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The results of our study show that whole-cell and nuclear levels of NADPH oxidase-1 (NOX1) are similar in human vascular endothelial cells (hVECs) and smooth muscle cells (hVSMCs), but lower in human endocardial endothelial cells (hEECs). NOX2 levels were higher in hVECs and lower in hVSMCs. NOX3 levels were the same in hVECs and hVSMCs, but lower in hEECs. NOX4 levels were similar in all of the cell types. NOX4 levels were higher in hVECs than in hVSMCs. NOX5 was also present throughout the 3 cell types, including their nuclei, in the following order: hEECs > hVSMCs > hVECs. The level of basal reactive oxygen species (ROS) was highest in hVECs and lowest in hVSMCs. However, the Ca2+ level was highest in hVSMCs and lowest in hVECs. These findings suggest that all types of NOXs exist in hEECs, hVECs, and hVSMCs, although their density and distribution are cell-type dependent. The density of the different NOXs correlated with the ROS level, but not with the Ca2+ level. In conclusion, NOXs, including NOX3, exist in cardiovascular cells and their nuclei. The nucleus is a major source of ROS generation. The nuclear NOXs may contribute to ROS and Ca2+ homeostasis, which may affect cell remodeling, including the formation of nuclear T-tubules in vascular diseases and aging. [ABSTRACT FROM AUTHOR]

It has been well-established that polarized migrating cells exhibit a stable and transient gradient of intracellular calcium concentration ([Ca2+]i), increasing from front-to-rear, that is thought to be responsible for rear retraction. The paradox that arises is how calcium at the front of a cell catalyzes critical high-threshold calcium-dependent processes during cell migration and particularly in decision-making for a cell to turn. In this brief review, we discuss the recent discovery of flickering high-[Ca2+]i microdomains (“calcium flickers”) at the front of migrating fibroblasts and their common role in transducing local membrane mechanical stress (via TRPM7, a stretch-activated calcium-permeating transient receptor potential channel) and chemoattractant-elicited signals (via type 2 inositol 1,4,5-trisphosphate receptor in the endoplasmic reticulum). Furthermore, we present a new model for patterned calcium flicker activity as the mechanism for steering the turning of a migrating cell. [ABSTRACT FROM AUTHOR]

Paeonia emodi (peony) is a well known plant used medicinally to treat hypertension, palpitations, and asthma. Despite its popularity, there are few reports in the scientific literature examining its use in such conditions. We prepared a 70% ethanolic extract of peony root (Pe.Cr) and applied it to segments of guinea pig atria and trachea and rat aorta suspended separately in tissue baths. Activity against arachidonic acid (AA)-induced platelet aggregation was measured in human platelet-rich plasma. Airway relaxant effect was evaluated against acetylcholine (ACh)-induced airway contraction in mouse lung slices loaded with fluo-4. Pe.Cr (0.3-10 mg/mL) showed an atropine-resistant negative inotropic effect in atria. In rat aorta, an endothelium-independent vasodilatory effect (0.3-10 mg/mL) was seen in phenylephrine- and high-K+-induced contractions. Pe.Cr (0.01-1 mg/mL) also inhibited AA-induced platelet aggregation. In isolated trachea, Pe.Cr (0.3-10 mg/mL) relaxed carbachol- and histamine-induced contractions independently of β-adrenergic receptors. In mouse lung slices, Pe.Cr (0.3-1 mg/mL) inhibited ACh-induced airway narrowing and oscillations of intracellular Ca2+ in airway smooth muscle cells. The results showed cardiosuppressant, vasodilatory, antiplatelet, and tracheal and airway relaxant activities of peony, providing potential justification for its medicinal use in different hyperactive cardiovascular and respiratory disorders. [ABSTRACT FROM AUTHOR]

Asthma is a chronic disease characterized by inflammation and hypersensitivity of airway smooth muscle cells (ASMCs) to different spasmogens. The past decade has seen increased use of herbal treatments for many chronic illnesses. Ginger (Zingiber officinale) is a common food plant that has been used for centuries in treating respiratory illnesses. In this study, we report the effect of its 70% aqueous methanolic crude extract (Zo·Cr) on acetylcholine (ACh)-induced airway contraction and Ca2+ signalling in ASMCs using mouse lung slices. Airway contraction and Ca2+ signalling, recorded via confocal microscopy, were induced with ACh, either alone or after pretreatment of slices with Zo·Cr and (or) verapamil, a standard Ca2+ channel blocker. ACh (10 μmol/L) stimulated airway contraction, seen as decreased airway diameter, and also stimulated Ca2+ transients (sharp rise in [Ca2+]i) and oscillations in ASMCs, seen as increased fluo-4-induced fluorescence intensity. When Zo·Cr (0.3-1.0 mg/mL) was given 30 min before ACh administration, the ACh-induced airway contraction and Ca2+ signalling were significantly reduced. Similarly, verapamil (1 μmol/L) also inhibited agonist-induced airway contraction and Ca2+ signalling, indicating a similarity in the modes of action. When Zo·Cr (0.3 mg/mL) and verapamil (1 μmol/L) were given together before ACh, the degree of inhibition was the same as that observed when each of these blockers was given alone, indicating absence of any additional inhibitory mechanism in the extract. In Ca2+-free solution, both Zo·Cr and verapamil, when given separately, inhibited Ca2+ (10 mmol/L)-induced increase in fluorescence and airway contraction. This shows that ginger inhibits airway contraction and associated Ca2+ signalling, possibly via blockade of plasma membrane Ca2+ channels, thus reiterating the effectiveness of this age-old herb in treating respiratory illnesses. [ABSTRACT FROM AUTHOR]

