Bioactive LH Levels Research Articles

Changes in the bioactivity to immunoreactivity ratio of circulating luteinizing hormone in impotent men treated with testosterone undecanoate

Testosterone undecanoate was administered orally (80 mg twice daily) for 30 days to 10 impotent men with mild Leydig cell failure, age 28 to 42 years. Placebo was administered for 30 days both before and at the end of testosterone undecanoate therapy. Serum levels of bioactive LH, immunoreactive LH and testosterone were determined in basal conditions (day zero), 30 days after the first placebo administration, at the 15th and 30th day of testosterone undecanoate therapy, and at the end of the second treatment with placebo (90th day). Bioactive LH was measured by a sensitive and specific in vitro bioassay based on testosterone production by mechanically dispersed mouse Leydig cell preparations. Immunoreactive LH and testosterone were determined by a double-antibody RIA technique. The results were compared with those obtained in 30 untreated normal young men. In the basal state, serum concentrations of immunoreactive LH were significantly higher in the patients (P less than 0.02) than in control subjects, whereas testosterone levels were significantly lower (P less than 0.001) in the impotent men. In contrast, bioactive LH levels and the bioactive LH to immunoreactive LH ratios were similar in the two groups. In the patients, at the 15th day of treatment with testosterone undecanoate, serum levels of testosterone and bioactive LH were significantly higher (P less than 0.01) than basal values, whereas immunoreactive LH concentrations showed no significant changes. Consequently, the bioactive LH to immunoreactive LH ratios rose significantly (P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Gonadal and pituitary responses to progesterone antagonist ZK 98.299 during the follicular phase of the menstrual cycle in bonnet monkeys

ZK 98.299 is a potent progesterone antagonist. Its effects on folliculogenesis, bioactive LH, ovulation and menstrual cycle (m.c.) length were studied in adult bonnet monkeys. ZK 98.299 (20 mg/day) was administered s.c., once daily on days 5 to 15 of m.c., to ten animals. The pretreatment m.c. was of 26.5 days (25 to 28 days, mean with 95% confidence limits) and on treatment it was significantly (p< 0.001) prolonged to 46.9 days (39 to 54 days). The anticipated midcycle rise in estradiol and bioactive LH levels was completely blocked in six and in three animals during the treatment period. However the levels did not drop below the early follicular phase levels. In one animal (#90), though the cycle length was prolonged by 5 days the midcycle rise in estradiol and bioactive LH levels was observed during the treatment period and this animal had normal luteal function. Seven animals had delayed ovulation whereas, two had anovulatory treatment cycles. The rise in estradiol and bioactive LH levels, prior to ovulation in the treatment cycles, was compatible with the midcycle rise observed in the pretreatment cycles. Serum progesterone levels during the luteal phase of the treatment cycles were normal in six animals whereas, in two they were indicative of luteal insufficiency. In two animals, the treatment cycles were anovulatory. ZK 98.299 had no effect on the duration of menses. The post-treatment cycles were of normal duration. This study suggests that the administration of ZK 98.299 during the follicular phase blocks estradiol and bioactive LH release and terminates the follicular phase in most of the animals. The follicular phase is reinitiated after the treatment is stopped.

QUANTITATIVE AND QUALITATIVE CHANGES IN LH SECRETION FOLLOWING PULSATILE GnRH THERAPY IN A MAN WITH IDIOPATHIC HYPOGONADOTROPHIC HYPOGONADISM

The pattern of bioactive and immunoreactive LH secretion before and during pulsatile GnRH therapy (18 micrograms/90 min) in a hypogonadotrophic hypogonadal male has been studied. Before treatment the patient was azoospermic and had low testosterone (1.2 nmol/l) with low and apulsatile immunoreactive LH (1.9 +/- 0.2 IU/l) and FSH (1.4 +/- 1.9 IU/l) levels. There was no detectable LH bioactivity. During the first 24 h of GnRH therapy there was a small increase in immunoreactive (5.4 +/- 0.8 IU/l) and bioactive (6.7 +/- 1.3 IU/l) LH, with an irregular pattern and little effect on testosterone production (2.2 nmol/l). Within 1 week of treatment both bioactive (30.5 +/- 6.8 IU/l) and immunoreactive (13.6 +/- 1.5 IU/l) LH levels were above the normal range and the pattern of secretion was pulsatile. The bioactive to immunoreactive (B:I) LH ratios within the pulses (2.6 +/- 0.3) were higher (P less than 0.01) than between pulses (1.97 +/- 0.1) and the testosterone concentration (17.8 +/- 2.1 nmol/l) was now normal. At one month LH secretion was similar and testosterone pulses of high amplitude were evident corresponding to high-amplitude bioactive LH pulses. By 3 months mature spermatozoa (1.3 x 10(6)/ml) were seen in the patient's semen. The pattern of LH secretion was pulsatile but the levels of bioactive (13.1 +/- 3.6 IU/l) and immunoreactive (9.5 +/- 1.3 IU/l) LH decreased towards the normal range reflecting maturation of the testicular feedback control at the pituitary level. This effect was more pronounced on bioactive rather than immunoreactive LH secretion (57% vs 32% relative decrease). At 6 months LH levels were similar and the sperm count was normal (34 x 10(6)/ml).

