Luteinizing Hormone Binding Research Articles

Luteinizing Hormone Dynamics in Menstruation.

Menstruation is a finely-controlled cycle that responds to the prevailing endocrine and paracrine environment. However, social stigma has led to inadequate menstrual literacy, both among academics and the larger public. The poorly understood mechanisms of menstruation ultimately lead to suboptimal healthcare treatment and services for biological females, culminating in a physical, financial, and emotional burden. Various hormones signal the beginning and end of each stage of menstruation. In particular, luteinizing hormone (LH) is a major player in ovulation, corpus luteum function, and the stimulation of other key hormones. A LH model could be used to understand the larger control system of menstruation if analyzed in conjunction with models for other major hormones (e.g., FSH, progesterone, and GnRH). Thus, exploring a smaller subsection of LH dynamics within the larger control system of menstruation can lead to a greater understanding of menstruation, contributing towards therapeutics and research for women's health. Using parameters and kinetic equations in the existing body of literature, a transfer function was derived to model LH dynamics. Analysis of system stability reveals overdamped dynamics in LH sensitization at baseline, and underdamped mildly resonant dynamics at the peak of the menstrual cycle, the strength of which depends on the values of the rate constants of LH receptor formation, binding, and desensitization.

Mechanism of Action of an Environmentally Relevant Organochlorine Mixture in Repressing Steroid Hormone Biosynthesis in Leydig Cells.

Within Leydig cells, steroidogenesis is induced by the pituitary luteinizing hormone (LH). The binding of LH to its receptor increases cAMP production, which then activates the expression of genes involved in testosterone biosynthesis. One of these genes codes for the steroidogenic acute regulatory (STAR) protein. STAR is part of a complex that shuttles cholesterol, the precursor of all steroid hormones, through the mitochondrial membrane where steroidogenesis is initiated. Organochlorine chemicals (OCs) are environmental persistent organic pollutants that are found at high concentrations in Arctic areas. OCs are known to affect male reproductive health by decreasing semen quality in different species, including humans. We previously showed that an environmentally relevant mixture of OCs found in Northern Quebec disrupts steroidogenesis by decreasing STAR protein levels without affecting the transcription of the gene. We hypothesized that OCs might affect STAR protein stability. To test this, MA-10 Leydig cell lines were incubated for 6 h with vehicle or the OCs mixture in the presence or absence of 8Br-cAMP with or without MG132, an inhibitor of protein degradation. We found that MG132 prevented the OC-mediated decrease in STAR protein levels following 8Br-cAMP stimulation. However, progesterone production was still decreased by the OC mixture, even in the presence of MG132. This suggested that proteins involved in steroid hormone production in addition to STAR are also affected by the OC mixture. To identify these proteins, a whole cell approach was used and total proteins from MA-10 Leydig cells exposed to the OC mixture with or without stimulation with 8Br-cAMP were analyzed by 2D SDS-PAGE and LC-MS/MS. Bioinformatics analyses revealed that several proteins involved in numerous biological processes are affected by the OC mixture, including proteins involved in mitochondrial transport, lipid metabolism, and steroidogenesis.

Leydig cell clustering and Reinke crystal distribution in relation to hormonal function in adult patients with testicular dysgenesis syndrome (TDS) including cryptorchidism.

