Calcium is a highly positively charged ionic species. It regulates all cell types’ functions and is an important second messenger that controls and triggers several mechanisms, including membrane stabilization, permeability, contraction, secretion, mitosis, intercellular communications, and in the activation of kinases and gene expression. Therefore, controlling calcium transport and its intracellular homeostasis in physiology leads to the healthy functioning of the biological system. However, abnormal extracellular and intracellular calcium homeostasis leads to cardiovascular, skeletal, immune, secretory diseases, and cancer. Therefore, the pharmacological control of calcium influx directly via calcium channels and exchangers and its outflow via calcium pumps and uptake by the ER/SR are crucial in treating calcium transport remodeling in pathology. Here, we mainly focused on selective calcium transporters and blockers in the cardiovascular system.

The present study was designed to verify if human (h) Angiotensin II (Ang II) type-1 receptor (hAT₁R) undergoes internalization, nuclear translocation, and de novo synthesis in primary culture of human aortic vascular smooth muscle cells (hVSMCs) and if overexpression of this receptor modulates sustained free cytosolic ([Ca]_c) and nuclear ([Ca]_n) calcium. 3-dimensional (3-D) confocal microscopy was used to monitor free intracellular Ca²⁺ and hAT₁R-green fluorescence protein (GFP) fusion protein in cultured hVSMCs. Immunofluorescence studies showed the presence of hAT₁R and the absence of hAT₂R in normal hVSMCs. Using 3-D imaging technique, hAT₁ receptors were localized at the sarcolemma and in the cytosolic and nuclear compartments. In native as well as in normal hAT₁R or hAT₁R –GFP overexpressing hVSMCs, Ang II (10^–9 and 10^–4 M) induced internalization and nuclear translocation of this type of receptor. The internalization of hAT₁Rs is mediated via clathrin-coated pits and vesicles pathway. This phenomenon of trancellular trafficking of receptors was associated with an increase of hAT₁R. The Ang II induced increase of hAT₁R density was prevented by the protein synthesis inhibitor cycloheximide. Overexpression of hAT₁R and hAT₁R–GFP decreased both basal cytosolic and nuclear Ca²⁺. In normal hVSMCs and low hAT₁R–GFP overexpressing hVSMCs, Ang II (10^–15 to 10^–4 M) induced a dose-dependent sustained increase of [Ca]_c and [Ca]_n with an EC₅₀ near 5 × 10^–11 and 5 × 10^–9 M, respectively. Our results suggest that hAT₁Rs are the predominant type of Ang II receptors in aortic hVSMCs and are present in the sarcolemma, the cytosolic and the nuclear compartments. Ang II rapidly induces hAT₁R internalization, nuclear translocation, as well as nuclear de novo synthesis of this receptor. The hAT₁R overexpression in hVSMCs modulates sustained [Ca]_c and [Ca]_n.Key words: angiotensin, calcium, protein synthesis, nucleus, AT₁ receptor, nuclear de novo synthesis.

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.

The vascular endothelium plays a vital role during embryogenesis and aging and is a cell monolayer that lines the blood vessels. The immune system recognizes the endothelium as its own. Therefore, an abnormality of the endothelium exposes the tissues to the immune system and provokes inflammation and vascular diseases such as atherosclerosis. Its secretory role allows it to release vasoconstrictors and vasorelaxants as well as cardio-modulatory factors that maintain the proper functioning of the circulatory system. The sealing of the monolayer provided by adhesion molecules plays an important role in cardiovascular physiology and pathology.

The use of confocal microscopy has shown that the nucleus plays an important role in excitation–contraction and excitation–secretion coupling of several excitable and nonexcitable cardiovascular cells. It has shown that the nuclear membranes, like the sarcolemmal membrane, possess ionic transporters as well as G protein-coupled receptors (GPCRs), which play a major role in modulating both cytosolic and nuclear ionic homeostasis and nuclear signalling. During spontaneous contraction of heart cells, the increase in cytosolic Ca²⁺was immediately followed by a transient increase in nuclear Ca²⁺. The nuclear Ca²⁺rise during excitation–contraction and excitation–secretion coupling was both dependent and independent of changes in cytosolic Ca²⁺. Nuclear membrane GPCRs, such as those of angiotensin II, neuropeptide Y, and ET-1, were functional and contributed to modulation of nuclear ionic homeostasis via direct and (or) indirect modulation of nuclear membrane ionic transporters such as channels, pumps, and exchangers. The signalling of nuclear membrane GPCRs may also contribute to modulation of gene expression, which may regulate proliferation and remodelling of cells and, indeed, life and death. Direct or indirect targeting of nuclear membrane ionic transporters and GPCRs may constitute a new target for drug action.

Plasma membrane endothelin type A (ET_A) receptors are internalized and recycled to the plasma membrane, whereas endothelin type B (ET_B) receptors undergo degradation and subsequent nuclear translocation. Recent studies show that G protein-coupled receptors (GPCRs) and ion transporters are also present and functional at the nuclear membranes of many cell types. Similarly to other GPCRs, ET_Aand ET_Bare present at both the plasma and nuclear membranes of several cardiovascular cell types, including human cardiac, vascular smooth muscle, endocardial endothelial, and vascular endothelial cells. The distribution and density of ET_ARs in the cytosol (including the cell membrane) and the nucleus (including the nuclear membranes) differ between these cell types. However, the localization and density of ET-1 and ET_Breceptors are similar in these cell types. The extracellular ET-1-induced increase in cytosolic ([Ca]_c) and nuclear ([Ca]_n) free Ca²⁺is associated with an increase of cytosolic and nuclear reactive oxygen species. The extracellular ET-1-induced increase of [Ca]_cand [Ca]_nas well as intracellular ET-1-induced increase of [Ca]_nare cell-type dependent. The type of ET-1 receptor mediating the extracellular ET-1-induced increase of [Ca]_cand [Ca]_ndepends on the cell type. However, the cytosolic ET-1-induced increase of [Ca]_ndoes not depend on cell type. In conclusion, nuclear membranes' ET-1 receptors may play an important role in overall ET-1 action. These nuclear membrane ET-1 receptors could be targets for a new generation of antagonists.