Estrogen dependence of the periovulatory plasma prolactin response to gonadotropin-releasing hormone in normal women

Estrogen Dependence of The Periovulatory Plasma Prolactin Response to Gonadotropin-Releasing Hormone in Normal Women A. Barbarino, L. De Marinis, A. Mancini, C. D’Amico, and S. Minnielli

It has been previously reported that GnRH is capable of inducing a PRL response in intact and castrated men treated with estradiol benxoate for 9-9 days. To further support the hypothesis of an estrogen-dependence of the PRL response to GnRH. GnRH was administered, either as a bolus or as a continuous infusion, to 45 normal women during various phases of their menstrual cycles. Synthetic GnRH (100 pg intravenous bolus) elicited a significant increase (mean 175%) in circulating PRL levels in nine women studied during the periovulatory phase of the menstrual cycle (days 14-17). Similarly, GnRH infusion (0.2 pg/min x 3 h) induced a PRL response (mean 149%) in six women studied during the same period. In contrast, saline infusion induced a modest decrease (37%) in plasma PRL levels in five women studied during the periovulatory period. No significant changes in circulating PRL levels were found after GnRH administration as a bolus or a continuous infusion, in 13 women during the late follicular phase (days 10-13) and in 12 women during the midluteal phase (days 21-24). Synthetic GnRH elicited the expected increase in gonadotropin levels during all phases of the cycle studied. The maximal response was found for both LH and FSH during the periovulatory phase of the cycle. In conclusion, the data confirm that GnRH is capable of stimulating a PRL response in humans and again suggest that this response is estrogen-dependent. Finally, a temporal correlation between the midcycle gonadotropin peak and the positive PRL response to exogenous GnRH has been established.

I

NTRAVENOUS INJECTIONS of gonadotropinreleasing hormone (GnRH) are commonly used as a test for pituitary gonadotropin reserve. In normal women the effect of this hypothalamic hormone has been considered specific since it stimulates the release of luteinizing hormone (LH), and, to a lesser extent, of follicle-stimulating hormone (FSH), while plasma levels of other pituitary hormones remain unaltered.lm3 Recent studies, however, have demonstrated that GnRH is capable of inducing a prolactin (PRL) release in patients with active acromegaly4 and in some normoprolactinemic anovulatory women.’ Such PRL responses were considered “nonspecific” or “paradoxical” since they occurred in pathologic conditions. However, a PRL increment after GnRH has also been described in normal and hypogonadal women,6 and this effect seemed independent of the ovarian steroid environment. Previously, we have observed a PRL response to GnRH in intact and castrated men after 8 to 9 days of treatment with estradiol benzoate (E2).’ We have now evaluated the pattern of PRL secretion after GnRH administration (as a bolus or a continuous infusion) in normal women during different phases of the menstrual cycle, in order to further assess the hypothesis that the PRL response to GnRH is estrogen dependent.

blood samples. These and basal body temperature were used to document the precise period of the cycle. The gonadotropin and PRL responses to an intravenous bolus of synthetic GnRH (100 rg) (Relisorm L 100, Serono, Rome, Italy) were evaluated in 24 women during the following menstrual phases: late follicular (days 9-12; eight women), periovulatory (days 14-17; nine women), and midluteal (days 22-24; seven women). During each study, blood samples were obtained via an indwelling cannula at - IO and 0 minutes and at 10,20,30,45,60.90, 120, 150. and 180 minutes after administration of the GnRH. The GnRH was infused at a rate of 0.2 rg/min for a three-hour period in five women during the late follicular phase, in six women during the periovulatory phase, and in five women during the luteal period. The GnRH was administered via a Holter pump through an indwelling needle in one arm, and blood samples were obtained from a needle inserted in the other arm I5 minutes before starting the infusion, every 15minutes for the first hour after infusion began, and every 30 minutes in the following two hours of infusion. Saline infusion for three hours was performed in five women during the pcriovulatory phase of the cycle, as a control. The modality and timing of samples during saline infusion were similar to those of GnRH infusion.