Testicular dysgenesis syndrome (TDS) comprises testicular germ cell cancer, cryptorchidism and some cases of male infertility and hypospadias, which can be linked to impairment of intrauterine gonadal development. Among histological signs of TDS, large Leydig cell (LC) clusters (micronodules) are frequently present. This study aimed to investigate possible associations of LC micronodules with the presence of Reinke crystals and hormonal function of LCs, the latter primarily reflected by serum concentrations of luteinising hormone (LH) and testosterone, in patients with TDS. A retrospective study of 101 andrological patients with TDS (infertility with and without a history of cryptorchidism or presence of germ cell neoplasia in situ) and 20 controls with normal testis histology and LC-function. Archived testicular biopsies were re-evaluated for the presence of LC micronodules and Reinke crystals and the findings were correlated with testis size and serum concentrations of LH, follicle-stimulating hormone (FSH), testosterone, inhibin B, estradiol and sex hormone binding globulin (SHBG). TDS patients with bilateral LC micronodules had significantly lower concentrations of LH, FSH and inhibin B, a lower testosterone/LH-ratio and smaller testis sizes compared to TDS-patients lacking this feature. Presence of LC micronodules was correlated with a lower number of Reinke crystals, while cryptorchid testes had a significantly higher number of crystals than normally descended TDS testes. LC micronodules appear to be a compensatory mechanism caused by androgenic failure and are presumably driven by high concentrations of LH. A relative paucity of Reinke crystals in LCs within micronodules in normally descended TDS testes may be a feature of recently renewed immature Leydig cells. The increased number of Reinke crystals in LCs in testes that were either undescended at birth or are persistently undescended could indicate an impairment of LC renewal in cryptorchidism.

Exploration of Steroidoginic Activity of Ketorolac on Isolated Leydig Cells of Goat (Capra hircus) In Vitro: Part II

In the present thesis was conducted to clarify the effect of ketorolac (NSAID) on the Leydig cells of goats to achieve the ketorolac event, by the evaluation steroidoginic activity of ketorolac series on Leydig cell concentrations in vitro. Mitochondrial defects reduction of functional processes of sterodoginc activity reduction of testosterone levels via mitotracker probe indicator, ketorolac may be affected on the steroidogenic acute regulatory (STAR) protein and translocator protein (TSPO). Ketorolac might mutilation and impair the cells were via the uncoupling of oxidative phosphorylation in the cells followed by uncouple mitochondrial respiratory chain as well as ATP depletion. Testosterone level displays in all treated maneuvers LH, LH-GnRH and GnRH in ketorolac challenge decrease testosterone levels at a concentration of ketorolac from 0.6-0.15 mM with the exception in GNRH – ketorolac maneuvers result showed an increase in testosterone level in 0.03-0.06 mM, then fall up down other maneuvers of treatment. The testosterone synthesis in Leydig cells of mammalian is achieved almost motivated by the binding of luteinizing hormone (LH) to the plasma membrane of Leydig cell receptors efficaciously, that lead to creation of c.AMP Inhibition of LTB4 that due to direct inhibition of lipoxygenase action, and had a direct effect to reduce LTB4 and may be various eicosanoids also 12 (S) - hydroxyeicosatetraenoic acid and 15 (S) - hydroxyeicosatetraenoic acid binds to and activates LTB.

Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles

Meiosis in mammalian oocytes is paused until luteinizing hormone (LH) activates receptors in the mural granulosa cells of the ovarian follicle. Prior work has established the central role of cyclic GMP (cGMP) from the granulosa cells in maintaining meiotic arrest, but it is not clear how binding of LH to receptors that are located up to 10 cell layers away from the oocyte lowers oocyte cGMP and restarts meiosis. Here, by visualizing intercellular trafficking of cGMP in real-time in live follicles from mice expressing a FRET sensor, we show that diffusion of cGMP through gap junctions is responsible not only for maintaining meiotic arrest, but also for rapid transmission of the signal that reinitiates meiosis from the follicle surface to the oocyte. Before LH exposure, the cGMP concentration throughout the follicle is at a uniformly high level of ∼2-4 μM. Then, within 1 min of LH application, cGMP begins to decrease in the peripheral granulosa cells. As a consequence, cGMP from the oocyte diffuses into the sink provided by the large granulosa cell volume, such that by 20 min the cGMP concentration in the follicle is uniformly low, ∼100 nM. The decrease in cGMP in the oocyte relieves the inhibition of the meiotic cell cycle. This direct demonstration that a physiological signal initiated by a stimulus in one region of an intact tissue can travel across many layers of cells via cyclic nucleotide diffusion through gap junctions could provide a general mechanism for diverse cellular processes.

