The role of periodic changes in cyclase and protein kinase activity in the mechanism of the proliferative effect of ACTH

A prolonged effect of ACTH on the state of adenylate and guanylate cyclase systems in the adrenal glands of experimental animals was investigated. It was found that in guinea pigs injected with ACTH (4 units daily for 1-50 days) the weight of adrenal glands and the DNA content in these organs increased 2.0-2.5-fold by the end of experiment; the increase in both values was stepwise. The corticosteroid level in the blood varied throughout the experiment: the changes in the DNA content in adrenals and in the corticosteroid content in the blood were oppositely directed. This was accompanied by cyclic changes in the basal and stimulated activities of adenylate and guanylate cyclases and proteinases in the adrenal glands occurring with a periodicity of 6-15 days. The activity peaks for cyclases and protein kinases preceded the rise in the DNA content in the adrenals. A clearcut correlation between the changes in the enzyme activity and the hormone dose was observed. The changes in the basal and stimulated activities of guanylate cyclase seem to be due to the control of cAMP level in the cell (stimulation of cGMP-dependent cAMP phosphodiesterase). Apparently, the periodic changes in the activity of cAMP-dependent protein kinases in the cytoplasmic and nuclear fractions and a relatively high activation of nuclear protein kinases (by 30-60%) in comparison of cytoplasmic ones (8-10%) are related to stimulation of DNA synthesis. It is concluded that the changes in the activity of cyclases and protein kinases play a role in the mechanism of proliferative effect of ACTH.     

ACTH-induced refractoriness in cultured adrenal cell line (Y1).

Treatment of cultured mouse adrenal cells Y₁ with ACTH induced cell refractoriness to further hormonal stimulation. When ACTH was added to the cells every 2 hours the first addition increased the levels of 20αOH-progesterone and cAMP secreted into the medium. Upon the second and third additions of ACTH the levels of 20αOH-progesterone and cAMP secreted were greatly diminished and upon the fourth addition of ACTH were absent. Prolonged incubation (14 hours) with different concentrations of ACTH (5 × 10^?11 M to 10^?6 M) induced a dose-related steroidogenic refractoriness to further ACTH stimulation, 10^?8 M ACTH inducing complete refractoriness. The number of ACTH binding sites of cell particles prepared from desensitized cells was similar to that of the control but ACTH failed to stimulate the adenylate cyclase of desensitized cells, whereas the enzyme responded fully to NaF and Gpp(NH)p. The cAMP phosphodiesterase activity was similar in both desensitized and control cells. In addition the steroidogenic response to dibutyryl cAMP of desensitized cells was abolished. Thus, ACTH-induced adrenal cell desensitization seems to be related to at least two phenomena : a defect in the “coupling” between the hormone-receptor sites and the adenylate cyclase and an alteration of certain steps beyond cAMP formation.     

Adenylate cyclase activity and cyclic AMP production in the outer and inner zones of the adrenal cortex

It has been reported that cells isolated from the inner zone of the guinea pig adrenal cortex fail to have a steroidogenic response to ACTH. To further explore this, adenylate cyclase activity of membrane particles and cAMP production by cells prepared from the inner and outer adrenocortical zones were determined. The cAMP response to ACTH and forskolin was similar for cells from both zones. Basal adenylate cyclase activity was significantly higher in the inner zone; and while absolute responses to ACTH, GppNHp, GTP, NaF, and forskolin were greater for the inner zone, relative responses were similar for the two zones. These observations suggest that the inner zone of the guinea pig adrenal cortex may have a defect in ACTH action at a step(s) beyond cAMP formation.     

