Actions of growth hormone-releasing factor and somatostatin on adenylate cyclase and growth hormone release in rat anterior pituitary

The interaction of growth hormone-releasing factor (GRF) and somatostatin (SRIF) on adenylate cyclase activity and growth hormone release was investigated in pituitary homogenates and 2-day cultured rat anterior pituitary cells. GRF stimulated growth hormone release by about 3-fold (ED50 1.6 X 10(-12) M) and caused a rapid 15-fold increase in cyclic AMP production (ED50 6.0 X 10(-12) M). The increase in cyclic AMP was due to direct stimulation of adenylate cyclase by GRF, which caused a 4-fold increase in the activity of the enzyme measured in anterior pituitary homogenates. GRF-induced cyclic AMP formation and GRF-stimulated adenylate cyclase activity were maximally inhibited to the extent of about 50% by 10(-8) M somatostatin. In contrast, GRF-stimulated growth hormone release was completely inhibited by somatostatin (ID50 3.2 X 10(-11) M), suggesting a second site of action of somatostatin. These studies demonstrate that GRF stimulates growth hormone release via activation of adenylate cyclase and a rise in intracellular cyclic AMP. In addition, these findings indicate that the inhibitory action of somatostatin on growth hormone release is exerted at two levels, one at the level of adenylate cyclase affecting the production of cyclic AMP, and the other beyond the formation of the nucleotide, at a site which modulates the release of growth hormone from the cell.     

STIMULATION OF ANTERIOR PITUITARY ADENYL CYCLASE ACTIVITY AND ADENOSINE 3′:5′-CYCLIC PHOSPHATE BY HYPOTHALAMIC EXTRACT AND PROSTAGLANDIN E1

Hypothalamic extract, containing the releasing factors for anterior pituitary hormones, within minutes stimulated adenyl cyclase activity and adenosine 3':5'-cyclic phosphate (cyAMP) concentrations in rat anterior pituitary in vitro. Cerebral cortical extract was ineffective and hypothalamic extract had no effect on these parameters in posterior pituitary or thyroid. Prostaglandin E(1) also increased adenyl cyclase activity and cyAMP levels in anterior pituitary tissue. Although NaF augmented adenyl cyclase activity, it did not elevate cyAMP. Epinephrine, norepinephrine, histamine, serotonin, dopamine, and vasopressin did not increase either adenyl cyclase or cyAMP. The increased adenyl cyclase and cyAMP produced by hypothalamic extract was associated with greater luteinizing hormone release from anterior pituitary in vitro.     

Stimulation of prostaglandin accumulation in the rat anterior pituitary gland by luteinizing hormone releasing hormone in vitro.

The addition of luteinizing hormone releasing hormone releasing hormone (LH-RH) to cultures of monolayers of rat anterior pituitary cells was shown to increase both the concentrations of prostaglandins E1 and E2 (PGE) in the cells and the release of LH over similar ranges of concentrations of LH-RH (10(-6) to 10(-10) mol/l). The peak concentration of PGE was observed after 2.5 h. The stimulation of the level of PGE in the cells by LH-RH was completely inhibited by two inhibitors of prostaglandin synthetase, which only partially inhibited the stimulation of LH release. Therefore the increased concentration of PGE was not obligatory for the effect of LH-RH on LH release. It was also shown that monobutyryl cyclic AMP stimulated the intracellular concentration of PGE and it is suggested that the stimulation of PGE levels may be mediated by increased levels of cyclic AMP in the cells after the addition of LH-RH.     

Stimulation of Adenosine 3′:5′-Cyclic Monophosphate Accumulation in Anterior Pituitary Gland In Vitro by Synthetic Luteinizing Hormone-Releasing Hormone

A near-maximal dose (20 ng/ml) of synthetic luteinizing hormone(LH)-releasing hormone/follicle-stimulating hormone(FSH)-releasing hormone added to incubated anterior pituitary tissue of male rats leads to concomitant increases of intracellular concentrations of adenosine 3':5'-monophosphate and of release of both LH and FSH. The stimulatory effect of LH-releasing hormone/FSH-releasing hormone is observed after a lag period of about 90 min and is progressive at later time intervals; a 3-fold stimulation of cAMP accumulation over control is seen after 210 min of incubation. Half-maximal stimulation of cAMP accumulation is observed between 0.1 and 1.0 ng/ml (0.1-1 nM) of LH-releasing hormone/FSH-releasing hormone. In the presence of 10 mM theophylline, the stimulatory effect of LH-releasing hormone/FSH-releasing hormone on cAMP accumulation is similar to that observed in the absence of the inhibitor of cyclic nucleotide phosphodiesterase, indicating that the releasing hormone exerts its effect by specific activation of adenylate cyclase in LH- and FSH-secreting cells rather than by inhibition of cyclic nucleotide phosphodiesterase. Since the release of growth hormone, thyrotropin, prolactin, and adrenocorticotropic hormone is not affected by LH-releasing hormone/FSH-releasing hormone, and since cAMP stimulates the release of all six adenohypophyseal hormones. the observed changes of cAMP concentrations indicate specific stimulation of adenylate cyclase activity in LH-and FSH-secreting cells of the adenohypophysis.     

