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T Reisine

Publications and source records attributed to T Reisine.

At least 109 records · Page 6Linked to original sources
PubMed1988

Somatostatin receptor subtypes in the clonal anterior pituitary cell lines AtT-20 and GH3.

The functional and biochemical characteristics of somatostatin (somatotropin release-inhibiting factor) (SRIF) receptor subtypes were examined in the clonal pituitary cell lines AtT-20 and GH3. SRIF inhibits evoked calcium influx into each of these cell lines. The rank order of potencies of structural analogues of SRIF to inhibit calcium influx into GH3 versus AtT-20 cells was different. Inhibitory actions of SRIF on calcium influx desensitized in AtT-20 cells but not GH3 cells. The biochemical properties of the SRIF receptor subtypes in AtT-20 and GH3 cells were assessed by photoaffinity labeling of each receptor with the nonreducible SRIF analogue [125I]CGP 23996 and the photocrosslinking agent n-hydroxysuccinimidyl-4-azidobenzoate. The covalently labeled receptors in both cell lines had the same size, 55 +/- 5 kDa, as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The covalent binding of [125I]CGP-23996 to GH3 and AtT-20 cell membranes was blocked by 1 microM SRIF, somatostatin 28, Trp8-SRIF and was GTP sensitive. Analysis of the labeled receptors in GH3 and AtT-20 cell membranes by two-dimensional polyacrylamide gel electrophoresis indicated that they were of similar charge (pI = 6-6.5) and that they comigrate when applied together. Proteolysis of the GH3 and AtT-20 cell SRIF receptors with Staphylococcus aureus V-8 and thermolysin revealed similar peptide maps. Pretreatment of AtT-20 cells with different stable SRIF analogues abolished the subsequent equilibrium or covalent labeling of the SRIF receptor with [125I]CGP-23996. Similar treatment of GH3 cells did not reduce the covalent labeling of the SRIF receptor by [125I]CGP 23996. These studies indicate that the functional characteristics of SRIF receptors in GH3 and AtT-20 cells are different. However, clear differences in the biochemical properties of these receptor subtypes were not observed. Subtle variations in the structure of the SRIF receptors may therefore be responsible for the functional differences.

Affinity Labels↗
PubMed1988

Somatostatin inhibits cAMP-dependent and cAMP-independent calcium influx in the clonal pituitary tumor cell line AtT-20 through the same receptor population.

The characteristics of somatostatin (SRIF) inhibition of calcium influx stimulated by corticotropin releasing factor (CRF), an activator of adenylate cyclase, and K+, a membrane depolarizing agent, in AtT-20 cells were assessed. Changes in cytosolic calcium levels were measured using the fluorescence probe Quin 2. Both CRF and K+ raise cytosolic calcium levels by stimulating calcium influx. SRIF induced an immediate inhibition of CRF and K+-stimulated calcium influx. This effect was concentration-dependent with IC50 values for SRIF's blockade of CRF and K+ stimulation of 64 +/- 13 pM and 100 +/- 15 pM, respectively. The SRIF analogs, SRIF 28, Trp8-SRIF and Tyr11-SRIF had the same rank order of potency to block CRF and K+-induced calcium influx. The inhibitory effects of SRIF on AtT-20 cells were abolished by pertussis toxin pretreatment. SRIF inhibition of both CRF and K+-induced calcium influx desensitized. The desensitization was rapid (T1/2 approximately 2.5 min), dependent on the concentration of SRIF and not due to the degradation of the peptide. The ability of SRIF to block CRF (cyclic AMP-dependent) and K+ (cyclic AMP-independent)-stimulated calcium influx into AtT-20 cells cannot be separated.

Adenylate Cyclase Toxin↗
PubMed1988

Pertussis toxin modifies the characteristics of both the inhibitory GTP binding proteins and the somatostatin receptor in anterior pituitary tumor cells.