Assays Blood samples were centrifuged within two hours after collection, and separate plasma aliquots for steroid and gonadotropin determinations were stored at -20 “C until assayed. Plasma PRL,8 LH, FSH,9 and estradiol’O were measured by radioimmunoassay methods previously described, using kits by Biodata (Rome. Italy). Concentrations of progesterone (P) in plasma were measured by a specific radioimmunoassay with the use of polyethylene glycol

MATERIALS AND METHODS Subjects Forty-five normal female students (aged 21-28 years) from our University volunteered for this study. All of the subjects had regular menstrual cycles with a usual cycle length of 27 to 32 days, and none were taking any medications. The women were studied between 9 AM and 10 AM in a fasting state. The subjects were fully awake during the experiments and were resting in bed in a quiet room. Gonadotropm and ovarian steroid levels were determined in appropriate daily

Metabolism, Vol. 30, No. 11 (November),

1983

From the Institute of Internal Medicine, Division of Endocrinology, the Catholic University School of Medicine. Rome, Italy. Receivedfor publication March I, I983. Address reprint requests to A. Barbarino MD, L. De Marks. MD, Institute of Internal Medicine, Division of Endocrinology. the Catholic University School of Medicine, L. A. Gemelli 8. 00168 Rome, Italy. Q I983 by Grune & Stratton, Inc. 0026%0495/83/3211~008%01.00/0

1059

1060

BARBARINO ET AL

(PEG) to separate free and bound hormone.” All samples from a serial study were measured in a single assay. The one-way analysis of variance (F test) was used for statistical

PERIOVULATORY o---o LATE FOLLICULAR c.4

evaluation. The normal ranges for basal hormone concentrations in our laboratory are follicular phase: E, = 20-70 pg/mL, P = 0.3-0.8 ng/mL, FSH = S-20 mIU/mL, LH = 5-15 mlU/mL; periovulatory phase: E, = 200-500 pg/mL, P = l-2 ng/mL, FSH = 15-40 mIU/mL, LH = 40-120 mlU/mL; luteal phase: E, = 150-400 pg/mL, P = 6-18 ng/mL, FSH = 5-15 mIU/mL, LH = S-15 mlU/mL; during all phases: PRL = 5-25 ng/mL. 5-

RESULTS

Basal Hormone Levels

The mean (i- SEM) basal concentrations of plasma FSH, LH, PRL, El, and P found in normal women during different phases of the menstrual cycle are reported in Table 1. Mean E, periovulatory values are not statistically different from the late follicular levels since the midcycle E, peak had occurred before the gonadotropin surge. Plasma P concentrations were low in the late follicular phase, slightly elevated (although not significantly) in the periovulatory phase, and significantly elevated in the midluteal phase of the cycle. Basal LH levels presented a highly significant increment during the periovulatory phase. The FSH levels showed a nonsignificant increase during the same period. Plasma PRL levels did not change significantly during the various phases of the cycle. Prolactin and Gonadotropin

MID-LUTEAL

Responses to GnRH

The administration of 100 pg GnRH as an intravenous bolus induced an increment in circulating PRL levels during the periovulatory phase of the cycle (Fig. 1 and Table 2). A peak concentration was reached betweeen 30 and 60 minutes following GnRH administration in nine subjects (Table 2). Similarly, during the periovulatory phase, GnRH infusion induced a progressive increment in PRL levels with a maximum rise above basal levels occurring at three hours (Fig. 2, Table 2). Thus, the pattern of PRL response was

I, , , , , , ,

0 -10

0

20 30

10

45

60

90

120

150

minutes Fig. 1. The effects of an intravenous bolus of GnRH (100 pg at time 0) on plasma PRL levels in normal women during various phases of the menstrual cycle. l * = P -c 0.01: **+ = P < 0.005, statistical significance using one-way analysis of variance (F test).

different depending on the modality of GnRH administration. In five subjects studied during the same period of the cycle a decrement (mean 37%) in PRL levels in response to saline infusion was observed. No significant changes in the PRL concentration were observed after GnRH administration, given either as a bolus or a continuous infusion during the late follicular and midluteal phases of the cycle (Figs. 1 and 2). In subjects with a positive PRL response to GnRH, the basal levels of LH were found elevated (Tables 1 and 2), with levels compatible with the concentrations seen during the midcycle LH peak. Administration of GnRH induced the expected increase in circulating LH and FSH levels. The mean increments of LH and FSH were greater during the periovulatory phase than the late follicular and midluteal phases of the cycle (Table 2). DISCUSSION

The results of the present investigation clearly demonstrated that, in normal women, GnRH administered as a bolus or a continuous infusion stimulated PRL

Table 1. Mean Basal Plasma Estradiol (E,), Progesterone (PI, LH, FSH. and Prolactin (PRL) Levels in Normal Women During the Late Follicular, Periovulatory, and Midluteal Phases of the Menstrual Cycle Mean IrSEM) BasalPlasma Level Phase of

Cycle

Number

of Woman

LH

P

E* (pg/mLI

(ng/mL)

FSH

lmlU/mL)

(mlU/mLl

PRL bg/mL)

Late follicular (Days 9- 12)

112 + 20

0.04

+ 0.2

12.5 + 1.4

11.6 + 1.07

9.04

k 1.03

103 f

2.59

+ 0.93

51.5

15.5 f 3.64

9.61

+ 0.65

7.5 + 0.59

11.91

Periovulatory (Days 14-17) Midluteal (Days 22-24) F test* (P)

13

83 + 12 NS

*Statistical significance using one-way analysis of variance.

