Mass, Spectroscopy, and Two-Neutron Decay of

The structure and decay of the most neutron-rich beryllium isotope, ^{16}Be, has been investigated following proton knockout from a high-energy ^{17}B beam. Two relatively narrow resonances were observed for the first time, with energies of 0.84(3) and 2.15(5) MeV above the two-neutron decay threshold and widths of 0.32(8) and 0.95(15) MeV, respectively. These were assigned to be the ground (J^{π}=0^{+}) and first excited (2^{+}) state, with E_{x}=1.31(6) MeV. The mass excess of ^{16}Be was thus deduced to be 56.93(13) MeV, some 0.5 MeV more bound than the only previous measurement. Both states were observed to decay by direct two-neutron emission. Calculations incorporating the evolution of the wave function during the decay as a genuine three-body process reproduced the principal characteristics of the neutron-neutron energy spectra for both levels, indicating that the ground state exhibits a strong spatially compact dineutron component, while the 2^{+} level presents a far more diffuse neutron-neutron distribution.

Quasifree Neutron Knockout Reaction Reveals a Small s-Orbital Component in the Borromean Nucleus

A kinematically complete quasifree (p,pn) experiment in inverse kinematics was performed to study the structure of the Borromean nucleus ^{17}B, which had long been considered to have a neutron halo. By analyzing the momentum distributions and exclusive cross sections, we obtained the spectroscopic factors for 1s_{1/2} and 0d_{5/2} orbitals, and a surprisingly small percentage of 9(2)% was determined for 1s_{1/2}. Our finding of such a small 1s_{1/2} component and the halo features reported in prior experiments can be explained by the deformed relativistic Hartree-Bogoliubov theory in continuum, revealing a definite but not dominant neutron halo in ^{17}B. The present work gives the smallest s- or p-orbital component among known nuclei exhibiting halo features and implies that the dominant occupation of s or p orbitals is not a prerequisite for the occurrence of a neutron halo.

Surface Localization of the Dineutron in

The formation of a dineutron in the ^{11}Li nucleus is found to be localized to the surface region. The experiment measured the intrinsic momentum of the struck neutron in ^{11}Li via the (p,pn) knockout reaction at 246 MeV/nucleon. The correlation angle between the two neutrons is, for the first time, measured as a function of the intrinsic neutron momentum. A comparison with reaction calculations reveals the localization of the dineutron at r∼3.6 fm. The results also support the density dependence of dineutron formation as deduced from Hartree-Fock-Bogoliubov calculations for nuclear matter.

Application of inorganic phosphate limitation to efficient isoprene production in Pantoea ananatis.

AIMS: Establishment of an efficient isoprene fermentation process by adopting inorganic phosphate limitation as the trigger to direct metabolic flux to the isoprene synthetic pathway. METHODS AND RESULTS: We constructed isoprene-producing strains of Pantoea ananatis (a member of the Enterobacteriaceae family) by integrating a heterologous mevalonate pathway and a metabolic switch that senses external inorganic phosphate (Pi) levels. This metabolic switch enabled dual-phase isoprene production, where the initial cell growth phase under Pi-saturating conditions was uncoupled from the subsequent isoprene production phase under Pi-limiting conditions. In fed-batch fermentation using our best strain (SWITCH-PphoC/pIspSM) in a 1-l bioreactor, isoprene concentration in the off-gas was maintained between 300 and 460 ppm during the production phase and at 20 ppm during the cell growth phase, respectively. The strain SWITCH-PphoC/pIspSM produced totally 2·5 g l-1 of isoprene from glucose with a 1·8% volumetric yield in 48 h. CONCLUSIONS: This proof-of-concept study demonstrated that our Pi-dependent dual-phase production system using a P. ananatis strain as a producer has potential for industrial-scale isoprene fermentation. SIGNIFICANCE AND IMPACT OF THE STUDY: This Pi-dependent dual-phase fermentation process could be an attractive and economically viable option for the production of various commercially valuable isoprenoids.

