Testosterone and its precursors and metabolites enhance guanylate cyclase activity.

Both testosterone and cyclic GMP stimulate DNA synthesis. Because cyclic GMP and testosterone seem to have similar actions, the objective of this investigation was to determine if testosterone and its precursors might have part of their mechanism of action through stimulation of guanylate cyclase [GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2], the enzyme that catalyzes the formation of cyclic GMP from GTP. The precursors--namely, progesterone, pregnenolone, 17 alpha-progesterone, 17 alpha-hydroxypregnenolone, androstenedione, and dehydroepiandrosterone--caused a 2- to 3 1/2-fold enhancement of guanylate cyclase activity in rat liver, kidney, skeletal muscle, and ventral prostate at a concentration of 1 microM. These precursors are generated from cholesterol, which had no effect itself on guanylate cyclase activity. Testosterone, 19-nortestosterone, 17-methyltestosterone, and 5 alpha-dihydrotestosterone enhanced guanylate cyclase activity 2- to 5-fold in the same tissues at 1 microM. Etiocholanolone, androsterone, and epiandrosterone, metabolites of testosterone metabolism, enhanced guanylate cyclase activity 1 1/2- to 2-fold at this same concentration. Dose-response relationships revealed that testosterone and its precursors and metabolites had their maximal effect at 1 microM but still had some effect at 0.001 microM. The data in this investigation suggest that the guanylate cyclase-cyclic GMP system plays a role in the mechanism of action of testosterone and its precursors.     

Adenylate and guanylate cyclase activities and cellular differentiation in rat small intestine.

Adenylate and guanylate cyclase activities were measured in rat small intestinal villus and crypt cells to determine possible correlations with cellular differentiation. Isolated intestinal cells were prepared by a method which effectively separates differentiated villus cells from undifferentiated crypt cells (J Biol Chem 248:2542, 1973). Crypt cells were found to have a significantly lower guanylate cyclase activity than villus cells. Adenylate cyclase activity was higher in crypt cells than villus cells, although the difference was less striking than the reverse gradient observed for guanylate cyclase. There was no gradient of activity for cyclic guanosine 3':5'-monophosphate phosphodiesterase. However, cyclic adenosine 3':5'-monophosphate phosphodiesterase activity was lower in villus cells. No villus to crypt gradient of cyclic adenosine 3':5'-monophosphate concentration was detected in mucosa frozen rapidly in liquid nitrogen. The properties and subcellular localization of the cyclases were also evaluated, and of particular interest was the localization of guanylate cyclase to the microvillus membrane and the confirmation of adenylate cyclase activity in the lateral-basal membrane. The villus to crypt gradient of guanylate cyclase suggests that this enzyme has a specialized role in the differentiated villus cell. The contrasting subcellular localization of the cyclases suggests that the cyclases may be interrelated, possibly reflecting the epithelial cell polarity for absorption and secretion.     

Hemin induces neuroglobin expression in neural cells

Neuroglobin is a newly identified vertebrate globin that binds O(2) and is expressed in cerebral neurons. We found recently that neuronal expression of neuroglobin is stimulated by hypoxia and ischemia and protects neurons from hypoxic injury. Here we report that, like hemoglobin and myoglobin, neuroglobin expression can also be induced by hemin. Induction was concentration dependent and time dependent, with maximal (about 4-fold) increases in neuroglobin mRNA and protein levels occurring with 50 microM hemin and at 8 to 24 hours. The inductive effect of hemin was attenuated by the protein kinase G inhibitor KT5823 and the soluble guanylate cyclase inhibitor LY83583, was mimicked by treatment with 8-bromo-cyclic guanosine 3',5'-monophosphate, and was accompanied by a greater than 10-fold increase in cGMP levels, suggesting that it is mediated through protein kinase G and soluble guanylate cyclase. In contrast, hypoxic induction of neuroglobin was blocked by the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase inhibitor PD98059, indicating that hemin and hypoxia regulate neuroglobin expression by different mechanisms. These results provide evidence for regulation of neuroglobin expression by at least 2 signal transduction pathways.     

