The Builders of the Junction: Roles of Junctophilin1 and Junctophilin2 in the Assembly of the Sarcoplasmic Reticulum–Plasma Membrane Junctions in Striated Muscle

Contraction of striated muscle is triggered by a massive release of calcium from the sarcoplasmic reticulum (SR) into the cytoplasm. This intracellular calcium release is initiated by membrane depolarization, which is sensed by voltage-gated calcium channels CaV1.1 (in skeletal muscle) and CaV1.2 (in cardiac muscle) in the plasma membrane (PM), which in turn activate the calcium-releasing channel ryanodine receptor (RyR) embedded in the SR membrane. This cross-communication between channels in the PM and in the SR happens at specialized regions, the SR-PM junctions, where these two compartments come in close proximity. Junctophilin1 and Junctophilin2 are responsible for the formation and stabilization of SR-PM junctions in striated muscle and actively participate in the recruitment of the two essential players in intracellular calcium release, CaV and RyR. This short review focuses on the roles of junctophilins1 and 2 in the formation and organization of SR-PM junctions in skeletal and cardiac muscle and on the functional consequences of the absence or malfunction of these proteins in striated muscle in light of recently published data and recent advancements in protein structure prediction.

Altered Calcium Permeability of AMPA Receptor Drives NMDA Receptor Inhibition in the Hippocampus of Murine Obesity Models

Evidence has accumulated that higher consumption of high-fat diets (HFDs) during the juvenile/adolescent period induces altered hippocampal function and morphology; however, the mechanism behind this phenomenon remains elusive. Using high-resolution structural imaging combined with molecular and functional interrogation, a murine model of obesity treated with HFDs for 12 weeks after weaning mice was shown to change in the glutamate-mediated intracellular calcium signaling and activity, including further selective reduction of gray matter volume in the hippocampus associated with memory recall disturbance. Dysregulation of intracellular calcium concentrations was restored by a non-competitive α-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) antagonist, followed by normalization of hippocampal volume and memory recall ability, indicating that AMPARs may serve as an attractive therapeutic target for obesity-associated cognitive decline.

On the Role of the Excitation/Inhibition Balance of Homeostatic Artificial Neural Networks

Homeostatic models of artificial neural networks have been developed to explain the self-organization of a stable dynamical connectivity between the neurons of the net. These models are typically two-population models, with excitatory and inhibitory cells. In these models, connectivity is a means to regulate cell activity, and in consequence, intracellular calcium levels towards a desired target level. The excitation/inhibition (E/I) balance is usually set to 80:20, a value characteristic for cortical cell distributions. We study the behavior of these homeostatic models outside of the physiological range of the E/I balance, and we find a pronounced bifurcation at about the physiological value of this balance. Lower inhibition values lead to sparsely connected networks. At a certain threshold value, the neurons develop a reasonably connected network that can fulfill the homeostasis criteria in a stable way. Beyond the threshold, the behavior of the artificial neural network changes drastically, with failing homeostasis and in consequence with an exploding number of connections. While the exact value of the balance at the bifurcation point is subject to the parameters of the model, the existence of this bifurcation might explain the stability of a certain E/I balance across a wide range of biological neural networks. Assuming that this class of models describes the self-organization of biological network connectivity reasonably realistically, the omnipresent physiological balance might represent a case of self-organized criticality in order to obtain a good connectivity while allowing for a stable intracellular calcium homeostasis.

A turquoise fluorescence lifetime-based biosensor for quantitative imaging of intracellular calcium

The most successful genetically encoded calcium indicators (GECIs) employ an intensity or ratiometric readout. Despite a large calcium-dependent change in fluorescence intensity, the quantification of calcium concentrations with GECIs is problematic, which is further complicated by the sensitivity of all GECIs to changes in the pH in the biological range. Here, we report on a sensing strategy in which a conformational change directly modifies the fluorescence quantum yield and fluorescence lifetime of a circular permutated turquoise fluorescent protein. The fluorescence lifetime is an absolute parameter that enables straightforward quantification, eliminating intensity-related artifacts. An engineering strategy that optimizes lifetime contrast led to a biosensor that shows a 3-fold change in the calcium-dependent quantum yield and a fluorescence lifetime change of 1.3 ns. We dub the biosensor Turquoise Calcium Fluorescence LIfeTime Sensor (Tq-Ca-FLITS). The response of the calcium sensor is insensitive to pH between 6.2–9. As a result, Tq-Ca-FLITS enables robust measurements of intracellular calcium concentrations by fluorescence lifetime imaging. We demonstrate quantitative imaging of calcium concentrations with the turquoise GECI in single endothelial cells and human-derived organoids.

Excessive Accumulation of Intracellular Ca2+ After Acute Exercise Potentiated Impairment of T-cell Function

Ca2+ is an important intracellular second messenger known to regulate several cellular functions. This research aimed to investigate the mechanisms of exercise-induced immunosuppression by measuring intracellular calcium levels, Ca2+-regulating gene expression, and agonist-evoked proliferation of murine splenic T lymphocytes. Mice were randomly assigned to the control, sedentary group (C), and three experimental groups, which performed a single bout of intensive and exhaustive treadmill exercise. Murine splenic lymphocytes were separated by density-gradient centrifugation immediately (E0), 3h (E3), and 24h after exercise (E24). Fura-2/AM was used to monitor cytoplasmic free Ca2+ concentration in living cells. The combined method of carboxyfluorescein diacetate succinimidyl ester (CFSE) labeling and flow cytometry was used for the detection of T cell proliferation. The transcriptional level of Ca2+-regulating genes was quantified by using qPCR. Both basal intracellular Ca2+ levels and agonist (ConA, OKT3, or thapsigargin)-induced Ca2+ transients were significantly elevated at E3 group (p<0.05 vs. control). However, mitogen-induced cell proliferation was significantly decreased at E3 group (p<0.05 vs. control). In parallel, the transcriptional level of plasma membrane Ca2+-ATPases (PMCA), sarco/endoplasmic reticulum Ca2+-ATPases (SERCA), TRPC1, and P2X7 was significantly downregulated, and the transcriptional level of IP3R2 and RyR2 was significantly upregulated in E3 (p<0.01 vs. control). In summary, this study demonstrated that acute exercise affected intracellular calcium homeostasis, most likely by enhancing transmembrane Ca2+ influx into cells and by reducing expression of Ca2+-ATPases such as PMCA and SERCA. However, altered Ca2+ signals were not transduced into an enhanced T cell proliferation suggesting other pathways to be responsible for the transient exercise-associated immunosuppression.