A new look to the andropause: Altered function of the gonadotrophs

After being controversial for a long time, the age associated decline in plasma T levels and secretion in normal men is now generally accepted. Although most data point towards a primary testicular origin for the decline in Leydig cell function, other data point towards alterations at the hypothalamo-pituitary levels: —T levels in elderly men are decreased notwithstanding an important secretory reserve of both LC and gonadotrophs. —Nycthemeral variations in plasma T levels are significantly decreased in elderly men. —The increase in immunoreactive LH levels is dysproportionate to the increase of bioactive LH levels. Study of the activity of the LHRH pulse generator in 27 young and 21 elderly men revealed a significant decrease in frequency of high amplitude pulses for both LH and T. Moreover the sensitivity of the gonadostat to sex hormone feed back was significantly greater in the elderly men. Acute administration of an antiopoid (Naloxone), 2 mg i.v. both in young and elderly men at rest showed that in young and elderly men, Naloxone induced a comparable increase of LH levels. Naltrexone on the other hand increased LH pulse frequency in elderly but not in young men. The data show that opioids maintain their inhibitory effect on LHRH secretion in elderly men and suggest either that the opioid tone in the elderly men is decreased or that the response of the gonadotrophs to LHRH is decreased.

Bioactive LH in women with polycystic ovaries and the effect of gonadotrophin suppression.

Discrepancies between levels of bioactive LH (B-LH) and immunoreactive LH (I-LH) in polycystic ovarian syndrome (PCO) have been reported previously. Serum levels of I-LH, B-LH (by dispersed Leydig cell assay), FSH, oestradiol (E2) and progesterone (Prog) were measured once to three times weekly over 4 weeks in 13 women with classical clinical, ultrasound and endocrine features of PCO. Eleven women attending for infertility but whose profiles when studied three times weekly by combined endocrine and ultrasound assessment were normal and ovulatory served as controls. Seven of the women with PCO were evaluated during and after 3 weeks suppression with ethinyloestradiol (30 micrograms) plus 150 micrograms either of desogestrel or levonorgestrel; two were given both treatments. Both I-LH and B-LH levels were higher in PCO patients (20 +/- SD 5 U/l and 46 +/- 9 U/l respectively, P less than 0.0001), compared with all phases of the normal cycles except the mid-cycle peak. The B-LH to I-LH (B:I LH) ratio in PCO patients (2.5 +/- 0.7) was higher than in all the control cycle phases (P less than 0.05). I-LH, B-LH, B:I LH ratio, FSH and E2 were all suppressed from the second week of oestrogen-progestogen treatment (P less than 0.01) and returned gradually to pretreatment levels by the third or fourth week after suppression. The LH and FSH levels and B:I LH ratio in PCO patients during suppression were comparable with levels in the early and mid-follicular phases of control cycles but the LH/FSH ratio remained significantly raised (P less than 0.01) at 2.3 +/- 0.7.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in the qualitative and quantitative secretion of luteinizing hormone (LH) following orchidectomy in man.