In this study, we report for the first time concurrent measurements of membrane potential and dynamics and respiratory chain activities in rat heart mitochondria, as well as calcium transients in the hearts of rats in an early phase of streptozotocin diabetes, not yet accompanied with diabetes-induced complications. Quantitative relationships among these variables were assessed. The mitochondria from diabetic rats exhibited decreased fluorescence anisotropy values of diphenylhexatriene. This indicates that hydrophobic core of the membranes was more fluid compared with controls (p < 0.05). We discuss the changes in fluidity as having been associated with augmented energy transduction through the diabetic membranes. Reduced ratio of JC-1 fluorescence (aggregates to monomers) in the mitochondria from diabetic hearts reflected descendent transmembrane potential. A significant negative association between membrane fluidity and potential in the diabetic group was found (p < 0.05; r = 0.67). Further, we observed an increase in calcium transient amplitude (CTA) in the diabetic cardiomyocytes (p = 0.048). We conclude that some of the calcium-induced regulatory events that dictate fuel selection and capacity for ATP production in diabetic heart occur at the membrane level. Our findings offer new insight into acute diabetes-induced changes in cardiac mitochondria. [ABSTRACT FROM AUTHOR]

The 3-dimensional confocal microscopy technique has allowed us to identify the presence of yet another cardioactive factor and its receptor, namely neuropeptide Y (NPY) and its Y1 receptor, at the level of vascular smooth muscle cells and heart cells including endocardial endothelial cells (EECs). Using this technique, we also demonstrated that NPY is able to induce an increase in both cytosolic and nuclear calcium in all these cell types. Furthermore, besides being expressed at the level of EECs, NPY is also released from these cells following a sustained increase of intracellular Ca2+. This suggests the ability of NPY to contribute to the regulation of the excitation–secretion coupling of EECs and the excitation–contraction coupling of cardiomyocytes and vascular smooth muscle cells. [ABSTRACT FROM AUTHOR]

It is now well accepted that the heart is a multifunctional organ in which endothelial cells, and more particularly endocardial endothelial cells (EECs), seem to play an important role in regulating and maintaining cardiac excitation–contraction coupling. Even if major differences exist between vascular endothelial cells (VECs) and EECs, all endothelial cells including EECs release a variety of auto- and paracrine factors such as nitric oxide, endothelin-1, angiotensin II, and neuropeptide Y. All these factors were reported to affect cardiomyocyte contractile performance and rhythmicity. In this review, findings on the morphology of EECs, differences between EECs and other types of endothelial cells, interactions between EECs and the adjacent cardiomyocytes, and effects of NPY on the heart will be presented. We will also show evidence on the presence and localization of NPY and the Y1 receptor in the endocardial endothelium and discuss their role in the regulation of cytosolic and nuclear free calcium. [ABSTRACT FROM AUTHOR]

Nuclear tubules (NTs) were found in the nucleus of HeLa cells. Although no function has been ascribed to these structures, our previous data has shown that they are the sites of Ca2+ release with mitochondria shuttled around. In the present study, we further characterized these NTs through different fluorescent dye-labeling and red fluorescent protein transfection experiments. We found that doxorubicin (Dox) is a good indicator to demonstrate the NTs since Dox is fluorescent and DNA is able to quench its fluorescence. By using confocal and electron microscopy, we show that the number and nature of the NTs in HeLa vary from cell to cell, ranging from tubular to intricately branched structures. Additionally, these NTs are double-membrane invaginations of the nuclear envelope and usually lie close to nucleolus. At rest, NTs appeared to be stable and their mouths are always closed. Upon Ca2+ ionomycin stimulation, various forms of dynamism, including membrane protrusion to the nucleus, enlargement and shrinkage of the NTs, and distortion of the nuclear envelope and NTs were observed over a time scale of minutes. These observations suggest that the NT represents a specialized and dynamic compartment inside the nucleus under the control of Ca2+. [ABSTRACT FROM AUTHOR]

The action of several peptides and drugs is thought to be primarily dependent on their interactions with specific cell surface G-protein-coupled receptors and ionic transporters such as channels and exchangers. Recent development of 3-D confocal microscopy allowed several laboratories, including ours, to identify and study the localization of receptors, channels, and exchangers at the transcellular level of several cell types. Using this technique, we demonstrated in the nuclei of several types of cells the presence of Ca2+ channels as well as Na+–H+ exchanger and receptors such as endothelin-1 and angiotensin II receptors. Stimulation of these nuclear membrane G-protein-coupled receptors induced an increase of nuclear Ca2+. Our results suggest that, similar to the plasma membrane, nuclear membranes possess channels, exchangers and receptors such as those for endothelin-1 and angiotensin II, and that the nucleus seems to be a cell within a cell. This article will emphasize these findings. [ABSTRACT FROM AUTHOR]

Asynchronous Ca2+ waves or wave-like [Ca2+]i oscillations constitute a specialized form of agonist-induced Ca2+ signaling that is observed in a variety of smooth muscle cell types. Functionally, it is involved in the contractile regulation of the smooth muscle cells as it signals for tonic contraction in certain smooth muscle cells while causing relaxation in others. Mechanistically, repetitive Ca2+ waves are produced by repetitive cycles of sarcoplasmic reticulum Ca2+ release followed by Ca2+ uptake. Plasmalemmal Ca2+ entry mechanisms are important for providing the additional Ca2+ necessary to maintain proper refilling of the sarcoplasmic reticulum Ca2+ store and support ongoing Ca2+ waves. In this paper, we will review the phenomenon of asynchronous Ca2+ waves in smooth muscle and discuss the scientific and clinical significance of this new understanding. [ABSTRACT FROM AUTHOR]