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Biomedical subjects

E Haug

Publications and source records attributed to E Haug.

At least 127 records · Page 7Linked to original sources
PubMed1989

Protein kinase C stimulates adenylate cyclase activity in prolactin-secreting rat adenoma (GH4C1) pituicytes by inactivating the inhibitory GTP-binding protein Gi.

The phorbol ester 12-O-tetradecanoyl-phorbol 13-acetate (TPA) and thyroliberin exerted additive stimulatory effects on prolactin release and synthesis in rat adenoma GH4C1 pituicytes in culture. Both TPA and thyroliberin activated the adenylate cyclase in broken cell membranes. When combined, the secretagogues displayed additive effects. TPA did not alter the time course (time lag) of adenylate cyclase activation by hormones, guanosine 5'-[beta,gamma-imino]triphosphate or forskolin, nor did it affect the enzyme's apparent affinity (basal, 7.2 mM; thyroliberin-enhanced, 2.2 mM) for free Mg2+. The TPA-mediated adenylate cyclase activation was entirely dependent on exogenously added guanosine triphosphate. ED50 (dose yielding half-maximal activation) was 60 microM. Access to free Ca2+ was necessary to express TPA activation of the enzyme, however, the presence of calmodulin was not mandatory. TPA-stimulated adenylate cyclase activity was abolished by the biologically inactive phorbol ester, 4 alpha-phorbol didecanoate, by the protein kinase C inhibitor polymyxin B and by pertussis toxin, while thyroliberin-sensitive adenylate cyclase remained unaffected. Experimental conditions known to translocate protein kinase C to the plasma membrane and without inducing adenylate cyclase desensitization, increased both basal and thyroliberin-stimulated enzyme activities, while absolute TPA-enhanced adenylate cyclase was maintained. Association of extracted GTP-binding inhibitory protein, Gi, from S49 cyc- murine lymphoma cells with GH4C1 cell membranes yielded a reduction of basal and hormone-stimulated adenylate cyclase activities, while net inhibition of the cyclase of somatostatin was dramatically enhanced. However, TPA restored completely basal and hormone-elicited adenylate cyclase activities in the Gi-enriched membranes. Finally, TPA completely abolished the somatostatin-induced inhibition of adenylate cyclase in both hybrid and non-hybrid membranes. These data suggest that, in GH4C1 cells, protein kinase C stimulation by phorbol esters completely inactivates the n alpha i subunit of the inhibitory GTP-binding protein, leaving the n beta subunit functionally intact. It can also be inferred that thyroliberin conveys its main effect on the adenylate cyclase through activation of the stimulatory GTP-binding protein, Gs.

Adenylate Cyclase Toxin↗
PubMed1989

Vasoactive intestinal peptide (VIP) may reduce the removal rate of cytosolic Ca2+ after transient elevations in clonal rat lactotrophs.

The prolactin-producing rat anterior pituitary GH4C1 cells possess Ca2+-activated K channels which are activated by physiological elevations of the cytosolic Ca2+ concentration even at membrane potentials more negative than the normal level of about -50 mV. Whole-cell current recordings showed a marked outward tail current following depolarizing voltage steps to 0 mV from a holding potential close to the normal membrane potential. The half-time of this tail current was about 1.3 s after a 4-s depolarization step. The GH4C1 cells also possess voltage-activated Ca channels, and we conclude that this tail current is a Ca2+-activated K+ current for the following reasons: (1) The reversal potential for the tail current was close to the K+ equilibrium potential for a range of transmembrane K+ gradients. (2) The tail current was blocked by a Ca2+ antagonist, and the voltage dependence of this current closely mirrored the voltage dependence of the isolated Ca2+ current. The time-course of the decline of the tail current thus reflects the removal rate of the Ca2+ entering the cytosol through voltage-dependent Ca channels during the depolarizing voltage step. VIP stimulates prolactin secretion from GH4C1 cells, and this peptide prolonged the half-time of the tail current by about 47% in 63% of the cells. This indicates that VIP may prolong the transient cytosolic Ca2+ elevations following the action potentials in these cells. Such a mechanism might be an important factor for the control of the cytosolic Ca2+ level, and hence hormone secretion.