Ring-deleted analogs of atrial natriuretic factor inhibit adenylate cyclase/cAMP system. Possible coupling of clearance atrial natriuretic factor receptors to adenylate cyclase/cAMP signal transduction system

We have recently shown that atrial natriuretic factor (ANF) inhibits adenylate cyclase activity in rat platelets where only one population of ANF receptors (ANF-R2) is present, indicating that ANF-R2 receptors may be coupled to the adenylate cyclase/cAMP system. In the present studies, we have used ring-deleted peptides which have been reported to interact with ANF-R2 receptors also called clearance receptors (C-ANF) without affecting the guanylate cyclase/cGMP system, to examine if these peptides can also inhibit the adenylate cyclase/cAMP system. Ring-deleted analog C-ANF4-23 like ANF99-126 inhibited the adenylate cyclase activity in a concentration-dependent manner in rat aorta, brain striatum, anterior pituitary, and adrenal cortical membranes. The maximal inhibition was about 50-60% with an apparent Ki between 0.1 and 1 nM. In addition, C-ANF4-23 also decreased the cAMP levels in vascular smooth muscle cells in a concentration-dependent manner without affecting the cGMP levels. The maximal decrease observed was about 60% with an apparent Ki of about 1 nM. Furthermore, C-ANF4-23 was also able to inhibit cAMP levels and progesterone secretion stimulated by luteinizing hormone in MA-10 cell line. Other smaller fragments of ANF with ring deletions were also able to inhibit the adenylate cyclase activity as well as cAMP levels. Furthermore, the stimulatory effects of various agonists such as 5'-(N-ethyl)carboxamidoadenosine, dopamine, and forskolin on adenylate cyclase activity and cAMP levels were also significantly inhibited by C-ANF4-23. The inhibitory effect of C-ANF4-23 on adenylate cyclase was dependent on the presence of GTP and was attenuated by pertussis toxin treatment. These results indicate that ANF-R2 receptors or so-called C-ANF receptors are coupled to the adenylate cyclase/cAMP signal transduction system through inhibitory guanine nucleotide regulatory protein.     

Steroidogenesis in adrenal glands of young and adult female mice with hyperexpression of the Aguoti gene

Dominant mutation Agouti yellow (AY) leads to ectopic overexpression of the Agouti gene and yellow coat color in mice. Furthermore, the mutation Ay increased adrenal response to emotional stress. The study assessed whether pleiotropic effect of the mutation Ay on adrenals function was dependent on sex and age. 3- and 15-week old female C57B1/6J mice of two agouti-genotypes: Ay/a (ectopic Agouti-gene overexpression) and a/a (absence of Agouti-protein), were investigated. Cyclic AMP level (adenylate cyclase activity) and corticosterone production in adrenal isolated cells stimulated by ACTH and dibutyrul cAMP (db-cAMP) were measured. ACTH increased cAMP accumulation to the same extent in Ay/a- and a/a-mouse adrenal cells of both ages. The dibutyrul cAMP-induced corticosterone production was higher in Ay/a than in a/a-mouse adrenal cells of both ages. The ACTH-induced corticosterone production in 3-week- old Ay/a-m/CQ was lower and in 15-week old Ay/a-mice was higher than in a/a-mice of the respective ages. The ACTH- and db-cAMP-induced steroidogenesis was not changed in Ay/a-mice and decreased in a/a-mice with age. Thus, in females as well as in males, the mutation Agouti yellow did not affect adenylate cyclase activity, increased db-cAMP-induced corticosterone production and disturbed development of adrenal cortex.     

Adrenal adenylate cyclase and steroidogenic activities of 63 day old ovine fetuses: in vitro effects of ACTH1-24 and forskolin

