Products second messengers

Recently, an additional IP3 isomer, inositol 1,3,4-trisphosphate (1,3,4-IP3), which is ineffective in releasing calcium from the ER, has been identified. Unlike 1,4,5-IP3, 1,3,4-IP3 is thought not to be a product of the direct hydrolysis of an isomer of PIP2, but rather a result of the action of a 5-phosphatase on a more polar inositol phosphate, inositol 1,3,4,5-tetrakisphosphate (IP4) [34], At this time neither 1,3,4-IP3 nor its precursor IP4, which is formed as a result of a 3-kinase-catalysed phosphorylation of 1,4,5-IP3, has a clear physiological role, although IP4 has been implicated in the regulation of plasma membrane calcium influx (see Rasmussen and Barrett, Chapter 4). 

Although in the hepatocyte IP4 and the two IP3 isomers are known to be formed in response to certain hormones, the effect of All on these isomers has yet to be investigated. There is also little information available to date concerning changes in the levels of these inositol phosphates following All stimulation of vascular smooth muscle cells. 

The addition of All to the target tissues under discussion leads to three changes in cellular calcium metabolism (1) a transient increase in cytosolic calcium (2) a reduction in total cell calcium and (3) a sustained increase in the rate of calcium influx across the plasma membrane. Each of these changes in cellular calcium metabolism is discussed below. 

Although slightly attenuated, the rise in cytosolic calcium proceeds in the absence of extracellular calcium (as mentioned above), indicating that upon All stimulation calcium is released into the cytosol from an internal pool. The identity of this mobilized internal pool was initially inferred from cellular studies in which treatment with dantrolene inhibited the redistribution of intracellular calcium [39,40], Be- 

Once mobilized, a large proportion of the cytosolic calcium load is extruded from the cell across the plasma membrane. This extrusion can be demonstrated by measuring the efflux of radiocalcium from previously labeled cells. All stimulation of adrenal, hepatic and vascular smooth muscle cells preloaded with radioactive calcium induces a marked increase in efflux of the radiolabel. This increased efflux is transient, peaking between 4 and 5 minutes after All addition and is observed in the absence of extracellular calcium [39]. Furthermore, under conditions of zero calcium, treatment of cells with dantrolene prior to hormonal stimulation abolishes the All-induced calcium efflux [40], confirming that the radiocalcium lost from the cell is mobilized from a component of the ER. 

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In addition to the mechanism involving cycHc AMP, nonsugar sweeteners, eg, saccharin and a guanidine-type sweetener, have been found to enhance the production of another second messenger, inositol 1,4,5-trisphosphate (IP3), causing the closure of potassium channels and the release of... 

FIGURE 2.6 Production of cyclic AMP from ATP by the enzyme adenylate cyclase. Cyclic AMP is a ubiquitous second messenger in cells activating numerous cellular pathways. The adenylate cyclase is activated by the a subunit of Gs-protein and inhibited by the a-subunit of Gj-protein. Cyclic AMP is degraded by phosphodiesterases in the cell. 

FIGURE 2.7 Production of second messengers inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG) through activation of the enzyme phospholipase C. This enzyme is activated by the a- subunit of Gq-protein and also by Py subunits of Gj-protein. IP3 stimulates the release of Ca2+ from intracellular stores while DAG is a potent activator of protein kinase C. 

FIGURE 5.15 Different modes of response measurement, (a) Real time shows the time course of the production of response such as the agonist-stimulated formation of a second messenger in the cytosol, (b) The stop-time mode measures the area under the curve shown in panel A. The reaction is stopped at a designated time (indicated by the dotted lines joining the panels) and the amount of reaction product is measured. It can be seen that in the early stages of the reaction, before a steady state has been attained (i.e., a plateau has not yet been reached in panel A), the area under the curve is curvilinear. Once the rate of product formation has attained a steady state, the stop-time mode takes on a linear character. 

Screening. See High-throughput screening Second messenger systems calcium ion, 83 description of, 24 production of, 25f Series hyperbolae, 38 Serotonin, 150, 151 f Seven transmembrane receptors, 3-4 Shennong Herbal, 147 Short interfering RNA duplex molecules, 184... 

Second Messengers Their Production and Their Action... 

Inositol trisphosphate Receptor/G-protein cascades. As discussed above, IP3 is one of the products of the hydrolysis of PIP2. To say that it acts as a second messenger means that a rise in its concentration occurs as a result of some meaningful event and that the rise causes some other significant event. In terms of information flow, the signal immediately preceding the rise in IP3 is a rise in the concentration of active PLC. This rise is due to the binding of a subset of G-proteins... 

Figure 10. The G-protein cascades in smooth muscle catalyze the exchange GDP for GTP on G-protein. Following the binding of GTP, the trimeric G-protein splits into an a-GTP part and a P-y part. The a-GTP part ordinarily then combines with its specific apoenzyme to constitute the active enzyme. For the activation of the contractile activation path, the enzyme is phospholipase C and the second messenger products are IP3 and DAG. The IP3 in the myoplasm binds to Ca channels in the SR membrane, opening them. Other second messengers include the inhibitors of contractile activity, cGMP and cAMP.

The exact process(es) by which a2-adrenoceptors blunt release of transmitter from the terminals is still controversial but a reduction in the synthesis of the second messenger, cAMP, contributes to this process. a2-Adrenoceptors are negatively coupled to adenylyl cyclase, through a Pertussis toxin-sensitive Gi-like protein, and so their activation will reduce the cAMP production which is vital for several stages of the chemical cascade that culminates in vesicular exocytosis (see Chapter 4). The reduction in cAMP also indirectly reduces Ca + influx into the terminal and increases K+ conductance, thereby reducing neuronal excitability (reviewed by Starke 1987). Whichever of these releasecontrolling processes predominates is uncertain but it is likely that their relative importance depends on the type (or location) of the neuron. 

Several groups have shown that hormonal stimuli which lead to NaCl secretion in normal tissues fail to do so in CF tissues (reviewed in [17,18,60]). The production of the second messenger cAMP was unimpeded in these CF tissues. Hence it was... 

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