Animals↗
PubMed1989

Plasma gastrointestinal hormones during spontaneous and induced menstrual cycles.

Plasma levels of secretin, vasoactive intestinal polypeptide (VIP), somatostatin (SRIH), motilin, and/or pancreatic polypeptide, as well as serum estradiol, progesterone, PRL, LH, FSH, and/or GH were measured during the follicular phase, midcycle, and luteal phase of a spontaneous menstrual cycle in eight women and during ovarian stimulation with clomiphene citrate/human menopausal gonadotropin and hCG for in vitro fertilization in nine women. Plasma SRIH concentrations were significantly (P less than 0.02) higher in the luteal phase of spontaneous menstrual cycles than in follicular phase and midcycle. Serum GH levels, however, did not change. Plasma motilin concentrations also were higher in the luteal phase than at mid-cycle (P less than 0.04). Plasma secretin, VIP, and pancreatic polypeptide concentrations did not change during the cycle. Throughout the spontaneous menstrual cycle we found significant positive correlations between plasma SRIH and serum progesterone (P less than 0.007; r = 0.5869), plasma motilin and serum progesterone (P less than 0.02; r = 0.5331), plasma secretin and serum estradiol (P less than 0.04; r = 0.4711), and plasma secretin and serum PRL (P less than 0.02; r = 0.5507). During ovarian stimulation both plasma secretin and VIP gradually increased to a peak on cycle days 0 and 1, respectively (day 0 = the day of hCG injection), whereas plasma SRIH did not change. Serum estradiol and PRL increased significantly, and both peaked on cycle day 1. During ovarian stimulation plasma secretin correlated significantly with serum estradiol (P less than 0.00001; r = 0.9333), serum PRL (P less than 0.03; r = 0.6521), and plasma VIP (P less than 0.03; r = 0.6534). In addition, plasma VIP and serum PRL both correlated significantly with serum estradiol (P less than 0.05; r = 0.6024 and P less than 0.04; r = 0.6384, respectively). These results indicate 1) a possible effect of progesterone on the release of SRIH and motilin during the spontaneous menstrual cycle; 2) the unaltered serum GH concentrations in the luteal phase of the spontaneous menstrual cycle despite elevated plasma SRIH levels are probably due to a stimulatory effect of both progesterone and motilin on GH release; and 3) the increase in plasma secretin and VIP concentrations during ovarian stimulation is probably secondary to the concomitant increase in serum estradiol and/or PRL. We suggest that estradiol and/or PRL beyond a certain threshold level stimulate the release of secretin, and possibly also VIP, into plasma.

Adult↗
PubMed1989

Serum procollagen III peptide in alcoholic and other chronic liver diseases.

Increased levels of serum procollagen III peptide (P-III-P) have been found in patients with alcoholic hepatitis and cirrhosis. Serum P-III-P was increased (greater than 15 micrograms/l) in 38 of 44 (86%) patients with alcoholic liver cirrhosis, in 6 of 20 (30%) with fatty liver, in 1 of 13 (8%) with non-alcoholic fatty liver, and in 3 of 14 (21%) with other chronic liver diseases. Median serum P-III-P was almost three times higher in alcoholic liver cirrhosis than in alcoholic fatty liver (p less than 0.001). Serum P-III-P was increased in three of six patients with alcoholic fatty liver and periportal fibrosis. In the total material (n = 91), a statistically significant negative correlation between serum P-III-P and albumin (r = -0.71, p less than 0.001) and Normotest (r = -0.63, p less than 0.001), respectively, and a positive correlation between serum P-III-P and bilirubin (r = 0.65, p less than 0.001) were found. The serum level of P-III-P had no prognostic value concerning the mortality in patients with alcoholic cirrhosis.

Adult↗
PubMed1989

GnRH-agonist therapy in women with the polycystic ovarian syndrome: effects on ovarian volume and on gonadotropin and sex steroid levels.

