Studies on the microheterogeneity of anterior pituitary follicle-stimulating hormone in the female rat. Isoelectric focusing pattern throughout the estrous cycle

Anterior pituitary (AP) glands were removed from adult female rats at different times throughout the estrous cycle, and the isohormones of follicle-stimulating hormone (FSH) present within them were separated by isoelectric focusing in polyacrylamide gels (PAGE-IEF; pH range 3.0-8.0). Gel eluents were analyzed for FSH content by radioimmunoassay (RIA) and radioreceptor assay (RRA). All AP samples exhibited several peaks of FSH immunoactivity within a pH range of 6.2-4.0; the major peak constantly exhibited an isoelectric point (pI) of 4.9-4.5. To quantify differences in the IEF pattern of AP-FSH between the pituitaries collected during the different days of the cycle, each PAGE-IEF profile was divided into 7 regions (pI 7.0-6.3 = Area 1, 6.2-5.5 = Area 2, 5.4-5.0 = Area 3, 4.9-4.5 = Area 4, 4.4-4.0 = Area 5, 3.9-3.5 = Area 6, and less than 3.5 = Area 7), and the amount of FSH present within each was determined. In all APs collected at 0900 h of diestrus 1 (D1) and 2 (D2), proestrus (P), and estrus (E); at 1300 h of D1, D2 and E; at 2200 h of P; and at 0200 h of E, the majority of FSH immunoactivity (37-55% of total FSH on gel) focused within Area 4, whereas Areas 2 and 3 contained minor amounts of FSH activity (11-26% and 14-24%, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)     

Seasonal variation in pituitary gonadotropin in the adult male newt, Cynops pyrrhogaster pyrrhogaster, revealed by isoelectric focusing technique and radioreceptor assay

The seasonal variation in pituitary gonadotropin in the adult male newt, Cynops pyrrhogaster pyrrhogaster was investigated by means of isoelectric focusing (IEF) coupled with radioreceptor assay (RRA), which employed Anolis or Xenopus testicular homogenates as receptors and 125I-rat FSH as radioligand. In the Anolis RRA system, the standard curve was obtained with 0.125-16 ng/tube of NIAMDD rat FSH I-3. Purified preparations, chicken LH IEF-1, chicken FSH AGCHD11113A and bullfrog basic gonadotropin-IV competitively inhibited the binding of the radioligand, but NIAMDD rat LH I-4 and human chorionic gonadotropin did not crossreact. The autoradiographic study revealed that 125I-rat FSH bound to the constituent cells of the seminiferous tubules in the Anolis testis, but scarcely to Leydig cells. In the IEF pattern of gonadotropin in February obtained by Anolis RRA, distinct peaks were observed at pH 9.05 (component B) and 8.55 (component C), and less distinct peaks were observed at pH 9.80 (component A), 7.55 (component D) and 7.05 (component E). When the same fractions were assayed by Xenopus RRA, five components were found in the alkaline region, which corresponded to those observed with Anolis RRA. Similar results were obtained with pituitary extracts in May. In July, the IEF pattern obtained by Anolis RRA indicated two additional components at pH 6.30 (component F) and 5.27 (component G) in the acidic region, which were not found by Xenopus RRA. The relationship between the testicular function and the nature of pituitary gonadotropin in the reproductive cycle was discussed.     

Species specificity of radioreceptor assay and radioimmunoassay for rat FSH

Species specificity of the radioreceptor assay (RRA) for rat FSH, in which pregnant mare serum gonadotropin (PMSG)-treated immature rat ovary was employed as the receptor, was compared with that of NIAMDD rat FSH radioimmunoassay (RIA). In the RIA system, pituitary preparations from mammals only showed significant crossreaction. Their inhibition curves, however, were not always parallel to the standard curve. On the other hand, in the RRA system, the pituitary preparations from mammals, avians, lizard and amphibians competitively inhibited the binding of radioactive rat FSH to the ovarian receptor. Only the pituitary preparation from dog salmon failed to show any crossreaction in the RRA system. These results indicated that this RRA system would be useful for the measurement of FSH or gonadotropins of the pituitaries from mammals to amphibians.     

