Cytosolic calcium

Another mechanism in initiating the contraction is agonist-induced contraction. It results from the hydrolysis of membrane phosphatidylinositol and the formation of inositol triphosphate (IP3)- IP3 in turn triggers the release of intracellular calcium from the sarcoplasmic reticulum and the influx of more extracellular calcium. The third mechanism in triggering the smooth muscle contraction is the increase of calcium influx through the receptor-operated channels. The increased cytosolic calcium enhances the binding to the protein, calmodulin [73298-54-1]. 

Another major second messenger in cells is calcium ion. Virtually any mammalian cell line can be used to measure transient calcium currents in fluorescence assays when cells are preloaded with an indicator dye that allows monitoring of changes in cytosolic calcium concentration. These responses can be observed in real time, but a characteristic of these responses is that they are transient. This may lead to problems with hemi-equilibria in antagonist studies whereby the maximal responses to agonists may be depressed in the presence of antagonists. These effects are discussed more fully in Chapter 6. 

Snowdowne, K. W., Ertel, R. J., and Borle, A. B. (1985). Measurement of cytosolic calcium with aequorin in dispersed rat ventricular cells. J. Mol. Cell. Cardiol. 17 233-241. 

Smooth Muscle Tone Regulation. Figure 2 Membrane mechanisms leading to an increases in cytosolic calcium concentration, depolar, depolarisation of the membrane see text for abbreviations. 

Neutrophils represent an ideal system for studying osmotic effects on exocytosis. Stimulation of cytochalasin-B-treated neutrophils with the chemotactic peptide Jlf-formylmethionyl-leucyl-phenyl-alanine (FMLP) results in a rapid compound exocytosis up to 80% of lysosomal enzymes are released within 30 s (9-14). Secretion appears to be triggered by a rise in the level of cytosolic free calcium (15-18) promoted in part by entry of extracellular calcium through receptor-gated channels and in part by release of calcium that is sequestered or bound at some intracellular site (19-21). In this presentation, we augment our previously published data (22,23), which demonstrates that lysosomal enzyme release from neutrophils is inhibited under hyperosmotic conditions and that the rise in cytosolic calcium preceding secretion is inhibited as well. 

Figure 2. Reporting of cytosolic free calcium levels by indo-1. Increases in cytosolic calcium, due either to entry of extracellular calcium via calcium channels or to release of intracellular calcium sequestered in organelles such as smooth endoplasmic reticulum, results in formation of the indo-l-calcium complex. Fluorescence intensity at 400 nm (excitation at 340 nm) is proportional to the concentration of this complex the dissociation constant for this complex is about 250 nff (24), making this probe useful for detecting calcium activities in the range of 25 to 2500 nJ. ...

Pasti L, Zonta M, Pozzan T, Vicini S, Carmignoto G (2001) Cytosolic calcium oscillations in astrocytes may regulate exocytotic release of glutamate. J Neurosci 21 477 84 Fenner R, Neher E (1988) The role of calcium in stimulus-secretion coupling in excitable and non-excitable cells. J Exp Biol 139 329-345... 

KAiiYA H, OKABE K, OKAMOTO F, TSUZUKi T and SOEDA H (2000) Proteiu tyrosiue kinase inhibitors increase cytosolic calcium and inhibit actin organization as resorbing activity in rat osteoclasts. J Cell Phys 183, 83-90. 

If some of the electrophysiological effects of oxidant stress occur secondary to an elevation in intracellular calcium, it is important to consider the possible factors that may underlie the initial elevation of calcium. In the simplest analysis, elevation of cytosolic calcium may be due to (1) redistribution of intracellular calcium stores (2) increased calcium influx or (3) decreased calcium efflux. 

Recent studies by Crompton et al. have shown that oxidant stress may open a Ca-sensitive, non-selective pore in the inner mitochondrial membrane that is blocked by cyclosporin A (Crompton, 1990 Crompton and Costi, 1990). This pore opening results in massive mitochondrial swelling, dissipation of the transmembrane proton gradient and disruption of mitochondrial energy production (Crompton et al., 1992). Since mitochondria may play a role as a slow, high-capacity cytosolic calcium buffer (Isenberg et al., 1993), disruption of mitochondrial function may also contribute to calcium overload and cell injury. 

Increased levels of cytosolic calcium could potentiate ischaemia-reperfusion injury in several ways. For example, conversion of xanthine dehydrogenase to xanthine oxidase may be catalysed by a calcium-dependent protease (McCord, 1985). However, because it has been so difficult to demonstrate the presence of xanthine... 

Another mechanism to maintain CO when contractility is low is to increase heart rate. This is achieved through sympathetic nervous system (SNS) activation and the agonist effect of norepinephrine on P-adrenergic receptors in the heart. Sympathetic activation also enhances contractility by increasing cytosolic calcium concentrations. SV is relatively fixed in HF, thus HR becomes the major determinant of CO. Although this mechanism increases CO acutely, the chronotropic and inotropic responses to sympathetic activation increase myocardial oxygen demand, worsen underlying ischemia, contribute to proarrhythmia, and further impair both systolic and diastolic function. 

The amount of tension developed by a muscle fiber during tetanic contraction can be as much as three to four times greater than that of a single muscle twitch. The mechanism involved with this increased strength of contraction involves the concentration of cytosolic calcium. Each time muscle fiber is stimulated by an action potential, Ca++ ions are released from the sarcoplasmic reticulum. However, as soon as the these ions are released, a... 

