Véronique Maupoil

Catecholaminergic automatic activity in the rat pulmonary vein: electrophysiological differences between cardiac muscle in the left atrium and pulmonary vein

Ectopic activity in cardiac muscle within pulmonary veins (PVs) is associated with the onset and the maintenance of atrial fibrillation in humans. The mechanism underlying this ectopic activity is unknown. Here we investigate automatic activity generated by catecholaminergic stimulation in the rat PV. Intracellular microelectrodes were used to record electrical activity in isolated strips of rat PV and left atrium (LA). The resting cardiac muscle membrane potential was lower in PV [-70 +/- 1 (SE) mV, n = 8] than in LA (-85 +/- 1 mV, n = 8). No spontaneous activity was recorded in PV or LA under basal conditions. Norepinephrine (10(-5) M) induced first a hyperpolarization (-8 +/- 1 mV in PV, -3 +/- 1 mV in LA, n = 8 for both) then a slowly developing depolarization (+21 +/- 2 mV after 15 min in PV, +1 +/- 2 mV in LA) of the resting membrane potential. Automatic activity occurred only in PV; it was triggered at approximately -50 mV, and it occurred as repetitive bursts of slow action potentials. The diastolic membrane potential increased during a burst and slowly depolarized between bursts. Automatic activity in the PV was blocked by either atenolol or prazosine, and it could be generated with a mixture of cirazoline and isoprenaline. In both tissues, cirazoline (10(-6) M) induced a depolarization (+37 +/- 2 mV in PV, n = 5; +5 +/- 1 mV in LA, n = 5), and isoprenaline (10(-7) M) evoked a hyperpolarization (-11 +/- 3 mV in PV, n = 7; -3 +/- 1 mV in LA, n = 6). The differences in membrane potential and reaction to adrenergic stimulation lead to automatic electrical activity occurring specifically in cardiac muscle in the PV.

An Increased Regional Blood Flow Precedes Mesenteric Inflammation in Rats Treated by a Phosphodiesterase 4 Inhibitor

The study was undertaken to assess the hemodynamic effects induced by a single dose of the phosphodiesterase 4 (PDE4) inhibitor, CI-1044, which is known to cause mesenteric vascular alterations in rats. In the present study, an administration of 160 mg/kg of CI-1044 caused perivascular and interstitial inflammation, with infiltrates of admixed neutrophils and macrophages but without evidence of vascular necrosis (ileum, 15/20 rats; duodenum + jejunum, 7/20 rats). Four hours after administration, blood pressure was decreased (- 13%). A fluorescent microsphere technique demonstrated that, in these conditions, cardiac output was doubled (+ 100%) and total peripheral resistance was decreased (- 54%). The largest increases in blood flow were measured in the duodenum (+ 101%), in the jejunum (+ 110%), and in the ileum (+ 192%). Therefore, the mesentery was the most sensitive organ affected by the drug and, within this area, parts with the highest incidence of vascular alteration were those which had shown the highest increase in flow. In addition, isolated precontracted mesenteric resistance arteries dissected from untreated animals were fully relaxed when incubated with increasing concentrations of CI-1044 up to 2.5 x 10(-5)M. At this latter concentration, contractile abilities and sensitivities to the physiological agonist noradrenaline (NA) and to the thromboxane analogue U46619 were significantly attenuated (- 28 and - 27%, respectively). This effect could lead to a decreased response to NA and possibly to other agonists in vivo consistent with the vasodilation observed with the microsphere technique. These data provide evidence that the PDE4 inhibitor CI-1044 induces changes of vascular tone that could lead to histological alterations in the mesenteric area.

A TTX-Sensitive Resting Na+ Permeability Contributes to the Catecholaminergic Automatic Activity in Rat Pulmonary Vein

Ectopic activity arising from pulmonary veins (PV) plays a prominent role in the onset of atrial fibrillation in humans. Rat PV cardiac muscle cells have a lower resting membrane potential (RMP) than the left atria (LA) and presents in the presence of norepinephrine an automatic activity, which occurs in bursts. This study investigated the role of Na channels upon the RMP and the catecholaminergic automatic activity (CAA) in PV cardiac muscle.

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.

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.

Structural heterogeneity of the rat pulmonary vein myocardium: consequences on intracellular calcium dynamics and arrhythmogenic potential

Mechanisms underlying ectopic activity in the pulmonary vein (PV) which triggers paroxysmal atrial fibrillation are unknown. Although several studies have suggested that calcium signalling might be involved in these arrhythmias, little is known about calcium cycling in PV cardiomyocytes (CM). We found that individual PV CM showed a wide range of transverse tubular incidence and organization, going from their virtual absence, as described in atrial CM, to well transversally organised tubular systems, like in ventricular CM. These different types of CM were found in groups scattered throughout the tissue. The variability of the tubular system was associated with cell to cell heterogeneity of calcium channel (Cav1.2) localisation and, thereby, of Cav1.2-Ryanodine receptor coupling. This was responsible for multiple forms of PV CM calcium transient. Spontaneous calcium sparks and waves were not only more abundant in PV CM than in LA CM but also associated with a higher depolarising current. In conclusion, compared with either the atrium or the ventricle, PV myocardium presents marked structural and functional heterogeneity.

