Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0043167 (pertussis)
 19,595 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Under control incubation conditions, gonadotropin-releasing hormone (GnRH) binds only a fraction of its receptors in rat-cultivated pituitary cells. Unmasking of the remaining receptors, which have been termed 'cryptic', requires drug- or peptide-induced protein kinase activation. Spontaneous masking however is not observed on pituitary cells sampled from castrated male rats, suggesting the presence of an intrinsic unmasking factor. Many endogenous factors could theoretically account for the effect. Here we attempted to identify the factor involved by taking advantage of their differential dependency upon second messengers and transduction cascades. Spontaneous unmasking of GnRH binding was found reversed by pertussis toxin (PTX), an inhibitor of alphai and alphao subunits of heterotrimeric G proteins, and by U73122, a phospholipase C (PLC) inhibitor. In contrast, desensitization of protein kinase C (PKC) or inhibition of tyrosine kinase by herbimycin were ineffective. Among endogenous pituitary factors able to unmask GnRH receptors in pituitary cells from normal male rats, as EGF, NPY or opiate peptides, only the latter were found to correspond to this transduction profile. In an attempt to characterize the pharmacology of opiate effects, naloxone (10 microM), a poorly selective opiate antagonist, restored masking of GnRH binding in cells from castrates. Only the delta antagonist naltrindole (1 microM) was able to mimick the action of naloxone. Conversely, when tested on cells from intact animals, morphine (10 microM), as well as dslet (1 microM) and met-ENK (10 nM), preferential delta agonists, but not dago and beta-endorphin or U50488 H and dynorphin, respectively micro and kappa agonists, were able to suppress masking. Among opioid peptides endogenous to the pituitary, only met-ENK was able to unmask cryptic receptors, an effect antagonized by naltrindole. We conclude that an opiate delta receptor subtype is endogenously activated in the pituitary of castrated male rats to prevent masking of GnRH binding.
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PMID:Delta opiate receptors account for the castration-induced unmasking of gonadotropin-releasing hormone binding sites in the rat pituitary. 987 2

Evidence has accumulated indicating that LHRH might behave as an autocrine/paracrine growth inhibitory factor in some peripheral tumors. However, LHRH receptors in tumor cells have not been fully characterized, so far. The present experiments were performed to analyze: 1) the messenger RNA expression; 2) the molecular size; and 3) the signal transduction pathway of LHRH receptors in prostate cancer. For these studies, the human androgen-dependent LNCaP and androgen-independent DU 145 prostate cancer cell lines were used. 1) By RT-PCR, a complementary DNA product, which hybridized with a 32P-labeled oligonucleotide probe specific for the pituitary LHRH receptor complementary DNA, was found both in LNCaP and in DU 145 cells. 2) Western blot analysis, using a monoclonal antibody raised against the human pituitary LHRH receptor, revealed the presence of a protein band of approximately 64 kDa (corresponding to the molecular mass of the pituitary receptor) in both cell lines. 3) In LNCaP and DU 145 cells, pertussis toxin completely abrogated the antiproliferative action of a LHRH agonist (LHRH-A). Moreover, LHRH-A substantially antagonized the pertussis toxin-catalyzed ADP-ribosylation of a Galpha(i) protein. Finally, LHRH-A significantly counteracted the forskolin-induced increase of intracellular cAMP levels in both cell lines. These data demonstrate that the LHRH receptor, which is present in prostate cancer cells, independently of whether they are androgen-dependent or not, corresponds to the pituitary receptor, in terms of messenger RNA expression and protein molecular size. However, at variance with the receptor of the gonadotrophs, prostate cancer LHRH receptor seems to be coupled to the Galpha(i) protein-cAMP signal transduction pathway, rather than to the Galpha(q/11)-phospholipase C signaling system. This might be responsible for the different actions of LHRH in anterior pituitary and in prostate cancer.
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PMID:The luteinizing hormone-releasing hormone receptor in human prostate cancer cells: messenger ribonucleic acid expression, molecular size, and signal transduction pathway. 1053 55

