0021-972X/91/7201-0151$02.00/0 Journal of Clinical Endocrinology and Metabolism Copyright© 1991 by The Endocrine Society
Vol. 72, No. 1 Printed in U.S.A.
Acceleration of Luteinizing Hormone Pulse Frequency in Functional Hypothalamic Amenorrhea by Dopaminergic Blockade* S. L. BERGAt, A. B. LOUCKS, W. G. ROSSMANITH, L. M. KETTEL, G. A. LAUGHLIN, AND S. S. C. YEN Department of Reproductive Medicine (T-002) and the General Clinical Research Center, University of California-San Diego School of Medicine, La Jolla, California 92093
ABSTRACT. A constellation of neuroendocrine secretory aberrations, including reduced LH pulse frequency and PRL concentrations, has been documented in women with functional hypothalamic amenorrhea (FHA). As pituitary function was preserved, these aberrations were attributed to an alteration in hypothalamic neuromodulation. To investigate the participation of the dopaminergic system in the genesis of the reduced LH pulse frequency and suppressed PRL levels in FHA, we studied six women with FHA and six cyclic women in the early follicular phase by obtaining blood samples at 15-min intervals for 48 h during sequential 24-h infusions of saline and a dopamine receptor blocker, metoclopramide (MCP). A hypothalamic vs. pituitary site of action was inferred from the pulsatility characteris-
tics. MCP consistently elicited an increase in the LH pulse frequency in the women with FHA [7.3 ± 1.2 (±SE) to 10.5 ± 1.3 pulses/24 h; P< 0.005]. In contrast, the eumenorrheic women did not show a significant change in LH pulse frequency in response to MCP (15.2 ± 1.0 to 14.3 ± 0.9 pulses/24 h). While the PRL concentrations were significantly lower in the FHA group during the infusion of saline (P < 0.001) and MCP (P < 0.005), the relative increases in PRL during MCP were similar in both groups. The acceleration of LH pulse frequency by blockade of dopamine receptors implies that there is increased hypothalamic dopaminergic inhibition of GnRH pulse frequency in women with FHA. (J Clin Endocrinol Metab 72: 151-156, 1991)
I
T IS NOW well established that decreased LH pulse frequency and, by inference, slowing of the hypothalamic GnRH pulse generator represent the underlying cause of ovarian acyclicity in women with functional hypothalamic amenorrhea (FHA) (1-4). Attempts to discern the mechanisms that account for the slowing of GnRH pulse frequency have revealed an opioidergic inhibition at the hypothalamic level in some, but not all, women with FHA (5-7). A dopaminergic mechanism also has been implicated, since a prompt rise in LH levels occurred in response to a bolus injection of the dopamine receptor blocker metoclopramide (MCP) (7). In that study, a causal relationship between dopaminergic inhibition and slowing of the GnRH pulse frequency could
not be ascertained because LH pulse characteristics were not determined. The present study was undertaken to further elucidate the role of dopaminergic neuromodulation of GnRH and FHA. The response of the 24-h LH pulse pattern to MCP (30 jiig/kg-h) was determined so that a hypothalamic, as distinct from a pituitary, site of action could be discerned. The PRL release elicited by MCP also was examined to determine whether the profound suppression of PRL concentrations observed in women with FHA (7) was mediated by a dopaminergic mechanism. Accordingly, the secretory patterns of LH and PRL during the 24-h rest-activity cycle were determined in six eumenorrheic women in their early follicular phase and in six women with FHA by obtaining blood samples at 15-min intervals for 48 h during sequential 24-h infusions of saline and MCP.
Received May 22,1990. * This work was supported by NIH Grants HD-21198, HD-1230312, and GCRC RR-00827, and the American Fertility Society-Ortho Distinguished Fellowship in Reproduction, and was conducted by the Clayton Foundation for Research. Presented in part at the 36th Annual Meeting of the Society for Gynecological Investigation, San Diego, CA, 1989. t Recipient of the Bank of America-Giannini Foundation Fellowship, the American Fertility Society-Ortho Distinguished Fellowship in Reproduction, and the Clinical Associate Physician Award from the NIH. To whom all correspondence should be addressed. Current address: University of Pittsburgh, Magee-Womens Hospital, Department of Obstetrics and Gynecology, Pittsburgh, Pennsylvania 15213.
