The G-protein–gated K+ channel, IKACh, is required for regulation of pacemaker activity and recovery of resting heart rate after sympathetic stimulation

Parasympathetic regulation of sinoatrial node (SAN) pacemaker activity modulates multiple ion channels to temper heart rate. The functional role of the G-protein–activated K+ current (IKACh) in the control of SAN pacemaking and heart rate is not completely understood. We have investigated the functional consequences of loss of IKACh in cholinergic regulation of pacemaker activity of SAN cells and in heart rate control under physiological situations mimicking the fight or flight response. We used knockout mice with loss of function of the Girk4 (Kir3.4) gene (Girk4−/− mice), which codes for an integral subunit of the cardiac IKACh channel. SAN pacemaker cells from Girk4−/− mice completely lacked IKACh. Loss of IKACh strongly reduced cholinergic regulation of pacemaker activity of SAN cells and isolated intact hearts. Telemetric recordings of electrocardiograms of freely moving mice showed that heart rate measured over a 24-h recording period was moderately increased (10%) in Girk4−/− animals. Although the relative extent of heart rate regulation of Girk4−/− mice was similar to that of wild-type animals, recovery of resting heart rate after stress, physical exercise, or pharmacological β-adrenergic stimulation of SAN pacemaking was significantly delayed in Girk4−/− animals. We conclude that IKACh plays a critical role in the kinetics of heart rate recovery to resting levels after sympathetic stimulation or after direct β-adrenergic stimulation of pacemaker activity. Our study thus uncovers a novel role for IKACh in SAN physiology and heart rate regulation

Rôle fonctionel des canaux ioniques dans la génération de l'automatisme et de la conduction cardiaque

La génération et la régulation du rythme cardiaque nécessite une coordination précise entre l'activité électrique du nœud sinoatrial (NSA), à l'origine de l'activité pacemaker cardiaque, et le tissu de conduction (TC), composé du nœud atrioventriculaire (NAV) et du réseau de His-Purkinje. Lors de pathologies cardiaques, il arrive que le centre primaire pacemaker (NSA) soit défaillant et ne génère plus d'impulsions électriques. Le NAV, en plus de sa fonction de conduction, est doté d'automatisme et impose sa fréquence au cœur. La connaissance de la physiologie du rythme cardiaque nécessite l'identification et la description du rôle des gènes des différents canaux ioniques impliqués dans la génération et la régulation de l'activité pacemaker. L'approche via les souris génétiquement modifiées nous donnent l'opportunité de mieux comprendre le mécanisme exact de génération de l'automatisme cardiaque. L'approche expérimentale était jusqu'à présent considérée comme difficile du fait du manque d'accessibilité à ce type de cellules chez la souris. Pendant ma thèse, j'ai développé une technique d'isolement des différentes cellules automatiques de la souris adulte (cellules du NSA, cellules du NAV et cellules des FP). Une comparaison des différents courants ioniques dépendants du voltage sur ces cellules automatiques montrent que le courant pacemaker (If) et les courants calciques (ICa,L et ICa,T) sont présents dans les cellules de conduction en moins forte densité que dans les cellules du SAN. Les souris invalidées pour les gènes Cav1.3 codant pour ICa,L et Cav3.1 codant pour ICa,T montrent une absence totale de ces courants dans les cellules du NAV et du SAN. Au niveau cellulaire, les cellules du SAN et du NAV présentent une bradycardie associée à des dysrythmies impliquant que ces courants calciques sont primordiaux pour l'activité pacemaker. Par contre, les courants potassiques (IK,r et IK1) sont davantage exprimés dans les cellules du NAV et les cellules de Purkinje que dans les cellules du SAN. Cette forte expression de ces courants pourrait expliquer ce ralentissement de l'automatisme des cellules pacemaker du NAV et des cellules des FPThe generation and regulation of the heartbeat is a phenomenon, which requires the precise coordination between the electrical activity of the sino-atrial node (SAN) and the specialized conduction tissue. The heart conduction system (CS) is composed by the atrioventricular node (AVN) and the his-Purkinje fibers (PFs) network system. Automaticity is initiated in the SAN by primary pacemaker cells which generate spontaneous periodic oscillation of their membrane potential. The AVN is also able to generate automaticity , and can drive the heartbeat in case of SAN failure. The PFs network can pace the heart, in case of atrioventricular block. The spontaneous activity of the pacemaker cells is due to the diastolic depolarisation, a slow phase of the pacemaker cycle which drives the membrane voltage from the end of repolarisation phase of an action potential toward the threshold of the following action potential. At the cellular level, pacemaker activity requires the intervention of several families of ionic channels. The molecular basis of AVN and PF automaticity are poorly understood and the role of different ionic channel genes in controlling the diastolic depolarisation phase (DD) in CS have not been investigated. We have isolated spontaneously beating cells from the mouse AVN region and studied ionic channels and pacemaker activity by employing the whole-cell configuration of the patch-clamp technique. ). Compared to SAN, the AVN cells (AVNC) were characterized by a higher expression of the inward rectifier current (IK1) and of the fast delayed-rectifier current (IKr). In contrast, the densities of the hyperpolarization-activated current (If), that of the T- (ICa,T) and L-type (ICa,L) Ca2+ currents were lower AVNC than in SAN cells (SANC). Inhibition of If by 3 M ZD-7228 slowed automaticity of AVNC indicating that If participates to AVNC automaticity in spite of a more negative activation curve in AVNC. The Cav1.3 mediated ICa,L plays an obligatory role in automaticity of AVNC and accounted for almost the totality of ICa,L. TTX-sensitive (INas) and TTX-resistant (INar) Na+ currents had a differential role in automaticity of mouse AVNC, because block of INas slowed pacemaker activity by specifically reducing the rate of the diastolic depolarization (DD), while inhibition of INar showed that this current is the key contributor of the action potential (AP) threshold and upstroke velocity. Modelling of AVN cell automaticity indicates that the lower densities of If, ICa,L, ICa,T, and INas, as well as the higher expression of IK1 and IKr in AVNC than in SANC can explain the intrinsically slower automaticity of AVNCMONTPELLIER-BU Sciences (341722106) / SudocSudocFranceF

