CAMP accumulation

Figure 4 Dose-response curves for the potentiation by brucine of ACh wholecell Mi muscarinic receptor responses. (A) Brucine (10-4 M) enhanced the potency of ACh to increase cAMP accumulation in Mi CHO cells by 2.6-fold. (B) Brucine (100 pM) produced a 3.0-fold increase in ACh potency in Ca2+ response to ACh. (From Ref. 9.)...

Rosenberg, P. A. 8r Dichter, M. A. (1989). Extracellular cAMP accumulation and degradation in rat cerebral cortex in dissociated cell culture. J. Neurosci. 9, 2654-63. 

The multiplicity of G proteins coupled to opiate receptors may explain how different opiates can bind to the same receptor yet induce different cellular responses. For example, morphine binds to the cloned rat fi receptor expressed in HEK 293, CHO and COS-7 cells and inhibits cAMP accumulation [80-82]. Morphine can be continuously applied to the cells for up to 16 h, and the potency and magnitude of morphine inhibition of adenylyl cyclase does not diminish [80, 81]. In contrast, the opiate sufentanil can bind to the same cloned fi receptor in HEK 293 cells to inhibit cAMP accumulation. However, sufentanil s actions rapidly desensitize [83]. Since both compounds bind to the same receptor, and the fi receptor is the only receptor these drugs can interact with in these cells, the ability of these two full agonists to differentially regulate the fi receptor must be due to their abilities to affect separate adaptive processes in these cells. 

Mutagenesis studies have shown that morphine and sufentanil bind differently to the jj, receptor [83, 85]. Mutation of an aspartic acid at residue 114 of the // receptor to an asparagine resulted in a mutant that did not bind morphine and morphine was ineffective in inhibiting adenylyl cyclase via that receptor. In contrast, sufentanil bound to the mutant and wild-type receptors equally well and it effectively inhibited cAMP accumulation via the mutant receptor. These findings demonstrate that morphine and sufentanil have different requirements for binding to the // receptor. By binding differentially, these two agonists may induce the ft receptor to interact with different G proteins to induce distinct cellular effects. 

While chronic morphine treatment uncouples the // receptor from K+ channels, it did not affect the coupling of ft receptors to adenylyl cyclase. Pretreatment of the cloned ft receptor expressed in HEK 293, AtT-20, CHO and COS cells with morphine or DAMGO for up to 16h did not alter the subsequent ability of fi agonists to inhibit cAMP accumulation [25, 65, 80-82]. These findings suggest that morphine treatment induces a selective desensitization of the coupling of the fi receptor to K+ channels. 

Ohno T, Kato N, Ishii C, Shimizu M, Ito Y, Tomono S and Kawazu S. 1993. Genistein augments cyclic adenosine 3 5 -monophosphate(cAMP) accumulation and insulin release in MIN6 cells. Endocr Res 19(4) 273-285. 

The effects of Li+ upon this system have been reviewed in depth by Mork [131]. Animal studies originally demonstrated that Li+ inhibits cAMP formation catalyzed by adenylate cyclase in a dose-dependent manner [132]. The level of cAMP in the urine of manic-depressive patients changes with mental state, being abnormally elevated during the switch period between depression and mania it is proposed that Li+ s inhibitory effect upon adenylate cyclase activity may correct this abnormality. Subsequent research, in accord with the initial experiments, have shown that Li+ s interference with this second messenger system involves more than one inhibitory action. At therapeutic levels, Li+ inhibits cAMP accumulation induced by many neurotransmitters and hormones, both in... 

Li+, at therapeutically relevant concentrations, is a potent inhibitor of norepinephrine-stimulated adenylate cyclase activity ex vivo in both rat [133] and human brain [134], and it inhibits norepinephrine-stimulated cAMP accumulation in Li+-treated patients. Li+ also inhibits dopamine-stimulated cAMP accumulation in rat brain [135]. These inhibitory effects of Li+ have been shown to be region specific within rat brain, a fact that has obvious significance for a therapeutic mechanism of action. It is interesting that other antimanic drugs may also have dampening effects on dopaminergic neurotransmission. 