Although gonadotropin-releasing hormone agonists (GnRHa) have been used in the therapy of the endocrine-dependent cancers, their biological mechanism remained obscure. We have studied the roles of mitogen-activated protein kinase family in the antiproliferative effect of GnRHa on the Caov-3 human ovarian cancer cell line. Reverse transcription-PCR assays confirmed mRNA for GnRH receptor in Caov-3 cells. In the presence of 1 microM GnRHa, the proliferation of cells was significantly reduced to 76% of controls after 24 h, and the effect was sustained up to 4 days. Although GnRHa had no effect on the activation of the Jun N-terminal kinase (JNK), treatment of Caov-3 cells with GnRHa activated extracellular signal-regulated protein kinase (ERK), and its effect was more than that induced by GnRH. Activation of ERK by GnRHa occurred within 5 min, with the maximum occurring at 3 h and sustained until 24 h. GnRHa also activated ERK kinase (mitogen-activated protein/ERK kinase) and resulted in an increase in phosphorylation of son of sevenless (Sos), and Shc. Furthermore, we examined the mechanism by which GnRHa induced ERK activation. Both pertussis toxin (10 ng/ml), which inactivates Gi/Go proteins, and expression of a peptide derived from the carboxyl terminus of the beta-adrenergic receptor kinase I, which specifically blocks signaling mediated by the betagamma subunits of G proteins, blocked the GnRHa-induced ERK activation. Phorbol 12-myristate 13-acetate (PMA) also induced the ERK activity, but pretreatment of the cultured cells with PMA to down-regulate protein kinase C did not abolish the activation of ERK by GnRHa. Elimination of extracellular Ca2+ by EGTA also did not abolish the activation of ERK by GnRHa. To examine the role of ERK cascade in the antiproliferative effect of GnRHa, PD98059, an inhibitor of mitogen-activated protein/ERK kinase, was used. This inhibitor canceled the antiproliferative effect of GnRHa and apparently reversed the GnRH-induced dephosphorylation of the retinoblastoma protein, the hyperphosphorylation of which is a hallmark of G1-S transition in the cell cycle. These results provide evidence that GnRHa stimulation of ERK activity may be mediated by Gbetagamma protein, not by PMA-sensitive protein kinase C nor extracellular Ca2+ in the Caov-3 human ovarian cancer cell line, suggesting that this cascade may play an important role in the antiproliferative effect of GnRHa.
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PMID:Role of mitogen-activated protein kinase/extracellular signal-regulated kinase cascade in gonadotropin-releasing hormone-induced growth inhibition of a human ovarian cancer cell line. 1053 88

The agonist-bound gonadotropin-releasing hormone (GnRH) receptor engages several distinct signaling cascades, and it has recently been proposed that coupling of a single type of receptor to multiple G proteins (G(q), G(s), and G(i)) is responsible for this behavior. GnRH-dependent signaling was studied in gonadotropic alphaT3-1 cells endogenously expressing the murine receptor and in CHO-K1 (CHO#3) and COS-7 cells transfected with the human GnRH receptor cDNA. In all cell systems studied, GnRH-induced phospholipase C activation and Ca(2+) mobilization was pertussis toxin-insensitive, as was GnRH-mediated extracellular signal-regulated kinase activation. Whereas the G(i)-coupled m2 muscarinic receptor interacted with a chimeric G(s) protein (G(s)i5) containing the C-terminal five amino acids of Galpha(i2), the human GnRH receptor was unable to activate the G protein chimera. GnRH challenge of alphaT3-1, CHO#3 and of GnRH receptor-expressing COS-7 cells did not result in agonist-dependent cAMP formation. GnRH challenge of CHO#3 cells expressing a cAMP-responsive element-driven firefly luciferase did not result in increased reporter gene expression. However, coexpression of the human GnRH receptor and adenylyl cyclase I in COS-7 cells led to clearly discernible GnRH-dependent cAMP formation subsequent to GnRH-elicited rises in [Ca(2+)](i). In alphaT3-1 and CHO#3 cell membranes, addition of [alpha-(32)P]GTP azidoanilide resulted in GnRH receptor-dependent labeling of Galpha(q/11) but not of Galpha(i), Galpha(s) or Galpha(12/13) proteins. Thus, the murine and human GnRH receptors exclusively couple to G proteins of the G(q/11) family. Multiple GnRH-dependent signaling pathways are therefore initiated downstream of the receptor/G protein interface and are not indicative of a multiple G protein coupling potential of the GnRH receptor.
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PMID:Gonadotropin-releasing hormone receptor initiates multiple signaling pathways by exclusively coupling to G(q/11) proteins. 1073 55

