Local activation of focal adhesion kinase orchestrates the positioning of presynaptic scaffold proteins and Ca2+ channel function to control glucose dependent insulin secretion.

A developing understanding suggests that spatial compartmentalisation of components the glucose stimulus?secretion pathway in pancreatic β cells are critical in controlling insulin secretion. To investigate the mechanisms, we have developed live-cell sub-cellular imaging methods using the organotypic pancreatic slice. We demonstrate that the organotypic pancreatic slice, when compared with isolated islets, preserves intact β cell structure, and enhances glucose dependent Ca2+ responses and insulin secretion. Using the slice technique, we have discovered the essential role of local activation of integrins and the downstream component, focal adhesion kinase, in regulating β cells. Integrins and focal adhesion kinase are exclusively activated at the β cell capillary interface and using in situ and in vitro models we show their activation both positions presynaptic scaffold proteins, like ELKS and liprin, and regulates glucose dependent Ca2+ responses and insulin secretion. We conclude that focal adhesion kinase orchestrates the final steps of glucose dependent insulin secretion within the restricted domain where β cells contact the islet capillaries.

Numerical methods for the detection of phase defect structures in excitable media

Electrical waves that rotate in the heart organize dangerous cardiac arrhythmias. Finding the region around which such rotation occurs is one of the most important practical questions for arrhythmia management. For many years, the main method for finding such regions was so-called phase mapping, in which a continuous phase was assigned to points in the heart based on their excitation status and defining the rotation region as a point of phase singularity. Recent analysis, however, showed that in many rotation regimes there exist phase discontinuities and the region of rotation must be defined not as a point of phase singularity, but as a phase defect line. In this paper we use this novel methodology and perform comparative study of three different phase definitions applied to in-silico data and to experimental data obtained from optical voltage mapping experiments on monolayers of human atrial myocytes. We introduce new phase defect detection algorithms and compare them with those that appeared in literature already. We find that the phase definition is more important than the algorithm to identify sudden spatial phase variations. Sharp phase defect lines can be obtained from a phase derived from local activation times observed during one cycle of arrhythmia. Alternatively, similar quality can be obtained from a reparameterization of the classical phase obtained from observation of a single timeframe of transmembrane potential. We found that the phase defect line length was 35.9(62)mm in the Fenton-Karma model and 4.01(55)mm in cardiac human atrial myocyte monolayers. As local activation times are obtained during standard clinical cardiac mapping, the methods are also suitable to be applied to clinical datasets. All studied methods are publicly available and can be downloaded from an institutional web-server.

Association of Reduced Free T3 to Free T4 Ratio with Lower Serum Creatinine in Japanese Hemodialysis Patients

Purpose: Low T3 syndrome is defined by a fall in free triiodothyronine (FT3) in spite of normal serum thyroid-stimulating hormone (TSH) and often normal free thyroxin (FT4). A low FT3/FT4 ratio, a relevant marker for low T3 syndrome, is known as a risk of mortality in hemodialysis (HD) patients, as well as low muscle mass in the general population. Because of the local activation of T4 to FT3 in muscle tissue, we examined the association of FT3/FT4 ratio with serum creatinine, a marker of muscle mass and strength in HD patients to investigate the significance of muscle tissue in the development of low T3 syndrome in HD patients. Methods: This was a cross-sectional study derived from our prospective cohort study, named DREAM, of Japanese HD patients. After the exclusion of patients with treated and untreated thyroid dysfunction, 332 patients were analyzed in the study. Results: The serum FT4 and TSH of HD patients (n = 332) were 0.9 ± 0.1 ng/dL. and 2.0 ± 0.9IU/mL, which were within the respective normal range, while serum FT3 was 2.2 ± 0.3 pg/mL. As many as 101 out of 332 (30.4%) HD patients exhibited a serum FT3 less than the normal lower limit of 2.2 pg/mL. The serum FT3/FT4 ratio correlated significantly positively with serum creatinine, and inversely with serum log CRP and total cholesterol, while it exhibited a tendency towards positive correlation with serum albumin. Multiple regression analysis, which included serum creatinine, albumin, and log CRP, simultaneously, in addition to sex, age, diabetic kidney disease or not, log HD duration, body mass index, systolic blood pressure, and Kt/V, as independent variables, revealed an independent and significant positive association of serum creatinine, but not serum albumin or CRP, with the serum FT3/FT4 ratio. Conclusions: The present study demonstrated an independent and positive correlation of serum creatinine with the serum FT3/FT4 ratio in HD patients. The lack of association of the serum FT3/FT4 ratio with serum albumin and CRP suggested the presence of a creatinine-specific mechanism to associate with serum FT3/FT4 ratio. Because of the local activation of T4 to T3 at muscle tissue, a lower muscle mass may be causatively associated with low T3 syndrome.