If current inhibition: cellular basis and physiology

The slow diastolic depolarization phase in cardiac pacemaker cells is the electrical basis of cardiac automaticity. The hyperpolarization-activated current (I(f)) is one of the key mechanisms underlying diastolic depolarization. Particularly, I(f) is unique in being activated on membrane hyperpolarization following the repolarization phase of the action potential. I(f) has adapted biophysical properties and voltage-dependent gating to initiate pacemaker activity. I(f) possibly constitutes the first voltage-dependent trigger of the diastolic depolarization. For these reasons, I(f) is a natural pharmacological target for controlling heart rate in cardiovascular disease. In this view, I(f) inhibitors have been developed in the past, yet the only molecule to have reached the clinical development is ivabradine. At the cellular level, the remarkable success of ivabradine is to be ascribed to its relatively high affinity for f-channels. Furthermore, ivabradine is the most I(f)-specific inhibitor known to date, since moderate inhibition of other voltage-dependent ionic currents involved in automaticity can be observed only at very high concentrations of ivabradine, more than one order of magnitude from that inhibiting I(f). Finally, the mechanism of block of f-channels by ivabradine has particularly favorable properties in light of controlling heart rate under variable physiological conditions. In this article, we will discuss how I(f) inhibition by ivabradine can lead to reduction of heart rate. To this aim, we will comment on the role of I(f) in cardiac automaticity and on the mechanism of action of ivabradine on f-channels. Some aspects of the cardiac pacemaker mechanism that improve the degree of security of ivabradine will also be highlighted

Alveolar ridge augmentation with 3D‐printed synthetic bone blocks: A clinical case series

Abstract This report documents the clinical and histological outcome of 3D‐printed calcium phosphate blocks placed in two‐stage procedures to successfully rehabilitate atrophic alveolar ridges. This approach yielded a functionally favorable result. Histological evaluations were performed after healing periods of 6 months and showed ongoing bone regeneration and sprouting capillaries

Evaluation of the validity of digital optical microscopy in the assessment of marginal adaptation of dental adhesive interfaces

Analysis of marginal adaptation of dental adhesive interfaces using scanning electron microscopy has proven to be a powerful nondestructive method to evaluate the quality of adhesion. This methodology is, however, time-consuming and needs expensive equipment. The purpose of this study was to evaluate the possibility and efficiency of using a digital optical microscope (DOM) to perform marginal analysis and to compare it with the scanning electron microscope (SEM) analysis. Fifteen defect-free molars were selected for this study. Class V cavities were prepared and restored with resin composite, and epoxy replicas were obtained from silicone impressions of the restored teeth. Custom-made image analysis software was then used to measure the percentage of the noncontinuous margins (NCM) of each sample. To compare the DOM to the gold standard, SEM, each sample was analyzed 10 times using the DOM and three times using the SEM, by the same experienced operator. The repeatability coefficient and concordance were evaluated, and a Bland and Altman analysis was used for comparison of the two methods of measurements. To validate the DOM analysis method, an ANOVA approach (Gage R R) was used. Repeatability and reproducibility, which are two components of precision to validate the DOM analysis system, were calculated. For this, the same restorations were analyzed by two additional operators three times with the DOM. The duration of each step of the analysis using both methods was also recorded as a secondary outcome. Regarding the repeatability of each method, the Friedman test showed no statistically significant difference within the repetitions of measurements by SEM and DOM (p= 0.523 andp= 0.123, respectively). Moreover, the Bland-Altman analysis revealed a bias of 0.86 and concluded no statistically significant difference between the two methods, DOM and SEM. ANOVA evaluated DOM measurement system variation including the variances of repeatability and reproducibility that represented, respectively, 0.3% and 4% of the variance components. Reproducibility or inter-operator variability represented the principal source of variability with a statistically significant difference (p= 0.024). The time required for analysis with SEM was almost double that of DOM. The use of digital optical microscopy appears to be a valid alternative to the SEM for the analysis of marginal adaptation of dental adhesive interfaces. Further studies to evaluate the effect of training of operators in digital optical microscopy could reveal higher accuracy for this method and inter-operator agreement when experience is gained