The signaling pathway through which LHRH acts in endometrial and ovarian cancers is distinct from that in the anterior pituitary. The LHRH receptor interacts with the mitogenic signal transduction of growth factor receptors, resulting in down-regulation of expression of c-fos and proliferation. Only limited data are available on the cross-talk between LHRH receptor signaling and inhibition of mitogenic signal transduction. The present experiments were performed to analyze in endometrial and ovarian cancer cells: 1) whether mutations or splice variants of the LHRH receptor are responsible for differences in LHRH signaling, 2) the coupling of G protein subtypes to LHRH receptor, 3) the phosphotyrosine phosphatase (PTP) activation counteracting growth factor receptor tyrosine kinase activity. For these studies, the well characterized human Ishikawa and Hec-1A endometrial cancer cell lines and human EFO-21 and EFO-27 ovarian cancer cell lines were used, which express LHRH and its receptor. 1) Sequencing of the complementary DNA of the LHRH receptor from position 31 to position 1204, covering the complete coding region (position 56 to position 1042) showed that there are neither mutations nor splice variants of the LHRH receptor transcript in Ishikawa and Hec-1A endometrial cancer cells or in EFO-21 and EFO-27 ovarian cancer cells. 2) All analyzed cell lines except for the ovarian cancer cell line EFO-27 expressed both G proteins, alpha(i) and alpha(q), as shown by RT-PCR and Western blotting. In the EFO-27 cell line only G protein alpha(i), not G protein alpha(q), expression was found. Cross-linking experiments using disuccinimidyl suberate revealed that in the cell lines expressing G protein alpha(i) and G protein alpha(q), both G proteins coupled to the LHRH receptor. Inhibition of epidermal growth factor (EGF)-induced c-fos expression by LHRH, however, was mediated through pertussis toxin (PTX)-sensitive G protein alpha(i). Moreover, LHRH substantially antagonized the PTX-catalyzed ADP-ribosylation of G protein alpha(i). 3) Using a phosphotyrosine phosphatase assay based on molybdate-malachite green, treatment of quiescent EFO-21 and EFO-27 ovarian cancer cells and quiescent Ishikawa and Hec-1A endometrial cancer cells with 100 nM of the LHRH agonist triptorelin resulted in a 4-fold increase in PTP activity (P < 0.001). This effect was completely blocked by simultaneous treatment with PTX, supporting the concept of mediation through G protein alpha(i). As shown by quantitative Western blotting, EGF-induced tyrosine autophosphorylation of EGF receptors was reduced 45-63% after LHRH (100 nM) treatment (P < 0.001). This effect was completely blocked using the PTP inhibitor vanadate (P < 0.001). These results demonstrate that mutations or splice variants of the LHRH receptor in human endometrial and ovarian cancer cells are not responsible for the different signal transduction compared with that in pituitary gonadotrophs. We provide evidence that the tumor LHRH receptor couples to multiple G proteins, but the antiproliferative signal transduction is mediated through the PTX-sensitive G protein alpha(i). The tumor LHRH receptor activates a PTP counteracting EGF-induced tyrosine autophosphorylation of EGF receptor, resulting in down-regulation of mitogenic signal transduction and cell proliferation.
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PMID:Antiproliferative signaling of luteinizing hormone-releasing hormone in human endometrial and ovarian cancer cells through G protein alpha(I)-mediated activation of phosphotyrosine phosphatase. 1135 84

The activation of pituitary GABA(B) receptors by the specific agonist baclofen inhibits pituitary hormone secretion in vitro. Here we studied the mechanism of action of GABA(B) receptors in rat adenohypophysis. Anterior pituitary cells were obtained by trypsinization and were either plated for hormonal studies and cAMP determination or incubated in FURA 2AM for calcium measurements. Baclofen (BACL: 1 x 10(-5) M) significantly inhibited basal and thyrotropic releasing hormone (TRH)-stimulated (1 x 10(-7) M) PRL secretion in anterior pituitary cells from proestrous rats. In the presence of pertussis toxin (PTX: 150 ng/ml, 20 h), which leads to the uncoupling of the G(i/o)-protein from the receptor, both effects of BACL were abolished while the effect of dopamine (DA: 1 x 10(-8) M), used as an inhibitory control, was reduced from 70 to 25%. PTX also reversed BACL-induced inhibition of gonadotropin-releasing hormone (GnRH)-elicited luteinizing hormone (LH) secretion in anterior pituitary cells from 15-day-old female rats. In addition, though working in a pituitary mixed cell population, in which only some cell types possess GABA(B) receptors, BACL (1 x 10(-5) M) attenuated the forskolin-induced (0.5 microM) increase in cAMP. This effect was prevented by co-incubation with the antagonist 2 hydroxysaclofen and by preincubation with PTX. BACL (5 x 10(-5) M) and DA (5 x 10(-7) M) inhibited basal intracellular calcium concentrations ([Ca(2+)](i)) in pituitary cells and the effect of the latter was significantly stronger. The effect of BACL on [Ca(2+)](i) was abolished after preincubation with PTX. In the presence of the potassium channel blocking agents barium (200 microM and 1 mM) and tetraethylammonium (10 mM), BACL was still able to inhibit [Ca(2+)](i). Blockade of voltage-sensitive calcium channels (VSCC) with either verapamil (5 x 10(-6) M) or nifedipine (1 x 10(-6) M) completely abolished the effect of BACL on [Ca(2+)](i). In the presence of 12.5 mM potassium concentration baclofen significantly inhibited [Ca(2+)](i). In conclusion, our results describe the negative coupling of adenohypophyseal GABA(B) receptors to VSCC through PTX-sensitive G-proteins. These characteristics suggest a resemblance of these receptors to the typical presynaptic GABA(B) sites described in the central nervous system.
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PMID:GABA(B) receptors in anterior pituitary cells. Mechanism of action coupled to endocrine effects. 1139 6

