Effects of calcium ionophore (A-23187) on glucose oxidation and iodide transport in dog thyroid slices.

A calcium ionophore (A-23187, 20 mug/ml) stimulted 14C-1-glucose oxidation in dog thyroid slices to an extent equivalent to that obtained by the optimal concentration of dibutyryl cyclic AMP (1mM). Furthermore, the ionophore augmented the stimulation by dibutyryl cyclic AMP much more than the simple additive effect. The ionophore also enhanced the effect of TSH, but to a lesser extent. Under conditions where organic binding was blocked, T/M ratio of radioiodine concentration was lowered in slices by the ionophore; the findings similar to those obtained with TSH and dibutyryl cyclic AMP. The ionophore exhibited a slightly depressive effect on the basal cyclic AMP level. The elevation by TSH of cyclic AMP levels was also slightly depressed by the ionophore, but statistically insignificant in most cases. These results indicate that calcium ion may play an important role in the TSH regulation of iodide transport and glucose metabolism in the thyroid, in some cases by augmenting the effects of cyclic AMP.     

Thyroid cell responses to thyrotropin and 12-O-tetradecanoyl-phorbol-13-acetate: translocation of protein kinase C and phosphorylation of thyroid cell polypeptide substrates

Not all of the effects of thyroid-stimulating hormone (TSH) on the thyroid are mediated by activation of the adenylate cyclase-cyclic AMP system, indicating that other control systems must also exist. Although a calcium-phospholipid-dependent protein kinase (protein kinase C) and specific substrates had been identified in thyroid tissue, their responsiveness to TSH and other stimulators has not been determined. In thyroid cells which had been preloaded with [32P]orthophosphate, TSH and 12-O-tetradecanoyl-phorbol-13-acetate (TPA) increased the phosphorylation of a 33K polypeptide substrate within 5 min in a dose-dependent fashion. The effect was observed with 1 mU/ml TSH and 3 nM TPA and was maximal with 100 mU/ml TSH and 100 nM TPA. The biologically inactive analog of TPA, 4 alpha-phorbol, had no effect. Isobutylmethylxanthine (IBMX) decreased the phosphorylation of the 33K polypeptide and inhibited the effect of TSH and TPA, indicating that the phosphorylation is not mediated by cyclic AMP. TSH and IBMX, but not TPA, augmented phosphorylation of a 38K polypeptide, suggesting involvement of cyclic AMP. In contrast TPA, but not TSH, increased the phosphorylation of 58K and 28K polypeptides. TSH, but not TPA or 4 alpha-phorbol, elevated the cyclic AMP level of thyroid slices. Incubation of thyroid slices with TSH or TPA significantly decreased protein kinase C activity in the 100,000g cytosol fraction and increased it in an extract of plasma membranes. The effect was present within 5 min and was maximal by 30 min. The effect was observed with 100 mU/ml TSH or 1 nM TPA. The stimulation by TSH or TPA of protein kinase C and its translocation from the cytosol to the plasma membranes of thyroid tissue may provide another mechanism for control of thyroid cell metabolism.     

Effect of thyroid-stimulating hormone and norepinephrine on cyclic AMP levels in human normal thyroids and human thyroid tumors

The effect of TSH (100mU/ml) and norepinephrine (100 muM) on the cyclic AMP levels was studied in 10 human normal tissues, 10 thyroid adenomas and 4 thyroid carcinomas (3 papillary and 1 follicular). Normal tissues responded to TSH with a marked elevation of the cyclic AMP level. Response patterns of 10 thyroid adenomas to TSH were variable; the patterns of 6 cases resembled those of normal tissues, 3 responded mildly, and one had no response to TSH. Thyroid carcinomas had a higher basal level of cyclic AMP than those of normal tissues, although they responded only slightly to TSH. Two among 4 thyroid carcinomas had no response to TSH. Norepinephrine stimulated the accumulation of cyclic AMP in 4 thyroid adenomas and 3 thyroid carcinomas, while it had little effect on normal tissues. Responses to norepinephrine was observed only in thyroid tumors, although they had low response to TSH. It is suggested from these results that tumor cells originating from thyroid follicular cells have a modified response to hormones due to neoplastic alterations.     