In skeletal muscle, calcium binds to troponin and causes the repositioning of tropomyosin. As a result, the myosin-binding sites on the actin become uncovered and crossbridge cycling takes place. Although an increase in cytosolic calcium is also needed in smooth muscle, its role in the mechanism of contraction is very different. Three major steps are involved in smooth muscle contraction ... 

All of these factors (ANS stimulation, blood-borne and locally produced substances) alter smooth muscle contractile activity by altering the intracellular concentration of calcium. An increase in cytosolic calcium leads to an increase in crossbridge cycling and therefore an increase in tension... 

NO is a gaseous neurotransmitter implicated in signaling in the central and peripheral nervous system as well as in the immune system and the vasculature. NO is formed from L-arginine by nitric oxide synthase (NOS). There are three isoforms of NOS. All isoforms require NADPH as a cofactor, use L-arginine as a substrate, and are inhibited by Nw-nitro-L-arginine methyl ester (L-NAME). The three isoforms are separate gene products. One isoform of NOS is a cytosolic, calcium/calmodulin-independent, inducible enzyme (iNOS). It is found in macrophages, neutrophils, vascular smooth muscle, and endothelia. The iNOS... 

PBAN binding to a receptor results in signal transduction events to stimulate the pheromone biosynthetic pathway (Fig. 5). Receptor activation results in the influx of extracellular calcium and has been demonstrated in a number of moths [163-168]. The increase in cytosolic calcium can directly stimulate pheromone biosynthesis in some moths [165-168] or it will stimulate the production of cAMP [169,170]. So far cAMP has only been implicated in signal... 

Fig. 10.3. Acceptor photobleaching analysis of interaction between barley MLO and calmodulin. Barley MLO is a plant-specific integral membrane protein that associates with the cytosolic calcium sensor protein Calmodulin...

Ionized calcium (Ca2+) is the most common signal transduction element in cells [66], Excitable cells, like neurons, contain voltage-dependent Ca2+ channels, which enable these cells to drastically increase cytosolic calcium levels. Rapid fluctuations in presynaptic... 

Increases in the concentration of calcium in the cytosol provides a signal that can initiate muscle contraction, vision, and other signaling pathways. The response depends on the cell type. In muscle, a transient rise in the cytosolic calcium levels (from opening calcium channels in the sarcoplasmic reticulum) causes contraction. This signaling in contraction is a direct consequence of electrical activation of the voltage-gated channel. 

Calcium signaling is also involved in the response to growth factors. Normally, cells maintain low calcium levels in the cytosol. A low cytosolic calcium level is maintained by pumps that use ATP hydrolysis to move Ca2+ out of the cytosol. Ca2+ concentration in the cytosol increases by activating a calcium channel that lets Ca2+ flow back into the cytosol. 

Calcium signaling can be activated directly by regulating Ca2+ channels. However, there is an indirect way to cause cytosolic calcium to rise. 

In addition to the well-known iron effects on peroxidative processes, there are also other mechanisms of iron-initiated free radical damage, one of them, the effect of iron ions on calcium metabolism. It has been shown that an increase in free cytosolic calcium may affect cellular redox balance. Stoyanovsky and Cederbaum [174] showed that in the presence of NADPH or ascorbic acid iron ions induced calcium release from liver microsomes. Calcium release occurred only under aerobic conditions and was inhibited by antioxidants Trolox C, glutathione, and ascorbate. It was suggested that the activation of calcium releasing channels by the redox cycling of iron ions may be an important factor in the stimulation of various hepatic disorders in humans with iron overload. 

Gelperin, D., Mann, D., Del Valle, J. and Wiley, J. Bradykinin (BK) increases cytosolic calcium in cultured rat myenteric neurons via BK-2 type receptors coupled to mobilization of extracellular and intracellular sources of calcium Evidence that calcium influx is prostaglandin dependent. /. Pharmacol. Exp. Ther. 271 507-514,1994. 

Tyrosine phosphorylation plays an important role in synaptic transmission and plasticity. Evidence for this role is that modulators of PTKs and PTPs have been shown to be intimately involved in these synaptic functions. Among the various modulators of PTKs, neuro-trophins have been extensively studied in this regard and will be our focus in the following discussion (for details of growth factors, see Ch. 27). BDNF and NT-3 have been shown to potentiate both the spontaneous miniature synaptic response and evoked synaptic transmission in Xenopus nerve-muscle cocultures. Neurotrophins have also been reported to augment excitatory synaptic transmission in central synapses. These effects of neurotrophins in the neuromuscular and central synapses are dependent on tyrosine kinase activities since they are inhibited by a tyrosine kinase inhibitor, K-252a. Many effects of neurotrophins on synaptic functions have been attributed to the enhancement of neurotransmitter release BDNF-induced increase in neurotransmitter release is a result of induced elevation in presynaptic cytosolic calcium. Accordingly, a presynaptic calcium-depen-dent phenomenon - paired pulse facilitation - is impaired in mice deficient in BDNF. 

The exact mechanisms by which BDNF enhances the induction of LTP remain obscure. Nevertheless, both pre-and postsynaptic mechanisms appear to be possible. As mentioned above, BDNF elevates presynaptic cytosolic calcium level and thus increases vesicular neurotransmitter release. When a postsynaptic neuron is injected with a Trk tyrosine kinase inhibitor (K252a), BDNF-augmented LTP is curtailed this suggests that a postsynaptic mechanism is adopted by BDNF in the manifestation of LTP. It has been postulated that neurotrophins may act as... 

---