The hypothalamic hormone gonadotropin-releasing hormone (GnRH) stimulates the synthesis and release of the pituitary gonadotropins. GnRH acts through a plasma membrane receptor that is a member of the G protein-coupled receptor (GPCR) family. These receptors interact with heterotrimeric G proteins to initiate downstream signaling. In this study, we have investigated which G proteins are involved in GnRH receptor-mediated signaling in L beta T2 pituitary gonadotrope cells. We have shown previously that GnRH activates ERK and induces the c-fos and LH beta genes in these cells. Signaling via the G(i) subfamily of G proteins was excluded, as neither ERK activation nor c-Fos and LH beta induction was impaired by treatment with pertussis toxin or a cell-permeable peptide that sequesters G beta gamma-subunits. GnRH signaling was partially mimicked by adenoviral expression of a constitutively active mutant of G alpha(q) (Q209L) and was blocked by a cell-permeable peptide that uncouples G alpha(q) from GPCRs. Furthermore, chronic activation of G alpha(q) signaling induced a state of GnRH resistance. A cell-permeable peptide that uncouples G alpha(s) from receptors was also able to inhibit ERK, c-Fos, and LH beta, indicating that both G(q/11) and G(s) proteins are involved in signaling. Consistent with this, GnRH caused GTP loading on G(s) and G(q/11) and increased intracellular cAMP. Artificial elevation of cAMP with forskolin activated ERK and caused a partial induction of c-Fos. Finally, treatment of G alpha(q) (Q209L)-infected cells with forskolin enhanced the induction of c-Fos showing that the two pathways are independent and additive. Taken together, these results indicate that the GnRH receptor activates both G(q) and G(s) signaling to regulate gene expression in L beta T2 cells.
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PMID:Involvement of both G(q/11) and G(s) proteins in gonadotropin-releasing hormone receptor-mediated signaling in L beta T2 cells. 1205 Jan 61

The involvement of G proteins in the transduction mechanism of M current (Im) inhibition by extracellular ligands in bullfrog sympathetic neurons was examined using the hydrolysis resistant nucleotide analogues GTPgammaS and GDPbetaS. Im was recorded in large (40 - 60 microm) isolated neurons using the patch-clamp technique in the whole-cell configuration, as well as in neurons from the intact ganglion impaled with conventional microelectrodes. In whole-cell recordings Im could be recorded without significant loss for 1 h or more provided ATP was present in the patch pipette. Muscarine, D-Ala6-LHRH, substance P and UTP reversibly inhibited Im in isolated control neurons, with full and rapid recovery of the current following agonist washout. Dialysis of isolated neurons with various concentrations of GTPgammaS (1 - 100 microM) affected, in a dose-dependent manner, the recovery of Im after its inhibition by brief agonist application. With 50 microM GTPgammaS, Im inhibition became completely irreversible. Similarly, the reversibility of Im inhibition by muscarine was reduced or abolished by the iontophoretic injection of GTPgammaS through a second microelectrode into neurons of the intact ganglion. GTPgammaS by itself caused a slow, agonist-independent suppression of Im in dialysed neurons, thus mimicking agonist action. Dialysis of isolated neurons with GDPbetaS (100 - 500 microM) attenuated by half or more the magnitude of Im inhibition by agonist as compared to control neurons. In addition, GDPbetaS attenuated the response of a given neuron to muscarine and D-Ala6-LHRH, and caused slow increase of Im, as a function of dialysis time. Incubation (2 - 72 h, 4 - 36 degrees C) of isolated neurons or intact ganglions with activated pertussis toxin had no effect on the response to muscarine. Toxin injections to experimental animals were equally ineffective. In contrast to Im, the additional inward current with increase in conductance induced by muscarine and D-Ala6-LHRH reversed with agonist washout in GTPgammaS-dialysed neurons, although more slowly than in control neurons. The results in this study indicate that a G protein, possibly pertussis toxin-insensitive, provides a common coupling step linking muscarinic, substance P, D-Ala6-LHRH and UTP receptors to the inhibition of M current.
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PMID:A G Protein Mediates the Inhibition of the Voltage-Dependent Potassium M Current by Muscarine, LHRH, Substance P and UTP in Bullfrog Sympathetic Neurons. 1210 39