In vitro effects of arachidonic acid and of inhibitors of its metabolism on the dog thyroid gland

Incubation of dog thyroid tissue with arachidonic acid (10 to 200 microM) led to the following events: --low conversion to prostaglandins E2 and F2 alpha: 0.07% and 0.02% per hour and 100 mg tissue, respectively --inhibition of the stimulatory effect of low concentrations of TSH on thyroid secretion: the secretory effect of supra-maximal concentrations of TSH and of dB-cAMP was unaffected --inhibition of the cyclic AMP accumulation induced by TSH: this effect was inhibited neither by indomethacin nor by ETYA; cyclic AMP accumulation in response to cholera toxin or PGE1 was unaffected --no effect on cyclic GMP level --stimulation of thyroid proteins iodination. ETYA, but not indomethacin, depressed the iodination of thyroid proteins in resting and stimulated tissue. These data show that arachidonic acid-or a metabolite-can modulate thyroid responsiveness to TSH and suggest that lipoxygenase-products of arachidonic acid metabolism could be involved in thyroid proteins iodination.     

Large effects of preparative techniques on lymphocyte cyclic AMP content.

When a cell suspension is formed by disruption of a pig lymph node into medium, large and transient increases in intracellular cyclic AMP occur. Similar effects are observed when pig lymphocytes are centrifuged and the cell pellets resuspended, or when the cells are subjected to rapid temperature changes. These observations define the conditions required to manipulate the cells while maintaining a stable cyclic AMP concentration. Under these conditions, neither concanavalin A nor ionophore A23187 at mitogenic concentrations have any early effect on cyclic AMP in pig lymphocytes, but small increases in cyclic AMP (less than 2-fold) were observed at supramitogenic concentrations of concanavalin A (50 microgram/ml) or A23187 (500nM). Mouse thymocytes show qualitatively similar but much smaller changes in cyclic AMP concentration in response to experimental manipulations, and no response to mitogenic or supramitogenic concentrations of concanavalin A below the cytotoxic value.     

"In vitro" study on release of cyclic AMP and thyroid hormone in autonomously functioning thyroid nodules

The TSH effect on slice and the incubation medium cyclic AMP levels and T3 and T4 released from 8 autonomously functioning thyroid nodules (AFTN) and their respective perinodular (PN) tissues were examined. The thyroid slices were incubated in Eagle's Medium containing TSH (5 to 100 mU/ml) for 60 min and 300 min for tissue cyclic AMP generation and for cyclic AMP, T3 and T4 release, respectively. Basal cyclic AMP levels were not different either in AFTN and in PN slices or into the incubation medium. In both tissues TSH produced a similar cyclic AMP generation. In contrast, cyclic AMP released into the incubation medium was significantly higher in AFTN than in PN tissues, after TSH stimulation. Basal T3 values and TSH-stimulated T3 release in AFTN were not different from PN tissue. However, basal T4 levels were significantly higher in AFTN than in PN tissue as well as T4 released in response to TSH. In addition, T3/T4 ratio was lower in AFTN than in PN tissues. The cyclic AMP released into the incubation medium correlated with both T3 and T4 release in PN tissue but in the AFTN tissue no correlations were found. These findings suggest that the adenylate cyclase-cyclic AMP system is more sensitive to TSH-stimulation in AFTN when compared with PN tissue and that AFTN tissue has a preferential T4 secretion.     

Effects of cholera toxin on thyroid cyclic AMP-dependent protein kinase and ornithine decarboxylase activities.

Cholera toxin activated beef thyroid cyclic AMP-dependent protein kinase in a dose (0.2 to 8 microgram/ml)-related fashion. Thus, when beef thyroid slices were incubated with toxin (8 microgram/ml) for 90 minutes and then assayed for protein kinase, the activity ratio (i.e. -cyclic AMP/+cyclic AMP) increased from 0.32 +/- 0.02 to 0.77 +/- 0.06. The toxin (5 microgram/ml)-induced increase was abolished by inclusion of ganglioside GM1 in the incubation medium (I50, 0.7 microgram/ml), whereas, gangliosides GD1a and GT1 were without effect. In contrast, TSH-activated protein kinase was unaffected by ganglioside addition. Cholera toxin increased rat thyroid ornithine decarboxylase (ODC) activity in-vitro in a dose (0.1 to 10 microgram/ml)-related fashion [basal, 100 cf cholera toxin (10 microgram/ml), 1500 pmol 14CO2/g tissue/30 min]. The toxin (1 microgram/ml)- (but not TSH-) induced increase in ODC was abolished by inclusion of ganglioside Ga and GT1 were without effect. Cholera toxin stimulation of ODC was inhibited by indomethacin or iodide as are the stimulatory effects of TSH or dibutyryl cyclic AMP. These results demonstrate that although there are differences in the TSH and cholera toxin responses with respect to receptor (ganglioside) interaction, they nevertheless elicit similar intracellular responses in thyroid.     