Materials and Methods Participants Six women with FHA and six eumenorrheic women were studied. Before enrollment, organic causes of amenorrhea, including hyperprolactinemia, thyroid and adrenal disease, premature ovarian failure, and hyperandrogenism, were excluded. 151
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BERGA ET AL.
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Women who met the Diagnostic and Statistical Manual of Mental Disorders criteria for an eating disorder (8) or the Beck Depression Inventory criteria for depression (9) were excluded, as were those engaged in excessive exercise (10). This study was approved by the Human Subjects Committee of the University of California-San Diego. Before participation, written consent was obtained from each woman. The clinical characteristics of the six women with FHA are shown in Table 1. The eumenorrheic women had documented menstrual cycle lengths of 27-31 days and were studied in the early follicular phase (days 2-5) of their cycle. The percent ideal body weight, based on the 1983 Metropolitan Life height and weight tables for women, was comparable in both groups. While all participants were within the reproductive age range of 18-35 yr, the women with FHA were younger than the control group. Study protocol Each woman was admitted to the Clinical Research Center between 0800-1000 h. Starting 30 min after the placement of an indwelling iv catheter in the forearm, blood samples were collected at 15-min intervals for 48 h during sequential 24-h infusions of saline and MCP (30 ^g/kg/h). The volume of each infusion was 240 mL. There were no observed side-effects during the MCP infusion. Serum samples were stored at —20 C until assay. Sleeping was permitted only from 2300-0700 h and was monitored by observation through a one-way mirror. During sleep, blood samples were collected by extending the iv line outside the room through a port in the wall. Standard mixed meals free of caffeine were provided at 0800-0830,12001230, and 1700-1730 h. Hormone measurements Serum LH, FSH, PRL, and estradiol (E2) were determined by previously described RIA methods (11-14). FSH and E2 were determined in the initial sample only. LH and PRL were determined from each sample, and all samples from a given individual were run in the same assay.
JCE & M • 1991 Vol 72 • No 1
Analyses The number of pulses per 24 h and pulse amplitude of LH were determined by the computerized algorithm Cluster analysis (15). A cluster width of 2 for nadirs and 1 for peaks was used. A t statistic of 2.1 for both up-stroke and down-stroke was applied to limit the false positive rate to 5%. Mean values were analyzed, and the SD for a given mean was estimated by an independently derived quadratic equation developed from serial RIA determinations of LH in serum at 10 dose levels (16). The pulse number per 24 h, pulse amplitude, and transverse mean of LH and PRL before and during MCP in each group were analyzed by mixed two-factor analysis of variance (ANOVA). To evaluate the peak response of PRL to MCP, the PRL concentration just before the initiation of the MCP infusion and the maximal PRL concentration during the infusion were compared in the FHA and eumenorrheic women by a mixed two-factor ANOVA. Post-hoc comparisons were performed by group t tests for between-group comparisons and paired t tests for within-group comparisons. P < 0.05 was considered significant. Results Mean (±SE) FSH and E2 concentrations determined in the initial blood sample did not differ significantly in the FHA and control groups, but the mean LH and PRL levels were lower in the FHA group (Table 1). Table 2 shows the mean pulse number per 24 h, pulse amplitude, and 24-h transverse mean of LH and the 24-h transverse mean of PRL during the saline and MCP infusions in each group. During the saline and MCP infusions, only the LH pulse frequency (P < 0.005) and the 24-h transverse mean of PRL (P < 0.001) were significantly lower in the FHA women compared to those in the cyclic women. In both groups, a prompt increase in PRL was evident in all women after the initiation of the MCP infusion (P
TABLE 1. Clinical characteristics and baseline hormone concentrations of six women with FHA and six cyclic women in the early follicular phase Age (yr) FHA women 19 20 20 23 19 20 Mean 20.2 SE0.6 Cyclic women Mean 30.3 SEl.2 P value
Vol. 72, No. 1 Printed in U.S.A.