The effects of pertussis toxin treatment on the characteristics of somatostatin receptors in the anterior pituitary tumor cell line AtT-20 were examined. Pertussis toxin selectively catalyzed the ADP ribosylation of the alpha subunits of the inhibitory GTP binding proteins in AtT-20 cells. Toxin treatment abolished somatostatin inhibition of forskolin-stimulated adenylyl cyclase activity and somatostatin stimulation of GTPase activity. To examine the effects of pertussis toxin treatment on the characteristics of the somatostatin receptor, the receptor was labeled by the somatostatin analog [125I]CGP 23996. [125I]CGP 23996 binding to AtT-20 cell membranes was saturable and within a limited concentration range was to a single high affinity site. Pertussis toxin treatment reduced the apparent density of the high affinity [125I]CGP 23996 binding sites in AtT-20 cell membranes. Inhibition of [125I]CGP 23996 binding by a wide concentration range of CGP 23996 revealed the presence of two binding sites. GTP predominantly reduced the level of high affinity sites in control membranes. Pertussis toxin treatment also diminished the amount of high affinity sites. GTP did not affect [125I]CGP 23996 binding in the pertussis toxin-treated membranes. The high affinity somatostatin receptors were covalently labeled with [125I] CGP 23996 and the photoactivated crosslinking agent n-hydroxysuccinimidyl-4-azidobenzoate. No high affinity somatostatin receptors, covalently bound to [125I]CGP 23996, were detected in the pertussis toxin-treated membranes. These results are most consistent with pertussis toxin uncoupling the inhibitory G proteins from the somatostatin receptor thereby converting the receptor from a mixed population of high and low affinity sites to only low affinity receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclase Inhibitors↗
PubMed1988

Characteristics of somatostatin desensitization in the pituitary tumor cell line AtT-20.

The molecular mechanisms of somatostatin (SRIF) desensitization were investigated in the anterior pituitary tumor cell line AtT-20. Previous studies have shown that pretreatment of AtT-20 cells with SRIF analogs desensitizes the cells to SRIF inhibition of hormone release, cyclic AMP formation and calcium influx. This desensitization may involve a change in the properties of the SRIF receptors. Pretreatment of AtT-20 cells with Trp8-SRIF reduced the binding of the SRIF analog [125I]CGP 23996 (des-Alal, Gly2-[desamino-Cys3, Tyr11]-3, 14-dicarbasomatostatin) to AtT-20 cell membranes. The loss of [125I]CGP 23996 binding was dependent on the time of Trp8-SRIF treatment and was reversible. The ability of GTP analogs to inhibit [125I]CGP 23996 binding was reduced after Trp8-SRIF treatment, suggesting that the SRIF receptor and the inhibitory G proteins become uncoupled during desensitization. This is indicated further by the decrease in SRIF stimulation of GTPase activity and SRIF inhibition of forskolin-stimulated adenylyl cyclase activity in desensitized membranes. The reduction and recovery of SRIF inhibition of adenylyl cyclase activity after Trp8-SRIF pretreatment has a similar time course as the changes in [125I]CGP 23996 binding. GTP inhibition of forskolin-stimulated adenylyl cyclase activity is also reduced in SRIF-desensitized membranes. The loss of the GTP effect occurs rapidly and does not fully recover after Trp8-SRIF pretreatment. The levels of ADP-ribosylation of inhibitory GTP binding protein, the relative quantity of the alpha subunits of the inhibitory G proteins and their electrophoretic mobility after 2-dimensional gel electrophoretic analysis, are not altered in SRIF-desensitized membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclase Inhibitors↗
PubMed1987

Hormone receptor regulated proopiomelanocortin gene expression.