This study examines the activity of the adenylate cyclase system and that of some enzymes of the steroidogenic pathway of adrenal cells from 62-63 day old ovine fetuses. Synthetic corticotropin (ACTH1-24), cholera toxin and forskolin stimulated both cAMP and corticoid productions by freshly isolated adrenal cells. The cAMP response to ACTH1-24 was lower than that to forskolin. However, forskolin-induced steroidogenesis was significantly lower than the ACTH1-24-induced steroid output. Freshly isolated cells metabolized quickly [14C]-labeled pregnenolone mainly through the 17-deoxy pathway. The amounts of cortisol and of corticosterone formed, in the presence of exogenous pregnenolone, were roughly 15-fold higher than under maximal stimulation by ACTH1-24. When the cells were cultured for 6 days in the absence or presence of ACTH1-24 (10(-8) M) or forskolin (10(-5) M), a small development of the cAMP response to these factors was observed in the course of the experiment. However, the mechanism of this development appeared different, according to the conditions of culture. The amounts of corticosterone secreted on day 6 by ACTH1-24- or forskolin-treated cells were 2- to 4-fold higher than on day 1, whereas cortisol outputs were much lower on day 6 than on day 1. The response to ACTH1-24 of cells maintained in ACTH-free media decreased dramatically during the culture in terms of both cortisol and of corticosterone. On day 6 of the experiment, the metabolism of [14C]pregnenolone was lower than on day 1 under all 3 conditions of culture. Only the 3 beta-hydroxysteroid dehydrogenase/isomerase activity could be maintained by continuous treatment with forskolin. However, both ACTH1-24 and forskolin enhanced the production of pregnenolone from an endogenous substrate. In conclusion, these results present evidence that: 1) the adenylate cyclase system is not a bottleneck in the steroidogenic response to ACTH1-24 of freshly isolated adrenal cells from 62-63 day old ovine fetuses; 2) the main rate-limiting step for steroidogenesis by these cells is the availability of pregnenolone; 3) neither ACTH1-24 nor forskolin is able to maintain the activity of most enzymes involved in the metabolization of pregnenolone by cultured cells while increasing pregnenolone availability; 4) some inhibiting factors are involved in the loss of adrenal cells responsiveness to ACTH between days 50 and 100 of gestation, and they probably act mainly on the adenylate cyclase system.     

Hormonal control of the adenyl cyclase activity of adipose cell membranes prepared from badger, rabbit, fox and rat adipose tissues]

1. We have shown differences in hormonal regulation of adenylate cyclase activity in fat cell ghosts prepared from rat, rabbit, fox and badger adipose tissue, under the influence of catecholamines, ACTH and insulin. a) In the rat, catecholamines induced a large stimulation (+315%) of adenylate cyclase. b) In the rabbit, ACTH was the most effective hormone. c) In the fox and the badger, only catecholamines could stimulate adenylate cyclase. d) In both rat and rabbit, insulin did not reduce spontaneous enzymatic activity. Moreover, the activation of adenylate cyclase by ACTH in the rabbit was not altered by insulin, while in the rat, this hormone slightly decreased epinephrine stimulation. 2. Hormonal regulation of adenylate cyclase correlated with the lipolytic response.     

Effects of protease inhibitors on adenylate cyclase activation and aldosterone production in rat adrenal zona glomerulosa cells

It has been shown that serine proteases are involved in aldosterone and 18-hydroxycorticosterone production by the rat adrenal zona glomerulosa in response to a variety of stimulants. From evidence presented for various tissues, including the rat adrenal cortex, the observation that adenylate cyclase can be activated by proteolytic enzymes and inhibited by protease inhibitors has led to the suggestion that serine proteases may also be involved in the hormonal stimulation of adenylate cyclase. In studies designed to test this hypothesis using protease inhibitors, only high concentrations (greater than 10(-4) M) of TAME (p-tosyl-L-arginine methyl ester) inhibited ACTH stimulated steroid and cAMP production in rat adrenal glomerulosa cells. TPCK (tosyl-L-phenylalanine chloromethylketone) and TLCK (tosyl-L-lysine chloromethylketone) were found to have a similar effect at very high concentrations (10(-2) M) but had no effect at the serine protease inhibitory concentration of 5 X 10(-6) M. Other protease inhibitors tested had no effect on ACTH-stimulated cAMP but the inhibitory effect of high concentrations of protease inhibitors on ACTH-stimulated adenylate cyclase was duplicated by the polyanion dextran sulphate. The results suggest that the inhibitors act through non-specific membrane effects and that proteases are not involved in the activation of zona glomerulosa adenylate cyclase by ACTH. In view of these findings it is concluded that a more rigorous approach should be applied to the use of protease inhibitors in whole cell systems, and that the concept of hormonal activation of adenylate cyclase via proteolytic events, which is based on studies with such inhibitors, should be reconsidered.     

