Chamber Preparation Research Articles

An Effective Surface Passivation Assay for Single-Molecule Studies of Chromatin and Topoisomerase II.

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Eliciting Presynaptic Homeostatic Potentiation at the Drosophila Larval Neuromuscular Junction.

The Drosophila melanogaster neuromuscular junction (NMJ) is an easily accessible synapse and an excellent model for understanding synapse development, function, and plasticity. A form of plasticity called presynaptic homeostatic potentiation (PHP) operates at the NMJ and keeps synapse excitation levels stable. PHP can be induced rapidly in 10 min by application of a pharmacological antagonist of glutamate receptors (philanthotoxin-433) or chronically by deletion of the gene encoding the postsynaptic glutamate receptor subunit GluRIIA. To assess PHP, electrophysiological recordings of spontaneous miniature excitatory postsynaptic potentials and evoked excitatory postsynaptic potentials are usually performed at the NMJ of muscle 6 at abdominal segments A2 and A3. This protocol describes steps for larval dissection to access the NMJ, use of mutant lines to assess PHP, application of philanthotoxin-433 to the NMJ, and electrophysiological recordings following drug application. Collectively, these steps allow for analysis of the acute induction and expression of PHP. Recording chamber preparation, electrophysiology rig setup, larval dissection, and current clamp recording steps have been described elsewhere.

Single-molecule magnetic tweezers to probe the equilibrium dynamics of individual proteins at physiologically relevant forces and timescales.

The reversible unfolding and refolding of proteins is a regulatory mechanism of tissue elasticity and signalling used by cells to sense and adapt to extracellular and intracellular mechanical forces. However, most of these proteins exhibit low mechanical stability, posing technical challenges to the characterization of their conformational dynamics under force. Here, we detail step-by-step instructions for conducting single-protein nanomechanical experiments using ultra-stable magnetic tweezers, which enable the measurement of the equilibrium conformational dynamics of single proteins under physiologically relevant low forces applied over biologically relevant timescales. We report the basic principles determining the functioning of the magnetic tweezer instrument, review the protein design strategy and the fluid chamber preparation and detail the procedure to acquire and analyze the unfolding and refolding trajectories of individual proteins under force. This technique adds to the toolbox of single-molecule nanomechanical techniques and will be of particular interest to those interested in proteins involved in mechanosensing and mechanotransduction. The procedure takes 4 d to complete, plus an additional 6 d for protein cloning and production, requiring basic expertise in molecular biology, surface chemistry and data analysis.

Exploring the free energy landscape of proteins using magnetic tweezers.

Proteins fold to their native states by searching through the free energy landscapes. As single-domain proteins are the basic building block of multiple-domain proteins or protein complexes composed of subunits, the free energy landscapes of single-domain proteins are of critical importance to understand the folding and unfolding processes of proteins. To explore the free energy landscapes of proteins over large conformational space, the stability of native structure is perturbed by biochemical or mechanical means, and the conformational transition process is measured. In single molecular manipulation experiments, stretching force is applied to proteins, and the folding and unfolding transitions are recorded by the extension time course. Due to the broad force range and long-time stability of magnetic tweezers, the free energy landscape over large conformational space can be obtained. In this article, we describe the magnetic tweezers instrument design, protein construct design and preparation, fluid chamber preparation, common-used measuring protocols including force-ramp and force-jump measurements, and data analysis methods to construct the free energy landscape. Single-domain cold shock protein is introduced as an example to build its free energy landscape by magnetic tweezers measurements.

The impact of extended pulp chamber preparations on the clinical performance of endocrowns in Indian patients: A 1-year observational study

PurposeThe limitations of foundation restorations have been reduced with the advent of endocrowns. Despite their success, the extent of endocrowns tooth preparation into the pulp chamber has not been fully explored in the literature. The study aimed to evaluate the clinical performance of endocrowns with 5 mm extended pulp chamber tooth preparations in Indian patients over a 1- year period using modified USPHS criteria. MethodsThe observational study recruited 36 participants with definite criteria. Ceramic crown was planned for all the participants. Conventional endocrown tooth preparation was made with 5 mm pulpal extension. Clinical assessments were made at 0-, 6- and 12-months intervals using modified USPHS criteria. The data was statistically analyzed using Friedman test and Wilcoxon signed rank test. ResultsOne patient (2.8%) experienced loss of retention, marginal adaptation, and loss of anatomical form during 1-year follow up period. The majority of the participants (97.3%) reported no discomfort and high satisfaction. (P < .05) No opposing tooth wear was observed at 0- and 6-months. The results showed statistical significance in the parameters of retention, marginal adaptation, loss of anatomical form, and patient satisfaction. (P < .05). ConclusionThe clinical performance of endocrowns with extended pulp chamber preparation was satisfactory for a period of 12 months.