The activity of the POMC gene is regulated by both stimulatory and inhibitory hormones. Presumably, some balance exists in the influence of these hormones in order to maintain a steady-state level of activity of this gene. Physiological insults, such as stress, may upset this balance and change the rate of production of POMC and its biologically active peptides. The relative strength of these different hormones may therefore determine the long-term expression of this gene. Chronic administration of CRF to rats, primates and humans produces prolonged increases in plasma ACTH levels. This long-term effect is most likely due to an activation of the POMC gene. Interestingly, chronic treatment of anterior pituitary cells with CRF desensitizes CRF receptors. Thus, despite the corticotrophs becoming refractory to the acute stimulatory actions of CRF, the POMC gene remains stimulated. These findings suggest that corticotrophs do not have to be continuously activated by CRF to maintain a long-term increase in POMC gene expression. This contrasts with the actions of glucocorticoids whose effects are abruptly terminated following their removal from the target tissue. The molecular basis of this form of cellular memory to the actions of CRF may involve cAMP regulatory phosphoproteins binding to and activating the POMC gene. If this phenomenon is shown to occur and the phosphorylation state of these nuclear proteins is found to govern their level of interaction with POMC gene, then it would represent a novel mechanism of gene regulation. Proof for this mechanism and the elucidation of how other second messengers such as protein kinase C and calcium regulate the POMC gene will greatly aid our understanding of the precise molecular mechanisms controlling opioid peptide expression.

Animals↗
PubMed1987

Interactions among lithium, calcium, diacylglycerides, and phorbol esters in the regulation of adrenocorticotropin hormone release from AtT-20 cells.

Interactions among lithium, calcium, and phorbol esters in the regulation of adrenocorticotropin hormone (ACTH) release were examined in a tumor cell line (AtT-20) of the anterior pituitary. Lithium, which blocks the phosphatase that converts inositol phosphates (IPs) to inositol, increases the levels of IPs in these cells and stimulates ACTH release. This ion potentiates the ability of calcium, an activator of phospholipase C, to raise levels of IPs in these cells and to stimulate ACTH secretion. Pretreatment of AtT-20 cells with calcium specifically abolishes the ACTH release response to lithium or calcium, a result suggesting that these secretagogues may act through a common mechanism to induce hormone secretion. Prior exposure of AtT-20 cells to either lithium or calcium also attenuates the ACTH release induced by phorbol ester, an activator of protein kinase C. To examine the link among lithium, calcium, phosphatidylinositol (PI) turnover, and phorbol ester-evoked ACTH secretion, AtT-20 cells were treated with 1-oleoyl-2-acetoyl-sn-3-glycerol (OAG), an analogue of the diacylgylcerols that are formed by phospholipase C during PI metabolism and that also activate protein kinase C. OAG itself does not alter ACTH release or the levels of IPs in AtT-20 cells. Pretreatment of AtT-20 cells with OAG, however, selectively blocks the ACTH release response to lithium, calcium, or phorbol ester. Furthermore, such pretreatment reduced the ability of lithium to increase levels of IPs. The results suggest that one mechanism of action of lithium is to potentiate selectively an action of calcium, possibly the stimulation of phospholipase C activity.(ABSTRACT TRUNCATED AT 250 WORDS)

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1987

Translocation of protein kinase C in anterior pituitary tumor cells.

Previous studies have shown that phorbol esters and lithium each stimulate the secretion of adrenocorticotropic hormone (ACTH) by the anterior pituitary tumor cell line AtT20/D16-16. Pretreatment with either lithium or phorbol ester desensitizes the cells to subsequent stimulation by phorbol ester. An early consequence of phorbol ester action in other systems is the translocation of protein kinase C from cytosol to membranes. We have assayed protein kinase C activity in cytosol and membranes of AtT20 cells after treatment with phorbol dibutyrate, lithium, or other agents that stimulate secretion of ACTH in these cells. Phorbol dibutyrate clearly induced translocation of protein kinase C, but lithium treatment did not cause translocation itself, nor did pretreatment with lithium affect the translocation induced by phorbol dibutyrate. These results are consistent with a role for translocation of protein kinase C in the stimulatory and desensitizing effects of phorbol esters but fail to implicate translocation in the actions of lithium on AtT20 cells.

Adrenocorticotropic Hormone↗
PubMed1987

Activators of protein kinase C and cyclic AMP-dependent protein kinase regulate intracellular calcium levels through distinct mechanisms in mouse anterior pituitary tumor cells.