Eleven patients with infertility and polycystic ovarian syndrome were treated with the GnRH agonist buserelin, 100 micrograms administered intranasally 6 times daily. The ovarian volume and morphology were monitored by vaginal sonography, and serum levels of hormones were measured by immunoassay. The mean ovarian volume was reduced from 13.0 +/- 4.1 cm3 to 9.1 +/- 3.1 cm3 after 42 days of treatment, p less than 0.001. The number of discernible ovarian microcysts was reduced from 11.9 +/- 2.1 to 9.6 +/- 2.9, p less than 0.05. The gonadotropin levels were reduced in all patients; however, there was no direct relationship between the reduction in ovarian volume loss and the concomitant decrease in LH or FSH levels. The levels of testosterone and androstenedione were normalized, whilst estradiol was suppressed to postmenopausal levels during the treatment period. The decrease in ovarian volume and the suppressed levels of ovarian steroids therefore seem to be related to the inhibition of the pituitary-ovarian axis with the GnRH agonist.

Administration, Intranasal↗
PubMed1988

Validation of a radioreceptor assay for 1,25-dihydroxyvitamin D using selected ion monitoring GC-MS.

This report describes two methods for the measurement of 1,25-dihydroxyvitamin D [1,25(OH)2D] in serum: A modified radio receptor assay (RRA), employing a 1,25(OH)2D receptor from calf thymus, and selected ion monitoring (SIM) with combined capillary gas chromatography (GC)-mass spectrometry (MS). The intra-assay coefficient of variation was close to 13% for both methods, and the inter-assay coefficients of variation were 14.0 and 6.5% for RRA and SIM (GC-MS), respectively. Aliquots of 2 ml (RRA) and 20 ml (SIM) serum were used, and the limits of detection were 10 and 6 pmol/l, respectively. The analytical recovery of each method was assessed, and a maximum deviation from the expected value of 10 and 2% was found for RRA and SIM, respectively. A correlation coefficient of 0.93 (slope 0.97) was obtained when 27 different serum samples were analyzed by both methods. Included in this study were serum samples from healthy subjects and patients with subnormal as well as supranormal 1,25(OH)2D levels. This result showed that the RRA accurately measured the serum levels of 1,25(OH)2D and therefore should be useful in the diagnosis and control of vitamin D dependent diseases.

Adult↗
PubMed1988

Phorbol esters and thyroliberin have distinct actions regarding stimulation of prolactin secretion and activation of adenylate cyclase in rat pituitary tumour cells (GH4C1 cells).

The phorbol ester 12-O-tetradecanoyl phorbol 13-acetate (TPA) enhances the effects of TRH on phase II of prolactin secretion as well as on hormone synthesis at both low and high TPA receptor occupancy. Furthermore TPA, but not the biologically inactive substance 4 alpha-phorbol 12,13-didecanoate (4 alpha-PDD), stimulates the particulate bound adenylate cyclase with a time course paralleling that of TRH activation. However, the combined additions of TRH and TPA activate this cyclase in an additive manner while the Gpp(NH)p- and the forskolin-sensitive enzyme are unaffected by TPA addition. Polymyxin B, which inhibits protein kinase C, abolishes activation of adenylate cyclase by TPA without interfering with the stimulatory action of TRH. Also, when phosphatase activity is preferentially inhibited by pretreatment of the cells with sodium vanadate, the TRH-sensitive cyclase is unaltered, while TPA activation is obliterated. Maximal stimulation of adenylate cyclase by cholera toxin pretreatment, obliterated the actions of TRH and TPA. Cells pretreated with pertussis toxin retained their TRH-sensitive cyclase, however, TPA-responsiveness was lost. We therefore suggest that the action of TPA as it relates to activation of adenylate cyclase, is probably mediated via the Gi component of the adenylate cyclase complex, while TRH stimulates the enzyme via the classical pathway involving the stimulatory GTP binding protein (Gs).