Competitive networks using chaotic circuits with hierarchical structure.

Coupled oscillatory systems are good models that are able to describe a variety of higher dimensional nonlinear phenomena. Coupled chaotic circuits produce many kinds of interesting synchronization phenomena. In recent years, research studies on complex networks related to the synchronization of coupled oscillators have attracted much attention. In the real world, there are a variety of different network structures. We focus on the competitive interaction network that includes conflict between two networks. Here, we propose a new paradigm for this competitive interaction network using coupled chaotic circuits.

Estradiol and raloxifene induce the proliferation of osteoblasts through G-protein-coupled receptor GPR30.

BACKGROUND: Although G-protein-coupled receptor, GPR30, has been considered as a G-protein-coupled estrogen receptor, conflicting results have been reported and the function of GPR30 in bone remains unresolved. The aim of this study was to clarify the functional role of GPR30 in osteoblasts using its derived cell line. METHODS AND RESULTS: Immunohistochemical study revealed that GPR30 is expressed in human osteoblasts. Human fetal osteoblast cell lines, hFOB cells, which express GPR30 but lack estrogen receptor, were used for the in vitro experiments. Estradiol or raloxifene induced the proliferation of hFOB cells, which was accompanied by the activation of mitogen-activated protein (MAP) kinase. Those proliferative effects were completely abrogated by the transfection of GPR30 small interfering RNA, while the transfection alone did not affect the cell viability. CONCLUSION: GPR30 is required for the proliferation of hFOB cells induced by estradiol or raloxifene. This proliferative effect was at least partly mediated via MAP kinase activation. These findings revealed a novel function of GPR30 in osteoblasts and might lead to a better understanding of how estrogen and selective estrogen receptor modulators show their osteoprotective effects.

Counting-backward test for executive function in idiopathic normal pressure hydrocephalus.

OBJECTIVES: The aim of this study was to develop and validate a bedside test for executive function in patients with idiopathic normal pressure hydrocephalus (INPH). MATERIALS AND METHODS: Twenty consecutive patients with INPH and 20 patients with Alzheimer's disease (AD) were enrolled in this study. We developed the counting-backward test for evaluating executive function in patients with INPH. Two indices that are considered to be reflective of the attention deficits and response suppression underlying executive dysfunction in INPH were calculated: the first-error score and the reverse-effect index. Performance on both the counting-backward test and standard neuropsychological tests for executive function was assessed in INPH and AD patients. RESULTS: The first-error score, reverse-effect index and the scores from the standard neuropsychological tests for executive function were significantly lower for individuals in the INPH group than in the AD group. The two indices for the counting-backward test in the INPH group were strongly correlated with the total scores for Frontal Assessment Battery and Phonemic Verbal Fluency. The first-error score was also significantly correlated with the error rate of the Stroop colour-word test and the score of the go/no-go test. In addition, we found that the first-error score highly distinguished patients with INPH from those with AD using these tests. CONCLUSION: The counting-backward test is useful for evaluating executive dysfunction in INPH and for differentiating between INPH and AD patients. In particular, the first-error score may reflect deficits in the response suppression related to executive dysfunction in INPH.

Pharmacokinetics of cyclosporine A at a high-peak concentration of twice-daily infusion and oral administration in allogeneic haematopoietic stem cell transplantation.

