Selective Inhibition of Pulmonary Vein Excitability by Constitutively Active GIRK Channels Blockade in Rats.

Pulmonary veins (PV) are the main source of ectopy, triggering atrial fibrillation. This study investigated the roles of G protein-coupled inwardly rectifying potassium (GIRK) channels in the PV and the left atrium (LA) of the rat. Simultaneous intracellular microelectrode recording from the LA and the PV of the rat found that in the presence or absence of acetylcholine, the GIRK channel blocker tertiapin-Q induced AP duration elongation in the LA and the loss of over-shooting AP in the PV, suggesting the presence of constitutively active GIRK channels in these tissues. Patch-clamp recordings from isolated myocytes showed that tertiapin-Q inhibited a basal inwardly rectified background current in PV cells with little effect in LA cells. Experiments with ROMK1 and KCa1.1 channel blockers ruled out the possibility of an off-target effect. Western blot showed that GIRK4 subunit expression was greater in PV cardiomyocytes, which may explain the differences observed between PV and LA in response to tertiapin-Q. In conclusion, GIRK channels blockade abolishes AP only in the PV, providing a molecular target to induce electrical disconnection of the PV from the LA.

Spiky: An ImageJ Plugin for Data Analysis of Functional Cardiac and Cardiomyocyte Studies.

INTRODUCTION AND OBJECTIVE: Nowadays, investigations of heart physiology and pathophysiology rely more and more upon image analysis, whether for the detection and characterization of events in single cells or for the mapping of events and their characteristics across an entire tissue. These investigations require extensive skills in image analysis and/or expensive software, and their reproducibility may be a concern. Our objective was to build a robust, reliable and open-source software tool to quantify excitation-contraction related experimental data at multiple scales, from single isolated cells to the whole heart. METHODS AND RESULTS: A free and open-source ImageJ plugin, Spiky, was developed to detect and analyze peaks in experimental data streams. It allows rapid and easy analysis of action potentials, intracellular calcium transient and contraction data from cardiac research experiments. As shown in the provided examples, both classical bi-dimensional data (XT signals) and video data obtained from confocal microscopy and optical mapping experiments (XYT signals) can be analyzed. Spiky was written in ImageJ Macro Language and JAVA, and works under Windows, Mac and Linux operating systems. CONCLUSION: Spiky provides a complete working interface to process and analyze cardiac physiology research data.

Aqueous Fraction from Hibiscus sabdariffa Relaxes Mesenteric Arteries of Normotensive and Hypertensive Rats through Calcium Current Reduction and Possibly Potassium Channels Modulation.

BACKGROUND/OBJECTIVES: Hibiscus sabdariffa L. (H. sabdariffa (HS)) extract has a vascular relaxant effect on isolated rat thoracic aorta, but data on small resistance arteries, which play an important role on the development of hypertension, are still missing. The purposes of this study were (1) to assess the effect on isolated mesenteric arteries (MA) from normotensive (Wistar and Wistar-Kyoto (WKY)) and spontaneous hypertensive rats (SHR); (2) to elucidate the mechanism(s) of action underling the relaxant effect in light of bioactive components. METHODS: Vascular effects of HS aqueous fraction (AF) on isolated MA rings, as well as its mechanisms of action, were assessed using the contractility and intracellular microelectrode technique. The patch clamp technique was used to evaluate the effect of HS AF on the L-type calcium current. Extraction and enrichment of AF were carried out using liquid-liquid extraction, and the yield was analyzed using HPLC. RESULTS: The HS AF induced a concentration-dependent relaxant effect on MA rings of SHR (EC50 = 0.83 ± 0.08 mg/mL), WKY (EC50 = 0.46 ± 0.04 mg/mL), and Wistar rats (EC50 = 0.44 ± 0.08 mg/mL) pre-contracted with phenylephrine (10 µM). In Wistar rats, the HS AF maximum relaxant effect was not modified after endothelium removal or when a guanylate cyclase inhibitor (ODQ, 10 µM) and a selective ß2-adrenergic receptor antagonist (ICI-118551, 1 µM) were incubated with the preparation. Otherwise, it was reduced by 34.57 ± 10.66% when vascular rings were pre-contracted with an 80 mM [K+] solution (p < 0.001), which suggests an effect on ionic channels. HS AF 2 mg/mL significantly decreased the peak of the L-type calcium current observed in cardiac myocytes by 24.4%. Moreover, though the vasorelaxant effect of HS, AF was reduced by 27% when the nonselective potassium channels blocker (tetraethylammonium (TEA) 20 mM) was added to the bath (p < 0.01). The extract did not induce a membrane hyperpolarization of smooth muscle cells, which might suggest an absence of a direct effect on background potassium current. CONCLUSION: These results highlight that the antihypertensive effect of HS probably involves a vasorelaxant effect on small resistance arteries, which is endothelium independent. L-type calcium current reduction contributes to this effect. The results could also provide a link between the vasorelaxant effect and the bioactive compounds, especially anthocyanins.

SarConfoCal: simultaneous sarcomere length and cytoplasmic calcium measurements for laser scanning confocal microscopy images.

Summary: Simultaneous recordings of myocytes contractility and their cytoplasmic calcium concentration allow powerful studies, particularly on heart failure and other cardiac dysfunctions. Such studies require dedicated and expensive experimental devices that are difficult to use. Thus we propose SarConfoCal, the first and only software to simultaneously analyse both cytoplasmic calcium variations (from fluorescence signal) and myocytes contractility (from sarcomere length measurement) on laser scanning confocal microscopy images. SarConfoCal is easy to set up and use, especially by people without programming skills. Availability and implementation: The software is freely distributed under the GNU General Public License. Download and setup instructions are available at http://pccv.univ-tours.fr/ImageJ/SarConfoCal . It is provided as a toolset for ImageJ (the open-source program for image analysis provided by the National Institutes of Health). SarConfoCal has been tested under Windows, Mac and Linux operating systems. Contact: come.pasqualin@univ-tours.fr. Supplementary information: Supplementary data are available at Bioinformatics online.

SarcOptiM for ImageJ: high-frequency online sarcomere length computing on stimulated cardiomyocytes.

Accurate measurement of cardiomyocyte contraction is a critical issue for scientists working on cardiac physiology and physiopathology of diseases implying contraction impairment. Cardiomyocytes contraction can be quantified by measuring sarcomere length, but few tools are available for this, and none is freely distributed. We developed a plug-in (SarcOptiM) for the ImageJ/Fiji image analysis platform developed by the National Institutes of Health. SarcOptiM computes sarcomere length via fast Fourier transform analysis of video frames captured or displayed in ImageJ and thus is not tied to a dedicated video camera. It can work in real time or offline, the latter overcoming rotating motion or displacement-related artifacts. SarcOptiM includes a simulator and video generator of cardiomyocyte contraction. Acquisition parameters, such as pixel size and camera frame rate, were tested with both experimental recordings of rat ventricular cardiomyocytes and synthetic videos. It is freely distributed, and its source code is available. It works under Windows, Mac, or Linux operating systems. The camera speed is the limiting factor, since the algorithm can compute online sarcomere shortening at frame rates >10 kHz. In conclusion, SarcOptiM is a free and validated user-friendly tool for studying cardiomyocyte contraction in all species, including human.