Biological and immunological properties of zebra pituitary gonadotropins: comparison with horse and donkey gonadotropins

Previous studies from this laboratory have described the properties of purified luteinizing hormone (LH) and follicle-stimulating hormone (FSH) from horse and donkey anterior pituitary glands. The present study afforded the opportunity to further characterize these previously purified hormone preparations and to compare them with enriched gonadotropin fractions from zebra pituitary glands. Although a single LH and FSH fraction was usually obtained for each pool of pituitaries, two separate zebra LH and two donkey FSH preparations were generated. Purified hormone preparations from the horse were designated eLH and eFSH. Preparations zLH-A, zLH-B, and zFSH were obtained from zebra pituitaries, and fractions dLH, dFSH-A, and dFSH-B were prepared from donkey pituitary glands. These preparations were analyzed by LH and FSH radioimmunoassays (RIAs), radioreceptor assays (RRAs), LH bioassay, and chromatofocusing. Clear immunological differences were observed between equid gonadotropins. Homologous RIAs for eLH and eFSH did not cross-react similarly, or in a parallel fashion, with gonadotropins from the donkey and zebra. In contrast, RIAs capable of assessing LH or FSH in a wide number of species showed all equid gonadotropin preparations to have considerable activity and to produce parallel dilution curves. Relative to eLH (1.00), zLH-A was found to have higher LH bioactivity:LH RIA (2.50), LH RRA:LH RIA (1.42), and LH bioactivity: LH RRA (2.21) activity ratios. The dLH and zLH-B fractions only differed from eLH in LH RRA:LH RIA activity (0.69 and 0.62, respectively). Only LH from the horse possessed clear intrinsic FSH-receptor-binding activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prolactin messenger ribonucleic acid concentrations throughout the ovine estrous cycle: assessment relative to prolactin serum and pituitary amounts

Prolactin (PRL) mRNA concentrations were assessed by nucleic acid hybridization assays in pituitaries of ewes representing the defined stages of the ovine estrous cycle. Concomitantly, pituitary and serum PRL concentrations were measured in these ewes using radioimmunoassays. It was observed that PRL serum, pituitary and mRNA concentrations tended to increase near the time of the gonadotropin preovulatory surge, particularly between 24 hrs before behavioral estrus to 5 hours after estrus. However, the changes in PRL mRNA, serum and pituitary concentrations were shown not to be statistically significant. These data suggest that PRL production during the sheep estrous cycle is maintained without dramatic changes in synthesis or secretion.     

Gonadotropin-releasing hormone binding sites in ovaries of normal cycling and persistent-estrus rats

The gonadotropin-releasing hormone (GnRH) binding capacity in ovaries and pituitaries of normal cycling rats at different stages of the estrous cycle and in ovaries of persistent-estrus rats was measured using radioligand-receptor assay (RRA). Persistent estrus was induced either by neonatal administration of testosterone propionate (1.25 mg s.c.) on the second day of life or by a hypothalamic suprachiasmatic frontal cut made with Halász' knife. All animals were killed during the critical period (1400-1600 h), and GnRH receptor was assayed. GnRH receptor levels in both ovaries and pituitaries changed during the estrous cycle. The total number of ovarian GnRH binding sites was significantly higher in proestrus than in diestrus 1, the stage in which the lowest level was found. When binding sites were expressed in fmol/mg ovary, the highest level was observed in diestrus 2; however, no changes were observed during the estrous cycle when GnRH binding sites were expressed as fmol/mg protein. Changes noted were very similar to those demonstrated in pituitary GnRH receptors in our present and previous experiments. Higher levels of pituitary binding sites were found in diestrus 2 and proestrus than in estrus and diestrus 1. The changes in the GnRH receptor levels were more striking in the pituitary than in the ovaries. It appears that the total number of ovarian GnRH binding sites was not altered in either of the two persistent-estrus groups, but that their concentration was significantly higher (expressed in fmol/mg ovary or fmol/mg protein) than on any day during the estrous cycle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hormone secretion in the asian elephant (Elephas maximus): characterization of ovulatory and anovulatory luteinizing hormone surges.