WHAT IS KNOWN AND OBJECTIVE: The most appropriate immunosuppressive strategy with calcineurin inhibitors for the prevention of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation (alloHSCT) has not yet been established. To estimate the safety and efficacy of a new strategy, we investigated the pharmacokinetics of cyclosporine A (CyA) delivered by twice-daily infusion and oral administration maintained with a peak level above 1000 ng/mL to keep 24 h area under the concentration-time curve (AUC0-24) higher than 10 000 ng·h/mL in 12 patients. METHODS: Cyclosporine A was started as a twice-daily infusion at 1·5 mg/kg and then orally administered at twice the infusion dose to maintain the trough blood concentration between 200 and 500 ng/mL, and with a peak level above 1000 ng/mL. Serial blood samples were collected at 0, 1, 2, 3, 5, 8 and 12 h after CyA dosing (C0, C1, C2, C3, C5, C8 and C12) on days 14-21 after transplantation and on days 7-14 after switching to oral administration, and the AUC was calculated. RESULTS: In all patients, the AUC0-24 for both twice-daily infusion and oral administration was higher than 10 000 ng·h/mL. Two close relationships were observed between AUC0-12 and the C3 for infusion and between AUC0-12 and the C8 for oral administration. None of the patients had grades 3-4 aGVHD or other serious complications. WHAT IS NEW AND CONCLUSION: This strategy was well tolerated, and the C3 for twice-daily infusion and the C8 for oral administration were the optimal points for monitoring of CyA concentration in the early phase of transplantation.

Transcription factor AP-2beta inhibits expression and secretion of leptin, an insulin-sensitizing hormone, in 3T3-L1 adipocytes.

BACKGROUND: We have previously reported an association between the activator protein-2beta (AP-2beta) transcription factor gene and type 2 diabetes. This gene is preferentially expressed in adipose tissue, and subjects with a disease-susceptible allele of AP-2beta showed stronger AP-2beta expression in adipose tissue than those without the susceptible allele. Furthermore, overexpression of AP-2beta led to lipid accumulation and induced insulin resistance in 3T3-L1 adipocytes. RESULT: We found that overexpression of AP-2beta in 3T3-L1 adipocytes decreased the promoter activity of leptin, and subsequently decreased both messenger RNA (mRNA) and protein expression and secretion. Furthermore, knockdown of endogenous AP-2beta by RNA-interference increased mRNA and protein expression of leptin. Electrophoretic mobility shift and chromatin immunoprecipitation assays revealed specific binding of AP-2beta to leptin promoter regions in vitro and in vivo. In addition, site-directed mutagenesis of the AP-2-binding site located between position +34 and +42 relative to the transcription start site abolished the inhibitory effect of AP-2beta. Our results clearly showed that AP-2beta directly inhibited insulin-sensitizing hormone leptin expression by binding to its promoter. CONCLUSION: AP-2beta modulated the expression of leptin through direct interaction with its promoter region.

Corticolimbic gray matter loss in Parkinson's disease without dementia.

BACKGROUND: The relationship between corticolimbic involvement and cognitive dysfunction in non-demented Parkinson's disease (PD) patients has not yet been elucidated. OBJECTIVES: To delineate involvement of the cerebral cortex and limbic structures in non-demented PD and to clarify distributional differences of gray matter loss between non-demented PD with impaired cognition (PD-CI) and without cognitive impairment (PD-NC). METHODS: Operational criteria based on the Clinical Dementia Rating were used to identify PD-CI. Of 40 consecutive non-demented patients with PD, 13 were classified as PD-CI and 27 as PD-NC. Comparisons of regional gray matter volume (rGMV) were made amongst the PD-CI, PD-NC, and control groups using voxel-based morphometry. RESULTS: Gray matter loss was found extensively in the frontal, temporal, parietal, and occipital cortices in the present non-demented patients with PD. rGMV in the medial frontal and medial occipital cortices was reduced comparably in the PD-NC and PD-CI groups. The severity of gray matter loss in the perisylvian cortices increased in order from the control, to the PD-NC, to the PD-CI groups. rGMV reduction in the lateral and orbital frontal, medial and lateral temporal, medial and lateral parietal, and lateral occipital cortices and cerebellum was found specifically in PD-CI. CONCLUSIONS: Our results suggest that corticolimbic degeneration occurs in non-demented patients with PD, and extensive involvement of the limbic and posterior cortical regions as well as the frontal cortices is associated with cognitive impairment in PD.

L-isoleucine and L-leucine: tumor promoters of bladder cancer in rats.