Adenylyl Cyclases↗
PubMed1988

Thyrotropin-releasing hormone-stimulated inositol trisphosphate formation is liable to thyrotropin-releasing hormone-induced desensitization by a calcium-dependent mechanism.

In cultured rat pituitary cells (GH4C1 cells) the ability of thyrotropin-releasing hormone (TRH) to stimulate phosphodiesteratic cleavage of phosphatidylinositol 4,5-bisphosphate (PIP2) by a phospholipase C-type reaction was confirmed. The dose-response relationship for the TRH-stimulated phospholipase C was elucidated as was the relationship between the various inositol phosphates formed during the first few seconds after stimulation. The TRH-stimulated phospholipase C was subject to desensitization by repeated TRH treatment of cell cultures. This desensitization was dependent on the dose of TRH during preincubation. Following desensitization no decline in the levels of PIP2 was detected, even in the presence of decreased levels of PIP2 precursors. The TRH-stimulated phospholipase C activity was not attenuated following pretreatment with 12-O-tetradecanoylphorbol 3-acetate (TPA) to stimulate protein kinase C activity, and TRH also induced desensitization in the presence of the protein kinase C inhibitor polymyxin B. Thus, regulation of protein kinase C activity seemed not to be involved in the desensitization process. It is suggested that the ability of TRH to desensitize its own receptors and their link to phospholipase C, is mediated by the rise in intracellular calcium that is initiated by the TRH-receptor interaction.

Animals↗
PubMed1988

Somatostatin inhibits prolactin secretion by multiple mechanisms involving a site of action distal to increased cyclic adenosine 3',5'-monophosphate and elevated cytosolic Ca2+ in rat lactotrophs.

The release of prolactin (PRL) from a clonal cell-line of anterior pituitary cells (GH4C1) was inhibited by somatostatin (SRIH) in a dose-dependent manner (ED50 nM). The inhibition (20% of control levels) was detectable within 50 s and maximal within 90 s. Thyroliberin (TRH) enhancement of PRL secretion was biphasic. SRIH inhibited both phases equally. Ionomycin in combination with the phorbol ester, TPA, mimics the TRH-elicited PRL release, and SRIH partly inhibited this effect. SRIH had no effect on TRH-stimulated formation of inositol trisphosphate, and only small effects on TRH-activated adenylate cyclase. Vasoactive intestinal peptide (VIP) and forskolin stimulated cAMP formation and PRL release potently. SRIH inhibited both effects of VIP and forskolin, and there was a close correlation between the inhibition of PRL secretion and cAMP accumulation. 8-Bromo-cAMP enhanced PRL release, an effect that was also partly reduced by SRIH. The Ca2+ channel activator, BAY-K-8644 and high extracellular K+ increased PRL release, and SRIH caused a partial reduction in the release response to both secretagogues. SRIH lowered [Ca2+]i, and markedly reduced the rise in [Ca2+]i elicited by TRH, VIP and K+. SRIH did not influence the Ca2+ spikes recorded in Na+-free solution, and had no effect on the TRH-induced membrane potential changes. Our results demonstrate that SRIH may inhibit PRL release from GH4C1 cells by (1) inhibiting hormone-sensitive adenylate cyclase, (2) blocking the effect of cAMP and (3) lowering [Ca2+]i. None of these effects is, however, sufficient to explain all the effects of SRIH, suggesting that SRIH also exerts a major action at a step subsequent to cAMP accumulation and [Ca2+]i elevation. Since the GH4C1 cells possess one single class of binding sites, this implies that the same SRIH receptor is coupled to several cellular signalling systems.

Animals↗
PubMed1987

On the involvement of cyclic AMP and extracellular Ca2+ in the regulation of hormone release from rat pituitary tumour (GH3) cells in culture.