The effects of the protein kinase C activator, phorbol myristate acetate (PMA), on cytosolic calcium levels and adrenocorticotropin (ACTH) release from the mouse anterior pituitary tumor cell line, AtT-20, were compared to those induced by the hormone, corticotropin-releasing factor (CRF), a stimulant of cAMP-dependent protein kinase activity. Cytosolic calcium levels were measured using the fluorescence probe Quin 2. PMA induced a time- and concentration-dependent rise in cytosolic calcium levels and ACTH release from AtT-20 cells that was blocked by verapamil and nifedipine, antagonists of voltage-regulated calcium channels, and tetraethylammonium (TEA), a K+ channel antagonist. The inactive phorbol ester, 4-phorbol 12,13-didecanoate, did not alter cytosolic calcium levels or ACTH release. Several minutes after the initial stimulation of calcium influx by PMA, cytosolic calcium levels returned to basal levels despite the continued presence of the phorbol ester. A short pretreatment (2-4 min) of AtT-20 cells with PMA abolished the ability of K+, CRF, and forskolin to raise intracellular calcium levels. These findings indicate that phorbol esters induce a secondary inhibition of calcium influx after an initial stimulation. In contrast to the effects of PMA, CRF induced a sustained rise in cytosolic calcium levels and did not reduce the subsequent stimulation of calcium influx by K+ or PMA. CRF-stimulated calcium influx was blocked by verapamil but not TEA. The ability of CRF to elevate cytosolic calcium levels was mediated by cAMP-dependent protein kinase because the insertion of a synthetic peptide inhibitor of cAMP-dependent protein kinase activity into AtT-20 cells attenuated the ability of CRF and forskolin but not PMA to raise cytosolic calcium levels. The results suggest that activators of protein kinase C and cAMP-dependent protein kinase regulate intracellular calcium levels in AtT-20 cells through different mechanisms.

Adrenocorticotropic Hormone↗
PubMed1987

Molecular mechanisms of corticotropin-releasing factor stimulation of calcium mobilization and adrenocorticotropin release from anterior pituitary tumor cells.

In a tumor cell line of the mouse anterior pituitary (AtT-20/D16-16) consisting of a homogeneous population of corticotrophs, corticotropin-releasing factor (CRF) activates adenylate cyclase and cAMP-dependent protein kinase. In addition, CRF induces a rise in cytosolic calcium levels in AtT-20/D16-16 cells and stimulates adrenocorticotropin hormone release. To determine whether activation of cAMP-dependent protein kinase is essential for CRF to stimulate calcium mobilization and trigger adrenocorticotropin hormone release, an inhibitor of cAMP-dependent protein kinase was inserted into AtT-20/D16-16 cells using a liposome technique. In control cells, CRF, forskolin (a direct activator of adenylate cyclase) and potassium increased cytosolic calcium levels. Insertion of the protein kinase inhibitor into AtT-20/D16-16 cells greatly attenuated CRF and forskolin-stimulated calcium mobilization although it did not alter the rise in cytosolic calcium induced by potassium. Treatment of the cells with liposomes lacking protein kinase inhibitor (but containing an equivalent amount of bovine serum albumin) had no effect upon the calcium mobilization elicited by any of the agents tested. These results reveal an essential role for cAMP-dependent protein kinase in mediating CRF-stimulated calcium mobilization and suggest that its activation may be an essential molecular event for CRF to evoke adrenocorticotropin hormone secretion.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1986

D-2 dopamine receptor-mediated inhibition of pro-opiomelanocortin synthesis in rat intermediate lobe. Abolition by pertussis toxin or activators of adenylate cyclase.