In the elephant, two distinct LH surges occur 3 wk apart during the nonluteal phase of the estrous cycle, but only the second surge (ovLH) induces ovulation. The function of the first, anovulatory surge (anLH) is unknown, nor is it clear what regulates the timing of these two surges. To further study this observation in the Asian elephant, serum concentrations of LH, FSH, progesterone, inhibin, estradiol, and prolactin were quantified throughout the estrous cycle to establish temporal hormonal relationships. To examine long-term dynamics of hormone secretion, analyses were conducted in weekly blood samples collected from 3 Asian elephants for up to 3 yr. To determine whether differences existed in secretory patterns between the anLH and ovLH surges, daily blood samples were analyzed from 21 nonluteal-phase periods from 7 Asian elephants. During the nonluteal phase, serum LH was elevated for 1-2 days during anLH and ovLH surges with no differences in peak concentration between the two surges. The anLH surge occurred 19.9+/-1.2 days after the end of the luteal phase and was followed by the ovLH surge 20.8+/-0.5 days later. Serum FSH concentrations were highest at the beginning of the nonluteal phase and gradually declined to nadir concentrations within 4 days of the ovLH surge. FSH remained low until after the ovLH surge and then increased during the luteal phase. Serum inhibin concentrations were negatively correlated with FSH during the nonluteal phase (r = -0.53). Concentrations of estradiol and prolactin fluctuated throughout the estrous cycle with no discernible patterns evident. In sum, there were no clear differences in associated hormone secretory patterns between the anLH and ovLH surge. However, elevated FSH at the beginning of the nonluteal phase may be important for follicle recruitment, with the first anLH surge acting to complete the follicle selection process before ovulation.     

Pituitary luteinizing hormone (LH) and follicle-stimulating hormone (FSH) responses to gonadotropin-releasing hormone during the rat estrous cycle: an increased ratio of FSH to LH is secreted during the secondary FSH surge

The hormonal interactions required for the generation of a secondary surge of FSH on the evening of proestrus have not been clearly defined. The role of GnRH in driving a surge of FSH has been questioned by findings in previous studies. In the current study, gonadotropin secretion was measured from pituitary fragments obtained from rats at 0900 and 2400 h on each day of the estrous cycle. Pituitary fragments were perifused in basal (unstimulated) conditions or in the presence of GnRH pulses to determine whether a selective increase in basal release of FSH and/or an increase in the responsiveness to GnRH occurs during the secondary FSH surge. Each anterior pituitary was cut into eighths and placed into a microchamber for perifusion. Seven pulses of GnRH (peak amplitude = 50 ng/ml; duration = approximately 2 min) were administered at a rate of one per hour starting at 30 min. Fractions of perfusate were collected every 5 min and frozen until RIA for LH and FSH. The mean total amount of LH or FSH secreted during the hour interval following each of the last six pulses of GnRH (or the corresponding basal hour) was calculated. Analysis of variance with repeated measures indicated that the evening secretion of LH on proestrus (2400 h) dropped significantly (p less than 0.05) from a maximum on the morning of proestrus (0900 h), whereas the FSH secretion remained elevated at this time. Therefore, the ratio of FSH to LH secreted in response to GnRH pulses was highest during the secondary FSH surge and lowest on the morning of proestrus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Subpopulations of luteinizing hormone (LH) possessing various ratios of bioactivity to immunoreactivity in the female rat pituitary glands and their changes during the estrous cycle

Twenty-four adult female Sprague-Dawley rats (3 from each of 8 litters), showing 4-day cycles, were used in the present study. Aqueous extracts of pools of 6 pituitary glands in each cycle date were fractionated with a column isoelectrofocusing (IEF) technique, pH range of 3.5-10. Biological and immunological LH activities were determined by an in vitro bioassay and a radioimmunoassay, respectively, in the original aqueous extracts of the pituitary glands and in the fractions separated by IEF. Pituitary content of LH was the highest in the proestrus before the preovulatory LH surge (1243.7 +/- 67.8 micrograms NIAMDD rat LH-RP-1/pituitary gland for the biological activity). In the estrus, after the LH surge, it was reduced to 688.9 +/- 51.2 micrograms/pituitary gland. The decreased pituitary content was recovered to the level in the proestrus during the metestrus and the diestrus (1047.0 +/- 53.8 and 1173.0 +/- 58.5 micrograms/pituitary gland, respectively). Rat LH in the pituitary aqueous extracts was separated into multiple subpopulations in terms of pI values by IEF; i.e. Subpopulations A (pI = 10.3), B (9.3), C (9.0), D (8.7), E (8.3), F (neutral LH), and G (acidic LH). Among them the most predominant one was Subpopulation A throughout the estrous cycle. Subpopulations A, B and C exhibited statistically significant cyclic changes as was observed in the pituitary LH content, whereas the remaining ones stayed at constant levels during the cycle. The highest ratio of biological to immunological LH activities (B/I ratio) was obtained in Subpopulation A (6.41), followed by G, C and B (5.15, 4.24 and 3.99, respectively). Depressed B/I ratios were revealed in D, E and F (2.59, 1.86 and 3.07, respectively). High alkaline LH subpopulations, i.e. A, B and C, preserving high biological potency and showing cyclic changes during the estrous cycle, seem to be the releasable types of the hormone and to be mainly discharged for the preovulatory LH surge. Although characteristic features of other types of the hormone are not known, it is possible that one of them, presumably the acidic LH, might be the newly-synthesized type of the hormone, which might attain releasability by certain molecular modifications involving a shift in the pI value.     