Design and Development of a Field Applicable Gold Nanosensor for the Detection of Luteinizing Hormone

In this paper, we describe a novel strategy for the fabrication of a nanosensor for detecting luteinizing hormone (LH) of sheep using a gold nanoparticle-peptide conjugate. A new peptide sequence "CDHPPLPDILFL" (leutinizing hormone peptide, LHP) has been identified, using BLAST and Clustal W analysis, to detect antibody of LH (sheep). LHP has been synthesized and characterized, and their affinity toward anti-LH was established using enzyme linked immunosorbant assay (ELISA) technique. The thiol group in LHP directly binds with gold nanoparticles (AuNPs) to yield AuNP-LHP construct. Detailed physicochemical analysis of AuNP-LHP construct was determined using various analytical techniques. Nanosensor using gold nanoparticle peptide conjugate was developed on the basis of competitive binding of AuNP-LHP and LH toward anti-LH. Nitrocellulose membrane, precoated with anti-LH, was soaked in the mixture of AuNP-LHP and sample of analysis (LH). In the absence of LH (sheep), anti-LH coated on the membrane binds with AuNP-LHP, leading to a distinctive red color, while in the presence of LH, no color appeared in the membrane due to the interaction of anti-LH with LH thereby preventing the binding of AuNP-LHP with membrane bound anti-LH. The sensor assay developed in this study can detect LH (sheep) up to a minimal concentration of ∼50 ppm with a high degree of reproducibility and selectivity. The gold-nanoparticle-peptide based nanosensor would be a simple, portable, effective, and low cost technique for infield applications.

Rescue of expression and signaling of human luteinizing hormone G protein-coupled receptor mutants with an allosterically binding small-molecule agonist

Naturally occurring mutations of G protein-coupled receptors (GPCRs) causing misfolding and failure to traffic to the cell surface can result in disease states. Some small-molecule orthosteric ligands can rescue such misfolded receptors, presumably by facilitating their correct folding and shuttling to the plasma membrane. Here we show that a cell-permeant, allosterically binding small-molecule agonist (Org 42599) rescues the folding and cell surface expression, and therefore target cell signaling, of mutant human luteinizing hormone (LH) receptors (A593P and S616Y) that cause Leydig cell hypoplasia in man. Both mutant receptors were retained in the cytoplasm whereas WT receptor localized at the cell membrane, and binding of LH to cells expressing the mutant receptors was markedly lower than to those expressing the WT receptor. Incubation with Org 42599 increased mutant receptor expression, cell surface localization, and the proportion of mutant receptor in the mature glycosylated form. Importantly, although LH stimulated little (S616Y) or no (A593P) activation of cells expressing mutant receptors, incubation of cells with Org 42599 facilitated rescue of expression and stimulation by the native ligand, LH. Although Org 42599 could activate these receptors, it could not displace (125)I-labeled human LH binding to the WT receptor, indicating that it acts in an allosteric manner. Here we demonstrate a small-molecule GPCR allosteric agonist that functionally rescues intracellularly retained mutant LH receptors by facilitating their cell surface expression. This approach may have application for treatment of infertile patients bearing such mutations and, more broadly, for other misfolded GPCR mutants resulting in human pathologic processes.

A calcium-induced calcium release mechanism supports luteinizing hormone-induced testosterone secretion in mouse Leydig cells

Leydig cells are responsible for the synthesis and secretion of testosterone, processes controlled by luteinizing hormone (LH). Binding of LH to a G protein-coupled receptor in the plasma membrane results in an increase in cAMP and in intracellular Ca(2+) concentration ([Ca(2+)](i)). Here we show, using immunofluorescence, that Leydig cells express ryanodine receptors (RyRs) and inositol 1,4,5-trisphosphate receptors (IP(3)Rs). Measurements of intracellular calcium changes using the fluorescent calcium-sensitive dye fluo-3 and confocal microscopy show that both types of receptors are involved in a calcium-induced calcium release (CICR) mechanism, which amplifies the initial Ca(2+) influx through plasma membrane T-type calcium channels (Ca(V)3). The RyRs and IP(3)Rs are functional, as judged from both their activation by caffeine and IP(3) and block by ryanodine and 2-aminoethoxydiphenyl borate (2-APB), respectively. RyRs are the principal players involved in the release of Ca(2+) from the endoplasmic reticulum, as evidenced by the fact that global Ca(2+) changes evoked by LH are readily blocked by 100 muM ryanodine but not by 2-APB or xestospongin C. Finally, steroid production by Leydig cells is inhibited by ryanodine but not by 2-APB. These results not only broaden our understanding of the role played by calcium in Leydig cells but also show, for the first time, that RyRs have an important role in determining testosterone secretion by the testis.