A soluble interaction between dibutyryl cyclic guanosine 3':5'-monophosphate and cholecystokinin: a possible mechanism for the inhibition of cholecystokinin activity

N2,O2'-dibutyryl cyclic guanosine 3':5'-monophosphate has been reported to inhibit the activity of cholecystokinin on several different end organ responses and in several species. Although the mechanism of this inhibition is unclear, competitive antagonism at the level of the receptor has been suggested. We have investigated an alternate possibility, that N2,O2'-dibutyryl cyclic guanosine 3':5'-monophosphate inhibits cholecystokinin action by interacting directly with the peptide while both are in solution. We used antisera with specificities for different regions of cholecystokinin-gastrin peptides and radioiodinated cholecystokinin-33, cholecystokinin-8, and gastrin-17. Our results support the possibility of a soluble interaction between N2,O2'-dibutyryl cyclic guanosine 3':5'-monophosphate and the peptides of cholecystokinin-gastrin family that is specific for the COOH-terminal (receptor binding) region of these peptides, that is dependent on the concentration of N2,O2'-dibutyryl cyclic guanosine 3':5'-monophosphate, and that correlates with concentrations that inhibit biologic activity. The proposed N2,O2'-dibutyryl cyclic guanosine 3':5'-monophosphate-cholecystokinin complex is dispersed by gel filtration chromatography, and is prevented from being formed by certain detergents.     

Cyclic GMP affecting the tracheal nonadrenergic noncholinergic inhibitory system

An association between guanosine 3',5'-monophosphate (cyclic GMP) and the nonadrenergic noncholinergic inhibitory system (NANCIS) has been demonstrated in the isolated bovine tissue (Bowman and Drummond, 1984). In order to investigate this association in the guinea pig trachea, we used cyclic GMP derivatives, guanylate cyclase activators (N-methylhydroxylamine (NMH) and nitroglycerin (NG)] and inhibitors [oxyhemoglobin (HbO2) and methylene blue (MB)]. Under general anesthesia paralysis, the animals were ventilated and hourly injected with atropine (0.2 mg/kg) and propranolol (1 mg/kg). Cervical segment of the trachea was converted to a closed tracheal pouch and then filled with Kreb's solution augmented with atropine (1 microM) and propranolol (3.5 microM). A decrease in the pouch pressure (Pp) reflected NANCIS nerve transmural stimulation (TS)--or drug-induced relaxation. Pharmacological agents were applied intravenously. At 2-11 min after injection, NMH and NG decreased baseline Pp and reduced TS-induced relaxation. NMH, which is more potent than NG in activating particulate guanylate cyclase activity, potentiated the TS-induced relaxation at high frequencies, but NG did not. HBO2 inhibited the TS-induced relaxation at high but not at low frequencies. In contrast, MB inhibited the relaxation at low but not high frequencies. The results suggest that activation of particulate or membrane bound guanylate cyclase potentiates NANCIS-induced decrease in Pp. Therefore, there is a possible association between cyclic GMP and the NANCIS in the guinea pig trachea.     

Regulation of low-density lipoprotein receptor activity by estrogens and phytoestrogens in a HepG2 cell model

BACKGROUND/AIMS: Estrogen treatment is thought to lower low-density lipoprotein (LDL) cholesterol levels by increasing clearance through hepatic LDL receptors. This study aimed to determine the effect of estrogens and phytoestrogens on LDL receptor activity in a human hepatoma cell line, HepG2. METHODS: HepG2 cells in culture were incubated for 24 h with estrogen or phytoestrogen and LDL receptor activity was measured by examining the cellular binding of colloidal gold-labelled LDL. RESULTS: 17Beta-estradiol significantly increased LDL receptor activity whereas estriol had negligible effects. Incubation with the isoflavonoids, formononetin, biochanin A and daidzein, caused significant elevations in receptor activity at concentrations above 40 microM. Coumestrol, a coumestan with a high level of estrogenic activity, caused a 3-fold increase in receptor activity at a concentration of 50 microM. Of the phytoestrogenic mammalian lignans enterolactone and enterodiol, only enterolactone displayed the ability to significantly upregulate LDL receptor activity at 50 microM. CONCLUSION: This study suggests that the LDL receptor-stimulating effect of natural estrogens is mainly due to estradiol and that the cholesterol-lowering effect of diets high in phytoestrogens may be due in part to their ability to increase hepatic LDL receptor activity.     