A 4-week assay for screening tumor promoters of bladder cancer has been developed in which increased agglutinability of isolated rat bladder cells with concanavalin A is used as an indicator. On the basis of this assay system, L-isoleucine and L-leucine were suspected of being possible tumor promoters. Results of 40- to 60-week carcinogenesis experiments in which N-butyl-N-(4-hydroxybutyl)nitrosamine was used as an initiator demonstrate that L-isoleucine and L-leucine promote bladder cancer in rats. This finding may be relevant to the high incidence of human bladder cancer in Western countries, where the diet is rich in protein.

Identification and characterization of a gene regulating enzymatic glycosylation which is induced by diabetes and hyperglycemia specifically in rat cardiac tissue.

Primary cardiac abnormalities have been frequently reported in patients with diabetes probably due to metabolic consequences of the disease. Approximately 2,000 mRNA species from the heart of streptozotocin-induced diabetic and control rats were compared by the mRNA differential display method, two of eight candidate clones thus isolated (DH1 and 13) were confirmed by Northern blot analysis. The expression of clone 13 was increased in the heart by 3.5-fold (P < 0.05) and decreased in the aorta by twofold (P < 0.05) in diabetes as compared to control. Sequence analysis showed that clone 13 is a rat mitochondrial gene. DH1 was predominantly expressed in the heart with an expression level 6.8-fold higher in the diabetic rats than in control (P < 0.001). Insulin treatment significantly (P < 0.001) normalized the expression of DH1 in the hearts of diabetic rats. DH1 expression was observed in cultured rat cardiomyocytes, but not in aortic smooth muscle cells or in cardiac derived fibroblasts. The expression in cardiomyocytes was regulated by insulin and glucose concentration of culture media. The full length cDNA of DH1 had a single open-reading frame with 85 and 92% amino acid identity to human and mouse UDP-GlcNAc:Gal beta 1-3GalNAc alpha R beta 1-6 N-acetylglucosaminyltransferase (core 2 GlcNAc-T), respectively, a key enzyme determining the structure of O-linked glycosylation. Transient transfection of DH1 cDNA into Cos7 cells conferred core 2 GlcNAc-T enzyme activity. In vivo, core 2 GlcNAc-T activity was increased by 82% (P < 0.05) in diabetic hearts vs controls, while the enzymes GlcNAc-TI and GlcNAc-TV responsible for N-linked glycosylation were unchanged. These results suggest that core 2 GlcNAc-T is specifically induced in the heart by diabetes or hyperglycemia. The induction of this enzyme may be responsible for the increase in the deposition of glycoconjugates and the abnormal functions found in the hearts of diabetic rats.

A nuclear factor for interleukin-6 expression (NF-IL6) and the glucocorticoid receptor synergistically activate transcription of the rat alpha 1-acid glycoprotein gene via direct protein-protein interaction.

The acute-phase reaction is accompanied by an increase in a variety of serum proteins, named acute-phase proteins. The synthesis of these proteins is synergistically controlled by glucocorticoids and inflammatory cytokines such as interleukin-1 (IL-1), IL-6, and tumor necrosis factor alpha. Recently, we have cloned nuclear factor-IL-6 (NF-IL6), a transcription factor that activates the IL-6 gene, and have demonstrated its involvement in the expression of acute-phase-protein genes. We report here an analysis of the molecular mechanisms by which inflammatory cytokines and glucocorticoid act synergistically to activate expression of the rat alpha 1-acid glycoprotein (AGP) gene. We found that NF-IL6 and ligand-activated rat glucocorticoid receptor acted synergistically to transactivate the AGP gene and that maximal transcriptional activation of the AGP gene required expression of both intact NF-IL6 and rat glucocorticoid receptor. Surprisingly, however, transcriptional synergism was still observed even when one of the two factors lacked either its DNA-binding or transcriptional-activation function. We present evidence for a direct protein-protein interaction between these two distinct transcription factors and propose that this may be responsible for the synergistic activation of the rat AGP gene.