Thyroliberin (TRH), dibutyryl cyclic AMP (db-cAMP), and 3-isobutyl-1-methylxanthine (MIX) had a stimulatory effect on prolactin (PRL) and growth hormone (GH) release from GH3 cells. Half-maximal and maximal effects were observed for TRH at 2.5 nM and 10 nM; for db-cAMP at 0.6 mM and 5 mM, respectively. MIX (0.1 mM-1 mM) induced a dose-dependent accumulation of cellular cyclic AMP, while the hormone release was already maximally stimulated at 0.1 mM MIX. The maximal effects on hormone release of TRH and db-cAMP, but not of TRH and MIX, were additive. The Ca2+ channel blockers Co2+ (5 mM) and verapamil (100 microM) and the Ca2+ chelator EGTA (4 mM) abolished the stimulatory effect of TRH (1 microM) on hormone release. Co2+ and verapamil, but not EGTA, inhibited the stimulatory effect of db-cAMP (5 mM) on hormone release. The inhibitory effects of Co2+ and verapamil on GH release were counteracted by the combination of TRH and db-cAMP. For PRL release Co2+, but not verapamil, was able to inhibit the combined action of TRH and db-cAMP. Co2+, verapamil, and EGTA eliminated the stimulatory effect of MIX (1 mM) on PRL release while only Co2+ and EGTA affected the GH release. Hormone release in the presence of MIX plus verapamil or EGTA, but not Co2+, was increased by TRH. The calmodulin antagonist trifluoperazine (TFP) at 30 microM inhibited basal hormone release and hormone release stimulated by TRH (1 microM), db-cAMP (5 mM), and MIX (1 mM). The Ca2+ ionophore A23187 (5 microM) had a stimulatory effect on basal hormone release which was abolished by 30 microM TFP.

1-Methyl-3-isobutylxanthine↗
PubMed1987

Corticosteroids and lung surfactant levels in adult male rats.

Following lung instillation in adult male rats of 3.4 mumol hexavalent chromium (K2Cr2O7) dissolved in 0.5 ml of 0.9% NaCl, increased levels of lung surfactant could be detected after 48 h. The blood serum concentration of corticosterone was elevated in these animals. Blood serum thyroxine and triiodothyronine showed an initial increase after lung instillation of hexavalent chromium followed by a decline. Metabolism of testosterone by the alveolar macrophages to 17 beta-hydroxy-5 alpha-androstane-3-one and 5 alpha-androstane-3 alpha, 17 beta-diol was reduced 6 and 12 h after the K2Cr2O7 instillation, which was also associated with damage of lung cell function and decreased uptake by the alveolar macrophages of Candida albicans particles. As early as 12 h after s.c. administration of 400 micrograms dexamethasone/100 g body wt, increased levels of lung surfactant could be measured. At this time the lungs showed no signs of cellular damage, and metabolism of testosterone as well as uptake of Candida albicans particles by the alveolar macrophages were normal. Lower s.c. doses of dexamethasone did not result in raising the levels of lung surfactant in 12 h. Within 12 h after s.c. administration of large doses of testosterone, dihydrotestosterone or dehydroepiandrosterone no measurable effects on the levels of lung surfactant could be measured. Since animals treated with dexamethasone (200 micrograms/100 g body wt) or long-acting synthetic ACTH (100 micrograms i.m. Synacthen Depot/100 g body wt) for 5 days after lung instillation of K2Cr2O7 had extremely high levels of lung surfactant, it is concluded that the corticosteroids in adult rats may help to create augmented surfactant levels following lung intoxication. This could proceed via stimulation of surfactant production and reduction of surfactant removal. Different aspects of lung surfactant metabolism are discussed.

Adrenal Cortex Hormones↗
PubMed1987

The mechanism of [3H]dexamethasone uptake into prolactin producing rat pituitary cells (GH3 cells) in culture.