The effect of orchidectomy and thus withdrawal of testicular hormones on the biological and immunological properties of plasma LH was studied. Plasma samples were obtained from five men (mean age 71, range 65-81 years) with advanced carcinoma of the prostate, before orchidectomy and 1, 4, 8 and 16 weeks after surgery. LH bioactivity was estimated by a mouse Leydig cell bioassay and immunoreactivity by radioimmunoassay, using the same human pituitary LH standard 68/40. FSH and testosterone were measured by radioimmunoassay. Similar baseline data were obtained from a group (n = 17) of normal adult men (26, 19-36 years). Baseline bioactive (40 IU/l, median) and immunoreactive (10.8 IU/l) LH levels in the patients were higher (P less than 0.01) than in the controls (15.1 and 5.7 IU/l respectively), but bioactive to immunoreactive (B:I) LH ratios (3.4 +/- 0.2 versus 2.8 +/- 0.7) and testosterone levels (15.3 vs 18.7 nmol/l) were no different, consistent with compensated Leydig cell failure in the elderly men. After orchidectomy there was a greater increase in immunoreactive (46.6 IU/l at 16 weeks) than bioactive (80.3 IU/l) LH levels i.e. a fourfold vs twofold increase from baseline values. Consequently the B:I LH ratio decreased significantly (1.8 +/- 0.4 at 16 weeks) from the baseline ratio (P less than 0.0001) and that of the controls (P less than 0.01). These data indicate that acute withdrawal of testicular sex steroids results not only in quantitative change in LH secretion but also in qualitative change that decreases the biopotency of the LH molecules.

Luteal function following ovarian stimulation in rhesus monkeys for in vitro fertilization: atypical response to human chorionic gonadotropin treatment simulating early pregnancy

This study determined if corpora lutea of hyperstimulated cycles in rhesus monkeys could be "rescued" by the pregnancy signal, chorionic gonadotropin (CG), given at the typical time of implantation. At menses, female monkeys received human follicle-stimulating hormone (hFSH, 60 IU, days 1 to 6) followed by human menopausal gonadotropin (hMG, 60 IU hFSH/60 IU luteinizing hormone [hLH], days 7 to 9). On day 10, human chorionic gonadotropin (hCG) was given to mimic the LH surge. Nine days later, a regimen of daily increasing doses of hCG (15 to 360 IU twice a day) was initiated to simulate rescue of the corpus luteum in early pregnancy. Serum levels of progesterone (P) increased through day 5 of the luteal phase but then declined. Circulating levels of bioactive LH were significantly less on days 7 to 9 of the luteal phase than at this stage in the natural cycle. The hCG regimen extended (P less than 0.05) the luteal phase in five of six animals. The hCG treatment elicited a persistent increase (P less than 0.05) in circulating P levels, rather than a transient rise typical of normal or simulated pregnancy in natural cycles. The authors conclude that (1) corpora lutea of hyperstimulated cycles can respond to CG, but (2) there are differences in luteal function during both the luteal phase and simulated early pregnancy that may be due to inadequate luteal development or the abnormal gonadotropin milieu existing after ovulation or both.

Maintenance of the ratio of bioactive to immunoreactive follicle-stimulating hormone in normal men during chronic luteinizing hormone-releasing hormone agonist administration.

Chronic administration of LHRH agonist analogs to humans reduces gonadal function through pituitary desensitization. Serum immunoreactive gonadotropin levels are modestly reduced, whereas serum bioactive LH levels are drastically suppressed. The effects on bioactive FSH levels, however, are not known. In this study, serum bioactive FSH was measured using an in vitro granulosa cell aromatase bioassay in four normal men given a LHRH agonist, [D-Trp6,Pro9-NEt]LHRH (LHRHA; 500 micrograms/day for 16 weeks), by sc infusion and testosterone enanthate (TE; 100 mg, im every 2 weeks) and in five men given 500 micrograms/day LHRHA by daily sc injection for 20 weeks and TE (100 mg every 2 weeks) from weeks 10 through 20. During the first study, serum immunoreactive FSH levels (IR-FSH) decreased by 56.5 +/- 4.8% (+/- SEM), and serum bioactive FSH (Bio-FSH) level decreased by 57.6 +/- 6.4%. The ratio of Bio-FSH to IR-FSH did not change. During the second study, both serum IR-FSH and Bio-FSH levels followed a triphasic pattern, decreasing slightly but not significantly immediately after initiation of LHRHA administration, progressively increasing to a peak (P less than 0.5 vs, baseline) at week 10, and then, after addition of TE to this regimen, decreasing slightly again. The Bio-FSH to IR-FSH ratio, as in the first study, did not change. When serum obtained at week 10 during the second study, just before initiation of TE, was chromatographed on a Sephadex G-100 column, IR-LH eluted in two distinct peaks, while IR-FSH eluted as a single peak. These results demonstrate that in normal men chronic LHRHA administration alone for up to 10 weeks or LHRHA plus TE for up to 16 weeks does not alter the qualitative characteristics of secreted FSH, since there was no dissociation between serum IR- and Bio-FSH levels.