Acceleration of Luteinizing Hormone Pulse Frequency in Functional Hypothalamic Amenorrhea by Dopaminergic Blockade* S. L. BERGAt, A. B. LOUCKS, W. G. ROSSMANITH, L. M. KETTEL, G. A. LAUGHLIN, AND S. S. C. YEN Department of Reproductive Medicine (T-002) and the General Clinical Research Center, University of California-San Diego School of Medicine, La Jolla, California 92093
ABSTRACT. A constellation of neuroendocrine secretory aberrations, including reduced LH pulse frequency and PRL concentrations, has been documented in women with functional hypothalamic amenorrhea (FHA). As pituitary function was preserved, these aberrations were attributed to an alteration in hypothalamic neuromodulation. To investigate the participation of the dopaminergic system in the genesis of the reduced LH pulse frequency and suppressed PRL levels in FHA, we studied six women with FHA and six cyclic women in the early follicular phase by obtaining blood samples at 15-min intervals for 48 h during sequential 24-h infusions of saline and a dopamine receptor blocker, metoclopramide (MCP). A hypothalamic vs. pituitary site of action was inferred from the pulsatility characteris-
tics. MCP consistently elicited an increase in the LH pulse frequency in the women with FHA [7.3 ± 1.2 (±SE) to 10.5 ± 1.3 pulses/24 h; P< 0.005]. In contrast, the eumenorrheic women did not show a significant change in LH pulse frequency in response to MCP (15.2 ± 1.0 to 14.3 ± 0.9 pulses/24 h). While the PRL concentrations were significantly lower in the FHA group during the infusion of saline (P < 0.001) and MCP (P < 0.005), the relative increases in PRL during MCP were similar in both groups. The acceleration of LH pulse frequency by blockade of dopamine receptors implies that there is increased hypothalamic dopaminergic inhibition of GnRH pulse frequency in women with FHA. (J Clin Endocrinol Metab 72: 151-156, 1991)
I
T IS NOW well established that decreased LH pulse frequency and, by inference, slowing of the hypothalamic GnRH pulse generator represent the underlying cause of ovarian acyclicity in women with functional hypothalamic amenorrhea (FHA) (1-4). Attempts to discern the mechanisms that account for the slowing of GnRH pulse frequency have revealed an opioidergic inhibition at the hypothalamic level in some, but not all, women with FHA (5-7). A dopaminergic mechanism also has been implicated, since a prompt rise in LH levels occurred in response to a bolus injection of the dopamine receptor blocker metoclopramide (MCP) (7). In that study, a causal relationship between dopaminergic inhibition and slowing of the GnRH pulse frequency could
not be ascertained because LH pulse characteristics were not determined. The present study was undertaken to further elucidate the role of dopaminergic neuromodulation of GnRH and FHA. The response of the 24-h LH pulse pattern to MCP (30 jiig/kg-h) was determined so that a hypothalamic, as distinct from a pituitary, site of action could be discerned. The PRL release elicited by MCP also was examined to determine whether the profound suppression of PRL concentrations observed in women with FHA (7) was mediated by a dopaminergic mechanism. Accordingly, the secretory patterns of LH and PRL during the 24-h rest-activity cycle were determined in six eumenorrheic women in their early follicular phase and in six women with FHA by obtaining blood samples at 15-min intervals for 48 h during sequential 24-h infusions of saline and MCP.
Received May 22,1990. * This work was supported by NIH Grants HD-21198, HD-1230312, and GCRC RR-00827, and the American Fertility Society-Ortho Distinguished Fellowship in Reproduction, and was conducted by the Clayton Foundation for Research. Presented in part at the 36th Annual Meeting of the Society for Gynecological Investigation, San Diego, CA, 1989. t Recipient of the Bank of America-Giannini Foundation Fellowship, the American Fertility Society-Ortho Distinguished Fellowship in Reproduction, and the Clinical Associate Physician Award from the NIH. To whom all correspondence should be addressed. Current address: University of Pittsburgh, Magee-Womens Hospital, Department of Obstetrics and Gynecology, Pittsburgh, Pennsylvania 15213.