Midcycle and luteal elevations of follicle stimulating hormone in squirrel monkeys (Saimiri boliviensis) during the estrous cycle

Follicle stimulating hormone (FSH) has fundamental importance in reproductive function, but its cyclic pattern has not previously been described in the squirrel monkey, due primarily to the lack of a suitable assay. An homologous radioimmunoassay (RIA) based on recombinant cynomolgus FSH measured changes in serum FSH relative to patterns of bioactive luteinizing hormone (LH), estradiol, and progesterone during the estrous cycle. FSH was observed to have a sharp peak during the late follicular phase coincident with the LH surge and then rose again during the luteal phase. Estradiol was low except for the midcycle rise, suggesting an inhibitory relationship. The rat granulosa cell in vitro FSH bioassay confirmed high levels of this hormone. Measurement of FSH in the squirrel monkey has found a pattern different from Old World primates in the luteal phase, which may provide insight into the reproductive mechanisms of this species.     

虎纹蛙促性腺激素含量随年龄及季节的变化(英文)

促性腺激素（ＧｔＨ;或ＬＨ和ＦＳＨ）在脊椎动物生殖调节中起中Ｃ作用;脑垂体和血浆ＧｔＨ水平在一定程度上反映着动物体的生殖生理状态。在蛙类,对脑垂体和血浆的ＬＨ及ＦＳＨ含量进行过较全面研究的,仅在牛蛙和日本蟾蜍有过报道,结果显示它们的ＬＨ和ＦＳＨ的含量存在有种类差异性。虎纹蛙属中国二级保护动物,也是唯一受保护的蛙类,对其生殖生物学的基础内容进行研究具有重要的意义。 为此,本文利用放射免疫测定法,测定了虎纹蛙幼蛙和不同性腺发育阶段（季节）成蛙脑垂体与血浆的ＬＨ及ＦＳＨ含量,以弄清这些激素的含量变化与年龄、季节（性腺发育阶段）变化的关系,以期为虎纹蛙的基础生殖生物学及蛙类的生殖内分泌学充实新的内容,为虎纹蛙的人工繁殖和保护提供理论依据。结果是：幼蛙血浆ＬＨ水平显著高于各期成蛙（Ｆｉｇ．Ａ）,血浆ＦＳＨ水平显著低于成熟前期成蛙,而和其它各性腺发育阶段成蛙相当（Ｆｉｇ．Ｂ）。而脑垂体ＬＨ或ＦＳＨ的含量显著低于各期成蛙（Ｆｉｇ．Ｃ＆Ｄ）。这说明,幼蛙脑垂体已具有一定的合成和释放ＬＨ及ＦＳＨ的能力。 成蛙脑垂体和血浆ＬＨ及ＦＳＨ水平随性腺发育阶段（季节）的不同而有一定的变化：雌蛙血浆ＬＨ水平在成熟期最高,性腺再发育期最低;     