A possible role of cyclic AMP in mediating the effects of thyrotropin-releasing hormone on prolactin release and on prolactin and growth hormone synthesis in pituitary cells in culture.

Thyrotropin-releasing hormone (TRH) has 3 effects on clonal strains of rat pituitary cells in culture (GH-cells). Two long-term effects of TRH on GH-cells, which are measurable after 3 h or longer, have been previously reported; these are an increase in prolactin synthesis and a decrease in growth hormone production. We report here that TRH also stimulates the rapid release of stored intracellular prolactin. We have investigated the role of cyclic AMP as a possible mediator of the effects of TRH on GH-cells. Cyclic AMP concentrations are higher in cells treated with TRH compared with paired controls; a maximum difference of greater than 150% of control values is detected at 15 min if the incubation is performed in serum-free medium in the presence of 1 mM theophylline. The concentration of TRH required to give half-maximum increases in both prolactin release and cyclic AMP accumulation is 0.3 nM; half-maximal increases in prolactin synthesis occur at 3 nM TRH. Exogenous cyclic AMP (1 mM) causes only a slight increase in prolactin release; 8-bromo-cyclic AMP and 8-methylthio-cyclic AMP (1 mM) do not cause significant release. Phosphodiesterase inhibitors (0.3 mM theophylline, 0.03 mM isobutyl-methylxanthine) increase prolactin release but their effects on hormone synthesis are more complicated. Isobutylmethylxanthine, 8-bromo-cyclic AMP and 8-methylthio-cyclic AMP (0.4 MM) increase prolactin synthesis, but do not significantly affect growth hormone synthesis. Theophylline increases the synthesis of both hormones. Dibutyryl cyclic AMP (0.5 mM or more) increases prolactin release and both growth hormone and prolactin synthesis, but equivalent amounts of sodium butyrate have the same effects. We conclude that in GH-cells under carefully defined experimental conditions: 1) TRH causes an increase in intracellular cyclic AMP concentrations; 2) the increase in endogenous cyclic AMP and the effects of phosphodiesterase inhibitors are consistent with a model with cyclic AMP as a mediator of the effects of TRH on prolactin release; however, they do not prove this model, because the interpretation of these results depends on assumptions which may not all be valid; and 3) none of the analogs of cyclic AMP or the phosphodiesterase inhibitors tested mimic the decrease in growth hormone production caused by TRH.     

Stimulation of Release of Adrenal Catecholamine by Adenosine 3′:5′-Cyclic Monophosphate and Theophylline in the Absence of Extracellular Ca2+

Stimulation of catecholamine release was studied in the isolated adrenal of the cat during retrograde perfusion. Theophylline, adenosine 3':5'-cyclic monophosphate (cyclic AMP), and dibutyryl-cyclic AMP stimulated catecholamine release in adrenal chromaffin tissue; adenosine, 2'-AMP, 3'-AMP, and 5'-AMP were ineffective. Addition of theophylline for 5-15 min had no effect on catecholamine release induced by KCl or nicotine, but it significantly increased the responses to cyclic AMP and its dibutyryl derivative. Glands perfused with Ca(2+)-free Locke's solution for 30-180 min rapidly lost their responsiveness to KCl or nicotine. In contrast, exposure to Ca(2+)-free medium for 180 min had no effect on secretory responses to either the cyclic nucleotides, themselves, or to methylxanthine-induced potentiation of cyclic AMP responses. Thus, dibutyryl-cyclic AMP, cyclic AMP, and theophylline do not require extracellular Ca(2+) to release adrenal catecholamines. They may act by translocating intracellular bound Ca(2+) or by a mechanism independent of calcium.     

