Computational models of cardiomyocyte dyads and perinuclear regions, and a sheep model of pressure overload
Computational simulation of IP3R activity and experimental pressure overload
Subcellular Ca2+ dynamics, spontaneous Ca2+ spark formation, and nuclear morphologysurrogate
Computational and experimental models demonstrate that IP3Rs play a key role in regulating subcellular calcium signaling, influencing cardiomyocyte function and hypertrophic growth.
The cyclic, coordinated contraction and relaxation of cardiomyocytes underlie the heart's pumping action. By linking electrical depolarisation of the sarcolemma during the action potential to myofilament contraction, calcium (Ca2+) is central to the excitation-contraction coupling (ECC) process underlying cardiomyocyte contraction. During ECC, a cell-wide rise in intracellular Ca2+ initiates contraction, while its subsequent clearance enables relaxation, resetting the cell for the next cycle. This global Ca2+ transient is spatially composed of synchronously evoked elementary Ca2+ release events, termed Ca2+ sparks, mediated by ryanodine receptors (RyRs), a class of sarcoplasmic reticulum Ca2+ channels, localised at specialised microdomains called dyads. Inositol 1,4,5-trisphosphate receptors (IP3Rs), another class of sarcoplasmic reticulum Ca2+ channels, have been shown to modulate ECC, an effect hypothesised to stem from their co-localisation with RyRs in the dyadic microdomain. To test this, we developed a spatial computational model of the dyad to investigate how functional IP3Rs influence Ca2+ dynamics within this microdomain. Simulations revealed that IP3Rs elevate dyadic Ca2+, sensitising proximal RyRs toward activation and thereby promoting spontaneous Ca2+ spark formation. The stochastic gating behaviour of IP3Rs is essential for eliciting this effect. RyRs are also expressed outside dyads in non-coupled clusters. Confocal and super-resolution imaging have indicated that non-coupled RyR clusters are more prevalent in pathological conditions, with IP3Rs preferentially co-localising with them. To investigate the functional implications of this spatial organisation, we extended the aforementioned computational model to simulate the Ca2+ release behaviour of non-coupled RyR clusters in the presence of neighbouring sparking dyads. Simulations revealed that IP3Rs increased spontaneous Ca2+ release events and enhanced the sensitivity of non-coupled RyRs to Ca2+ diffusing from nearby dyads, thereby improving the synchronicity of Ca2+ release between non-coupled and dyadic RyRs during ECC. In addition to ECC, Ca2+ also activates the pro-hypertrophic transcription factors NFAT and MEF2, driving gene expression programmes that underlie cardiomyocyte growth and cardiac hypertrophy. The necessity of nuclear Ca2+ for activating these transcription factors has been established, with Ca2+ release from IP3Rs expressed on the nuclear envelope (NE) shown to be sufficient for their activation. However, the nucleoplasm also exhibits Ca2+ fluctuations whose kinetics lag behind that of ECC-associated Ca2+ transients originating from the cytoplasm. The Ca2+-handling mechanisms hypothesised to underlie these ECC-associated nucleoplasmic Ca2+ transients have been indirectly supported, but remain to be biophysically demonstrated. Furthermore, it is unclear how NE-bound IP3Rs modulate nucleoplasmic Ca2+ amidst these transients to encode hypertrophic signalling. To address these gaps, we developed a spatial computational model of the perinuclear region that incorporates NE permeability, nuclear-specific Ca2+ buffering, and NE-bound IP3R activity to simulate nucleocytoplasmic Ca2+ dynamics during ECC. Simulations revealed that NE permeability primarily contributes to the kinetic delay of Ca2+ transients in the nucleoplasm relative to that of the cytoplasm, while NE-bound IP3R activity prolongs the transient's decay to baseline. Structural remodelling of the nucleus may also influence nucleoplasmic Ca2+ handling. Using transmission electron microscopy, we characterised age- and disease-associated changes in nuclear morphology of cardiomyocytes in a sheep model of pressure overload. Morphometric analyses revealed an increase in nuclear length with age while nuclear shape complexity decreases in young hearts subject to pressure overload, due to a reduction in NE invagination density. Together, this thesis provides mechanistic insights into the role of IP3Rs in regulating cardiomyocyte function and growth through new computational frameworks and experimental studies. These findings advance our understanding of subcellular Ca2+ signalling in cardiomyocyte pathophysiology.
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Joshua Jeen Tung Chung
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Joshua Jeen Tung Chung (Tue,) studied this question.