The pulsatile secretion of gonadotropin-releasing hormone (GnRH) from normal and immortalized hypothalamic GnRH neurons is highly calcium-dependent and is stimulated by cAMP. It is also influenced by agonist activation of the endogenous GnRH receptor (GnRH-R), which couples to G(q/11) as indicated by release of membrane-bound alpha(q/11) subunits and increased inositol phosphate/Ca(2+) signaling. Conversely, GnRH antagonists increase membrane-associated alpha(q/11) subunits and abolish pulsatile GnRH secretion. GnRH also stimulates cAMP production but at high concentrations has a pertussis toxin-sensitive inhibitory effect, indicative of receptor coupling to G(i). Coupling of the agonist-activated GnRH-R to both G(s) and G(i) proteins was demonstrated by the ability of nanomolar GnRH concentrations to reduce membrane-associated alpha(s) and alpha(i3) levels and of higher concentrations to diminish alpha(i3) levels. Conversely, alpha(i3) was increased during GnRH antagonist and pertussis toxin treatment, with concomitant loss of pulsatile GnRH secretion. In cholera toxin-treated GnRH neurons, decreases in alpha(s) immunoreactivity and increases in cAMP production paralleled the responses to nanomolar GnRH concentrations. Treatment with cholera toxin and 8-bromo-cAMP amplified episodic GnRH pulses but did not affect their frequency. These findings suggest that an agonist concentration-dependent switch in coupling of the GnRH-R between specific G proteins modulates neuronal Ca(2+) signaling via G(s)-cAMP stimulatory and G(i)-cAMP inhibitory mechanisms. Activation of G(i) may also inhibit GnRH neuronal function and episodic secretion by regulating membrane ion currents. This autocrine mechanism could serve as a timer to determine the frequency of pulsatile GnRH release by regulating Ca(2+)- and cAMP-dependent signaling and GnRH neuronal firing.
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PMID:An agonist-induced switch in G protein coupling of the gonadotropin-releasing hormone receptor regulates pulsatile neuropeptide secretion. 1259 45

Neonatal pituitary cells express MT1 and MT2 subtype of melatonin receptors that are coupled to pertussis toxin-sensitive G proteins. Their activation by melatonin leads to a decrease in cAMP production and activity of protein kinase A, and attenuation of gonadotropin-releasing hormone (GnRH)-induced gonadotropin secretion. Single cell calcium and electrophysiological recordings have revealed that a reduction in gonadotropin release results from melatonin-induced inhibition of GnRH-stimulated calcium signaling. Melatonin inhibits both calcium influx through voltage-dependent calcium channels and calcium mobilization from intracellular stores. Inhibition of calcium influx, probably in a cAMP/protein kinase C-dependent manner, and the accompanying calcium-induced calcium release from ryanodine-sensitive intracellular pools by melatonin results in a delay of GnRH-induced calcium signaling. Melatonin-induced attenuation of GnRH-induced and inositol (1,4,5)-trisphosphate-mediated calcium release from intracellular pools attenuates the amplitude of calcium signal. The potent inhibition of GnRH-induced calcium signaling and gonadotropin secretion by melatonin provides an effective mechanism to protect premature initiation of pubertal changes that are dependent on plasma gonadotropin levels. During the development, such tonic inhibitory effects of melatonin on GnRH action gradually decline due to a decrease in expression of functional melatonin receptors. In adult animals, melatonin does not have obvious direct effects on pituitary functions, whereas the connections between melatonin release and hypothalamic functions, including GnRH release, are preserved, and are critically important in synchronizing the external photoperiods and reproductive functions through still not well characterized mechanisms.
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PMID:Melatonin action in neonatal gonadotrophs. 1511 46


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