Prostaglandin E1 Effects on Adenosine 3′:5′-Cyclic Monophosphate Concentration and Phosphodiesterase Activity in Fibroblasts

Incubation of L-929 and L-2071 fibroblasts with prostaglandin E(1) (PGE(1)) caused a rapid increase in the cyclic AMP content of these cells. A maximal effect was produced with 0.2 mug PGE(1) per ml. At a concentration of 4 mug/ml, PGE(2) was almost equally effective, but PGF(2alpha) and PGA(2) were much less so. 2.6 muM epinephrine, 0.4 mM serotonin, and 0.2% ethanol were without effect. In L-929 cells, cyclic AMP concentrations remained elevated for 2-5 hr, and then declined, although even after a 24-hr incubation the medium contained PGE(1) in a concentration sufficient to increase maximally the cyclic AMP content of cells not previously exposed to this compound. A second addition of PGE(1) after 5 or 24 hr did not produce another increase in the concentration of cyclic AMP. After incubation with PGE(1) for 24 hr, cyclic AMP phosphodiesterase activity, assayed with 0.56 muM substrate, was increased 30-100%; the activity rose further between 24 and 48 hr. It is suggested that the increase in phosphodiesterase activity that appears to be a consequence of prolonged elevation of cyclic AMP concentration may account at least in part for the apparent "refractoriness" to PGE(1) that develops after incubation for several hours with this compound.     

Dopamine and somatostatin inhibit forskolin-stimulated prolactin and growth hormone secretion but not stimulated cyclic AMP levels in sheep anterior pituitary cell cultures

Forskolin, an activator of adenylate cyclase, has been used to investigate the effects of raising pituitary cell cyclic AMP concentrations on prolactin and growth hormone secretion and to examine the role of cyclic AMP in the inhibitory actions of dopamine and somatostatin. Incubation of cultured ovine pituitary cells with forskolin (0.1-10 microM; 30 min) produced a modest dose-related increase in prolactin release (120-140% of basal) but a much greater stimulation of growth hormone secretion (170-420% of basal). Cellular cyclic AMP concentrations were only increased in the presence of 1 and 10 microM forskolin (2-5.5 times basal). A study of the time course for forskolin (10 microM) action showed that stimulation of prolactin (1.5-fold) and growth hormone (4.7-fold) secretion occurred over 15 min; subsequently (15-60 min) the rate of prolactin secretion from forskolin-treated cells was equivalent to that measured in controls, while growth hormone release remained elevated. Cellular cyclic AMP concentrations were also rapidly stimulated by forskolin (10 microM); they reached a maximum (12 times control) within 15 min, and then declined (15-60 min) but remained elevated relative to those in untreated cells (4.9 times control at 60 min). Dopamine (0.1 microM) inhibited basal secretion of both prolactin and growth hormone. In the presence of forskolin (0.1-10 microM), dopamine (0.1 microM) inhibited prolactin secretion to below the basal level and considerably attenuated the stimulation of growth hormone secretion. Similarly, somatostatin suppressed both basal and forskolin-induced prolactin and growth hormone secretion. However, neither dopamine nor somatostatin significantly decreased the stimulatory effect of forskolin on cellular cyclic AMP accumulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of Thyrocalcitonin on Adenosine 3′:5′-Cyclic Phosphate Formation by Rat Kidney and Bone

Thyrocalcitonin (TCT) increased the rate of accumulation of adenosine 3':5'-cyclic phosphate (cyclic AMP) when added to incubations containing washed particles from whole rat kidney, adenosine triphosphate (ATP), MgSO(4), and caffeine. The maximum stimulatory effect of TCT, 44 +/- 6.7 per cent, was always less than the 150 to 250 per cent increase produced by parathyroid hormone (PTH). The effect of both hormones together was no greater than that of PTH alone when each was present at a maximally effective concentration. Since neither TCT nor PTH altered the rate of degradation of cyclic AMP by the kidney preparation, it may be inferred that their effects on cyclic AMP accumulation are the result of increased formation of cyclic AMP. Adenyl cyclase activity in homogenates of renal cortex was stimulated to a greater extent by TCT and PTH than was that of medulla, whereas, as reported earlier, the effect of vasopressin was much larger with homogenates of medulla. The accumulation of cyclic AMP in incubations of rat kidney cortex slices was increased 20 to 60 per cent by TCT and 50 to 140 per cent by PTH. The accumulation of cyclic AMP in incubations of rat calvaria was increased about threefold with TCT and nine to tenfold with PTH, while reduced and alkylated TCT had less than 10 per cent of the activity of TCT. These observations are consistent with the view that the physiological effects of TCT and PTH in kidney and bone are secondary to the enhanced formation of cyclic AMP.     