Identification of a calcium-binding protein as a calcium-dependent regulator of brain adenylate cyclase.

An activating factor of adenylate cyclase (EC 4.6.1.1) HAS BEEN OBTAINED FROM DETERGENT-DISPERSED PREPARATIONS OF PORCINE CEREBRAL CORTEX BY COLUMN CHROMATOGRAPHY ON ECTEOLA-cellulose. The factor was identified by acrylamide gel electrophoresis and by enzyme activation studies as the Ca2+-binding protein that regulates the activity of a brain cyclic nucleotide phosphodiesterase. This Ca2+-binding protein confers a Ca2+-dependent activation upon the adenylate cyclase, which is reversed by the subsequent addition of egta in excess of the free Ca2+. It is proposed that this Ca2+-dependent regulator controls enzymatic activities responsible for the synthesis of adenosine 3':5'-monophosphate and for the hydrolysis of guanosine 3':5'-monophosphate.     

Plant growth-promoting hormones activate mammalian guanylate cyclase activity

In vivo injections of plant growth-promoting hormones increase the growth of animals as well as plants. Plant growth-promoting hormones and positive plant growth regulators are known to increase RNA and protein synthesis. Since cyclic GMP also increases RNA and protein synthesis, the object of the present investigation was to determine whether physiological levels of plant growth-promoting hormones and positive plant growth regulators have part of their mechanism(s) of action through stimulation of the guanylate cyclase (EC 4.6.1.2)-cyclic GMP system. Representatives of the three classes of growth-promoting hormones were investigated. Thus, auxins (indole-3-acetic acid, indole-3-butyric acid, beta-naphthoxyacetic acid, and 2,4,5-trichlorophenoxy acetic acid), gibberellins (gibberellic acid), and cytokinins [N6-benzyl adenine, kinetin (6-furfuryl aminopurine), and beta-(2-furyl) acrylic acid] all increased rat lung, small intestine, liver, and renal cortex guanylate cyclase activity 2- to 4-fold at the 1 microM concentration. Dose response curves revealed that maximal stimulation of guanylate cyclase by these plant growth regulators was at 1 microM; there was no augmented cyclase activity at 1 nM. The guanylate cyclase cationic cofactor manganese was not essential for augmentation of guanylate cyclase by these plant growth-promoting regulators. The antioxidant butylated hydroxytoluene did not block the enhancement of guanylate cyclase by these plant growth-promoting factors. These data suggest that guanylate cyclase may play a role in the mechanism of action of plant growth-promoting hormones and even of positive plant regulators at the cellular level.     

Regulation of hepatic nuclear guanylate cyclase.

In immunohistochemical studies of rat liver tissue slices and purified nuclei, adenosine 3':5'-cyclic monophosphate (cAMP) and guanosine 3':5'-cyclic monophosphate (cGMP) immunofluorescence on the nuclear membrane are sequentially increased after glucagon administration. An explanation for the increased cGMP immunofluorescence was sought in experiments in which guanylate cyclase [GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2]activity of hepatic subcellular fractions was determined. The results showed that a nuclear guanylate cyclase exists which can be distinguished from the soluble and crude particulate guanylate cyclases. The activity of the nuclear enzyme was increased by 35% in nuclei isolated from rats 30 min after glucagon injection, the time at which maximal nuclear membrane cGMP immunofluorescence is observed. Because glucagon altered both cAMP location and levels prior to the observed changes in nuclear cGMP metabolism, the hypothesis that cAMP acted as the second messenger was tested. In vitro incubation of nuclei isolated from control rats with 10(-5) M cAMP produced a 25% increase in nuclear guanylate cyclase activity. With nuclei isolated from glucagon-treated rats, no significant increase in enzyme activity was observed; this indicates that maximal stimulation of nuclear guanylate cyclase by cAMP occurred at levels that are obtained in vivo after glucagon administration. These findings suggest that hepatic nuclear cGMP content may be regulated by a specific organelle guanylate cyclase and that cAMP may be one of the determinants of this enzyme's activity.     