Characterization of equine luteinizing hormone by chromatofocusing

Three equine luteinizing hormone (LH) preparations (eLH-A, -B, and -C) recently have been isolated in our laboratory and were shown to differ in average basicity (eLH-A greater than -B greater than -C). The present study further characterizes these preparations by chromatofocusing. Each of these preparations are comprised of a family of isohormones, with 5 major immunoreactive peaks in the pH range of 7 to 4 (approx. pIs = 6.6, 6.1, 5.7, 5.2, and 4.8), with varying amounts of material eluting to either side of the pH gradient. Although similar isoforms are seen in all three LH preparations, the relative proportions of different isoforms vary in a manner reflecting the average charge properties of eLH-A, -B, and -C. While eLH-A contains predominantly basic forms, eLH-C consists largely of acidic material, and eLH-B is composed mostly of isohormones with pIs intermediate to eLH-A and -C. Chromatofocusing of a crude extract from a single horse pituitary gland revealed isohormone peaks corresponding to those found in the highly purified LH preparations. Peak fractions of the various isoforms were used to generate a variety of activity ratios (LH bioactivity:LH radioimmunoassay (RIA), LH radioreceptorassay (RRA):LH RIA, LH bioactivity:LH RRA, follicle-stimulating hormone (FSH) RRA:LH RIA, and FSH RRA:LH RRA activity ratios). The LH bioactivity:LH receptor binding potency ratio showed a linear increase with increasing isohormone acidity (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential regulation of gonadotropin subunit messenger ribonucleic acids by gonadotropin-releasing hormone pulse frequency in ewes

Changes in the frequency of GnRH and LH pulses have been shown to occur between the luteal and preovulatory periods in the ovine estrous cycle. We examined the effect of these different frequencies of GnRH pulses on pituitary concentrations of LH and FSH subunit mRNAs. Eighteen ovariectomized ewes were implanted with progesterone to eliminate endogenous GnRH release during the nonbreeding season. These animals then received 3 ng/kg body weight GnRH in frequencies of once every 4, 1, or 0.5 h for 4 days. These frequencies represent those observed during the luteal and follicular phases, and the preovulatory LH and FSH surge of the ovine estrous cycle, respectively. On day 4, the ewes were killed and their anterior pituitary glands were removed for measurements of pituitary LH, FSH, and their subunit mRNAs. Pituitary content of LH and FSH, as assessed by RIA, did not change (P greater than 0.10) in response to the three different GnRH pulse frequencies. However, subunit mRNA concentrations, assessed by solution hybridization assays and expressed as femtomoles per mg total RNA, did change as a result of different GnRH frequencies. alpha mRNA concentrations were higher (P less than 0.05) when the GnRH pulse frequency was 1/0.5 h and 1 h, whereas LH beta and FSH beta mRNA concentrations were maximal (P less than 0.05) only at a pulse frequency of 1/h. Additionally, pituitary LH and FSH secretory response to GnRH on day 4 was maximal (P = 0.05) when the pulse infusion was 1/h.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of thymulin and GnRH on the release of gonadotropins by in vitro pituitary cells obtained from rats in each day of estrous cycle

The effects of thymulin and GnRH on FSH and LH release were studied in suspension cultures of anterior pituitary cells from female adult rats sacrificed on each day of the estrous cycle. The spontaneous release of gonadotropins by pituitaries, as well as their response to GnRH or thymulin addition, fluctuated during the estrous cycle. Adding thymulin to pituitary cells from rats in diestrus 1 increased the concentration of FSH; while in cells from rats in estrus, FSH level decreased. Thymulin had a stimulatory effect on the basal concentration of LH during most days of the estrous cycle. Adding GnRH increased FSH release in cells from rats in diestrus 1, diestrus 2, or proestrus, and resulted in higher LH levels in cells obtained from rats in all days of the estrous cycle. Compared to the GnRH treatment, the simultaneous addition of thymulin and GnRH to cells from rats in diestrus 1, diestrus 2, or proestrus resulted in lower FSH concentrations. Similar results were observed in the LH release by cells from rats in diestrus 1, while in cells from rats in proestrus or estrus, LH concentrations increased. A directly proportional relation between progesterone serum levels and the effects of thymulin on FSH release was observed. These data suggest that thymulin plays a dual role in the release of gonadotropins, and that its effects depend on the hormonal status of the donor's pituitary.     