Chemotaxis in Dictyostelium discoideum: effect of concanavalin A on chemoattractant mediated cyclic gmp accumulation anlight scattering decrease.

In cells of the cellular slime mold Dictyostelium discoideum concanavalin A (Con A), at a concentration of 100 microgram per ml, inhibits folic acid and cyclic AMP induced decrease in light scattering. Con A has no effect on folic acid mediated cyclic GMP accumulation and increases cyclic AMP mediated cyclic GMP accumulation two-fold. At a lower Con A concentration, 10 microgram per ml, changes in light scattering induced by folic acid are normal and cyclic AMP induces a monophasic instead of a biphasic response. The stimulatory effect of Con A on cyclic AMP mediated cyclic GMP accumulation is still observable at 10 microgram Con A per ml. When cells are repeatedly stimulated with cyclic AMP, a decrease in light scattering without being accompanied by changes in cyclic GMP concentration is observed. Based on these results a model for chemotaxis is proposed.     

Concentration of adenosine 3':5'-cyclic monophosphate in mouse pancreatic islets measured by a protein-binding radioassay

A protein-binding radioassay for cyclic AMP was modified to detect less than 0.025pmol of the nucleotide. The method was applied to the measurement of cyclic AMP in small numbers of mouse pancreatic islets (as little as 25μg of tissue) by use of barium acetate–H₂SO₄ for deproteinization. The concentration of cyclic AMP in mouse islets incubated in media containing 3.3 or 20mm-glucose was 0.016pmol/10 islets (approx. 1μm in intracellular water). Glucose concentration (3.3 or 20mm) had no detectable effect on islet concentrations of cyclic AMP with periods of incubation or perifusion ranging from 0.5 to 60min, although insulin release rate was rapidly increased by 20mm-glucose. Caffeine (5mm) or 3-isobutyl-1-methylxanthine (1mm), which are known inhibitors of islet cyclic AMP phosphodiesterase, produced marked and rapid increases in islet cyclic AMP concentration at 3.3 or 20mm-glucose, but only enhanced the insulin release rate at the higher glucose concentration. The role of cyclic AMP in insulin release induced by glucose is discussed.     

In vitro effects of arachidonic acid and of inhibitors of its metabolism on the dog thyroid gland

Incubation of dog thyroid tissue with arachidonic acid (10 to 200 μM) led to the following events:

- low conversion to prostaglandins E₂ and F_2α: 0.07% and 0.02% per hour and 100 mg tissue, respectively

- inhibition of the stimulatory effect of low concentrations of TSH on thyroid secretion: the secretory effect of supra-maximal concentrations of TSH and of dB-cAMP was unaffected

- inhibition of the cyclic AMP accumulation induced by TSH: this effect was inhibited neither by indomethacin nor by ETYA; cyclic AMP accumulation in response to cholera toxin or PGE₁ was unaffected

- no effect on cyclic GMP level

- stimulation of thyroid proteins iodination.

ETYA, but not indomethacin, depressed the iodination of thyroid proteins in resting and stimulated tissue. These data show that arachidonic acid-or a metabolite-can modulate thyroid responsiveness to TSH and suggest that lipoxygenase-products of arachidonic acid metabolism could be involved in thyroid proteins iodination.     

Impairment and restoration of response to TSH in dog thyroid slices after treatment with phospholipase A and lubrol PX.

Incubation of dog thyroid slices with phospholipase A (10-40 U/Ml) or Lubrol PX (0.08-0.4%) caused a diminution in the subsequent TSH effect on the tissue cyclic AMP level and glucose oxidation. The same treatment had no effect on the basal level of these parameters. When the phospholipase A or Lubrol PX-treated slices were rinsed intensively with a Krebs-Ringer bicarbonate buffer and then incubated at 37degreesC in the same buffer for a further 1 to 3 hours, responsiveness to TSH recovered progressively reaching almost completely that of the control slices. Again, these procedures were without any significant effect on the responsiveness of the control slices. The above results together with those reported previously suggest strongly that phospholipids are an essential component of the plasma membrane system by which TSH stimulates adenylate cyclase activity. In addition, these essential lipids in the membrane appear to be renewed rather efficiently in this tissue, thus securing the functional integrity of the thyroid in the face of various deleterious situations.     