Effects of atrial natriuretic factor and vasopressin on cyclic nucleotides in cultured kidney cells

The cellular mechanism of the action of atrial natriuretic factor (ANF) is thought to involve activation of guanylate cyclase. Increasing evidence shows a direct tubular effect of ANF. Part of the ANF-induced diuresis has been suggested to be due to inhibition of the action of arginine vasopressin (AVP) in the cortical collecting tubule. In this study we investigated the effect of ANF on cyclic nucleotide production in primary cultures of cortical collecting tubule cells immunodissected with a monoclonal antibody. ANF caused a dose-dependent stimulation in cyclic guanosine 3',5'-monophosphate (cGMP) production; the half-maximal stimulation was observed at approximately 1 nM of ANF. ANF (0.01-100 nM) had no effect on cyclic adenosine 3',5'-monophosphate (cAMP) accumulation in cortical collecting tubule cultures. AVP caused a dose-dependent increase in cAMP production, and this effect was not altered by the simultaneous addition of ANF (100 nM). Similarly, ANF-induced cGMP stimulation was not influenced by AVP (10 nM). We conclude that 1) ANF has a direct stimulatory action on cGMP production by cultured cortical collecting tubule cells and 2) any interaction between ANF and AVP is likely to occur at steps distal to cyclic nucleotide formation.     

Regulation of a Plant 5′(3′)-Ribonucleotide Phosphohydrolase by Cyclic Nucleotides and Pyrimidine, Purine, and Cytokinin Ribosides

A highly specific 5'(3')-ribonucleotide phosphohydrolase has been purified extensively from wheat seedling leaves. While catalyzing the hydrolysis of a wide variety of phosphomonoesters, the enzyme has K(m) values for adenosine 5'-monophosphate and adenosine 3'-monophosphate in the micromolar range and appears highly specific for 5'- and 3'-ribonucleoside monophosphates as substrates at pH 5.0. The cyclic nucleotides adenosine 3':5'-cyclic monophosphate, guanosine 3':5'-cyclic monophosphate, and 8-bromoadenosine 3':5'-cyclic monophosphate, as well as 8-bromoadenosine 5'-monophosphate, are powerful competitive inhibitors of the enzyme; the apparent K(i) values for these nucleotides are 3.4 muM, 1.6 muM, 26 muM, and 9.1 muM, respectively. The enzyme is inhibited noncompetitively by a variety of pyrimidine and purine (including cytokinin) ribosides and 2'-deoxyribosides. Since the cyclic nucleotide competitive inhibitors are also active growth promoters of higher plants, and, furthermore, since elevation of cyclic AMP levels appears to be a consequence of some phytohormone treatments that promote plant growth, it is suggested that negative control of this 5'(3')-ribonucleotide phosphohydrolase may be a significant component of growth regulation through the maximizing of the levels of nucleotide precursors for RNA and DNA synthesis.     