Glucocorticosteroids stimulate growth hormone (GH) synthesis and inhibit prolactin (PRL) synthesis and cell growth in cultured GH3 cells, a clonal cell strain derived from a rat pituitary tumour. This model system was used to study the mechanism by which glucocorticosteroids enter target cells. The cellular uptake of [3H]dexamethasone was temperature dependent and was further inhibited by addition of an excess amount of cold dexamethasone. Half maximal uptake was obtained after about 5 min at 37 degrees C. The initial rates of [3H]dexamethasone uptake were a linear function of the extracellular hormone concentration. The uptake of [3H]dexamethasone in intact cells studied at different temperatures resulted in linear Arrhenius plots, with a calculated energy of activation of 91.0 kJ x mole-1 x degree-1. Scatchard analysis of specifically cell bound [3H]dexamethasone at equilibrium (0 degrees C) showed a straight line with a calculated dissociation constant (Kd) of 1.6 x 10(-9) M and a maximal uptake of 180 x 10(-15) mole/mg cell protein. Specific binding of [3H]dexamethasone to cytosol proteins could only be demonstrated at 0 degrees C. These results indicate that [3H]dexamethasone diffuses passively into the cell, and binds to specific receptors in an energy dependent way.

Animals↗
PubMed1987

A permissive role for extracellular Ca2+ in regulation of prolactin production by 1,25-dihydroxyvitamin D3 in GH3 pituitary cells.

A clonal strain of rat pituitary tumor cells (GH3) that spontaneously synthesizes and secretes prolactin (PRL) and growth hormone (GH) was used as model system to study the mechanism of action of 1,25-(OH)2D3. We have previously demonstrated that these cells possess specific cytosol binding proteins for 1,25-(OH)2D3 (Haug and Gautvik, 1985). When the GH3 cells were incubated in a serum-free, chemically defined medium of low extracellular Ca2+ concentration, 1,25-(OH)2D3 stimulated PRL production in a dose-dependent manner. The stimulation was detectable at 10(-11) M, and the maximum effect (2-fold increase) was observed at 10(-9) M (ED50 = 2 x 10(-11) M). The dose-response curve was bell-shaped, and at 10(-6) M 1,25-(OH)2D3 even suppressed PRL production to about 75% of controls. The stimulatory effect was first seen after 2 days and was maximal after 4 days. On a molar basis 25-OHD3 and 1-OHD3 were at least 100 times less potent than 1,25-(OH)2D3, while 24,25-(OH)2D3 had no effect on PRL production. At an extracellular concentration of Ca2+ as low as 4 x 10(-5) M the stimulatory effect of 1,25-(OH)2D3 was small (1.3-fold). Increasing extracellular Ca2+ to 1.5 x 10(-4) M increased the 1,25-(OH)2D3-induced PRL response to 2.1-fold. In contrast to the biphasic effect of 1,25-(OH)2D3 on PRL production, GH production was decreased to about 60% of controls at 10(-8) M and above. These findings indicate that in serum-free medium the stimulatory effect of 1,25-(OH)2D3 on PRL production is critically dependent on the concentration of extracellular Ca2+.

Animals↗
PubMed1987

Bombesin stimulates prolactin secretion from cultured rat pituitary tumour cells (GH4C1) via activation of phospholipase C.

Bombesin (BBS) stimulated prolactin (PRL) secretion from monolayer cultures of rat pituitary tumour cells (GH4C1) in a dose-dependent manner with half maximal and maximal effect at 2 nM and 100 nM, respectively. No additional stimulatory effect on PRL secretion was seen when BBS was combined with thyroliberin (TRH) used in concentrations known to give maximal effects, while the effects of BBS and vasoactive intestinal peptide (VIP) were additive. Using a parafusion system, BBS (1 microM) was found to increase PRL secretion within 4 s and the secretion profiles elicited by BBS and TRH (1 microM) were similar. Both BBS and TRH increased inositoltrisphosphate (IP3) as well as inositolbisphosphate (IP2) formation within 2 s. BBS also induced the same biphasic changes in the electrical membrane properties of GH4C1 cells as TRH, and both peptides caused a rapid and sustained increase in intracellular [Ca2+]. These results suggest that BBS stimulates PRL secretion from the GH4C1 cells via a mechanism involving the immediate formation of IP3 thus resembling the action of TRH.

Animals↗
PubMed1987

Vasoactive intestinal peptide and peptide with N-terminal histidine and C-terminal isoleucine increase prolactin secretion in cultured rat pituitary cells (GH4C1) via a cAMP-dependent mechanism which involves transient elevation of intracellular Ca2+.