Gonadotropin and prolactin secretion increases during sleep during the puerperium in nonlactating women.

To investigate the neuroendocrine changes that regulate initiation of normal menstrual function after parturition, serum LH, FSH, and PRL concentrations were determined at 20-min intervals for 12-24 h in eight nonlactating postpartum women on a weekly basis between postpartum days 10-26. Sleep was monitored by EEG. On postpartum day 10, serum LH concentrations were similar to early follicular phase levels in normal cycling women, while FSH concentrations were lower than early follicular phase levels. Mean LH pulse frequency during each postpartum interval was 3.6 +/- 0.6 (+/- SE), 4.4 +/- 0.6, and 4.1 +/- 0.8 pulses/12 h on postpartum days 10-11, 17-21, and 24-26, respectively. Because mean serum LH levels and LH pulse frequency did not change significantly between postpartum days 10 and 26, the results from the two or three studies in each woman were combined for the purpose of comparing LH pulse characteristics during the waking and sleeping periods. During the waking hours, mean LH pulse frequency (6.1 +/- 0.5 pulses/12 h) was significantly greater than during sleep (4.1 +/- 0.4 pulses/12 h; P less than 0.02). The amplitude of the serum immunoreactive LH pulses (P less than 0.05) and bioactive LH levels (P less than 0.05) were significantly higher during sleep than during the waking period, with five of the eight women having higher sleep-associated immunoreactive LH and bioactive LH levels between postpartum days 17-26. These changes were associated with an increase in the bioactive to immunoactive LH ratio from 3.3 +/- 0.4 (awake) to 4.5 +/- 0.5 (sleep; P less than 0.05). Although serum PRL levels remained elevated during the puerperium, the diurnal pattern of PRL secretion was conserved. With each successive week postpartum, serum PRL concentrations declined. These results suggest that the increment in LH secretion (and, by inference, increased GnRH secretion) during sleep is a feature of postpartum pituitary-ovarian reactivation. Although the mechanism(s) responsible for the increase in GnRH secretion is not known, this hormonal pattern is analogous to that during early puberty and during recovery from anorexia nervosa and hypothalamic amenorrhea. Taken together, these findings provide evidence to support the concept of a centralized preprogrammed scheme for pituitary-gonadal reactivation.

The effects of aging in normal men on bioavailable testosterone and luteinizing hormone secretion: response to clomiphene citrate.

Serum testosterone (T) levels in men decline with age while serum LH levels, as measured by RIA, increase. To assess if the decline in serum T levels in healthy aging men is paralleled by an age-related decline in the bioavailable non-sex hormone-binding globulin (SHBG)-bound fraction of T and to determine whether there are age-related changes in LH secretion or LH control of T production, we studied 29 young (aged 22-35 yr) and 26 elderly (aged 65-84 yr) healthy men. All men had single random blood samples drawn, and 14 men in each age group underwent frequent blood sampling for 24 h, both before and after 7 days of clomiphene citrate (CC) administration. Both mean 24-h serum total T levels and non-SHBG-bound T were reduced in elderly men compared to those in young men (P less than 0.05), while estradiol and SHBG levels were similar in the 2 age groups. Serum FSH determined by RIA and LH by RIA and bioassay were higher in the elderly men compared to those in young men (P less than 0.05), but the ratios of LH bioactivity to immunoreactivity and the LH pulse frequency and amplitude were similar. After CC administration, mean serum total T and non-SHBG-bound levels in young men increased by 100% and 304%, respectively, while in older men these values increased by only 32% and 8%, respectively. However, CC-stimulated LH pulse characteristics and serum levels of estradiol, SHBG, FSH, and bioactive and immunoreactive LH were similar in the 2 groups. Thus, both at baseline and after CC stimulation, elderly men had significantly lower serum total T and non-SHBG-bound (bioavailable) T levels than did young men, despite similar or increased levels of bioactive LH and similar bioactive to immunoreactive LH ratios and LH pulse characteristics. These results suggest that major age-related changes in the hypothalamic-pituitary-testicular axis occur at the level of the testes and are manifested by decreased responsiveness to bioactive LH. Administration of CC to young and elderly men resulted in similar changes in LH pulse characteristics and LH bioactivity and immunoreactivity, suggesting preserved hypothalamic-pituitary responsiveness in the elderly.