Relationship between the inhibition of receptor-induced increase in cytosolic free calcium concentration and the vasodilator effects of nitrates in patients with congestive heart failure

Nitroglycerin, isosorbide dinitrate and sodium nitroprusside, like nifedipine, were found to inhibit the receptor-provoked increase of cytosolic free calcium concentration in human platelets loaded with 2-[(2-amino-5-methylphenoxy)methyl]-6-methoxy-8-aminoquinoline-N,N,N',N' - tetraacetate. Sodium nitroprusside and nitroglycerin induced elevation of cyclic guanosine 3',5'-monophosphate content in platelets which correlated with their calcium-blocking activity. Methylene blue and epinephrine decreased the calcium-blocking effect and the influence of nitroglycerin on cyclic guanosine 3'-5'-monophosphate content, but failed to suppress the inhibitory effect of sodium nitroprusside. Ascorbic acid increased the calcium blocking effect of sodium nitroprusside and its influence on cyclic guanosine 3'-5'-monophosphate content, but did not alter the inhibitory effect of nitroglycerin. In order to evaluate the relationship between the mode of action of nitrates at cellular level and their vasodilatory effectiveness, we studied the circulatory response of the forearm to isosorbide dinitrate and the influence of nitroglycerin on free calcium concentration in the platelets in 10 patients with chronic heart failure. We established a significant positive correlation between the basal values for venous tone and its peak decrease after administration of the 10-mg dose of isosorbide dinitrate. A correlation was also found between the deviation of maximal decrease of venous tone by this dose of isosorbide dinitrate from the regression line (the relationship between the basal venous tone and its lowering by the drug) and mean inhibitory concentration values for nitroglycerin in blocking that proportion of the rise of calcium ion concentration in platelets due to blocking of the platelet-activating factor. Thus, nitrates, like calcium antagonists, inhibit the receptor-provoked calcium supply to the contractile system of the cells so neutralizing the effects of increased concentrations of vasoconstrictors. This suggests that the effectiveness of nitrates appears to be positively related to the contribution of receptor-induced increase of cytosolic free calcium concentration in vasoconstriction together with their capacity to raise cyclic guanosine 3',5'-monophosphate.     

Adenosine 3',5'-monophosphate and the regulation of rat hepatic sterol synthesis: a reexamination based on Sutherland criteria.

Many investigators have reported that adenosine 3',5'-monophosphate (cyclic AMP) at supraphysiologic concentrations (10(-4) M) suppresses lipogenesis in rat livers in vitro. The data in this report demonstrate that the phosphodiesterase inhibitor theophylline does not suppress sterol synthesis by itself nor does it potentiate the cyclic AMP-mediated suppression. In addition the cyclic AMP-mediated effect was not specific as many structurally-related compounds including adenosine triphosphate, adenosine 5'-diphosphate, adenosine 5'-monophosphate, adenosine, coenzyme A, HMG coenzyme A, and guanosine 3',5'-monophosphate suppressed sterol synthesis. The phenomenon of cyclic AMP-suppression of sterol synthesis does not fulfill two of the four criteria defined by Sutherland to assess whether a given metabolic effect is mediated by cyclic AMP. Therefore, we conclude that the cyclic AMP mediated suppression of sterologenesis is not a physiologic effect of the nucleotide.     

Biochemical mechanism of organic nitrate action.

Increasing evidence suggests that organic nitrate action derives from their metabolic conversion to nitric oxide (NO) in the vascular smooth muscle cell. The primary catalytic activity of this process appears to reside at the cellular plasma membrane. There is no concrete evidence to indicate that NO formation is preceded by the production of inorganic nitrite ion or that the NO produced needs to form S-nitrosothiols before it can activate guanylate cyclase to produce cyclic guanosine 3',5'-monophosphate (cGMP). Although sulfhydryl donors can partially reverse nitroglycerin-induced tolerance in patients, this phenomenon (by itself) is not sufficient to implicate intracellular sulfhydryl depletion as an operating mechanism of clinical nitrate tolerance. This is because sulfhydryl donors can react with nitroglycerin extracellularly to form S-nitrosothiols, and nonsulfhydryl compounds, such as enalapril and hydralazine, can prevent the development of in vivo nitrate tolerance. In addition to the cellular biochemical reactions, organic nitrates also produce systemic biochemical effects through altering neurohormonal status. These systemic effects may contribute significantly to the development of nitrate tolerance in therapeutic situations.     