Effects of thyrotropin and N6,O2′-dibutyryl cyclic 3′,5′-adenosine monophosphate on prostaglandin levels in thyroid

We have shown that TSH increases PG levels in isolated bovine thyroid cells. We now report that TSH also increases PG levels in rat and mouse thyroid, and that these effects may be mediated via cyclic AMP. PG and cyclic AMP levels in intact rat and mouse thyroid lobes were measured by radioimmunoassay. During 60-min incubations at 37°C, 25 mU/ml TSH effected a 75–83% increase in PGE₁ and PGF_2α ”equivalents“ in rat thyroid; parallel measurements of endogenous cyclic AMP in these intact thyroid lobes revealed that maximal TSH-induced increase in cyclic AMP also required 60-min incubations. In mouse thyroid, 5 mU/ml TSH increased PGE₁ and PGF_2α levels 38–82% above basal; this TSH effect was evident within 15 min of incubation, thus mimicking the time-course of TSH-induced increase in mouse thyroid cyclic AMP. Exogenous DBcAMP, 0.5 to 3 mM, effected a dose-related increase in mouse thyroid PG levels. The stimulatory effects of both TSH and DBcAMP on mouse thyroid PG levels were abolished by aspirin and indomethacin. These studies suggest that TSH-induced increase in endogenous PG levels in thyroid may be mediated by cyclic AMP.     

Growth hormone control of glucose oxidation pathways in hypophysectomized rats.

An in vivo response of glucose oxidation to growth hormone has been demonstrated. Hypophysectomized rats were found to oxidize glucose at rates significantly higher than normal rats. Treatment with growth hormone 1 h before injection of 14C-U-glucose, 14C-6-glucose, or 14C-1-glucose caused a return to a normal oxidation pattern. This acute response was independent of insulin action but clearly time-dependent since no change from untreated hypophysectomized rats appeared when growth hormone was given at various times prior to administration of labeled glucose. The response observed for 14C-6-glucose was comparable to that observed for 14C-1-glucose with regard to dynamics but differed with respect to total 14C recovered as 14CO2. The cumulative percent 14CO2 recovered from oxidation of 14C-6-glucose 1 h after growth hormone injection exceeded that recovered from oxidation of 14C-1-glucose. These results suggest a change in glucose oxidation by a route that cannot be explained solely by changes in either the hexose monophosphate or Embden-Meyerhof pathways.     

Modification of radiation-induced division delay by caffeine analogues and dibutyryl cyclic AMP

The mitotic selection procedure for cell cycle analysis was utilized to investigate the concentration-dependent modification of radiation-induced division delay in Chinese hamster ovary (CHO) cells by methyl xanthines (caffeine, theophylline, and theobromine) and by dibutyryl cyclic AMP. The methyl xanthines (concentrations from 0.5 to 1000 micrograms/ml) all reduced radiation-induced division delay with the effect being linear between approximately 100 and 1000 micrograms/ml. After doses of 100-300 rad, delay was reduced by 75, 94 or 83 per cent at 1000 micrograms/ml for each drug, respectively. However, the addition of dibutyryl cyclic AMP had an opposite effect: radiation-induced delay was increased by the concentration range of 0.3 to 300 micrograms/ml. These results indicate that in mammalian cells the control of cell cycle progression and the modification of radiation-induced division delay are not simply related to intracellular levels of cyclic AMP. Rather, there appear to be at least two competing mechanisms which are differentially affected by caffeine analogues or by direct addition of dibutyryl cyclic AMP. The direct effect of caffeine and the methyl xanthines on membrane calcium permeability is considered.     

Regulation of coenzyme A biosynthesis by glucagon and glucocorticoid in adult rat liver parenchymal cells