Glucocorticoid-stimulated CCAAT/enhancer-binding protein alpha expression is required for steroid-induced G1 cell cycle arrest of minimal-deviation rat hepatoma cells.

By genetic correlation with the growth-suppressible phenotype and direct functional tests, we demonstrate that the glucocorticoid-stimulated expression of the CCAAT/enhancer-binding protein alpha (C/EBP alpha) transcription factor is required for the steroid-mediated G1 cell cycle arrest of minimal-deviation rat hepatoma cells. Comparison of C/EBP alpha transcript and active protein levels induced by the synthetic glucocorticoid dexamethasone in glucocorticoid growth-suppressible (BDS1), nonsuppressible receptor-positive (EDR1) and nonsuppressible receptor-deficient (EDR3) hepatoma cell proliferative variants revealed that the stimulation of C/EBP alpha expression is a rapid, glucocorticoid receptor-mediated response associated with the G1 cell cycle arrest. Consistent with the role of C/EBP alpha as a critical intermediate in the growth suppression response, maximal induction of transcription factor mRNA occurred within 2 h of dexamethasone treatment whereas maximal inhibition of [3H] thymidine incorporation was observed 24 h after steroid treatment. As a direct functional approach, ablation of C/EBP alpha protein expression and DNA-binding activity by transfection of an antisense C/EBP alpha expression vector blocked the dexamethasone-induced G1 cell cycle arrest of hepatoma cells but did not alter general glucocorticoid responsiveness. Transforming growth factor beta induced a G1 cell cycle arrest in C/EBP alpha antisense transfected cells, demonstrating the specific involvement of C/EBP alpha in the glucocorticoid growth suppression response. Constitutive expression of a conditionally activated form of C/EBP alpha caused a G1 cell cycle arrest of BDS1 hepatoma cells in the absence of glucocorticoids. In contrast, overexpression of C/EBP beta or C/EBP delta had no effect on hepatoma cell growth. Taken together, these results demonstrate that the steroid-induced expression of C/EBP alpha is necessary to mediate the glucocorticoid G1 cell cycle arrest of rat hepatoma cells and implicates a role for this transcription factor in the growth control of liver-derived epithelial tumor cells.

Crystallization and vitrification of electrons in a glass-forming charge liquid.

Charge ordering (CO) is a phenomenon in which electrons in solids crystallize into a periodic pattern of charge-rich and charge-poor sites owing to strong electron correlations. This usually results in long-range order. In geometrically frustrated systems, however, a glassy electronic state without long-range CO has been observed. We found that a charge-ordered organic material with an isosceles triangular lattice shows charge dynamics associated with crystallization and vitrification of electrons, which can be understood in the context of an energy landscape arising from the degeneracy of various CO patterns. The dynamics suggest that the same nucleation and growth processes that characterize conventional glass-forming liquids guide the crystallization of electrons. These similarities may provide insight into our understanding of the liquid-glass transition.

Actions anchored by concepts: defective action comprehension in semantic dementia.

OBJECTIVE: To study the ability of patients with semantic dementia to understand actions, in order to examine the contribution of semantic memory to action comprehension. METHODS: The ability to comprehend symbolic and instrumental actions was assessed in 6 patients with semantic dementia and 10 healthy controls. The patients were also given the imitation test of meaningful and meaningless actions. RESULTS: In all patients with semantic dementia, comprehension of both symbolic and instrumental actions was defective. The comprehension of symbolic actions was more impaired than that of instrumental actions. Their ability to imitate other's actions was well preserved. CONCLUSION: This study showed that comprehension of action was impaired in semantic dementia, suggesting that semantic memory has an important role in comprehension of human action.

Oral administration of tetrahydrobiopterin prevents endothelial dysfunction and vascular oxidative stress in the aortas of insulin-resistant rats.