Specialist aphids feeding causes local activation of salicylic and jasmonic acid signaling in Arabidopsis veins

Aphids, the phloem sap feeders, probe into leaf tissues and activate a complex network of plant defence responses. Phytohormonal signaling plays a major role in this network; however, the dynamics of the signals spreading is yet to be clarified. Despite the growing knowledge about transcriptomic changes upon infestation, results often differ due to sampling, varying strongly between the tissues collected at the single feeding site, individual leaves, pooled infested leaves, or whole plant rosettes. This study focuses on activation of salicylic and jasmonic acid signals in Arabidopsis leaves during infestation by cabbage aphid (Brevicoryne brassicae) in high spatio-temporal resolution. We used genetically encoded fluorescent biosensors, histochemistry and qRT-PCR to precisely map activation of distinct branches of phytohormonal signaling. We found a rapid induction of salicylic and jasmonic acid signaling markers in cells surrounding stylet puncture, co-localizing with callose deposition. For both PR1 and JAZ10 we detected activation at 24 hpi, increasing and spreading along the veins until 72 hpi and, to a lesser extent, within the epidermal pavement cells. The SA signaling wave appeared in parallel with JA-associated, and continued to increase in time. Our results first show a local activation of SA- and JA-related responses after stylet penetration of Arabidopsis leaves and bring a detailed insight into the spatio-temporal complexity of plant defence activation during specialist aphid attack.

A Phase Defect Framework for the Analysis of Cardiac Arrhythmia Patterns

During cardiac arrhythmias, dynamical patterns of electrical activation form and evolve, which are of interest to understand and cure heart rhythm disorders. The analysis of these patterns is commonly performed by calculating the local activation phase and searching for phase singularities (PSs), i.e., points around which all phases are present. Here we propose an alternative framework, which focuses on phase defect lines (PDLs) and surfaces (PDSs) as more general mechanisms, which include PSs as a specific case. The proposed framework enables two conceptual unifications: between the local activation time and phase description, and between conduction block lines and the central regions of linear-core rotors. A simple PDL detection method is proposed and applied to data from simulations and optical mapping experiments. Our analysis of ventricular tachycardia in rabbit hearts (n = 6) shows that nearly all detected PSs were found on PDLs, but the PDLs had a significantly longer lifespan than the detected PSs. Since the proposed framework revisits basic building blocks of cardiac activation patterns, it can become a useful tool for further theory development and experimental analysis.

KRAP tethers IP3 receptors to actin and licenses them to evoke cytosolic Ca2+ signals

Regulation of IP3 receptors (IP3Rs) by IP3 and Ca2+ allows regenerative Ca2+ signals, the smallest being Ca2+ puffs, which arise from coordinated openings of a few clustered IP3Rs. Cells express thousands of mostly mobile IP3Rs, yet Ca2+ puffs occur at a few immobile IP3R clusters. By imaging cells with endogenous IP3Rs tagged with EGFP, we show that KRas-induced actin-interacting protein (KRAP) tethers IP3Rs to actin beneath the plasma membrane. Loss of KRAP abolishes Ca2+ puffs and the global increases in cytosolic Ca2+ concentration evoked by more intense stimulation. Over-expressing KRAP immobilizes additional IP3R clusters and results in more Ca2+ puffs and larger global Ca2+ signals. Endogenous KRAP determines which IP3Rs will respond: it tethers IP3R clusters to actin alongside sites where store-operated Ca2+ entry occurs, licenses IP3Rs to evoke Ca2+ puffs and global cytosolic Ca2+ signals, implicates the actin cytoskeleton in IP3R regulation and may allow local activation of Ca2+ entry.

การสร้างภาพสามมิติ (3D Mapping) ในกลุ่มผู้ป่วยภาวะหัวใจเต้นผิดจังหวะชนิดเร็วที่ได้รับการรักษาโดยการจี้ด้วยกระแสไฟฟ้า (Ablation)

ในปัจจุบันการสร้างภาพสามมิติ (3D Mapping) ในกลุ่มผู้ป่วยภาวะหัวใจเต้นผิดจังหวะชนิดเร็วที่ได้รับการรักษาโดยการจี้ด้วยกระแสไฟฟ้า (RF Ablation) ได้รับความนิยมอย่างแพร่หลายมากขึ้น เนื่องจากการสร้างภาพสามมิติ มีการนำเทคโนโลยีสมัยใหม่เข้ามาช่วยในการสร้างภาพได้แก่ Magnetic Technology, Current-based technology, Hybrid technology ทำให้สามารถสร้างภาพสามมิติออกมาได้หลายรูปแบบอย่างเช่น Anatomical mapping, Local Activation Time mapping (LAT), Bipolar Voltage mapping, Complex Fractionated Atrial Electrogram (CFAEs) Map, Pace map, Merge หรือ Fusion, Reentrant map เป็นต้นโดยภาพที่ได้นอกจากจะแสดงเป็นภาพนิ่งแล้วยังสามารถแสดงเป็น Video Animation ได้อีกด้วยอย่างเช่น Propagation Map, Ripple Map เป็นต้น ทำให้มีความแม่นยำในการรักษา ผู้ป่วยได้รับปริมาณรังสีที่น้อยลง มีความปลอดภัย และลดภาวะแทรกซ้อน ซึ่งก่อให้เกิดประโยชน์สูงสุดแก่ผู้ป่วย คำสำคัญ: การสร้างภาพสามมิติ, การจี้ด้วยกระแสไฟฟ้า