Emerging roles of inositol 1,4,5-trisphosphate signaling in cardiac myocytes

Abstract

Inositol 1,4,5-trisphosphate (IP(3)) is a ubiquitous intracellular messenger regulating diverse functions in almost all mammalian cell types. It is generated by membrane receptors that couple to phospholipase C (PLC), an enzyme which liberates IP(3) from phosphatidylinositol 4,5-bisphosphate (PIP(2)). The major action of IP(3), which is hydrophilic and thus translocates from the membrane into the cytoplasm, is to induce Ca(2+) release from endogenous stores through IP(3) receptors (IP(3)Rs). Cardiac excitation-contraction coupling relies largely on ryanodine receptor (RyR)-induced Ca(2+) release from the sarcoplasmic reticulum. Myocytes express a significantly larger number of RyRs compared to IP(3)Rs (~100:1), and furthermore they experience substantial fluxes of Ca(2+) with each heartbeat. Therefore, the role of IP(3) and IP(3)-mediated Ca(2+) signaling in cardiac myocytes has long been enigmatic. Recent evidence, however, indicates that despite their paucity cardiac IP(3)Rs may play crucial roles in regulating diverse cardiac functions. Strategic localization of IP(3)Rs in cytoplasmic compartments and the nucleus enables them to participate in subsarcolemmal, bulk cytoplasmic and nuclear Ca(2+) signaling in embryonic stem cell-derived and neonatal cardiomyocytes, and in adult cardiac myocytes from the atria and ventricles. Intriguingly, expression of both IP(3)Rs and membrane receptors that couple to PLC/IP(3) signaling is altered in cardiac disease such as atrial fibrillation or heart failure, suggesting the involvement of IP(3) signaling in the pathology of these diseases. Thus, IP(3) exerts important physiological and pathological functions in the heart, ranging from the regulation of pacemaking, excitation-contraction and excitation-transcription coupling to the initiation and/or progression of arrhythmias, hypertrophy and heart failure.

Fig. 1

Elementary IP₃-induced Ca²⁺ release events in cardiac myocytes. (A) Confocal linescan image of Ca²⁺ sparks in a permeabilized cat atrial myocyte (control). Tetracaine (Tetrac) abolishes Ca²⁺ sparks. Addition of IP₃ elevates baseline Ca²⁺ concentration and causes the appearance of smaller Ca²⁺ release events, i.e. Ca²⁺ puffs, which are suppressed by heparin. (B) Averaged Ca²⁺ spark and Ca²⁺ puff. (C) Normalized Ca²⁺ spark (black) and Ca²⁺ puff (red) (top) and first derivative thereof (bottom), a measure for the underlying Ca²⁺ release flux. The Ca²⁺ puff exhibits slower kinetics and lower Ca²⁺ release flux than the Ca²⁺ spark. (D) Surface plots of averaged linescan images of Ca²⁺ sparks and Ca²⁺ puffs. The Ca²⁺ puff displays lower amplitude, longer rise time and duration, and similar width as compared to the Ca²⁺ spark. Modified from [70]. Copyright of the Journal of Physiology; used by kind permission.

Fig. 2

Colocalization of subsarcolemmal IP₃Rs and RyRs in atrial myocytes. (A) Confocal image of a rat atrial myocyte immunostained for the expression of IP₃Rs (red) and RyRs (green). N denotes the nucleus. The area marked by the dashed white rectangular is shown enlarged in (B). Colocalization of IP₃Rs and RyRs (yellow) occurs in some areas underneath the sarcolemma. (C) Schematic illustration of colocalization of subsarcolemmal IP₃Rs and RyRs. Modified from [63]. Copyright of the Journal of Physiology; used by kind permission.

Fig. 3

IP₃-dependent nuclear Ca²⁺ signaling in cardiac myocytes. (A) Confocal images of a permeabilized cat atrial myocyte. In the presence of 0.7 mM tetracaine, IP₃ increases nuclear Ca²⁺ concentration. The IP₃-induced nuclear Ca²⁺ increase is partly reversed by the addition of heparin (Hep). (B) Confocal images of an isolated rat cardiac nucleus. The nucleus and the solution surrounding the nucleus contained fluo-4 dextran. Fluorescence changes from the regions marked by the dashed black line (nucleus) and the area delimited by the nuclear border and the dashed white line (‘cytoplasm’) are shown below. The IP₃-induced Ca²⁺ increase is larger in the nucleus (black) than in the adjacent cytoplasm (red). (C) Confocal images of an isolated rat cardiac nucleus. The nuclear envelope (NE) has been loaded with the low affinity Ca²⁺ dye fluo-5N. Addition of IP₃ causes a reduction of Ca²⁺ stored in the NE. The Ca²⁺ ionophore A23187 further depletes NE Ca²⁺. Modified from [137]. Copyright of the Journal of Physiology; used by kind permission.