The pituitary-Leydig cell axis in men with severe damage to the germinal epithelium.

Eighteen men (mean age 27, range 18-30 years) treated for Hodgkin's disease with 6-8 courses of MVPP (Mustine, Vinblastine, Procarbazine and Prednisolone) have had Leydig cell function assessed by their steroidogenic responses to stimulation by a single bolus dose of HCG (1000 units intramuscularly). Normal age-matched men (n = 16) acted as controls. Baseline immunoreactive FSH was markedly raised in the patients (mean 18.1 +/- SD 6.9 vs 2.0 +/- 1.5 IU/l, P less than 0.0001) reflecting damage to the germinal epithelium. Immunoreactive LH was also greater in patients (10.3 +/- 3.9 IU/l) than in controls (3.9 +/- 1.9 IU/l, P less than 0.0001). There were no differences between the baseline testosterone, androstenedione, oestradiol, oestrone and sex hormone binding globulin (SHBG) concentrations. The testosterone/SHBG ratios were similar in the two groups and there was no correlation between baseline LH and testosterone concentrations or testosterone/SHBG ratios. Testosterone, androstenedione, oestradiol and oestrone secretion in response to HCG stimulation were similar at 24 h and 96 h in both groups. In order to explain the paradox of elevated immunoreactive LH in the face of normal testicular steroidogenesis in such patients, LH biological activity (B) as well as LH immunoreactivity (I) and FSH and testosterone were estimated in a second similar group of patients (n = 17, mean age 27, range 17-43 years) and in a further age-matched control group (n = 17). Bioactive and immunoreactive LH levels were significantly increased (P less than 0.005 and P less than 0.001, respectively) in the patient group.(ABSTRACT TRUNCATED AT 250 WORDS)

Reversibility of long-term effects of GnRH agonist administration on testicular histology and sperm production in the nonhuman primate.

The present investigation evaluates the long-term effects of GnRH agonist treatment on testicular histology, sperm production and the subsequent recovery of these parameters. Four adult rhesus monkeys (M. mulatta) were treated with the GnRH agonist nafarelin (D-Nal(2)6-GnRH), released from i.m.--injected poly-D,L-lactic-co-glycolide microspheres for 20 months. Monthly injection of the GnRH agonist preparation uniformly suppressed serum levels of bioactive LH and testosterone. The size of the testis was reduced to about 30% of pretreatment. Sperm counts were suppressed to azoospermia for a total period of 53 and 77 weeks, respectively, in two monkeys and the other two animals were extremely oligozoospermic. Evaluation of testicular biopsy material after 6, 12 and 20 months of treatment revealed decreased seminiferous tubule diameter, spermatogenic disruption at the level of spermatogonia or spermatocytes, accumulation of lipid droplets and secondary lysosomes in the Sertoli cell cytoplasm, and increased thickness of the tubular wall compared with pretreatment histology. Electron microscopic examination revealed that the increased wall thickness was due to an enlargement of the inner collagen layer. No evidence of fibrosis or calcification could be obtained. Leydig cells were atrophic. Serum hormones, testis size and sperm counts returned to pretreatment values within 5 to 8, 13 to 16, and 18 weeks, respectively, after termination of treatment. Testicular histology, assessed 8 months after cessation of treatment, was indistinguishable from pretreatment. It is concluded that GnRH agonist-containing microspheres are a feasible modality for sustained administration of GnRH agonists and GnRH agonist-induced suppression of pituitary and testicular function is reversible following withdrawal of treatment. Thus, GnRH agonists may have a potential for regulation of male fertility and, presumably, also for treatment of precocious puberty.

Dynamics of plasma gonadotropin and sex steroid release in polycystic ovarian disease after pituitary-ovarian inhibition with an analog of gonadotropin-releasing hormone.