Fig. 6.4 In vitro effects of mutation on desensitization and internalization of the dopamine receptor. Shown here are effects of mutation on dose-dependent intracellular cyclic adenosine monophosphate (cAMP) accumulation (A and B) and binding curves (C and D) for artificial ligand (SCH 23390) using three constructs controls (wild type, A and C) and the Thr360Ala mutant (360, B and D). In the desensitization experiments, cells were preincubated with 10 oA/ dopamine (o) or vehicle ( ) for 20min, and increasing concentrations of dopamine (10 to 10 (iM) were added to assess cAMP accumulation. Note that loss of efficacy and potency seen in wild-type cells (A) disappeared with the Thr360Ala mutation (B). Conversely, internalization, assessed by decrease in SCH23390 binding (C) after pretreatment with lOpM dopamine (o, compared to vehicle ), was essentially unchanged by the Thr360Ala mutation (D)...

Molecular studies explained this apparent paradox when the temperature-sensitive G a Ala366Ser mutation of the G a protein was identified. At 32°C, the G a 366Ser mutation results in the constitutive cAMP accumulation that causes the testosterone secretion that is the hallmark of the testotoxicosis phenotype. At 37°C, however, the G a 366Ser mutation results in loss of adenylyl cyclase signaling, causing PHP-Ia. As a result, a single mutation that performs differently in different tissues causes precocious puberty and abnormalities of PTH and TSH (91). 

Bioavailability of starch. Cooked rice was administered to colectomized rats by gastric intubation and the recovery of starch in the ileal digesta measured after 10 hours of ingestion. Significant starch (11-15%) was recovered from animals fed peas, lima beans, or kidney beans 0.2-0.4% of starch from rice. Oligosaccharide extraction, the size of the test meal, and the amount of starch did not affect starch biovailability . cAMP accumulation. Methanol extract of the grain, in cell culture at a concentration of 1 mg/mL, was active on mast cells " . 

The other major receptor-coupled second messenger system on which lithium exerts significant effects is the cAMP generating system. Forn and Valdecasas (1971] were the first to report that lithium in vitro attenuated NE-stimulated cAMP accumulation since then, numerous studies have... 

The pupils become dilated and there are associated signs of hyperactivity of the sympathetic nervous system, such as hypertension and pilomotor stimulation. The mechanism(s) underlying tolerance and dependence are poorly understood. While acute activation of Gi/o-coupled receptors leads to inhibition of adenylyl cyclase, chronic activation of such receptors produces an increase in cAMP accumulation, particularly evident upon withdrawal of the inhibitory agonist. This phenomenon, referred to as adenylyl cyclase superactivation, is believed to play an important role in opioid addiction. 

G protein-mediated responses to drugs and hormonal agonists often attenuate with time (Figure 2-12, top). After reaching an initial high level, the response (eg, cellular cAMP accumulation, Na+ influx, contractility, etc) diminishes over seconds or minutes, even in the continued presence of the agonist. This "desensitization" is often rapidly reversible a second exposure to agonist, if provided a few minutes after termination of the first exposure, results in a response similar to the initial response. 

The Di receptor is typically associated with the stimulation of adenylyl cyclase (Table 9-1) for example, Di-receptor-induced smooth muscle relaxation is presumably due to cAMP accumulation in the smooth muscle of those vascular beds in which dopamine is a vasodilator. D2 receptors have been found to inhibit adenylyl cyclase activity, open potassium channels, and decrease calcium influx. 

Receptor desensitization may also be mediated by second-messenger feedback. For example, adrenoceptors stimulate cAMP accumulation, which leads to activation of protein kinase A protein kinase A can phosphorylate residues on receptors, resulting in inhibition of receptor function. For the B2 receptor, phosphorylation occurs on serine residues both in the third cytoplasmic loop and in the carboxyl terminal tail of the receptor. Similarly, activation of protein kinase C by Gq-coupled receptors may lead to phosphorylation of this class of G protein-coupled receptors. This second-messenger feedback mechanism has been termed heterologous desensitization because activated protein kinase A or protein kinase C may phosphorylate any structurally similar receptor with the appropriate consensus sites for phosphorylation by these enzymes. 

ERK1/2 phosphorylation by involving G /o proteins, PKC and tyrosine kinase-dependent and -independent pathways. It has been found that Cl-IB-MECA produced a biphasic effect on cAMP accumulation with a stimulatory action starting at a concentration of 3 nM. This activity was triggered through PLC/PKC and not via direct Gs coupling (Germack and Dickenson 2004, 2005). 

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