Stimulation of the D-2 dopamine receptor inhibits pro-opiomelanocortin (POMC) synthesis in isolated rat intermediate lobe tissue. Intermediate lobe tissue was incubated in the absence or presence of various dopaminergic compounds, and then its capacity to incorporate [3H]tyrosine into POMC was tested. D-2 dopaminergic agonists caused a dose-dependent inhibition of POMC synthesis; the maximal inhibitory effect was approximately a 50% reduction in the amount of POMC synthesized. D-2 dopaminergic antagonists blocked the inhibitory effect of each agonist. Pretreatment of the tissue with pertussis toxin abolished the D-2 dopaminergic inhibition of POMC synthesis. The potency of pertussis toxin in abolishing the dopaminergic inhibition of POMC synthesis corresponded to its potency in abolishing the D-2 dopaminergic inhibition of adenylate cyclase activity. Cholera toxin, forskolin, and 8-bromo-cAMP, compounds that activate the cAMP pathway, enhanced the capacity of intermediate lobe tissue to synthesize POMC and counteracted the dopaminergic inhibition of POMC synthesis. Incubation of intermediate lobe tissue for 24 h with bromocriptine, a D-2 dopaminergic agonist, decreased the POMC mRNA content by 46% as determined by hybridization of RNA to a 32P-labeled probe. Incubation of intermediate lobe tissue with forskolin increased the level of POMC mRNA; incubation of the tissue with a combination of bromocriptine and forskolin also resulted in an increase in the level of POMC mRNA. It is proposed that Ni, the inhibitory guanyl nucleotide binding protein, and possibly adenylate cyclase mediate the dopaminergic inhibition of POMC synthesis.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1986

Liposome delivery of cyclic AMP-dependent protein kinase inhibitor into intact cells: specific blockade of cyclic AMP-mediated adrenocorticotropin release from mouse anterior pituitary tumor cells.

Insertion of a crude preparation of cyclic AMP (cAMP)-dependent protein kinase inhibitor (PKI) into a cloned mouse anterior pituitary cell line (AtT-20/D16-16) blocked cAMP-mediated hormone release. This was accomplished by developing a technique to incorporate PKI into multicellular cultures. The technique involved the encapsulation of the PKI into liposomes coupled to Protein A (a bacterial protein that binds to the Fc portion of antibodies). Application of such liposomes to AtT-20 cells targeted by pre-treatment with an antiserum against neural cell adhesion molecule (a cell surface glycoprotein expressed by these cells) resulted in the attachment of the liposomes onto the cell surface followed by the delivery of the liposome content into the cells. The AtT-20 cells respond to cAMP-promoting agents such as forskolin by secreting the hormone adrenocorticotropin (ACTH). Liposomes containing PKI and coupled to protein A specifically blocked cAMP-mediated ACTH release from cells treated with anti-N-CAM antibodies. In contrast, the ACTH release response to K+ or phorbol esters does not appear to involve cAMP and was not reduced by such manipulations. The specificity of PKI to block hormone release initiated by one but not by other secretagogues directly links cAMP-dependent protein kinase with the ACTH release process but suggests that there are other mechanisms also involved in stimulus-secretion coupling in corticotrophs.

Adrenocorticotropic Hormone↗
PubMed1986

Forskolin enhances basal and potassium-evoked hormone release from normal and malignant pituitary tissue: the role of calcium.

The release of immunoreactive ACTH (IR-ACTH) from AtT-20 pituitary tumor cells was transiently increased by exposure to an elevated concentration of potassium ion in an osmotically balanced extracellular medium. With the calcium-sensitive dye Quin 2, the concentration of free cytosolic calcium (Cai) in the AtT-20 tumor was determined to be 115 nM. Challenge of these cells with 60 mM potassium in an osmotically balanced salt solution raised the concentration of Cai to 246 nM. This is in accord with the view that agents promoting calcium entry into pituitary cells trigger hormone secretion. Addition of forskolin to the extracellular medium caused a sustained release of IR-ACTH from AtT-20 tumor cells. Challenge with forskolin (10 microM) increased the concentration of Cai to 149 nM. This observation is also in accord with the view that calcium entry is a necessary and sufficient stimulus to trigger hormone secretion from the anterior pituitary lobe. Exposure of cells to forskolin (10 microM) before a potassium challenge increased the quantity of IR-ACTH released in response to potassium, but did not alter the minute by minute time course of the response to this ion. Forskolin pretreatment did not alter the potassium-evoked rise in Cai concentration. This observation suggests that the magnitude of the secretory response of the pituitary gland can be enhanced by agents other than those promoting an increase in Cai. After exposure of the tumor cells to potassium for a sufficient time to permit the rate of release of hormone to return to the basal value, forskolin could still stimulate the release of hormone from the tumor cells. Under these circumstances, forskolin did not increase the concentration of Cai. This observation suggests that pituitary hormone secretion can be initiated by a factor(s) other than an acute change in the Cai concentration. Both forskolin and 8-bromo-cAMP stimulated hormone secretion from dispersed melanotrophs and potentiated the potassium-evoked secretory response of these cells. Neither compound affected the apparent time course of the response to potassium. These observations suggest that the effects of forskolin and potassium on the AtT-20 tumor cell may use mechanisms occurring in normal pituitary cells.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1986