Changes in expression of inhibin subunits in the cyclic golden hamster (Mesocricetus auratus) and the regulation of inhibin alpha subunit production by luteinizing hormone

In the present study, changes in localization of each inhibin subunit in the ovary were investigated during the estrous cycle of the golden hamster. The effect of LH surge on changes in localization in inhibin alpha subunit in the ovary was also investigated. Inhibin alpha subunit was localized in granulosa cells of various stages of follicles throughout the estrous cycle. Inhibin alpha subunit was also present in numerous interstitial cells on days 1 and 2 (day 1 = day of ovulation), but the number of positive interstitial cells was fewer on days 3 and almost disappeared on day 4 of the estrous cycle. Newly formed luteal cells were also positive for inhibin alpha subunit on days 1 and 2. On the other hand, positive reactions for inhibin beta A and beta B subunits were only present in the granulosa cells of healthy antral follicles. However, a positive reaction for inhibin beta B subunit in peripheral mural granulosa cells disappeared on days 3 and 4 of the estrous cycle. Treatment with LHRH-AS at 1100 h on day 4 completely blocked the luteinizing hormone (LH) surge and ovulation, although relatively high concentrations of plasma follicle-stimulating hormone (FSH) were maintained throughout the experiment. There were few positive reactions for inhibin alpha subunit in theca and interstitial cells 24 hr after LHRH-AS injection. The effect of LHRH-AS treatment was blocked by a single injection of 10 IU human chorionic gonadotropin. These results suggest that the major source of dimeric inhibin in the cyclic hamster was granulosa cells of healthy antral follicles. Different distribution pattern of inhibin beta A from beta B subunits in large antral follicles on days 3 and 4 of the estrous cycle suggests different secretion patterns of inhibin A from B on these days. Furthermore, the LH surge may be an important factor to induce production of inhibin alpha subunit in interstitial cells of the cyclic hamster.     

Superfused pituitary cell cultures: comparative responsiveness of cells derived from various stages of the estrous cycle to LHRH stimulation administered as short duration pulses

We have reinvestigated the question of maintenance of differential LHRH sensitivity in culture and further investigated the role of pulsatile LHRH in the in vitro release of pulsatile LH and FSH at different stages of the estrous cycle. Pituitaries were collected on each day of the 4 day cycle at 0800. In addition, pituitaries were also collected at 1500 and 1900 on proestrous. The cells were dispersed and exposed 48 hrs later to short duration 4 ng LHRH pulses; this dose was optimized for LH release and was applied at a frequency of 1 pulse/60 min. In terms of absolute magnitude of LH response, observed responsiveness was ranked in the following order: proestrous 1900 greater than estrous 0800 greater than diestrous 1 0800 greater than proestrous 1500 greater than diestrous 2 0800. Responsiveness was significantly greater at proestrous 1900 (p greater than 0.01), estrous 0800 (p greater than 0.05) and diestrous 1 0800 (p greater than 0.05) when compared to either of the other stages tested. The heightened LHRH sensitivity of proestrous was therefore maintained in cell culture indicating that the system should be valid for conducting studies on the control of gonadotropin secretion during this period. FSH did not respond in pulsatile manner to the LHRH levels employed further substantiating recent evidence that LHRH seems to function somehow less directly in FSH as compared to LH secretion.     

A similar distribution of gonadotropin isohormones is maintained in the pituitary throughout sexual maturation in the heifer.

Our working hypotheses for this study were that 1) the profile of intrapituitary LH and FSH isoforms would be shifted toward acidic forms as sexual maturation progresses in the bovine female; and 2) concentration of 17 beta-estradiol (E2) in circulation during sexual maturation would be a major factor modulating the percentage of the more acidic isoforms. In addition, the biological-immunoreactive (B:I) ratios of each isoform of LH were evaluated at selected stages of sexual maturation. Heifers (7 mo of age) were assigned to one of three treatment groups: 1) ovariectomized (OVX; n = 16); 2) OVX and administered E2 (OVXE; n = 16); or 3) ovary-intact (INTACT; n = 14). Pituitaries were collected from heifers in each group at an estimated 120 days (prepubertal) of 25 days before puberty (peripubertal). A fourth group of 6 heifers remained intact (postpubertal INTACT) to determine time of puberty during the experimental period. Pituitaries of heifers assigned to the postpubertal INTACT group were collected during the follicular phase of the first or second estrous cycle postpuberty. Pituitaries were used for determination of relative amounts of gonadotropin isohormones. Tissue extracts of the pituitaries were chromatofocused on pH 10.5-4.0 gradients. The LH of all pituitaries resolved into thirteen isoforms that were designated isoforms A-L, and S, with isoform A the most basic form. Isoforms F and G (basic pH range) were the predominant isoforms of each chromatofocusing profile and comprised 50-60% of the immunoreactive LH. Isoforms J and K were the major isoforms eluting in the acidic pH range.(ABSTRACT TRUNCATED AT 250 WORDS)     