Physiological and pharmacological insights into the role of ionic channels in cardiac pacemaker activity

The generation of cardiac pacemaker activity is a complex phenomenon which requires the coordinated activity of different membrane ionic channels, as well as intracellular signalling factors including Ca(2+) and second messengers. The precise mechanism initiating automaticity in primary pacemaker cells is still matter of debate and certain aspects of how channels cooperate in the regulation of pacemaking by the autonomic nervous system have not been entirely elucidated. Research in the physiopathology of cardiac automaticity has also gained a considerable interest in the domain of cardiovascular pharmacology, since accumulating clinical and epidemiological evidence indicate a link between an increase in heart rate and the risk of cardiac mortality and morbidity. Lowering the heart rate by specific bradycardic agents in patients with heart disease constitutes a promising way to increase cardioprotection and improve survival. Thus, the elucidation of the mechanisms underlying the generation of pacemaker activity is necessary for the development of new therapeutic molecules for controlling the heart rate. Recent work on genetically modified mouse models provided new and intriguing evidence linking the activity of ionic channels genes to the generation and regulation of pacemaking. Importantly, results obtained on genetically engineered mouse strains have demonstrated that some channels are specifically involved in the generation of cardiac automaticity and conduction, but have no functional impact on the contractile activity of the heart. In this article, we will outline the current knowledge on the role of ionic channels in cardiac pacemaker activity and suggest new potential pharmacological targets for controlling the heart rate without concomitant negative inotropism

Could 3D printing be the future for oral soft tissue regeneration ?

Oral soft tissue defects are a frequently encountered problem in dental praxis. Tooth loss, tooth root or implant recessions, infections or trauma require soft tissue reconstruction. The autologous graft remains the gold standard for gingiva and oral mucosa augmentation. However, prolonged pain, limited amount of harvested tissue, and increased risk of infection have prompted the search for off-shelf alternatives. Several acellularized dermal matrices have been studied without satisfactory results. A newly developed collagen-based sponge is currently in clinical studies for long term evaluation. In these approaches however, the matrix needs to be tailored chair-side for each specific defect. 3D printing technology represents a promising solution as it offers precise production of an individualized 3D graft based on a defined shape and inner structure via a specific computer-aided design using a biomaterial of choice. Combined with smart biocompatible polymers (bioinks) that can be co-printed with cells in a specific architectural design, a more natural-like tissues can be engineered. More natural oral mucosa and gingiva will find application in regenerative dental medicine, offer relevant organotypic cultures for basic research and provide testing platforms for drugs or chemical compounds. Tissue-engineered gingival equivalents comprising epithelial and connective tissues layers have been developed. 3D printing approaches have been applied for skin regeneration and the formation of vascular channels. Combining the gained knowledge from these studies may offer valuable cues on how to choose the best approach to create 3D printed patient-tailored gingival tissue to achieve the functionally and esthetically satisfying solution for the patient

Translational research on clinically failed zirconia implants

OBJECTIVES: To provide fractographic analysis of clinically fractured zirconia implants recovered with their cemented crown. To calculate bending moments, corresponding stress and crack onset location on the implant's fracture surface using a mathematical model integrating spatial coordinates of the crown-implant part and occlusal loading obtained from 2D and 3D images. METHODS: 15 fractured zirconia implants parts (11 posterior and 4 anterior) with their all- ceramic crowns still cemented on it were recovered. The implants were first generations from four manufacturers (AXIS Biodental, Z-Systems, Straumann, Swiss Dental Solutions). The time-to-failure varied between 2weeks and 9years. Fractography was performed identifying the failure origin and characteristic surface crack features. From 2D and 3D digital images of the crown-implant part, spatial coordinates anchoring the crown's occlusal contacts with the implant's central axis and reference plane were integrated in a mathematical model spreadsheet. Loads of 500 N in total were selectively distributed over identified occlusal contacts from wear patterns. The resultant bending and torsion moments, corresponding shear, tensile, maximum principal stress and von Mises stress were calculated. The fracture crack onset location on the implant's fracture surface was given by an angular position with respect to an occlusal reference and compared with the location of the fracture origin identified from fractographic analysis. RESULTS: Implants fractured from the periphery of the smaller inner diameter between two threads at the bone-entrance level except for one implant which failed half-way within the bone. The porous coating (AXIS Biodental) and the large grit alumina sandblasting (Z-System) created surface defects directly related to the fracture origin. The model spreadsheet showed how occlusal loading with respect to the implant's central axis affects bending moments and crack onset. Dominant loads distributed on contacts with important wear pattern provided a calculated crack onset location in good agreement with the fractographic findings of the fracture origin. SIGNIFICANCE: Recovered broken zirconia implant parts with their restorative crowns can provide not only information regarding the failure origin using fractography but also knowledge regarding occlusal crown loading with respect to the implant's axis. The mathematical model was helpful in showing how occlusal loading affects the location of the fracture initiation site on clinical zirconia implant fracture cases