13.2 -t 1.5 <0.0001

,

%fuJ

f 6.82

11.4 i

1.0

<0.0001


-c 1.69 NS

PERIOVULATORY PRL RESPONSE TO GnRH

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Table 2. Basal and GnRH-Stimulated (as a bolus or a continuous infusion) Levels of LH. FSH, and PRL in 15 Women During the Periovulatory Phase of the Menstrual Cycle Method of

LHlmlWmLl

GllRH

FSH(mlU/mL)

PRL(ng/mL)

Day of

Admnstration

CYCk

Subject

Bolus (100 /I9 IV)

Basal

Peak

Basal

Peak

Basal

Peak

I

17

68.0

190.0

25.0

61.0

8.0

18.0

2

I6

34.0

150.0

8.0

17.0

9.0

27.0

3

15

42.0

160.0

18.0

80.0

10.0

27.0

4

17

29.0

112.0

5.0

8.5

8.0

17.0

5

16

60.0

300.0

10.0

70.0

8.0

19.0

6

14

24.0

180.0

14.0

38.0

9.0

28.0

7

14

27.0

170.0

8.0

26.0

7.0

12.5

8

17

80.0

150.0

14.0

24.0

12.0

45.0

9

16

70.0

170.0

14.0

48.0

10.0

18.0

47.6 + 7.2

Mean + SEM

175.8

2 17.22

12.2 + 1.74

41.38

9.0 i 0.5

+ 8.3

23.5

t 3.23

Continuous

16

120.0

300.0

55.0

70.0

8.5

15.0

Infusion

16

60.0

125.0

15.0

42.0

16.0

50.0

(0.2 pg/min x 3 h)

14

20.0

100.0

7.0

19.0

7.0

12.0

15

48.0

120.0

17.0

40.0

8.0

40.0

I6

55.0

170.0

12.0

36.0

13.8

15.2

16

36.0

156.0

11.0

28.0

9.9

21.0

19.5 + 7.24

39.16 + 7.0

56.5 + 13.9

Mean + SEM

161.8

+ 28.5

It was originally reported that the administration of synthetic GnRH to men pretreated with estrogen, progesterone, or a combination of estrogen and progesterone, stimulated LH and FSH release without affecting PRL secretion.’ It was concluded that GnRH was one hypothalamic hormone that specifically controlled gonadotropin secretion and that ovarian steroids did not influence the PRL response to GnRH. Recent studies, however, have shown that GnRH was capable of stimulating PRL secretion in some normoprolactinemic anovulatory women’ in whom the PRL response was considered “paradoxical.” More recent-

release during the periovulatory phase of the menstrual cycle, (days 14-17) while it had no effect during the late follicular and midluteal phases of the cycle. It is unlikely that the PRL elevation observed after GnRH stimulation is due to spontaneous PRL fluctuations, since the spontaneous increments reported in normal women during the waking periods were of lesser magnitude than those observed after GnRH administration.‘” Moreover, the control tests performed with saline infusion did not show significant increases in PRL secretion during the periovulatory phase of the menstrual cycle. GnRH

INFUSION

(0.2

IO.53 zk 1.46 25.53 i 6.4

pug/min) i

_

PERIOVULAT~RY

O--o

LATE

A-*-A

MID-LUTEAL

FOLLICULAR

a 0.

OJr -15

I

I

0

30

I

60

I

90

120

.

150

,

180

210

240

minutes The effects of G&H infusion (0.2 pg/min x 3h) on plasma PRL levels in normal women during various phases of the Fig. 2. menstrual cycle. l = P < 0.05; l * = P -c 0.01: l ** = P -c 0.005; statistical significance using one-way analysis of variance (F test).