Measurement of human FSH by radioreceptor assay

We established a sensitive RRA system for human FSH, employing PMS-treated immature rat ovary. The dissociation constant of the binding of the receptor preparation to NIAMDD human FSH-2 was 1.15 x 10(-10) M. The standard curve was obtained with 0.2-25.6 ng/tube of NIAMDD hFSH-2. Purified hLH, hTSH, and HCG had no significant effect on the binding. When the anterior pituitary homogenates obtained from humans were assayed by this system, the intra-assay and inter-assay coefficients of variation were 11.9% and 13.4% respectively, and the assay values correlated well with those obtained by RIA. This assay is applicable for the measurement of FSH in serum, when the non-specific inhibitor effects of serum are compensated for by the addition of merthiolate and when FSH-free serum is used instead of the buffer for the standard curve. The intra-assay and inter-assay coefficients of variation were 9.31% and 19.7% respectively. The assay values correlated with those obtained by RIA under the same physiological state. The ratio of the assay values RRA/RIA, was dependent upon the physiological state, e.g. 6.29 in men, 3.84 and 4.18 in women at follicular and luteal phase respectively and 2.40 in menopausal women. During the menstrual cycle, our results showed that the value of RRA/RIA derived from serum did not change significantly.     

A change in basal FSH release accompanies the onset of the second or selective phase of increased serum FSH in the cyclic rat

Experiments were undertaken to investigate if changes occur at the level of the anterior pituitary gland to result in selective follicle-stimulating hormone (FSH) release during late proestrus in the cyclic rat. At 1200 h proestrus, prior to the preovulatory luteinizing hormone (LH) surge in serum and the accompanying first phase of FSH release, serum LH and FSH concentrations were low. At 2400 h proestrus, after the LH surge and shortly after the onset of the second or selective phase of FSH release, serum LH was low, serum FSH was elevated about 4-fold, pituitary LH concentration was decreased about one-half and pituitary FSH concentration was not significantly decreased. During a two hour $\text{in}$$\text{vitro}$ incubation, pituitaries collected at 2400 h released nearly two-thirds less LH and 2.5 times more FSH than did pituitaries collected at 1200 h. Addition of luteinizing hormone releasing hormone (LHRH) to the incubations caused increased pituitary LH and FSH release. However, the LH and FSH increments due to LHRH in the 2400 h pituitaries were not different from those in the 1200 h pituitaries. The results indicate that a change occurs in the rat anterior pituitary gland during the period of the LH surge and first phase of FSH release which results in a selective increase in the basal FSH secretory rate. It is suggested that this change is primarily responsible for the selective increase in serum FSH which occurs during the second phase of FSH release.     

Comparisons of estradiol,LH and FSH patterns in pregnant and nonpregnant beagle bitches

To characterize plasma estradiol, LH and FSH patterns of secretion during the bitch estrous cycle, blood samples were obtained daily from 15 days before until 135 days after the LH surge in 10 pregnant and 10 nonpregnant beagle bitches. After an initial increase between days 15 and 10 and an expected proestrous peak, estradiol concentrations increased again from days 9-12 (corresponding to cytological metestrus) from basal values observed around day 9 after the LH surge, and remained significantly elevated throughout the luteal phase both in pregnant and nonpregnant animals. Concomitantly with the end of the luteal phase, plasma concentrations of estradiol returned to basal values in both groups. During the mid- to late-luteal phase, mean basal LH secretion was significantly elevated throughout in the pregnant relative to the nonpregnant animals. However, in nonpregnant animals, pulsatility was increased and peaks of higher amplitude were observed. The plasma FSH profiles, determined by a specific homologous RIA, differed significantly between pregnant and nonpregnant bitches during the last two-thirds of the luteal phase with a mean FSH level more elevated during pregnancy. The FSH level then decreased around parturition and low concentrations during lactation period were observed. The FSH concentrations remained steady in nonpregnant luteal phases from early luteal phase through mid-anestrus. The differences in pregnant and nonpregnant LH and FSH concentrations suggest pregnancy differences in regulation of the corpus luteum. Finally, the elevated estradiol concentrations observed during the luteal phase of both pregnant and nonpregnant animals suggest that an ovarian production of estrogens may be involved in overall corpus luteum regulation in dogs as in other species.