Cholecystokinin-8 stimulates adrenocorticotropin release from anterior pituitary cells.

Cholecystokinin octapeptide (CCK-8) stimulated adrenocorticotropin hormone (ACTH) release from both rat anterior pituitary cells in culture and a tumor cell line of the mouse anterior pituitary (AtT-20/D16-16). The stimulation of ACTH release was dependent on the time of exposure to CCK-8 and the concentration of this peptide applied to anterior pituitary cells. Cerulein evoked ACTH release whereas human gastrin 1, CCK-4 and desulfated CCK-8 only produced minimal affects on ACTH release at concentrations of 10(-4) M. In contrast, these latter three peptides were as effective as CCK-8 in inducing the secretion of amylase from pancreatic acinar cells. Antagonists of CCK-8 receptors in the pancreas such as proglumide, benzotript and dibutyryl cyclic GMP did not affect the ACTH release response to CCK-8. The CCK-8 stimulation of ACTH release was calcium-dependent and blocked by glucocorticoid pretreatment. The mechanisms by which CCK-8 evoked ACTH release appears distinct from that of other ACTH secretagogues such as corticotropin releasing factor and vasopressin. The data suggest that CCK-8 is a corticotropin releasing factor-like agent acting through a putative novel receptor subtype in the anterior pituitary.

Adrenocorticotropic Hormone↗
PubMed1985

Corticotropin releasing factor increases proopiomelanocortin messenger RNA in mouse anterior pituitary tumor cells.

The ability of corticotropin releasing factor (CRF) to stimulate adrenocorticotropin (ACTH) synthesis in corticotrophs was assessed by measuring total cell content of ACTH and the levels of proopiomelanocortin (POMC) mRNA in a cloned tumor cell line of the mouse anterior pituitary (AtT-20/D16-16). CRF treatment caused a time-dependent increase in POMC mRNA levels (measured using a hybridization technique) as well as elevating total ACTH content in AtT-20 cells. The increase in POMC mRNA levels preceded changes in ACTH content and slowly returned toward control levels after CRF withdrawal. The rise in POMC mRNA levels following CRF stimulation appeared to be specific since beta-actin mRNA levels were not affected by CRF treatment. Both 8-bromo-cAMP and phorbol ester increased POMC mRNA levels in AtT-20 cells, suggesting that CRF may act through different protein kinases to regulate the POMC gene. CRF appears to activate the POMC gene since treatment of the AtT-20 cells with the peptide increased the levels of an RNA species in the nuclei having the expected molecular weight of the transcript of the POMC gene. The results indicate that continued exposure of corticotrophs to CRF induces long term increases in the ACTH synthetic capacity of those cells.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1985

Corticotropin-releasing factor-induced adrenocorticotropin hormone release and synthesis is blocked by incorporation of the inhibitor of cyclic AMP-dependent protein kinase into anterior pituitary tumor cells by liposomes.