Systematic shut-off of the hormone receptors in intraspecific adrenal x Leydig cell hybrids

The mouse Y1 adrenal cell line was fused with mouse Leydig cells in primary culture. The selected hybrids were examined for their response to gonadotropin (hCG) and ACTH. None of them bound specifically [125I]hCG, nor did they augment their cAMP production in response to gonadotropin or ACTH stimulation, whereas their adenylate cyclase remained responsive to forskolin and cholera toxin, thus indicating a repression of hCG receptor synthesis and probably a loss of ACTH receptors, rather than a lesion of the coupling between the hormone receptor complex and the adenylate cyclase. Basal pregnenolone production in 17 hybrids was close to that of Leydig and Y1 cells and was enhanced after 8-bromo adenosine 3',5'-monophosphate (8-Br-cAMP) stimulation in 11 of them. Therefore, the negative control leading to the extinction of both parental functions acts preferentially at the first step of steroidogenesis, i.e., the gene(s) coding for the hormone receptors.     

Adenylate cyclase and cAMP-dependent protein kinase activity in the adrenal glands of animals after chronic administration of ACTH

ACTH, a prolonged action hormone, in a dose of 2.5 mu. was injected into guinea pigs daily for 5-35 days. The adenylate cyclase activity of the crude adrenal membrane fraction and the activity of cAMP-dependent protein kinases in the cytoplasmic fraction were determined. Cyclic changes in the basal and stimulated adenylate cyclase activities occurring with 15-20-day intervals have been established for the first time. The sensitivity of adenylate cyclase to ACTH, NaF and GTP did not change in the course of two cycles. The activity of cAMP-dependent protein kinases increased during the first few days after ACTH administration and decreased after further injections of the hormone. The role of cyclic changes of the enzyme activity in the mechanism of proliferative effect of ACTH is discussed.     

Dynamics of adenylate cyclase desensitization during long-term exposure to the hormone

The dependence of adrenal gland adenylate cyclase desensitization on the dose of in vivo injected ACTH, the time of occurrence and duration of the enzyme refractory period and the dependence of desensitization on the number of ACTH injections were analyzed. The experiments were carried out on guinea pigs injected with prolonged action preparations of ACTH (4 and 6 units) daily for 1-6 days. Intramuscular injections of ACTH caused adenylate cyclase refraction to the repeated action of the hormone. The effect of desensitization was the most conspicuous within the first few hours after hormone injection. The decrease of adenylate cyclase sensitivity and the duration of this effect were found to depend on the ACTH dose as well as on the number of injections. It has been shown for the first time that a single in vivo injection of 0.9% NaCl causes short-term desensitization of adenylate cyclase to the repeated action of much higher doses of ACTH in vitro, presumably due to endogenous ACTH release in response to weak stress exposure. The periodicity of changes in adenylate cyclase sensitivity upon prolonged hormone administration is discussed. Sensitization of the enzyme upon daily short-term exposure to physiological doses of ACTH (administration of 0.9% NaCl for 6 days) was revealed.     

Cellular responsiveness to cyclic AMP in a phosphodiesterase-defective mutant of Dictyostelium discoideum

Responsiveness of Dictyostelium discoideum amoebae to cAMP, a chemotactic mediator, was investigated in a strain defective in cAMP-phosphodiesterase production. Cells were subjected to a high cAMP signal (10(-6) M) in the presence or absence of exogenous phosphodiesterase, and the changes of intracellular cAMP and cGMP concentrations and of adenylate cyclase activity were measured. In the presence of cAMP hydrolysis, both adenylate and guanylate cyclases are transiently activated. In the absence of hydrolysis, the high and constant extracellular cAMP concentration is sufficient to elicit a re-activation of adenylate cyclase a few minutes after the first transient response. In contrast, levels of cGMP remain basal for at least 20 min after termination of the initial response to the cAMP addition.     