The porcine ovarian follicle: III. Development of chorionic gonadotropin receptors associated with increase in adenyl cyclase activity during follicle maturation.

Specific binding of human chorionic gonadotropin (hCG), and the hCG-sensitive adenyl cyclase of granulosa cells from small (1-2 mm), medium (3-5 mm), and large (6-12 mm) porcine ovarian follicles have been studied. The number of hCG-binding sites per cell (n) increases during follicle maturation without a change in the binding affinity. The values for n were 300-400 for small, 1,000-1,600 for medium, and 8,200-10,000 for large-follicle cells. The dissociation constant is 2.4 X 10(-10)M for all cells. hCG-sensitive adenyl cyclase was demonstrated in porcine granulosa cells. The adenyl cyclase system of granulosa cells becomes increasingly responsive to hCG stimulation during follicle development. Maximal adenyl cyclase activation by hCG (1 mug/ml) was 240, 750, and 7,000 molecules of cyclic AMP formed/sec/cell, respectively, for small, medium, and large follicle cell. The concentration of hCG giving half-maximal stimulation (1.0 X 10(-9)M) was similar for both large and medium follicle cells. It is concluded that: 1) an increase in hCG receptor sites per cell occurs during maturation of the porcine ovarian follicle without change of binding affinity, and 2) the increase in the number of hCG receptors correlates well with hCG-sensitive adenyl cyclase activity during follicle development.     

Refractoriness of the pituitary gland after continuous exposure to luteinizing hormone releasing hormone.

The refractoriness of LH release by pituitary glands from intact female rats was studied during stimulation by luteinizing hormone releasing hormone (LH-RH), monobutyryl cyclic AMP+theophylline or potassium in vitro. Various concentrations of LH-RH (0.1, 0.3 and 10 ng/ml) all caused refractoriness within 24 h. Subsequent exposure to a supramaximally active concentration of LH-RH for 6 h also resulted in a depressed response; the degree of inhibition depended on the concentration of LH-RH to which the glands had been exposed previously. Glands made refractory to LH-RH also showed a depressed response to monobutyryl cyclic AMP+theophylline, although these agents by themselves were unable to induce refractoriness. Incubation in medium containing a high concentration of potassium also resulted in the release of LH, which in all respects was similar to that caused by LH-RH. Glands made refractory to LH-RH showed a decreased response to potassium and, conversely, the release of LH in response to LH-RH was reduced after exposure of glands to potassium. It is concluded that the LH releasing activity of LH-RH, which is mimicked by potassium, deteriorates during continuous exposure to the secretagogue.     

Dependence on protein synthesis of the N6-monobutyryl cyclic AMP plus theophylline mediated release of luteinizing hormone induced by luteinizing hormone releasing hormone from rat pituitary glands in vitro

The involvement of cyclic AMP in the action of LH releasing hormone (LH-RH) on LH secretion was studied by incubating pituitary glands from adult female rats on day 2 of dioestrus with 1 mN-N6-monobutyryl cyclic AMP (mbcAMP) and 10 mM-theophylline for periods of up to 10 h. This treatment induced a pattern of LH release similar to that observed in the presence of a low concentration of LH-RH (0.1 ng LH-RH/ml), i.e. an initial 4 h period during which the release of LH was minimal was followed subsequently by an increased rate of release. In this system inhibition of protein synthesis by cycloheximide (25 microgram/ml) did not impair the initial response of the pituitary tissue but the increase in the rate of LH release during the second phase of the response was blocked. Preincubation with mbcAMP and theophylline increased the responsiveness of the pituitary tissue to LH-RH. This action could be prevented by including cycloheximide during the preincubation period, whereas addition of this drug during the incubation with LH-RH no longer impaired the increased responsiveness. The size of the sensitizing action of mbcAMP and theophylline mediated through the induction of protein synthesis was comparable with that of a high concentration of LH-RH. From the absence of a significant change in total LH during the preincubation period, it was concluded that the increased responsiveness was not the result of newly synthesized LH. The present results suggest a role or roles for cyclic AMP in the secretion of LH induced by LH-RH. Besides an effect on the formation of a factor related to the synthesis of protein, other than LH which has a permissive role in the acute release of LH, cyclic AMP might also be concerned in the secretion process through a pathway which does not involve synthesis of protein.     

Regulation of thyrotropin (TSH) release and production in monolayer cultures of transplantable TSH-producing mouse tumors.