A calcium‐induced calcium‐release mechanism supports Luteinizing Hormone (LH) induced testosterone secretion in mice Leydig cells

Binding of LH to a G Protein Coupled Receptor in the plasma membrane, results in a primary increase in cAMP and in the intracellular calcium concentration ([Ca2+]i in Leydig cells. Immunocytochemical labeling with specific antibodies show the presence of three distinct isoforms of both ryanodine receptors (RYRs) and inositol 1,4,5‐trisphosphate receptors (IP3Rs). Measurements of intracellular calcium activity, using the fluorescent calcium sensitive dye Fluo3 and confocal microscopy, demonstrate that these receptors are involved in a calcium induced calcium release mechanism (CICR), responsible for the amplification of the initial Ca2+ influx through plasma membrane T‐type calcium channels (CaV3), induced by LH. The RYRs and IP3Rs are functional, as judged both from their activation by caffeine and IP3 and block by ryanodine and 2‐APB, respectively. Nevertheless, the intracellular global calcium changes resulting from treatment of the cells with LH are readily blocked 100 μM ryanodine but not by 2‐APB or xestospongin C. Besides this, blockage of the ryanodine receptor inhibits both the hormone‐induced [Ca2+]i transients and steroid secretion by Leydig cells. These results not only broaden the understanding of the role played by calcium in Leydig cells but also show, for the first time, that ryanodine receptors have a crucial role in determining secretion of testosterone by the testis.Financial Support: FAPESP, CNPq

Expression of follicle-stimulating hormone and luteinising hormone binding sites in the bitch ovary during the follicular phase

In the female dog, in contrast with most mammals, the growing follicle starts to luteinise several days before ovulation. Little is known about the physiological control of the final follicular growth in this species. In order to better understand the pituitary regulation of follicular growth, specific binding sites for FSH and LH were localised and quantified by autoradiography using [(125)I]-porcine (p) gonadotrophins on ovarian sections (7 microm) from adult Beagle bitches during the follicular phase. Follicles were analysed either before the LH surge (n = 4 bitches; n = 117 follicles) or after the LH surge and before ovulation (n = 5 bitches; n = 110 follicles). FSH binding sites were specifically and homogeneously expressed at high levels on granulosa cells of all healthy follicles from the preantral stage onwards. In contrast, LH binding sites were detected homogeneously and at high levels only on granulosa cells of follicles larger than 1 mm in diameter, including luteinised follicles. Theca binding of LH (but not FSH) was also observed, but only when using high concentrations of [(125)I]-pLH. The overall incidence of atresia was 45.8% and was dependent upon follicular diameter. Quantitative analysis of labelling showed that atretic follicles had reduced levels of both FSH and LH binding sites compared with healthy follicles. In healthy follicles, levels of both FSH and LH binding sites changed with follicle diameter. Compared with other mammals, the acquisition of LH binding on canine granulosa cells occurs in smaller sized follicles relative to the size of ovulation.