Corticotropin-releasing factor (CRF) is the most potent and effective natural stimulant of corticotropin (ACTH) secretion. In a tumor cell line of the mouse anterior pituitary (AtT-20/D16-16) consisting of a homogeneous population of corticotrophs, CRF is known to increase adenylate cyclase and cAMP-dependent protein kinase activities as well as to release ACTH. To determine whether activation of cAMP-dependent protein kinase is essential for CRF to evoke the secretion of ACTH, an inhibitor (PKI) of this kinase was inserted into AtT-20 cells. This was accomplished by first encapsulating PKI into liposomes and then covalently coupling them to protein A for binding to antibodies directed against an AtT-20 cell surface antigen, N-CAM (neural cell adhesion molecule). The binding of the liposomes to the anti-N-CAM antibodies led to the internalization of the PKI into the tumor cells. The PKI treatment greatly attenuated CRF-stimulated ACTH release as well as the secretory response to beta-adrenergic agonists. However, ACTH release in response to caerulein, an agonist of cholecystokinin 8 receptors, was not altered by the PKI treatment. CRF treatment also increased the levels of mRNA for proopiomelanocortin (POMC), the precursor for ACTH in AtT-20 cells. Application of liposomes containing PKI to AtT-20 cells blocked the ability of CRF and 8-bromo-cAMP, but not phorbol ester, to increase POMC mRNA levels. The results revealed an essential role for cAMP in mediating the effect of CRF on ACTH release and POMC gene expression.

Adrenocorticotropic Hormone↗
PubMed1985

Multiple mechanisms of somatostatin inhibition of adrenocorticotropin release from mouse anterior pituitary tumor cells.

The release of ACTH from a clonal cell line of the mouse anterior pituitary (AtT-20/D16-16) can be stimulated by forskolin, 8-bromo-cAMP, and K+. SRIF and its structurally related analogs are very potent inhibitors of the ACTH release response to these secretagogues. The potency of SRIF, its analogs, and somatostatin-28 to inhibit stimulated ACTH release is relatively the same for each of these three secretagogues. The mechanisms by which SRIF regulates the secretion of ACTH can be differentiated by various pharmacological manipulations. Pretreatment of AtT-20 cells with SRIF (10(-7) M) desensitizes SRIF's inhibition of forskolin but not K+ or 8-bromo-cAMP-stimulated ACTH release. Pertussis toxin pretreatment abolishes SRIF's inhibition of forskolin-stimulated ACTH release but not SRIF's inhibition of the ACTH release response to K+ or 8-bromo-cAMP. In contrast, increasing the calcium concentration in the medium reduces SRIF's inhibition of K+ but not forskolin or 8-bromo-cAMP-stimulated ACTH release. These results suggest that SRIF regulates ACTH release from AtT-20 cells through multiple mechanisms. If SRIF acts through a single receptor to produce its effects on ACTH release, then the data imply that the SRIF receptor is coupled to more than one second messenger system.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1985

Pertussis toxin treatment blocks the inhibition of somatostatin and increases the stimulation by forskolin of cyclic AMP accumulation and adrenocorticotropin secretion from mouse anterior pituitary tumor cells.

Somatostatin (SRIF) inhibits stimulated cyclic AMP accumulation and adrenocorticotropin (ACTH) release from mouse anterior pituitary tumor cells (AtT-20/D16-16). In order to determine whether guanine nucleotide inhibitory proteins (Ni) mediate these effects, AtT-20 cells were treated with pertussis toxin, an agent that inactivates Ni. Pertussis toxin catalyses the ADP-ribosylation of a 41,000 MW protein in membranes of AtT-20 cells. Pretreatment with pertussis toxin prevents the subsequent ability of toxin to catalyse the labeling of Ni. This effect is dependent on the time of pretreatment and is not reversible. The inhibition of SRIF of forskolin-stimulated cyclic AMP accumulation and ACTH release is prevented by pertussis toxin treatment. The blockade is dependent on the time and concentration of toxin used and is not reversible. Pertussis toxin treatment prevents SRIF from inhibiting corticotropin releasing factor and cholera toxin-stimulated cyclic AMP synthesis. The inhibition of K+ and 8-bromocyclic AMP-stimulated ACTH release by SRIF is attenuated partially by toxin treatment. The ability of forskolin and cholera toxin to stimulate cyclic AMP formation and ACTH release is enhanced by treatment of AtT-20 cells with pertussis toxin. The increased cyclic AMP response to forskolin is prevented by cycloheximide. The data indicate that Ni mediates the inhibition by SRIF of cyclic AMP formation and the ACTH release that results from adenylate cyclase stimulation.

8-Bromo Cyclic Adenosine Monophosphate↗