We studied the effects of glucagon, dibutyryl cyclic AMP and dexamethasone on the rate of [(14)C]pantothenate conversion to CoA in adult rat liver parenchymal cells in primary culture. The presence of 30nm-glucagon increased the rate by about 1.5-fold relative to control cultures (range 1.4-2.3) and 2.4-fold relative to cultures containing 1-3m-i.u. of insulin/ml. The half-maximal effect was obtained at 3nm-glucagon. Dibutyryl cyclic AMP plus theophylline also enhanced the rate by about 1.5-fold. Dexamethasone acted synergistically with glucagon; glucagon at 0.3nm had no effect when added alone, but resulted in a 1.7-fold enhancement when added in the presence of dexamethasone (maximum effect at 50nm). The 1.4-fold enhancement caused by the addition of saturating glucagon concentrations was increased to a 3-fold overall enhancement by the addition of dexamethasone. However, dexamethasone added alone over the range 5nm to 5mum had no effect on the rate of [(14)C]pantothenate conversion to CoA. The stimulatory effect of dibutyryl cyclic AMP plus theophylline was also enhanced by the addition of dexamethasone. Changes in intracellular pantothenate concentration or radioactivity could not account for the stimulatory effects of glucagon, dibutyryl cyclic AMP or dexamethasone. Addition of 18mum-cycloheximide, an inhibitor of protein synthesis, decreased the rate of incorporation of [(14)C]pantothenate into CoA and the enhancement of this rate by glucagon and dibutyryl cyclic AMP plus theophylline in a reversible manner. These results demonstrate an influence of glucagon, dibutyryl cyclic AMP and glucocorticoids on the intracellular mechanism regulating total CoA concentrations in the liver.     

Cyclic AMP release from perfused dog thyroid lobes as a sensitive indicator of TSH activation of the secretion process. Evidence for two types of thyroid secretion latency

It has been demonstrated in various types of thyroid tissue preparations that cyclic AMP (cAMP) released into the medium reflects the amount of cAMP in the cells. In the present study employing perfused dog thyroid lobes the dynamics of cAMP release were compared to those of thyroxine (T4) and triiodothyronine (T3) release. The experiments gave evidence that even the lowest concentrations of TSH which stimulate hormone release (in this study 1 microU/ml) also activate the cAMP system; the very high levels of cAMP obtained by stimulation with high concentrations of TSH (in this study 10,000 microU/ml) are not accompanied by corresponding high increases in hormone release. On the contrary the T4 and T3 release is lower than during stimulation with more moderate concentrations of TSH (100 microU/ml). Hence studies employing high concentrations of TSH and measurements of cAMP as indicator of activity of secretory processes should be interpreted very cautiously; the prolonged lag in thyroid hormone secretion observed after stimulation with low concentrations of TSH is accompanied by a corresponding lag in activation of the cAMP system. This pattern suggest that the duration of late secretory processes such as thyroglobulin pinocytosis and hydrolysis is independent of the degree of stimulation and not involved in the variations in secretion latency.     

Protein kinase C activating phorbolesters enhance the cyclic AMP response to parathyroid hormone,forskolin and choleratoxin in mouse calvarial bones and rat osteosarcoma cells

The protein kinase C-(PKC) activating phorbol esters 12-O-tetradecanoylphorbol-13-acetate (TPA; 100 nmol/l) and phorbol 12, 13-dibutyrate (PDBU; 100 nmol/l) enhanced basal cyclin AMP accumulation in cultured neonatal mouse calvaria. The cyclic AMP response to parathyroid hormone (PTH; 10 nmol/l) and the adenylate cyclase activators forskolin (1–3 mol/l) and choleratoxin (0.1 mg/ml) was potentiated in a more than additive manner by TPA and PDBU. In contrast, phorbol 13-monoacetate (phorb-13; 100 nmol/l), a related compound but inactive on PKC, had no effect on basal or stimulated cyclic AMP accumulation. In the presence of indomethacin (1mol/l), TPA and PDBU had no effect on cyclic AMP accumulation in calvarial bones per se, but were still able to cause a significant enhancement of the response to PTH, forskolin and choleratoxin. PTH-, forskolin- and choleratoxin-stimulated cyclic AMP accumulation in rat osteosarcoma cells UMR 106-01 was synergistically potentiated by TPA and PDBU, but not by phorb.-13. These data indicate that PKC enhances cyclic AMP formation and that the level of interaction may be at, or distal to, adenylate cyclase.     

Adenosine-Elicited Accumulation of Adenosine 3'', 5''-Cyclic Monophosphate in the Chick Embryo Retina