Intracellular calcium changes in mice Leydig cells are dependent on calcium entry through T‐type calcium channels

Luteinizing hormone (LH) regulates testosterone synthesis in Leydig cells by inducing an intracellular increase in cAMP concentration. LH also increases the intracellular calcium concentration ([Ca2+]i), dependent on the presence of Ca2+ in the extracellular medium ([Ca2+]e) for its effect. Despite these evidences, the identity of a pathway for calcium entry has remained elusive and the relationship between cAMP and [Ca2+]i has been questioned. Here we show that mice Leydig cells do have an inward Ca2+ current carried by T-type Ca2+ channels. In 10 mm [Ca2+]e, the currents start to be activated at -60 mV, reaching maximal amplitude of 1.8 +/- 0.3 pA pF(-1) at -20 mV. Currents were not modified by Ba2+ or Sr2+, were suppressed in Ca2+-free external solution, and were blocked by 100 microm nickel or 100 microm cadmium. The Ki for Ni2+ is 2.6 microm and concentrations of Cd2+ smaller than 50 microm have a very small effect on the currents. The calcium currents displayed a window centred at -40 mV. The half-voltage (V0.5) of activation is -30.3 mV, whereas the half-voltage steady-state inactivation is -51.1 mV. The deactivation time constant (taudeactivation) is around 3 ms at -35 mV. Confocal microscopy experiments with Fluo-3 loaded cells reveal that both LH and dibutyryl-cAMP (db-cAMP) increase [Ca2+]i. The db-cAMP induced calcium increase was dependent on Ca2+ influx since it was abolished by removal of extracellular Ca2+ and by 400 microm Ni2+. [Ca2+]i increases in regions close to the plasma membrane and in the cell nucleus. Similar effects are seen when Leydig cells are depolarized by withdrawing K+ from the extracellular solution. Altogether, our studies show that Ca2+ influx through T-type Ca2+ channels in the plasma membrane of Leydig cells plays a crucial role in the intracellular calcium concentration changes that follow binding of LH to its receptor.

Effects of vitamins C and E on steroidogenic enzymes mRNA expression in polychlorinated biphenyl (Aroclor 1254) exposed adult rat Leydig cells

Polychlorinated biphenyls (PCBs) are ubiquitous and persistent environmental contaminants that disturb normal endocrine functions including gonadal functions in humans and mammals. The present study was conducted to elucidate the protective role of vitamins C and E against Aroclor 1254-induced changes in Leydig cell steroidogenic acute regulatory (StAR) protein and steroidogenic enzymes mRNA expression. Adult male rats were dosed for 30 days with daily intraperitoneal (i.p.) injection of 2 mg/kg Aroclor 1254 or vehicle (corn oil). One group of rats was treated with vitamin C (100 mg/kg bw day) while the other group was treated with vitamin E (50 mg/kg bw day) orally, simultaneously with Aroclor 1254 for 30 days. One day after the last treatment, animals were euthanized and blood was collected for the assay of serum hormones such as luteinizing hormone (LH), follicle stimulating hormone (FSH), testosterone and estradiol. The serum androgen binding protein was also estimated. Testes were quickly removed and Leydig cells were isolated in aseptic condition. Purity of Leydig cells was determined by 3β-hydroxysteroid dehydrogenase (3β-HSD) staining methods. Purified Leydig cells were used for quantification of androgen and estrogen receptors. In addition, total RNA was isolated from control and treated Leydig cells to monitor the steady-state mRNA levels by RT-PCR for StAR protein, cytochrome P 450scc, 3β-HSD and 17β-HSD. Aroclor 1254 treatment significantly reduced the serum LH, FSH, testosterone, estradiol and androgen binding protein. In addition to this, Leydig cell androgen and estrogen receptors were markedly decreased. RT-PCR analysis of StAR mRNA level did not alter Aroclor 1254 treatment while steroidogenic enzymes such as cytochrome P 450scc, 3β-HSD and 17β-HSD mRNAs were drastically decreased in Aroclor 1254 treatment. However, the simultaneous administration of vitamins C or E in Aroclor 1254-exposed rats resulted a significant restoration of all the above-mentioned parameters to the control level. These observations suggest that vitamins C and E have ameliorative role against PCBs-induced testicular Leydig cells dysfunction.