HF_IDS_Cam: Fast Video Capture with ImageJ for Real-Time Analysis

Fast online video analysis is currently a key issue for dynamic studies in biology; however, very few tools are available for these concerns. Here we present an ImageJ plug-in: HF_IDS_Cam, which allows for video capture at very high speeds using IDS (Imaging Development Systems GmbH) cameras and image analysis software ImageJ. The software has been optimized for real time video analysis with ImageJ native function and other plug-ins and scripts. The plug-in was written in Java and requires ImageJ 1.47v or higher. HF_IDS_Cam offers a wide range of applications for exploration of dynamic phenomena in biology, from in vitro/ex vivo studies, such as fast fluorescent calcium imaging and voltage optical mapping in cardiac myocytes and neurons, to in-vivo behavioral studies.

0194 : Functional consequences of -adrenergic receptors activation in the rat pulmonary veins and left atria

The role of adrenergic stimulation on pulmonary veins (PV) ectopy and atrial fibrillation initiation is unclear. In the rat, left atrium (LA) and PV cardiomyocytes (CM) have different reactions to α-adrenergic receptor activation. Here, we examine the functional consequences of α-adrenergic receptors activation by cirazoline in rat LA and PV. PV, LA and right and left atria-PV preparations dissected from male Wistar rats were superfused with a physiological solution at 37°C. Electrical conduction within the LA and the PV was recorded with a linear array of 8 extracellular electrodes. Dual intracellular microelectrode recording used a WPI Duo 773 electrometer amplifier. Calcium transients (CaT) were recorded in isolated CM loaded with Fluo-4. Confocal microscopy was used to visualize the distribution of α-adrenergic receptors labeled with BODIPY-prazosin. In PV but not in LA strips stimulated at 0.1Hz, cirazoline induced a concentration-dependent negative inotropic effect (-79±5% at 1μM, p The author hereby declares no conflict of interest

0426 : SarcOptiM, an ImageJ plug-in for cardiomyocyte contractility recording

Accurate measurement of cardiomyocyte contractility is a critical issue for scientists working on cardiac physiology and physiopathology of diseases implying contraction impairment such as heart failure. The most reliable method for the evaluation of the cardiomyocyte contraction amplitude and kinetics consists in the measurement of sarcomere shortening. The few tools available for this are always tied to dedicated video cameras and none is freely distributed. Here, we describe SarcOptiM, a free open-source plug-in developed in Java for ImageJ/Fiji image analysis platform of the NIH, running under Windows, Mac and Linux operating systems. It computes sarcomere length via Fourier transform analysis of video frames captured or displayed in ImageJ and thereby allows real-time or offline sarcomere measurements. This plug-in has been validated with experimental recordings of rat ventricular cardiomyocyte in several conditions including beta adrenergic stimulation, and with synthetic videos of contracting cardiomyocyte with known features. SarcOptiM is not tied to a specific hardware contrary to marketed software, and works with all digital and analogical video cameras interfaced with ImageJ. Thus, the video camera can be chosen according to the experimental needs. The sampling frequency of the sarcomere length recording is set by the video camera frame rate; a sampling frequency of 1000 Hz is easily reachable with a mid-range costless video camera. Moreover, SarcOptiM overcomes some common problems such as displacements related artifacts and orientation of the cardiomyocyte in the visual field. In conclusion, SarcOptiM is a free, powerful and user-friendly tool for researchers studying cardiomyocyte contractility in all species including human. The author hereby declares no conflict of interest

0456 : Comparison of spontaneous calcium release events in pulmonary vein and left atria cardiomyocytes

Pulmonary veins (PV) have been involved in the onset of atrial fibrillation in humans. In rat, we reported a catecholaminergic automatic activity in PV cardiomyocytes (CM) and not in the left atria (LA). Our objective was to identify differences in calcium cycle between PV and LA CM which could explain the arrhythmogenic potential of PV since cytoplasmic calcium oscillations precede variations of membrane potential which are involved in arrhythmias. Confocal fluorescence imaging was performed in rat isolated CM with di- 8-ANEPPS to study tranverse tubule organization, with specific antibodies for ryanodine receptors and L-type calcium channels (RyR-Cav1.2) coupling, with fluo-4 for spontaneous calcium events (SCaE) frequency determination and calcium transient amplitude. Calcium current was recorded with a wholecell patch clamp technique. Spontaneous calcium events frequency was significantly (p In conclusion, the tranverse tubule organization associated with more important calcium exchanges support the higher spontaneous calcium events frequency observed in PV CM. This could be related to the arrhythmogenic potential of PV CM.