Fig. 4

Overview on IP₃ signaling and its involvement in excitation–contraction coupling, excitation–transcription coupling and arrhythmias in adult cardiac myocytes. Schematic drawing of an adult cardiac myocyte showing parts of the sarcolemma, the nucleus, the SR and the myofilaments. G protein-coupled receptors (GPCR), e.g. for angiotensin II, endothelin-1 or phenylephrine, as well as growth factor receptors (GFR) increase cytoplasmic IP₃ concentration via activation of phospholipase C (PLC). IP₃Rs are found in the SR (in atrial myocytes both in the subsarcolemmal and central SR) and in the nuclear envelope, where they face both the cytoplasm and the nucleoplasm. By contrast, RyR2s are found predominantly in the SR. IP₃ signaling is involved in excitation–contraction coupling, in excitation–transcription coupling and in the generation of arrhythmias. Excitation–contraction coupling is mediated by Ca²⁺-induced Ca²⁺ release (CICR) from the SR through RyR2. Cytoplasmic Ca²⁺ binds to the myofilaments and initiates contraction. IP₃-induced Ca²⁺ release from the SR through IP₃Rs may contribute to excitation–contraction coupling by increasing the Ca²⁺ transient and thus contraction. On the other hand, IP₃-induced Ca²⁺ release from (subsarcolemmal) SR may trigger further Ca²⁺ release through neighboring RyR2 with subsequent activation of sarcolemmal NCX. This induces arrhythmias via generation of delayed afterdepolarizations and triggered activity. IP₃-induced SR Ca²⁺ release also generates pro-arrhythmic Ca²⁺ alternans. Finally, IP₃ is involved in excitation–transcription coupling via modulation of nuclear Ca²⁺ concentration. Nuclear Ca²⁺ may be increased by IP₃ in three ways: (1) IP₃ may increase the cytoplasmic Ca²⁺ transient. Cytoplasmic Ca²⁺ then diffuses through the nuclear pores into the nucleoplasm to elicit a delayed nuclear Ca²⁺ transient. (2) Cytoplasmic IP₃ may itself diffuse through the nuclear pores into the nucleus to activate nuclear IP₃Rs facing the nucleoplasm. Ca²⁺ release from the nuclear envelope, a Ca²⁺ store continuous with the SR, into the nucleoplasm then increases the nuclear Ca²⁺ concentration directly. (3) Finally, IP₃ may also activate IP₃Rs on the nuclear envelope facing the cytoplasm. Ca²⁺ release from the nuclear envelope into the cytoplasm may then increase nuclear Ca²⁺ concentration indirectly via cytoplasmic Ca²⁺ diffusing into the nucleus through nuclear pores. A fourth alternative (not shown in the cartoon) is the generation of IP₃ in the nucleus via nuclear GPCR coupling to the nuclear PLC-IP₃ cascade. Abbreviations: CICR, Ca²⁺-induced Ca²⁺ release; GFR, growth factor receptor; GPCR, G protein-coupled receptor; LTCC, L-type Ca²⁺ channel; NPC, nuclear pore complex; NCX, Na⁺/Ca²⁺ exchanger; PLC, phospholipase C.

Fig. 5

Endothelin-1 enhances nuclear Ca²⁺ transients in atrial myocytes. (A) Two-dimensional confocal images of a rabbit atrial myocyte during an electrically-stimulated Ca²⁺ transient before (Control) and after the addition of 10 nM endothelin-1. The nucleus is visible as the oval area in the cell center with a delayed Ca²⁺ increase during the rising phase and with a delayed Ca²⁺ decrease during the decaying phase of the Ca²⁺ transient. (B) Ca²⁺ transients (from the cell shown in A) obtained from the entire cytoplasm (black) and the nucleus (red). Addition of 10 nM endothelin-1 (ET-1) increases the cytoplasmic and, even more pronounced, the nuclear Ca²⁺ transient. When a threshold concentration of 0.1 nM endothelin-1 is applied (right; different cell), the cytoplasmic Ca²⁺ transient remains unaltered, whereas the nuclear Ca²⁺ transient is augmented selectively. Modified from [72]. Copyright of the Journal of Cell Science; used by kind permission.d