To assess the dynamics of the suppression and recovery of plasma gonadotropins and sex steroids during and after inhibition of pituitary-ovarian function by a long-acting agonist GnRH-analog (GnRH-A), eight patients with polycystic ovarian disease were treated with 12 micrograms/kg X day GnRH-A for 56 consecutive days. In response to GnRH-A, these patients had a sharp and pronounced decline of their initially elevated immunoreactive LH and bioactive LH (bioLH) levels. Plasma immunoreactive FSH levels declined more rapidly than did bioLH, but the FSH decline was less sustained. Plasma testosterone, androstenedione, and estrone (E1) levels also declined during GnRH-A administration. The pattern of plasma androgen decrease resembled that of bioLH. There was a positive correlation between bioLH and the two androgens (r = 0.85; P less than 0.05, by Spearman's rank correlation, for both hormones). Cessation of GnRH-A administration was followed by prompt progressive increases in gonadotropin and androgen concentrations to pretreatment values. FSH recovered faster than bioLH. BioLH plasma concentrations reached pretreatment values by day 28. The recovery of plasma androstenedione and testosterone levels correlated positively with that of bioLH. Although plasma E1 levels were higher during the recovery period than during treatment, they never reached the concentrations found during the basal period, whereas estradiol concentrations were slightly but not significantly higher than those in the basal period. As a consequence, the E1 to estradiol ratio, very high in the basal period, approximated unity during recovery. These data indicate that hyperandrogenism in polycystic ovarian disease is gonadotropin dependent and accompanied by a relative abundance of LH bioactivity basally and during GnRH-A administration. Thus, the relative increase in bioLH secretion appears to be independent of the rate of gonadotropin secretion and the circulating sex steroid concentrations.

The effect of bromocriptine on gonadotropin and steroid secretion in polycystic ovarian disease.

Use of bromocriptine in some women with polycystic ovarian disease (PCO) has resulted in ovulation induction, although a mechanism has not been established. The purpose of this study was to determine the effect of bromocriptine on gonadotropin and steroid secretion in this disorder. Two groups of seven patients were given bromocriptine at a dose of either 5 mg/day for 2 months or 10 mg/day for 1 month. Ten normal ovulatory women served as controls. In PCO patients, mean serum levels of LH, bioactive LH, androstenedione, testosterone, unbound testosterone, dehydroepiandrosterone sulfate (DHEA-S), and estrone were significantly greater (P less than 0.05) than those of normal women, whereas FSH, PRL, dihydrotestosterone, 3 alpha-androstanediol, and estradiol were not different. Assessment of gonadotropin secretion before and during treatment revealed that basal levels, episodic secretion, and responses to GnRH (25 micrograms, iv) were unaltered by either dose of bromocriptine. Of the remaining hormones, PRL and DHEA-S significantly decreased in response to both doses. There were no changes in the clinical status of patients during treatment. These findings indicate that in PCO patients with normal PRL levels, gonadotropin secretion is unaltered by bromocriptine therapy. The concomitant declines of PRL and DHEA-S confirm previous data reported for this syndrome and suggest a role for PRL in the production of adrenal androgens.

Loss of luteinizing hormone bioactivity in patients with prostatic cancer treated with an LHRH agonist and a pure antiandrogen.

Chronic treatment of adult men with LHRH agonists causes a decrease in serum testosterone and 5 alpha-dihydrotestosterone to castrate levels. In the presence of such low levels of circulating testicular androgens, the concentration of serum LH measured by radioimmunoassay (RIA) sometimes remains normal or is only partially inhibited. In order to assess the biological activity of circulating LH, we have used the mouse interstitial cell assay. Blood samples were obtained from patients with prostatic carcinoma treated with the LHRH agonist [D-Trp6] LHRH ethylamide in combination with the pure antiandrogen Flutamide (Euflex). While serum LH levels measured by RIA were only partially reduced from 2.2 +/- 0.3 (SEM) to 1.1 +/- 0.1 ng/ml after 3 months of therapy, bioactive LH was markedly inhibited from 0.43 +/- 0.04 to 0.030 +/- 0.007 ng/ml, thus causing the ratio of biologically active to radioimmunoassayable LH to drop from 0.26 +/- 0.03 to 0.03 +/- 0.01. In the same patients, serum testosterone levels were decreased from 3.91 +/- 0.51 to 0.14 +/- 0.05 ng/ml after 3 months of treatment. In patients treated for 6 months, the bio/immuno ratio was still reduced at 0.032 +/- 0.005. These data show a marked loss of LH biological activity during treatment of adult men with an LHRH agonist and an antiandrogen. The close parallelism observed between serum testosterone and bioactive LH levels suggests that the loss of biological activity of the gonadotrophin is mainly, if not exclusively, responsible for the inhibition of testicular androgen secretion observed during chronic treatment with LHRH agonists.