Studies of TSH release and production were performed in short term monolayer cultures of transplantable, thyroid hormone responsive, thyrotropin (TSH) producing mouse pituitary tumors. These tumors contained large amounts of TSH, small amounts of growth hormone (GH) and no detectable luteinizing hormone (LH), indicating that the predominant hormone product of tumor cells was TSH. The TSH content per tumor cell was similar to that of the normal pituitary where thyrotrophs represent a small fraction of the total cells, suggesting that the TSH content per tumor cell was less than that of the normal thyrotroph. There was a time dependent release and production of TSH by tumor cells in monolayer culture. Thyrotropin releasing hormone (TRH) increased the release into the media and the production of TSH in a dose dependent manner. Maximum effects were noted at 0.2 ng/ml. Thyroid hormones and somatostatin inhibited both basal and TRH induced effects on both TSH release and production. TSH release as induced by TRH was calcium dependent. TSH release was stimulated by ouabain (10(-3)M) and potassium (57 mM), agents known to promote cellular calcium uptake in a calcium dependent manner. These studies indicate that tumor derived cells function in monolayer culture in a similar fashion to normal thyrotrophs. Studies were conducted to test the hypothesis that TRH action is mediated by adenosine 3',5' monophosphate (cAMP). Dibutyryl cAMP (6 mM) and theophylline (10 mM) increased TSH release suggesting that cAMP is involved in TSH release. However, TRH had no detectable effect on tumor cell adenylate cyclase activity or levels of cAMP. In contrast, PGE1 (1-10 mug/ml) stimulated adenylate cyclase activity and elevated cellular levels of cAMP without increasing TSH release. Thus, we are unable to confirm the postulate that cAMP is the intracellular mediator of TRH action.     

Luteinizing hormone secretion is enhanced by pertussis toxin, cholera toxin, and forskolin. Evidence for the involvement of the cyclic AMP-generating system

Pertussis toxin, cholera toxin and forskolin, all of which can increase adenylate cyclase activity, stimulated luteinizing hormone LH release from cultured rat anterior pituitary cells. Although cellular cyclic AMP and growth hormone were increased rapidly by cholera toxin and forskolin, enhanced LH release occurred significantly later with no change in total radioimmunoassayable LH (i.e., released plus stored). These data suggest that changes in cyclic AMP levels may regulate the tonic availability of releasable LH in the gonadotroph.     

Mechanisms involved in the effects of TRH on GHRH-stimulated growth hormone release from ovine and bovine pituitary cells

Incubation of cultured ovine pituitary cells with growth hormone-releasing hormone (GHRH) (10(-12)-10(-7) M) stimulated growth hormone secretion up to 3-fold. At a maximal stimulatory concentration of GHRH (10(-10) M), thyrotropin-releasing hormone (TRH) (10(-7) M) caused an inhibition of growth hormone release to approx. 50% of the response obtained with GHRH alone (during a 15 min incubation period). TRH also caused a small inhibition of the GHRH-stimulated cellular cyclic AMP level but this effect was only significant at a relatively high concentration of GHRH (10(-9) M). Incubation of cultured bovine pituitary cells with GHRH (10(-11)-10(-8) M) plus TRH (10(-7) M) caused a significant stimulation of growth hormone release by up to 40%, compared with the response obtained with GHRH alone (at all concentrations of GHRH). TRH (10(-7) M) had no effect on GHRH (10(-8) M)-stimulated cellular cyclic AMP levels in a partially purified bovine pituitary cell preparation. The effects of varying extracellular [Ca2+] (0.1-10 mM) on intracellular [Ca2+] and on the responsiveness to releasing hormones were also determined using ovine pituitary cells. GHRH (10(-10) M)-stimulated growth hormone release was inhibited when cells were incubated at both high (10 mM) and low (0.1 mM) [Ca2+] (compared with 1 mM or 3 mM Ca2+) with or without TRH (10(-7) M). At 1 mM Ca2+, TRH produced a synergistic effect with GHRH to stimulate growth hormone release. However, at 3 mM Ca2+ TRH inhibited GHRH-stimulated growth hormone release.(ABSTRACT TRUNCATED AT 250 WORDS)     

Relationship between stimulated prolactin release from GH cells and cyclic AMP degradation and formation