Sex-differences in adrenocortical responsiveness during development in rats

It is known that the stress hyporesponsive period (SHRP), which seems to be related to an immature hypothalamo-pituitary-adrenal (HPA) regulatory system, occurs during the first 2 weeks after birth in rats. In the present study, we investigated the effects of sex-steroid hormones on adrenocortical responsiveness to adrenocorticotropic hormone (ACTH) in neonatal rats. The levels of cyclic adenosine 3',5'-monophosphate (cAMP), corticosterone, and adenylate cyclase activity increased with the dose of ACTH in adrenal cells of males and females in vitro. The ACTH responsiveness in adrenal cells increased with age (7-35 days of age), that is, the loss in responsiveness to ACTH just after birth began to recover in 14-35-day-old rats, but the responsiveness in 14-day-old rats was attenuated in males compared with females. Although castration markedly augmented the responsiveness in male rats, testosterone-replacement in the castrated male rats inhibited the enhancement. Furthermore, the responsiveness in 14-day-intact female rats was suppressed by treatment with testosterone. Expression levels of ACTH receptor mRNA in adrenals increased with age in the female rat, but not in the male. Castration enhanced the level of ACTH receptor mRNA to three-fold of that in intact male rats at 14 days of age, but replacement treatment with testosterone in castrated male rats lowered the elevated levels. Testicular androgens are thought to evoke a gender-specific response in neonates, and the temporal decrease of adrenal ACTH-responsiveness might be due to the topically immature adrenal system as well as the central nervous system in mammals.     

The effects of furosemide on adenosine 3':5'-cyclic monophosphate, guanosine 3':5'-cyclic monophosphate and corticosterone production stimulated by adrenocorticotropin in monolayer cultured rat adrenal cells

Furosemide has been reported to have a suppressive effect on ADH-, PTH- and adrenaline-stimulated adenosine 3':5'-cyclic monophosphate (cAMP) production, but the effect on adrenocorticotropin (ACTH) action has not yet been elucidated. In the present study, therefore, the effects of furosemide on cAMP and also on guanosine 3':5'-cyclic monophosphate (cGMP) and corticosterone, stimulated by ACTH in monolayer cultured rat adrenal cells, were investigated. The intra- and extracellular cAMP stimulated by ACTH was dose-dependently suppressed by furosemide within the concentration range of 10(-3) M to 3 X 10(-3) M, and the suppressive effect of the drug was accompanied with decreased corticosterone production. However, non-stimulated basal corticosterone production was not influenced by the drug even at 3 X 10(-3) M. A similar suppressive effect of dibutyryl cAMP-stimulated corticosterone production by 3 X 10(-3) M furosemide was observed. The intracellular cAMP bound to its binding protein in sonicated adrenal cell extract was also suppressed in a very similar dose-dependent manner to total cAMP. However, though the effect on corticosterone production was also observed when the calcium concentration in the loading medium was changed, the magnitude of the effectiveness (percent of control) was relatively constant at each calcium concentration, suggesting that furosemide may not affect the site(s) at which calcium acts. Intracellular cGMP, on the other hand, was increased by 10(-3) M to 3 X 10(-3) M of furosemide, suggesting an intensifying effect of furosemide on guanylate cyclase activity. Dibutyryl cGMP-stimulated corticosterone production was also increased at the same concentration range. These results indicated that furosemide may act not only on adenylate cyclase but also on the additional step(s) to suppress the resultant corticosterone production. In contrast to the effects of furosemide on such cAMP-mediated processes, this drug treatment appeared to enhance cGMP-mediated corticosterone production.     

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.     

Effects of oxytocin and methacholine on cyclic nucleotide levels of rabbit myometrium

The effects of oxytocin and methacholine on cyclic nucleotide levels in estrogen-primed rabbit myometrium were studied in the presence and absence of 1-methyl-3-isobutyl xanthine (MIX), a phosphodiesterase inhibitor. In the absence of MIX, methacholine increased guanosine 3',5'-cyclic monophosphate (cGMP) levels at a time when contraction was decreasing, but had no influence on adenosine 3',5'-cyclic monophosphate (cAMP) levels. In contrast, oxytocin did not elevate cGMP, but rapidly decreased cAMP levels. MIX (1 mM) increased both cAMP and cGMP levels. Oxytocin or methacholine further increased cGMP, indicating activation of guanylate cyclase. Oxytocin- but not methacholine-induced stimulation of guanylate cyclase was abolished in Ca2+-free solution. Oxytocin increased cAMP over the levels produced by MIX alone, whereas methacholine decreased cAMP below the MIX control values; these effects were insensitive to indomethacin. Tissue levels of cGMP and cAMP did not directly correlate with isometric tension. The results also indicate that both oxytocin and methacholine stimulate guanylate cyclase but have opposing effects on adenylate cyclase of rabbit myometrium.     

