertil. & Steril. 16: 485, 1965. 13. Mishell, D. R.: Fertil. & Steril. 18: 102, 1.967.