Expression of follicle stimulating hormone and luteinizing hormone receptors during follicular growth in the domestic cat ovary

In order to better understand the pituitary regulation of follicular growth in the domestic cat, follicle stimulating hormone (FSH) and luteinizing hormone (LH) receptors (R) were localized and quantified in relation to follicle diameter and atresia using in situ ligand binding on ovarian sections. Expression of FSHR was homogeneous and restricted to follicle granulosa cells from the early antral stage onwards, whereas expression of LHR was heterogeneous on theca cells of all follicles from the early antral stage onward, and homogeneous on granulosa cells of healthy follicles larger than 800 microm in diameter and in corpora lutea. LHR were also widely expressed as heterogeneous aggregates in the ovarian interstitial tissue. Atretic follicles exhibited significantly reduced levels of both FSHR and LHR on granulosa cells, compared with healthy follicles whatever the follicular diameter, whereas levels of LHR on theca cells were lower only for atretic follicles larger than 1,600 microm in diameter. In healthy follicles, levels of FSHR and LHR in all follicular compartments increased significantly with diameter. Although generally comparable to that observed in other mammals, the expression pattern of gonadotropin receptors in the cat ovary is characterized by an early acquisition of LHR on granulosa cells of growing follicles and islets of LH binding sites in the ovarian interstitial tissue.

Zero-length cross-linking of the LH receptor reveals tight interactions between extracellular and transmembrane domains during receptor activation

Luteinizing hormone (LH) secreted by the anterior pituitary is critical for normal sexual development and function. Activation of the LH receptor in target tissues involves LH binding to the extracellular domain (ECD), thereby modifying interactions between the ECD and transmembrane domain (TMD). To analyse these intramolecular changes we developed a recombinant LH/CG receptor (LHR) bearing an intramolecular cleavage site at position 316 recognized by the proprotein convertase furin (LHR-F316). Western blot studies indicated normal synthesis, glycosylation and expression of the recombinant LHR with complete cleavage into both domains, ECD-316 and 316-TMD. Affinity binding for hCG as well as basal and hormone induced cAMP production were identical to the wildtype LHR, indicating intact functional activity of the cleaved LHR. Physical interaction between the ECD and TMD was studied by zero-length cross-linking on HEK 293 cells expressing LHR-F316. No difference in cross-linking intensity between both domains was observed after ligand binding or in constitutively active receptors. Incubation of cells expressing LHR-F316 with dithiotreitol led to a shedding of the ECD-316 from the 316-TMD without increase of the basal adenylate cyclase activity. Our results show that activation of the LH receptor does not result from a switch of an inverse agonist effect of the ECD-316 to an agonist effect but is related to subtle modifications of intramolecular interactions between ECD and TMD.

Progesterone secretion by luteinizing human granulosa cells: a possible cAMP-dependent but PKA-independent mechanism involved in its regulation

The corpus luteum formed after luteinization of follicular cells secretes progesterone under the control of luteinizing hormone (LH). Binding of LH to its G-protein-coupled receptor leads to the activation of the adenylate cyclase/ cyclic AMP (cAMP)/cAMP-dependent protein kinase (PKA) signalling pathway. The identification of a new class of cAMP-binding proteins termed 'guanine nucleotide exchange factors' (cAMP-GEFs) provides a means by which changes in cAMP could yield actions that are independent of PKA. Hence, in this study, we have explored the hypothesis that steroidogenesis in luteinizing cells is mediated in both a cAMP/PKA-dependent and cAMP-dependent, but PKA-independent, manner. Human granulosa cells were isolated from follicular aspirates of women undergoing assisted conception. Luteinizing human granulosa cells were cultured for up to 3 days in the presence of human (h)LH and the adenylate cyclase activator forskolin in the added presence or absence of increasing doses of the PKA inhibitors H89 (N-[2-(4-bromocinnamylamino)ethyl] 5-isoquinoline) and PKI (myristoylated protein kinase A inhibitor amide 14-22) or the cAMP antagonist, Rp-cAMP. Agonist-stimulated progesterone secretion was inhibited in a dose-dependent manner by the PKA inhibitors and the cAMP antagonist, with decreasing sensitivity as luteinization progressed. Pretreatment of granulosa cells for 4 h with human (h)LH reduced the effectiveness of H89 in inhibiting progesterone secretion. Under basal conditions, cAMP-GEFI expression increased progressively throughout culture, and this could be further enhanced when cells were incubated with increasing doses of LH and forskolin. Furthermore, incubation of cells in the presence of increasing concentrations of the novel cAMP-GEF-specific cAMP analogue, 8 CPT-2 ME-cAMP (8-(4-chloro-phenylthio)-2'-0-methyladenosine-3',5'-cyclic monophosphate), increased progesterone secretion in a dose-dependent manner. The results show that increases in cAMP generated by LH and forskolin, in addition to activating PKA, also induce increases in cAMP-GEFI protein expression in luteinizing human granulosa cells. In addition, activation of cAMP-GEFI results in increased progesterone secretion. Hence, increases in cAMP lead to the activation of PKA-dependent, as well as PKA-independent but cAMP-dependent (via cAMP-GEFI), signalling mechanisms. Since cAMP-GEFs have the capacity to activate the mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3-K)/protein kinase B (PKB) signalling pathways, these may provide the potential mechanisms by which cAMP-dependent but PKA-independent progesterone synthesis is regulated.