We have studied the relationship between the prolaction (PRL) release induced by thyroliberin (TRH) and theophylline and the formation and inactivation of adenosine 3', 5'-cyclic monophosphate (cyclic AMP) in cultured rat-pituitary cells (GH3 cells). TRH, which stimulated prolactin release, increased cyclic AMP formation and stimulated transiently both the low- and high-Km cyclic phosphodiesterases. The maximal effect on the phosphodiesterase was observed at 30 mM TRH. The stimulatory effect of TRH on the activity of the cyclic AMP phosphodiesterases was duplicated by incubation of the cells with cyclic AMP (2-10 mM). In washed particulate GH3 cell fractions, TRH increased the adenylyl cyclase activity up to 180%. Treatment of GH3 cells with theophylline stimulated the release of PRL and inhibited cyclic AMP degradation probably leading to the measured increase in cellular concentrations of the nucleotide. The effects of TRH and theophylline on cellular cyclic AMP concentrations and on PRL release were additive. There was a positive correlation between PRL release and cellular cyclic AMP concentration (r = 0.97). The elevations observed in cellular cyclic AMP concentration after TRH treatment are due to increased formation which in turn leads to phosphodiesterase activation. Therefore, cyclic AMP formation appears to be an intermediary step in the stimulus-secretion coupling caused by the tripeptide.     

Bradykinin-induced ACTH release from rat pituitary tissue in vitro

Bradykinin (10(-6) and 10(-5) M) stimulated ACTH-IR release from rat anterior pituitary tissue in vitro concentration-dependently. The onset of this effect was delayed in comparison to that of AVP or CRF. The combined treatment of bradykinin with AVP or CRF produced additive effects of ACTH-IR release. Bradykinin may represent another candidate involved in the regulation of ACTH release. In contrast to AVP, bradykinin did not stimulate prostaglandin E2 synthesis in the pituitary tissue. Bradykinin-induced ACTH-IR release remained unchanged following cyclooxygenase inhibition by indomethacin. It can be concluded that prostaglandins are not involved in the action of bradykinin on the anterior pituitary. Bradykinin did stimulate cyclic AMP accumulation in pituitary tissue. Inhibition of phosphodiesterase by 3-isobutyl-l-methylxanthine (IBMX) potentiated the ACTH-IR release evoked by bradykinin. From the results obtained, we concluded that cyclic AMP appears to be involved as a second messenger in the bradykinin-evoked ACTH-IR release.     

Cyclic AMP-dependent regulation of lipid mediators in white cells. A unifying concept for explaining the efficacy of theophylline in asthma

Activation of white cells, including the neutrophil, eosinophil, basophil, and mast cell, has long been known to be suppressed by high, nonphysiological levels of E-prostaglandins (PGE). In contrast, PGE at levels consistent with an interaction with the PGE receptor (5 X 10(-9) M) have recently been shown to suppress leukotriene (LT) and prostaglandin (PG) production by neutrophils and eosinophils. This occurs by cyclic AMP-dependent inhibition of release of substrate arachidonic acid (AA) from phospholipid pools. The additional observation that indomethacin (10(-9) M) enhances release of eicosanoids by suppressing endogenous PGE2 acting to increase cAMP levels in these cells. Theophylline and other phosphodiesterase inhibitors precisely duplicate the action of PGE2, and the combined effects of such phosphodiesterase inhibitors and adenylate cyclase stimulators are synergistic. The mechanism of action of theophylline in asthma is not know, although it is generally agreed that its effect is a direct one on the bronchial smooth muscle. The findings described in this report now permit the bronchial smooth muscle, but is primarily one of suppressing mediator release from relevant white cells by inhibition of cAMP phosphodiesterase, an action that is enhanced by the presence of inflammatory prostaglandins in the lung.     

Calcium requirement for stimulation of cyclic AMP accumulation in anterior pituitary gland by LH-RH.

Removal of Ca2+ from the incubation medium by addition of 2 mM ethylene glycol bis-(beta-aminoethyl ether)-N, N'tetraacetic acid (EGTA) leads to at least 75% inhibition of the luteinizing hormone-releasing hormone (LH-RH)-induced accululation of adenosine 3'5'-monophoshpate (cyclic AMP) in rat anterior pituitary gland in vitro. This inhibitory effect of EGTA is reversed by the addition of Ca2+. A half-maximal effect of Ca2+ on LH-RH--induced cyclic AMP accumulation is observed at 2-5 X 10-5 M free Ca2+. The LH-RH-induced LH and FSH release is completely dependent upon the presence of Ca2+ in the incubation medium, a half-maximal effect being measured at 1-2 X 10-4 M free Ca2+. The basal release release of LH is increased upon Ca2+ removal.