Effects of aging and illness on the pituitary testicular axis in men: qualitative as well as quantitative changes in luteinizing hormone.

Decreased testicular function occurs as a concomitant of aging in men and is accentuated by the presence of systemic illness. Previous studies identified an intrinsic Leydig cell defect, as reflected by high LH to testosterone ratios and impaired hCG responsiveness in older men. This study questioned whether a qualitative change in LH secretion, as reflected by secretion of LH with an altered ratio of biological to immunological LH (B/I) activity, might occur during aging. To examine this possibility, we measured the levels of plasma LH by RIA and rat interstitial cell testosterone bioassay in 67 men, ranging from 20-80 yr of age. Mean LH levels measured by immunoassay were similar in healthy men older than 40 yr [11.4 +/- 1.0 (+/- SE) mIU/ml; n = 22] and those younger than 40 yr (9.4 +/- 0.6; n = 18; P less than 0.01). Mean bioactive LH levels were also similar (30.0 +/- 4.8 vs. 36.7 +/- 3.3). LH B/I ratios, however, were significantly lower in older men (2.52 +/- 0.33) compared to those in younger (4.10 +/- 0.34; P less than 0.01) men. Regression analysis confirmed the expected inverse relationship of B/I ratio with age (r = -0.47; P less than 0.01) and plasma testosterone with age (r = -0.35; P less than 0.05). Systemic illness independently lowered B/I LH ratios. Systemically ill men over 40 yr of age had lower ratios (1.05 +/- 0.08; n = 27) than age-matched healthy men (2.52 +/- 0.33; n = 22; P less than 0.01). The significant changes in B/I ratio among subgroups reflected modest changes in LH immunoactivity and larger alterations in LH bioactivity in certain subgroups. These findings indicate that the qualitative nature of LH secreted by the pituitary, as reflected by altered LH B/I ratios, may vary as a function of aging and illness in men.

Bioassay and Radioimmunoassay of Serum Luteinizing Hormone in the Male Rat

Measurements of immunoactive and bioactive serum LH in the rat were performed after optimization of both RIA and rat interstitial cell-testosterone (RICT) assay to measure the low circulating LH levels of intact male rats. The effective sensitivity of the RIA was increased 3- to 5-fold to 6.2 or 4.7 ng RP1/ml by extraction of 1--1.5 ml serum, respectively. Serum LH levels of adult male rats were 48.5 +/- 16.3 ng RP-1/ml (n = 50), became undetectable after hypophysectomy, and were suppressed 50% after estrogen treatment. For RICT assay of serum rat LH, sensitivity was increased 5-fold by reduction of the incubation volume to 0.35 ml containing 2--4 x 10(6) interstitial cells/tube, with a detection limit of 3 ng RP-1 or 0.1 ng pure LH. The within-assay coefficient of variation for measurement of a pool of control male rat serum (126 +/- 19l6 ng RP-1) WAS +/- 12.5% and the between-assay variation was +/- 15%. Bioactive serum LH levels in adult male rats were 124.8 +/- 32.3 ng RP-1/ml or 2.74 +/- 0.71 ng rat LH/ml, with biopotency to immunopotency (B:I) ratios of 2.8 +/- 0.2 and 1.1 +/- 0.1, respectively, that were unchanged during elevations of serum LH by LHRH administration. Pituitary LH content was 726 +/- 183 micrograms RP-1 or 16.4 micrograms rat LH/gland, with B:I ratios of 2.3 +/- 0.4 and 1.0 +/- 0.2, respectively. The difference between B:I ratios in assays employing pure rat LH and assays using the RP-1 preparation was attributable to the presence of alpha-subunit and biologically inactive material in the RP-1 standard. Precise measurements of immunoactive and bioactive rat LH in male rat plasma can not be performed by these modified RICT and RIA procedures. The sensitive RICT assay can also be applied to the measurement of low levels of LH in the serum of primates and other species.