Nongonadal LH receptors, their involvement in female reproductive function and a new applicable approach

Luteinising hormone (LH) and human chorionic gonadotropin (hCG) share a common receptor in gonadal cells. The receptors have also been detected in several nongonadal but reproduction-associated tissues of pigs, cattle, and other species including the uterus (myometrium, endometrium), oviduct, cervix, blood vessels, mammary gland and other tissues. The main role of LH/hCG receptors in the myometrium is stimulation of growth and hyperplasia, and relaxation of uterine motility; hCG also boosts blood flow in the uterine artery. LH and hCG can increase production of prostaglandins in the endometrium, oviduct, and blood vessels. We suggest that the preovulatory surge of LH plays an important role in controlling oviductal contractions. Awareness of LH binding to many tissues of the female reproductive tract and integration with embryonic factors may lead to the elaboration of new strategies for improved reproductive efficiency in domestic species. Mammary glands also possess LH/hCG receptors through which gonadotropins can affect the metabolism of steroid hormones and could play an inhibitory role in mammary carcinogenesis and in the growth of breast tumours. A novel approach to target and ablate carcinoma cells through LH receptors is described.

Effects of exogenous testosterone on testicular luteinizing hormone and follicle-stimulating hormone receptors during aging.

During aging, the male Japanese quail exhibits a loss of fertility, increased morphological abnormalities in the testes, and a higher incidence of Sertoli cell tumors. Although there is a coincident loss of reproductive behavior, plasma androgen levels remain high until testicular regression occurs in association with senescence. The purpose of this study was to compare mean specific binding of chicken luteinizing hormone (LH) and follicle-stimulating hormone (FSH) as a measure of testicular receptors during identified stages during aging. Males were categorized according to age (young = 9 months, middle aged = 24 months, or old = 36+ months) and sexual behavior (active or inactive). Testicular samples were collected immediately after perfusion with 4% paraformaldehyde from the following groups: young active (n = 8), young photoregressed (n = 5), young photoregressed plus testosterone implant (n = 4), middle-aged active (n = 8), middle-aged inactive (n = 4), old inactive (n = 5), and old inactive plus testosterone implant (n = 6). A crude plasma membrane fraction was prepared from the testes of each bird and an aliquot deriving from 10 mg of testicular tissue was used for binding assay. Specific binding of labeled LH or FSH was expressed as percentage of total radioactive hormone. Results showed significant (P < 0.05) age-related decreases in both FSH and LH receptor numbers. The highest FSH binding was found in young and middle-aged active males, with low binding in old inactive males. Testicular LH binding decreased during aging, with a sharp decrease in middle-aged males, which was similar to old males. Testosterone implants weakly stimulated FSH and LH binding in old males. Both LH and FSH binding decreased in photoregressed young males. However, testosterone implants stimulated increased LH binding, but did not affect FSH binding in young photoregressed males. These results provide evidence for separate regulation of testicular LH and FSH receptors, with testosterone stimulation of LH receptor, but not FSH receptor number in young males. However, during aging there appears to be a loss of this response, potentially because of the reduced efficacy of testosterone stimulation, thereby implying a diminished capacity for response with aging.