1062

BARBARINO

ly, a PRL response to the infusion of GnRH has been described in hypogonadal women and normal women in various phases of their cycle.6 Although the ovarian steroid levels and the day of the cycle in which the test was performed were not reported, a correlation with the ovarian steroid environment was excluded on the basis of a similar time course and magnitude of PRL release in the two groups of subjects. In contrast, our present results show that the administration of GnRH has differential effects on PRL release during various phases of the cycle and elicits different patterns of PRL response during the periovulatory period depending on the modality of administration of the decapeptide. During GnRH infusion an increment of PRL levels followed a progressive course with a maximum rise above basal levels at three hours. In contrast, administration of GnRH as a bolus significantly increased plasma PRL levels at 30,45, and 60 minutes with a return to basal levels within 120 minutes. These patterns of response are similar to those previously reported in estrogen-treated intact and castrated men’ and support the hypothesis of an estrogen dependence of the PRL response to GnRH. However, the present data in women clearly show that the PRL response to GnRH is not temporally related to the midcycle estradiol peak, but rather to the gonadotropin peak. These observations suggest that during the normal menstrual cycle progressively

ET AL

increasing Ez levels exert a sensitization of the pituitary lactotropes to GnRH, which is manifest at midcycle. However, since a PRL response to GnRH can be elicited during the gonadotropin peak, it is conceivable that the estrogen-induced sensitization of the pituitary lactotropes is in some way mediated or potentiated by the gonadotropins. Our results, however, fail to explain why the potential augmentation of PRL release in response to GnRH is not manifest in physiologic states in women. It is possible that, in humans, the gonadotropin surge induced by estradiol during the periovulatory phase of the cycle is not preceded by an endogenous GnRH13.14 peak, or, alternatively, the estradiolinduced sensitization of the pituitary to GnRH is greater for the gonadotropic than the lactotropic cells, and the endogenous GnRH peak is sufficient for the release of FSH and LH, but not for PRL release. In conclusion, our present data confirm that GnRH is a substance capable of stimulating plasma PRL response in humans and further suggest’ that this response is estrogen dependent. Finally, a temporally related correlation between the midcycle gonadotropin peak and a positive PRL response to exogenous GnRH has been established. ACKNOWLEDGMENT The expert technical assistance of Dr. R. Finotti and Dr. M. Castellano is deeply appreciated. We wish to thank the students of our University who volunteered for this study.

REFERENCES 1. Kastin AJ, Gonzalez-Barcena D, Friesen H, et al: Unaltered plasma prolactin levels in men after administration of synthetic LH-releasing hormone. J Clin Endocrinol Metab 36:375-377, 1973 2. Rebar RW, Yen SSC, VandenBerg G, et al: Gonadotropin responses to synthetic LRF: dose-response relationship in men. J Clin Endocrinol Metab 36:1&14, 1973 3. Yen SSC, VandenBerg G, Rebar R, et al: Variation of pituitary responsiveness to synthetic LRF during different phases of the menstrual cycle. J Clin Endocrinol Metab 35:931-934, 1972 4. Catania A, Cantalamessa L, Reschini E: Plasma prolactin response to luteinizing hormone-releasing hormone in acromegalic patients. J Clin Endocrinol Metab 43:689-691, 1976 5. Giampietro 0, Moggi G, Chisci R, et al: Unusual prolactin response to luteinizing hormone-releasing hormone in some anovulatory women. J Clin Endocrinol Metab 49:141-143, 1979 6. Yen SSC, Hoff JD, Lasley BL, et al: Induction of prolactin release by LRF and LRF-agonist. Life Sci 26:1963-1967, 1980 7. Barbarino A, De Marinis L, Mancini A, et al: Estrogendependent plasma prolactin response to gonadotropin-releasing hormone in intact and castrated men. J Clin Endocrinol Metab 55:1212-1216,1982

8. Barbarino A, De Marinis L, Menini E, et al: Prolactinsecreting pituitary adenomas: prolactin dynamics before and after transsphenoidal surgery. Acta Endocrinol91:397-409, 1979 9. Barbarino A, De Marinis L: Estrogen induction of luteinizing hormone release in castrated adult human males. J Clin Endocrinol Metab 5 I :280-286, 1980 10. Barbarino A, De Marinis L. Lafuenti G, et al: Presence of positive feedback between estrogen and LH in patients with Klinefelter’s syndrome, and Sertoli-cell-only syndrome. Clin Endocrinol 10~235-242, 1979 11. De Villa 0, Roberts K, Wiest WG, et al: A specific radioimmunoassay of plasma progesterone. J Clin Endocrinol 35:458-463, 1972 12. Ehara Y, Siler T, VandenBerg G, et al: Circulating prolactin levels during the menstrual cycle: episodic release and diurnal variation. Am J Obstet Gynecol 1 I7:962-970, 1973 13. Neil1 JD, Freeman ME, Tillson SA: Control of the proestrus surge of prolactin and luteinizing hormone secretion by estrogen in the rat. Endocrinology 89:1448-1453, 1971 14. Knobil E: The neuroendocrine control of the menstrual cycle. Recent Prog Horm Res 36:53-87, 1980