Epidermal growth factor enhances guanylate cyclase activity in vivo and in vitro

Epidermal growth factor (EGF) increases DNA synthesis and cell division both in vivo and in vitro. The mechanism by which EGF increases growth and DNA synthesis is unknown. Since the intracellular messenger cGMP stimulates DNA synthesis, the present investigation was designed to determine if EGF might have part of its mechanism of action through activating guanylate cyclase [EC 4.6.1.2], the enzyme that catalyzes the formation of cGMP. EGF enhanced soluble and particulate guanylate cyclase activities as well as cGMP levels 2- to 3-fold in hypophysectomized and nonhypophysectomized tissues both in vivo and in vitro. EGF increased guanylate cyclase activity 0.5 h after ip injection in mice, and this increased activity was still present 12 h later. Guanylate cyclase activity was increased to a greater extent secondary to EGF in hypophysectomized cecum compared to nonhypophysectomized cecum. Dose-response curves revealed that maximal stimulation of guanylate cyclase by EGF occurred at 1 nM. There was no augmented guanylate cyclase activity when the concentration of EGF was decreased to 0.01 nM. The data in this investigation suggest that guanylate cyclase may play a role in the mechanism of action of EGF.     

Guanylate cyclase activity in fetal, neonatal and adult rat heart.

Guanosine 3′, 5′-monophosphate is thought to be an important factor in promoting cell growth. We, therefore, measured the activity of guanylate cyclase, the enzyme which in the developing rat heart catalyzes the production of guanosine 3′, 5′-monophosphate. Cardiac guanylate cyclase activity was found to be increased to 405 ± 44 pmol accumulated/10/min/mg protein in utero and maintained this level of activity for the first 3 weeks after birth. By 28 days, cardiac guanylate cyclase activity had decreased to 165 ± 7 pmol accumulated/10 min/mg protein which paralleled the adult mother cardiac guanylate cyclase activity of 173 ± 9 pmol accumulated/10 min/mg protein. This period of maximal activity of cardiac guanylate cyclase corresponds to the period of rapid growth of the heart and the decreased activity approximates the period of time when cardiac DNA synthesis declines.     

Cimetidine partially blocks gastrin's activation of gastric mucosal guanylate cyclase

The objective of the present investigation was to determine if gastrin at physiological concentrations has part of its mechanism(s) of action through stimulation of guanylate cyclase (EC 4.6.1.2). Human gastrin (I), pentagastrin, tetragastrin, and gastrin-related tetrapeptide all increased cyclic GMP levels and guanylate cyclase activity in rat gastric mucosa, whole stomach, and duodenum. Maximal stimulation was seen at 1 microM with all of the above. There was no further enhancement of guanylate cyclase with increasing the concentration to the millimolar range. The ED50 for human gastrin and pentagastrin was 0.01 microM, whereas the ED50 was 0.1 microM for tetragastrin and the tetrapeptide. No enhancement of guanylate cyclase activity was seen with decreasing the concentration to 1 nM of the respective gastrins. Cimetidine utilized at 1 microM or 1 mM concentrations partially blocked the augmentation by gastrin suggesting that part of this enhancement was through the histamine 2 receptor which has been shown to be important in pentagastrin-stimulated gastric acid release. Since the block was only partial these data would also indicate that some part of gastrin's activation of this enzyme is not mediated through the histamine 2 receptor.     

Selective inhibition of rat and human cardiac guanylate cyclase in vitro by doxorubicin (adriamycin): possible link to anthracycline cardiotoxicity.