Genetic variant of luteinizing hormone: impact on gonadal steroid sex hormones in women

A common variant of the LHb subunit has a varying prevalence in various ethnic groups. The consequences of the presence of mutated luteinizing hormone (LH) concern borderline alterations in pituitary/gonadal function that could be mediated by an altered action of variant LH on gonadal steroidogenesis. A comparison of plasma concentrations of gonadal steroid sex hormones was completed in women heterozygous for variant LH and in women with the wild type of LH in three different age ranges. The sample was a randomly selected group of 177 normal women 16 to 72 years old. Variant LH was determined by immunofluorimetric methods using two combinations of monoclonal antibodies. The ratios of LH measured by the two assays indicated whether the subject was wild type homozygote, heterozygote or homozygote for the variant LHb allele. The carriers of the variant LH allele in the group of postmenopausal women showed higher serum testosterone levels than those with the wild type LH. This is in agreement with the clinical observations made previously showing a slightly higher androgenic action in the population with variant LH. No differences were detected in serum LH, FSH, epitestosterone and sex hormone binding globulin (SHBG).

Biological function of the LH receptor is associated with slow receptor rotational diffusion

The biological activity of luteinizing hormone (LH) receptors can be affected by modifications to the receptor’s amino acid sequence or by binding of hormone antagonists such as deglycosylated hCG. Here we have compared rotational diffusion of LH receptors capable of activating adenylate cyclase with that of non-functional hormone-occupied receptors at 4°C and 37°C using time-resolved phosphorescence anisotropy techniques. Binding of hCG to the rat wild-type receptor expressed on 293 cells (LHR-wt cells) or to the LH receptor on MA-10 cells produces functional receptors which exhibit rotational correlation times longer than 1000 μs. However, modification of the LH receptor by substitution of Lys583→Arg (LHR-K583R) results in a receptor that is non-functional and which has a significantly shorter rotational correlation time of 130±12 μs following binding of hCG. When these receptors are treated with deglycosylated hCG, an inactive form of hCG, the rotational correlation times for the LH receptors on LHR-wt and MA-10 cells are also shorter, namely 64±8 and 76±14 μs, respectively. Finally, a biologically active truncated form of the rat LH receptor expressed in 293 cells (LHR-t631) has slow rotational diffusion, greater than 1000 μs, when occupied by hCG and a significantly shorter rotational correlation time of 103±12 μs when occupied by deglycosylated hCG. The effects of rat LH binding to LH receptors on these various cell lines were similar to those of hCG although the magnitude of the changes in receptor rotational diffusion were less pronounced. We suggest that functional LH receptors are present in membrane complexes that exhibit slow rotational diffusion or are rotationally immobile. Shorter rotational correlation times for non-functional hormone–receptor complexes may reflect the absence of essential interactions between these complexes and other membrane proteins.

Luteinizing hormone receptors are associated with non-receptor plasma membrane proteins on bovine luteal cell membranes

Biophysical studies of the bovine luteinizing hormone (LH) receptor on luteal cell membranes suggest that this receptor may be part of a larger molecular weight structure. We have used 5-iodonaphthyl-1-azide (INA) to identify plasma membrane proteins near LH receptors on plasma membranes from bovine corpora lutea. Following binding of eosin isothiocyanate-derivatized ovine LH or human chorionic gonadotropin (hCG), five proteins with molecular weights of 71, 57, 55, 49 and 36 kDa were selectively derivatized with [ 125I]-INA following 2 h exposure at 22°C to 514 nm light. However, there was no fluorescence energy transfer between LH receptors occupied by ovine LH or hCG indicating that LH receptors were not self-associated in these membrane preparations. Together these results suggest that, following hormone binding, single copies of the LH receptor may exist in large molecular weight structures that include non-receptor proteins.