Materials and Methods Participants Six women with FHA and six eumenorrheic women were studied. Before enrollment, organic causes of amenorrhea, including hyperprolactinemia, thyroid and adrenal disease, premature ovarian failure, and hyperandrogenism, were excluded. 151
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BERGA ET AL.
152
Women who met the Diagnostic and Statistical Manual of Mental Disorders criteria for an eating disorder (8) or the Beck Depression Inventory criteria for depression (9) were excluded, as were those engaged in excessive exercise (10). This study was approved by the Human Subjects Committee of the University of California-San Diego. Before participation, written consent was obtained from each woman. The clinical characteristics of the six women with FHA are shown in Table 1. The eumenorrheic women had documented menstrual cycle lengths of 27-31 days and were studied in the early follicular phase (days 2-5) of their cycle. The percent ideal body weight, based on the 1983 Metropolitan Life height and weight tables for women, was comparable in both groups. While all participants were within the reproductive age range of 18-35 yr, the women with FHA were younger than the control group. Study protocol Each woman was admitted to the Clinical Research Center between 0800-1000 h. Starting 30 min after the placement of an indwelling iv catheter in the forearm, blood samples were collected at 15-min intervals for 48 h during sequential 24-h infusions of saline and MCP (30 ^g/kg/h). The volume of each infusion was 240 mL. There were no observed side-effects during the MCP infusion. Serum samples were stored at —20 C until assay. Sleeping was permitted only from 2300-0700 h and was monitored by observation through a one-way mirror. During sleep, blood samples were collected by extending the iv line outside the room through a port in the wall. Standard mixed meals free of caffeine were provided at 0800-0830,12001230, and 1700-1730 h. Hormone measurements Serum LH, FSH, PRL, and estradiol (E2) were determined by previously described RIA methods (11-14). FSH and E2 were determined in the initial sample only. LH and PRL were determined from each sample, and all samples from a given individual were run in the same assay.
JCE & M • 1991 Vol 72 • No 1
Analyses The number of pulses per 24 h and pulse amplitude of LH were determined by the computerized algorithm Cluster analysis (15). A cluster width of 2 for nadirs and 1 for peaks was used. A t statistic of 2.1 for both up-stroke and down-stroke was applied to limit the false positive rate to 5%. Mean values were analyzed, and the SD for a given mean was estimated by an independently derived quadratic equation developed from serial RIA determinations of LH in serum at 10 dose levels (16). The pulse number per 24 h, pulse amplitude, and transverse mean of LH and PRL before and during MCP in each group were analyzed by mixed two-factor analysis of variance (ANOVA). To evaluate the peak response of PRL to MCP, the PRL concentration just before the initiation of the MCP infusion and the maximal PRL concentration during the infusion were compared in the FHA and eumenorrheic women by a mixed two-factor ANOVA. Post-hoc comparisons were performed by group t tests for between-group comparisons and paired t tests for within-group comparisons. P < 0.05 was considered significant. Results Mean (±SE) FSH and E2 concentrations determined in the initial blood sample did not differ significantly in the FHA and control groups, but the mean LH and PRL levels were lower in the FHA group (Table 1). Table 2 shows the mean pulse number per 24 h, pulse amplitude, and 24-h transverse mean of LH and the 24-h transverse mean of PRL during the saline and MCP infusions in each group. During the saline and MCP infusions, only the LH pulse frequency (P < 0.005) and the 24-h transverse mean of PRL (P < 0.001) were significantly lower in the FHA women compared to those in the cyclic women. In both groups, a prompt increase in PRL was evident in all women after the initiation of the MCP infusion (P
TABLE 1. Clinical characteristics and baseline hormone concentrations of six women with FHA and six cyclic women in the early follicular phase Age (yr) FHA women 19 20 20 23 19 20 Mean 20.2 SE0.6 Cyclic women Mean 30.3 SEl.2 P value