We have reported that a deficiency of tetrahydrobiopterin (BH(4)), an active cofactor of endothelial NO synthase (eNOS), contributes to the endothelial dysfunction through reduced eNOS activity and increased superoxide anion (O(2)(-)) generation in the insulin-resistant state. To further confirm this hypothesis, we investigated the effects of dietary treatment with BH(4) on endothelium-dependent arterial relaxation and vascular oxidative stress in the aortas of insulin-resistant rats. Oral supplementation of BH(4) (10 mg. kg(-1). d(-1)) for 8 weeks significantly increased the BH(4) content in cardiovascular tissues of rats fed high levels of fructose (fructose-fed rats). Impairment of endothelium-dependent arterial relaxation in the aortic strips of the fructose-fed rats was reversed with BH(4) treatment. The BH(4) treatment was associated with a 2-fold increase in eNOS activity as well as a 70% reduction in endothelial O(2)(-) production compared with those in fructose-fed rats. The BH(4) treatment also partially improved the insulin sensitivity and blood pressure, as well as the serum triglyceride concentration, in the fructose-fed rats. Moreover, BH(4) treatment of the fructose-fed rats markedly reduced the lipid peroxide content of both aortic and cardiac tissues and inhibited the activation of 2 redox-sensitive transcription factors, nuclear factor-kappaB and activating protein-1, which were increased in fructose-fed rats. The BH(4) treatment of control rats did not have any significant effects on these parameters. These results indicate that BH(4) augmentation is essential for the restoration of eNOS function and the reduction of vascular oxidative stress in insulin-resistant rats.

Sequential changes of mouse bladder epithelium during induction of invasive carcinomas by N-butyl-N-(4-hydroxybutyl)nitrosamine.

Invasive carcinoma of the bladder in humans shows aggressive growth with poor prognosis. Little is known about its preceding lesions. Sequential changes of the bladder epithelium following administration of N-butyl-N-(4-hydroxybutyl)nitrosamine (BHBN) were studied in mice. Female C3H/He mice were divided into 4 groups. Three groups were given 0.05, 0.01, and 0.005% concentrations of BHBN, respectively, in their drinking water, and the control group was given tap water. The mice were killed at regular intervals over a period of 26 weeks, and their bladder epithelium was examined histologically. Dysplasia, carcinoma in situ, and invasive carcinoma were observed sequentially in the groups treated with BHBN, and the incidences of dysplasia, carcinoma in situ, and invasive carcinoma were dependent on the dose of BHBN. The data indicate that bladder carcinoma in mice is a good model of invasive bladder carcinoma in humans, although it is not fully compatible with the human model because of the complete absence of metastases.

Estrogen activates telomerase.

Telomerase activity is present in most malignant tumors and provides a mechanism for the unlimited potential for division of neoplastic cells. Although telomerase is known to be a regulated enzyme, the factors and mechanisms involved in telomerase regulation are not well understood. In the present study, we examined the effects of estrogen on telomerase activity. Telomerase activity in estrogen receptor (ER)-positive MCF-7 cells was up-regulated by the treatment with 17beta-estradiol. This activation accompanied up-regulation of the telomerase catalytic subunit, hTERT mRNA. Gel shift assays revealed that the imperfect palindromic estrogen-responsive element in the hTERT promoter specifically binds to ER. Transient expression assays using luciferase reporter plasmids containing various fragments of hTERT promoter showed that this imperfect palindromic estrogen-responsive element is responsible for transcriptional activation by ligand-activated ER. We also found that estrogen activates c-Myc expression in MCF-7 cells and that E-boxes in the hTERT promoter that bind c-Myc/Max play additional roles in estrogen-induced transactivation of hTERT. Estrogen thus activates telomerase via direct and indirect effects on the hTERT promoter. These findings may help elucidate the mechanisms of hormonal control of telomerase activity and aid understanding of the roles of sex steroids in cellular senescence and aging as well as estrogen-induced carcinogenesis.

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.

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.