Bioassay of Circulating Luteinizing Hormone in the Rhesus Monkey: Comparison with Radioimmunoassay During Physiological Changes

The concentration of biologically active LH in rhesus monkey (Macaca mulatta) serum was measured by a highly sensitive bioassay based upon testosterone production by dispersed rat interstitial cells. The sensitivity of the in vitro bioassay was equal to or higher than that of radioimmunoassay, with detection limits of 0.1 mIU of human menopausal gonadotropin (hMG) or 10 ng of a rhesus pituitary gonadotropin preparation (LER-1909-2). Parallel dose-response curves were obtained for hMG and rhesus monkey pituitary gonadotropin. The method permits bioassy of LH in 20-100 micronl of serum from adult male monkeys, and from female monkeys during the follicular and luteal phases of the menstrual cycle. Bioactive LH concentrations could be assayed in 0.25 to 5 micronl of serum from mid-cycle, postmenopausal, and castrated female monkeys. Serum LH was undetectable in two hypophysectomized adult female monkeys and six intact immature animals, and was 13+/-6 (SD) mIU/ml in adult male monkeys. In adult females, follicular phase LH levels ranged from 17 to 169 mIU/ml, with a mean of 76+/-52 mIU/ml. The midcycle LH peak was 1738+/-742 mIU/ml and the luteal phase values ranged from 6-47 mIU/ml, with a mean of 35+/-5 mIU/ml. Serum LH concentrations ranged from 100 to 900 mIU/ml in two menopausal females, and from 590-1480 mIU/ml in castrated females. Treatment of castrated female monkeys with estrogen plus progesterone produced an initial two-fold rise in serum LH within 3 days, followed by a gradual decline to one-fourth to one-tenth of the initial levels after 10 days of treatment. Serum LH was suppressed to undetectable levels during the third week, and remained so for the duration of the 60-day treatment period. Bioactive serum LH levels were comparable to levels determined by radioimmunoassay during the follicular and luteal phases of the menstrual cycle, with increased bio-immunoratio at the midcycle peak. The concentrations of biologically active serum LH in rhesus monkeys were similar to those in the human female during the follicular and luteal phases of the menstrual cycle, and were higher at midcycle and after castration. Serum LH levels measured by the interstitial cell bioassay in the rhesus monkey showed appropriate physiological changes and responses to gonadal steroid administration. Furthermore, the bioassay did not detect the LH-like material measured by heterologous radioimmunoassay in the serum of hypophysectomized, immature and steroid-suppressed monkeys. Thus, the rat interstitial cell assay provides a sensitive and valid procedure for measurement of biologically active LH in the serum of these non-human primates.

Effects of luteinizing hormone releasing hormone (LHRH) upon bioactive and immunoreactive serum LH levels in normal subjects.

The effects of LHRH stimulation upon plasma LH levels measured by bioassay and radioimmunoassay were examined during the normal menstrual cycle, and in normal men and post-menopausal women. After administration of 100 mug LHRH by subcutaneous injection during the early follicular phase, a 2.7-fold rise in bioactive serum LH to 58 +/- 18 mIU/ml at 30-60 min was accompanied by an equivalent rise in immunoreactive LH, with unchanged bio:immuno (B:I) ratio of 0.9 +/ %/- 0.2 (SD). During the late follicular phase, bioactive serum LH rose 8-fold to 258 +/- 120 mIU/ml at 30-180 min, and the B:I ratio was significantly increased from 1.7 to 2.5. During the luteal phase, bioactive LH values rose 8.1-fold to 223 +/- 97 mIU/ml at 30-60 min, with increase in B:I ratio from 1.1 to 1.8. The LHRH-stimulated serum LH levels declined more rapidly in the early follicular phase than during the late follicular and luteal phases. Elevations of circulating LH concentrations following LHRH administration during the normal menstrual cycle were usually accompanied by a significant rise in B:I ratio, except during the early follic ular phase when the LH responses were small and the B:I ratio did not change. After LHRH stimulation of serum LH levels in men and post-menopausal women, relatively small and inconstant elevations of B:I ratio were observed above the basal value of 2.9. Although the B:I ratio was usually close to unity in cycling women, the ratio was from 2 to 3 in men and postmenopausal women, and during LHRH stimulation of normal women at the late follicular and luteal phases of the menstrual cycle. Thus, the increased LH secretion rate of post-menopausal women and LHRH-stimulated cycling women was frequently accompanied by a rise in the B:I ratio. These observations suggest that circulating LH molecules exhibit a relatively higher biological activity during states of increased biosynthesis and release of gonadotropins.