Vasoactive intestinal peptide (VIP) and peptide (P) with N-terminal histidine and C-terminal isoleucine (PHI) stimulated prolactin (PRL) secretion from GH4C1 cells equipotent with ED50 values of 30-50 nM. In a parafusion system optimized to give high time resolution both VIP and PHI increased PRL secretion with a delay of about 60 s and subsequent to the activation of the adenylate cyclase. Thyroliberin (TRH) increased PRL secretion within 4 s. The dose-response curves for VIP- and PHI-stimulated cAMP accumulation were superimposable on those for PRL secretion. At submaximal concentrations the effects of VIP and PHI on both cAMP accumulation and PRL secretion were additive, whereas the effects were not additive at concentrations giving maximal effects. VIP and PHI increased [Ca2+]i measured by quin-2 in a different way than TRH, without inducing changes in the electrophysiological membrane properties of the GH4C1 cells. We conclude that both VIP and PHI stimulate PRL secretion via a cAMP-dependent process involving an increase in [Ca2+]i.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1987

Effects of calmodulin antagonists on hormone release and cyclic AMP levels in GH3 pituitary cells.

In GH3 cells the calmodulin antagonists trifluoperazine and N-(6-aminohexyl)-5-chloro-1-napthalene sulphonamide hydrochloride (W-7) showed a dose-dependent, biphasic effect on the release of growth hormone (GH) and prolactin (PRL). Hormone release was inhibited with 15-30 microM trifluoperazine and with 30-80 microM W-7, while stimulation was observed with 50-100 microM trifluoperazine and with 150 microM W-7. Trifluoperazine (greater than or equal to 30 microM) and W-7 (greater than or equal to 80 microM) increased the concentration of cellular cyclic AMP. Sulphoxides of trifluoperazine and chlorpromazine (less than or equal to 150 microM were without effect on hormone release and cellular cyclic AMP. Hydrolysis of cyclic AMP by GH3 cytosol was reduced after incubation of intact GH3 cells with trifluoperazine (15-60 microM). When trifluoperazine was incubated with cytosol, both the high and low affinity forms of cyclic AMP phosphodiesterase were inhibited competitively with calculated Ki of 4.5 and 56 microM, respectively. Stimulation of cyclic AMP phosphodiesterase caused by endogenous calmodulin was blocked by trifluoperazine. Particulate bound adenylyl cyclase activity was inhibited by trifluoperazine, and this effect was counteracted by endogenous calmodulin.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1987

Binding and degradation of vasoactive intestinal peptide in prolactin-producing cultured rat pituitary tumour cells (GH4C1).

Vasoactive intestinal peptide (VIP) stimulates prolactin (PRL) secretion from cultured rat pituitary cells (GH cells) (Bjøro et al. 1984). This study demonstrates the presence of specific receptors for 125I-VIP on the GH4C1 cells. Specific binding was rapid and biphasic giving a transient plateau lasting from 7 to 30 min. Thereafter specific binding declined to about one-third after 90 min. This coincided with enhanced degradation of 125I-VIP. The degradation was mainly cell-mediated and only partly receptor dependent. Trichloroacetic acid precipitation and absorption chromatography indicated that the degradation products were either 125I- and/or small labelled peptide fragments. Bioassay, RIA and rebinding studies also demonstrated degradation of VIP. Pretreatment of GH4C1 cells with trypsin decreased the rate of degradation of 125I-VIP, but also reduced the amount of specific binding. Scatchard analysis of binding data indicated the existence of two independent classes of receptors, one with Kd = 2.2 nM and Bmax = 15 fmol per 10(6) cells and another with Kd = 180 nM and Bmax = 550 fmol per 10(6) cells. The IC50 for VIP, PHI and secretin were 4, 5 and 500 nM, respectively. We conclude that the high affinity receptor is the most probable mediator of VIP on PRL secretion. The effect of VIP and PHI on PRL secretion in GH4C1 cells is mediated through one common receptor.

Animals↗