ACTH sensitivity of isolated human pathological adrenocortical cells: variability of responses in aldosterone, corticosterone, deoxycorticosterone and cortisol production

In vitro aldosterone, deoxycorticosterone, corticosterone and cortisol production of human adrenocortical cells derived from adenomas (Conn's syndrome, Cushing's syndrome), from hyperplastic adrenals (Cushing's syndrome) and from adrenals surrounding aldosteronoma are described. Cells from adenomas causing either Cushing's syndrome or Conn's syndrome harboured the highest basal and ACTH-stimulated corticosteroid production. Adrenocortical cells derived from micronodular hyperplasia causing Cushing's syndrome and cells from cortisol producing adenoma displayed predominantly cortisol and corticosterone secretion both under basal conditions and following stimulation with ACTH. Aldosteronoma cells showed highly variable aldosterone, deoxycorticosterone, corticosterone and cortisol response to ACTH. However, in aldosteronoma cell suspensions, the basal and ACTH-stimulated ratios of aldosterone to cortisol were increased when compared to ratios of steroids produced by cells from other adrenal tissues. Chronic treatment with spironolactone of patients with Conn's syndrome before surgery was associated with a decreased ratio of aldosterone to corticosterone, revealing that 18-hydroxylase in aldosteronoma cells may be inhibited during long-term therapy. Non-tumorous cells isolated from adrenals surrounding aldosteronoma displayed less aldosterone prior to and after stimulation with ACTH than aldosteronoma cells.     

Changes in the activity of adenylate- and guanylate cyclase in different rat tissues after chronic administration of ACTH or cortisol

The state of adenylate and guanylate cyclases in the adrenals, suprarenal fat and ventricular myocardium of the rat was studied under conditions of chronic administration of ACTH or cortisol. By the end of ACTH injections the weight of the adrenals and the DNA content in them increased 6 and 3 times, respectively; both parameters showed a gradual increase. The corticosteroid level in the blood changed throughout the experiment. The changes in the DNA content in the adrenals and in the corticosteroid level in the blood were oppositely directed. This was paralleled with cyclic changes in the basal and stimulated activities of adenylate and guanylate cyclases occurring with a periodicity of 15 days. The peaks of the cyclase activity preceded the increase in the DNA content. Similar cyclic changes in the enzyme activity were observed in the adipose tissue and myocardium. It was supposed that periodic changes in the cyclase activity are the main prerequisites for the growth and replication of cells under conditions of changed hormonal status of the organism.     

Effect of preincubation irradiation on adenylate cyclase activity and cyclic nucleotide content of the thymus in chick ontogenesis

Adenylate cyclase activity and cAMP and cGMP content of thymus have been studied in intact and irradiated (0.029 Gy, prior to incubation) embryos and chickens. The enzyme activity is stimulated during the postnatal development. The changes in the cyclic nucleotide content are undulatory and oppositely directed. It is suggested that the observed radiation-induced stimulation of adenylate cyclase and the reciprocal changes in the cyclic nucleotide content after hatching are related to the increased specific differentiation of thymus cells.     

Activation of guanylate cyclase by E. coli heat-stable enterotoxin (STa). Modulation by NAD and pertussis toxin

The heat-stable enterotoxin (STa) of E. coli activates intestinal guanylate cyclase and leads to increased cGMP levels by an as yet undetermined mechanism. In comparing this cGMP system to other known toxin-mediated alterations in cAMP metabolism, we observed that pertussis toxin caused lower levels of intestinal cGMP synthesis in response to purified STa. Another participant in ADP-ribosylation reactions, NAD, enhanced the ability of STa to activate guanylate cyclase, yet had no effect on basal enzyme activity. Niacinamide and isoniacinamide also had no effect on basal activity, but attenuated the STa activation. These results are discussed in relation to current models of hormone/toxin-sensitive adenylate cyclase, and may suggest an involvement of guanine-nucleotide-binding proteins in intestinal cGMP metabolism.