The cyclic AMP level of 17-day-old chick embryo retina increased from 20 to 331 pmol/mg protein when the tissue was incubated for 20 min in the presence of 4-(3-butoxy-4-methoxybenzyl-2-imidozolinone) (RO 20-1724). The addition of 0.5 mM-3-isobutyl-1-methylxanthine (IBMX) or 0.5 units/ml of adenosine deaminase (EC 3.5.4.4) to the medium reduced the increase of cyclic AMP content from 20 to 100 pmol/mg protein. Dipyridamole did not interfere with the rise of the retinal cyclic AMP level observed with RO 20-1724. The EC50 of 6-amino-2-chloropurine riboside (2-chloroadenosine)-elicited accumulation of cyclic AMP of retinas incubated in the presence of RO 20-1724 plus adenosine deaminase was approximately 1 microM. When retina incubation was carried out in the presence of 0.5 mM-IBMX, the 2-chloroadenosine dose-response curve was shifted to the right two orders of magnitude. Maximal stimulation of the cyclic AMP level of 17-day-old chick embryo retina incubated in the presence of 0.5 mM-IBMX was observed at 1 mM-adenosine concentration. This effect was not blocked by dopamine antagonists. Guanosine and adenine did not affect the retinal cyclic AMP level. AMP and ATP had a slight stimulatory effect. Adenosine response of embryonic retina increased sharply from the 14th to the 17th embryonic day. A similar, but not identical adenosine effect was observed in cultured retina cells.     

Receptor-mediated gonadotropin action in the ovary. Inhibitory actions of concanavalin A and wheat-germ agglutinin on gonadotropin-stimulated cyclic AMP and progesterone responses in ovarian cells.

Pretreatment of ovarian cells with concanavalin A and wheat-germ agglutinin blocked the gonadotropin-induced cyclic AMP and progesterone responses and this effect was time- and concentration-dependent. Basal production of either cyclic AMP or progesterone, however, was not affected by treatment of cells with lectin. The effect of concanavalin A on gonadotropin-mediated cyclic AMP and progesterone responses was blocked by alpha-methyl D-mannoside and alpha-methyl d-glucoside. Similarly the inhibitory effect of wheat-germ agglutinin was reversed by N-acetyl-D-glucosamine. Pretreatment of ovarian cells with concanavalin A or wheat-germ agglutinin had no effect on protein synthesis in the ovary as monitored by [3H]proline incorporation studies. Concanavalin A and wheat-germ agglutinin did not affect steroid production in response to dibutyryl cyclic AMP and 8-bromo cyclic AMP, indicating that the inhibitory action of lectin was occurring at a step before cyclic AMP formation. Lectins specific for L-fucose, D-galactose and N-acetyl-D-galactosamine, gorse seed agglutinin, peanut agglutinin and Dolichos biflorus agglutinin respectively, did not interfere with gonadotropin-induced cyclic AMP and progesterone responses. The present studies suggest that gonadotropin receptors may be glycoprotein in nature or closely associated with glycoprotein structures with the carbohydrate chain containing N-acetyl-D-glucosamine, mannose and possibly N-acetylneuraminic acid.     

Are cyclic nucleotides involved in the initiation of mitogenic activation of human lymphocytes?

In order to obtain more insight into the possible role of cyclic AMP or cyclic GMP in modulating the initial cellular processes following activation of lymphocytes, we measured the effects of the T-cell mitogen concanavalin A and other substances including hormones on the cyclic nucleotide levels in human peripheral blood lymphocytes. The enzyme activities of the corresponding nucleotide cyclases, adenylate cyclase and guanylate cyclase were measured in both isolated plasma membranes or the cytosol of resting or concanavalin A stimulated rabbit thymocytes. Concanavalin A in a mitogenic concentration of about 5-10 micrograms/ml caused small, but consistent increases in cAMP but no changes in cGMP levels during the first hour of activation. Concomitantly, the specific activity of plasma membrane-bound adenylate cyclase was always increased at least 1.5-fold 30 min after stimulation of rabbit thymocytes with concanavalin A, but no effect could be detected on the specific activities of plasma membrane-bound or soluble guanylate cyclase. At high, supraoptimal concentrations of concanavalin A (more than 20 micrograms/ml) cAMP levels dramatically increased in human lymphocytes within minutes, but cGMP levels again were unaffected. Forskolin and beta-adrenergic hormones elevated cAMP in human lymphocytes, whereas cGMP levels were increased by the addition of sodium nitroprusside or alpha-adrenergic hormones. Sodium nitroprusside, in concentrations which elevated cGMP in human lymphocytes, had no influence on the incorporation of [3H]uridine into RNA of resting or concanavalin A stimulated human lymphocytes. Addition of forskolin resulted in an increase of cAMP levels and a dose-dependent decrease of [3H]uridine incorporation into RNA of concanavalin A-stimulated lymphocytes with no effect on resting lymphocytes. The data suggest that cGMP does not play a role in the initial phase of mitogenic activation of lymphocytes, whereas cAMP may be involved in the blast transformation process as an inhibitory signal.