The cardiotoxicity of anthracycline antibiotic anti-tumor agents is well-described but the molecular basis of the cardiotoxicity is not understood. We examined the effect of doxorubicin (Adriamycin) on the activity of guanylate cyclase (E.C. 4.6.1.2), the enzyme catalyzing the production of guanosine 3′,5′-monophosphate, from rat heart, liver, lung, kidney, and spleen. Doxorubicin produced a decrease in cardiac guanylate cyclase activity over the concentration range 0.4 to 2 mm but was without effect, or slightly stimulated, guanylate cyclase from the other tissues. Daunorubicin (Daunomycin), a related, cardiotoxic anthracycline antibiotic also decreased cardiac guanylate cyclase activity over the concentration range 0.8 to 4 mm. Other antibiotic anti-tumor agents which are not cardiotoxic, including streptonigrin, porfiromycin, and mitomycin C did not decrease cardiac guanylate cyclase activity. Doxorubicin, 1 mm and 2 mm, and daunorubicin, 4 mm decreased cardiac guanylate cyclase approximately 50% in experiments utilizing guanylate cyclase prepared from human heart. The data suggests that some aspects of anthracycline cardiotoxicity may be related to altered cardiac guanylate cyclase activity.     

Uterine estrogen sulfatase may play a more important role than the hepatic sulfatase in mediating the uterotropic action of estrone-3-sulfate

The estrogenic activity of sulfonated estrogens results from the release of active estrogens via desulfonation (hydrolysis) catalyzed by estrogen sulfatase. In this study, the relative importance of uterine or hepatic estrone (E₁)-3-sulfatase in mediating the uterotropic action of E₁-3-sulfate is evaluated by comparing its hormonal potency in animals that have comparable uterine E₁-3-sulfatase activity but markedly different hepatic enzyme activity. Liver microsomes from immature or adult female Sprague-Dawley rats contained 12-or 55-fold higher E₁-3-sulfatase activity, respectively, than the liver microsomes from immature or adult female CD-1 mice. In contrast, uterine whole homogenates from immature female Sprague-Dawley rats contained approx twofold higher E₁-3-sulfatase activity than was detected in the uterine whole homogenates from immature female CD-1 mice. It is estimated that the total E₁-3-sulfatase activity in the liver of an immature female rat or mouse is approx 1080-or 260-fold higher, respectively, than the activity in the uterus. The ED₅₀ values for the uterotropic effect of E₁-3-sulfate and E₁ in immature female CD-1 mice were 240 and 8 pmol/g body wt, respectively, and the corresponding ED₅₀ values in immature female Sprague-Dawley rats were 840 and 60 pmol/g body wt, respectively. The difference in the ratios of the uterotropic ED₅₀ for E₁-3-sulfate over that for E₁ in immature rats and mice (14 and 30, respectively) is 1.14-fold, which correlates very closely with their difference in the uterine E₁-3-sulfatase activity (approx twofold), but not their difference in the hepatic sulfatase activity (approx 12-fold). The results of this study provide evidence suggesting that E₁-3-sulfatase in the uterus (an estrogen target organ) may play a more important role than the hepatic sulfatase in mediating the uterotropic action of sulfonated estrogens.     

Effects of cyclic adenosine 3':5'-monophosphate and related agents on acid secretion by isolated rabbit gastric mucosa.

The effects of various adenosine phosphate compounds, theophylline, histamine,a nd metiamide, on steady rates of acid secretion by isolated fundic mucosa of the rabbit were measured by the pH stat method. Cyclic adenosine 3':5'-monophosphate (cyclic AMP), N(6), O(2')-dibutyryl adenosine 3':5'-monophosphate and theophylline increased the rate of acid secretion. Addition of theophylline in a concentration which has no stimulatory effect, reduces the effective concentrations cyclic AMP or histamine required for stimulation of acid secretion. Measurements of lactate, pyruvate, and CO2 appearances indicated that the increases in acid secretory rates were predominantly due to H+ and not organic acid accumulation in the luminal bath-secretion. Metiamide prevented the stimulatory effects of histamine and ATP. However, metiamide did not prevent the stimulatory effects of N(6),9(2')-dibutyryl adenosine 3':5'-monophosphate, theophylline, or 5'-AMP. The results provide further evidence for a role of cyclic AMP in governing the rate of acid secretion by rabbit stomach. The data also are consistent with histamine and ATP (at least in the concentration used) requiring adenylate cyclase activity for stimulation of acid secretion and 5'-AMP inhibiting phosphodiesterase activity.