3D-printed lightweight dorsal skin fold chambers from PEEK reduce chamber-related animal distress

The dorsal skinfold chamber is one of the most important in vivo models for repetitive longitudinal assessment of microcirculation and inflammation. This study aimed to refine this model by introducing a new lightweight chamber made from polyetheretherketone (PEEK). Body weight, burrowing activity, distress, faecal corticosterone metabolites and the tilting angle of the chambers were analysed in mice carrying either a standard titanium chamber or a PEEK chamber. Data was obtained before chamber preparation and over a postoperative period of three weeks. In the early postoperative phase, reduced body weight and increased faecal corticosterone metabolites were found in mice with titanium chambers. Chamber tilting and tilting-related complications were reduced in mice with PEEK chambers. The distress score was significantly increased in both groups after chamber preparation, but only returned to preoperative values in mice with PEEK chambers. In summary, we have shown that light chambers reduce animal distress and may extend the maximum dorsal skinfold chamber observation time. Chambers made of PEEK are particularly suitable for this purpose: They are autoclavable, sufficiently stable to withstand rodent bites, inexpensive, and widely available through 3D printing.

Connexin Hemichannel Activation by S-Nitrosoglutathione Synergizes Strongly with Photodynamic Therapy Potentiating Anti-Tumor Bystander Killing.

Simple SummaryBystander effects depend on direct cell-cell communication and/or paracrine signaling mediated by the release of soluble factors into the extracellular environment and may greatly influence therapy outcome. Although the limited data available suggest a role for intercellular gap junction channels, far less is known about the role of connexin hemichannels. Here, we investigated bystander effects induced by photodynamic therapy in syngeneic murine melanoma models in vivo. We determined that (i) photoactivation of a photosensitizer triggered calcium-dependent cell death pathways in both irradiated and bystander tumor cells; (ii) hemichannel activity and adenosine triphosphate release were key factors for the induction of bystander cell death; and (iii) bystander cell killing and antitumor response elicited by photodynamic therapy were greatly enhanced by combination treatment with S-nitrosoglutathione, which promoted hemichannel opening in these experimental conditions. Therefore, these findings in a preclinical model have important translational potential.In this study, we used B16-F10 cells grown in the dorsal skinfold chamber (DSC) preparation that allowed us to gain optical access to the processes triggered by photodynamic therapy (PDT). Partial irradiation of a photosensitized melanoma triggered cell death in non-irradiated tumor cells. Multiphoton intravital microscopy with genetically encoded fluorescence indicators revealed that bystander cell death was mediated by paracrine signaling due to adenosine triphosphate (ATP) release from connexin (Cx) hemichannels (HCs). Intercellular calcium (Ca2+) waves propagated from irradiated to bystander cells promoting intracellular Ca2+ transfer from the endoplasmic reticulum (ER) to mitochondria and rapid activation of apoptotic pathways. Combination treatment with S-nitrosoglutathione (GSNO), an endogenous nitric oxide (NO) donor that biases HCs towards the open state, greatly potentiated anti-tumor bystander killing via enhanced Ca2+ signaling, leading to a significant reduction of post-irradiation tumor mass. Our results demonstrate that HCs can be exploited to dramatically increase cytotoxic bystander effects and reveal a previously unappreciated role for HCs in tumor eradication promoted by PDT.

Lithium Disilicate Ceramic Endocrown Biomechanical Response According to Different Pulp Chamber Extension Angles and Filling Materials.

The purpose of this study is to evaluate the effect of pulp chamber extension angles and filling material mechanical properties on the biomechanical response of a ceramic endocrown. A 3D model of maxillary molar that underwent endodontically treatment was exported to computer aided design software to conduct finite element analysis (FEA). The endocrown model was modified considering different pulp chamber extension angles (right angle; 6°, 12° and 18° of axial divergence). The solids were imported into the computer aided engineering software in Standard for the Exchange of Product Data (STEP) format. Nine different filling materials were simulated to seal the orifice of the root canal system under each endocrown restoration (resin composite, bulk-fill resin composite, alkasite, flowable resin composite, glass ionomer cement, autocured resin-reinforced glass ionomer cement, resin cement, bulk-fill flowable resin composite, zinc oxide cement), totaling 36 models. An axial load (300 N) was applied at the occlusal surface. Results were determined by colorimetric graphs of von-Misses stress (VMS) and Maximum Principal Stress (MPS) on tooth, cement layer, and endocrown restorations. VMS distribution showed a similar pattern between the models, with more stress at the load region for the right-angled endocrowns. The MPS showed that the endocrown intaglio surface and cement layer showed different mechanical responses with different filing materials and pulp chamber angles. The stress peaks plotted in the dispersion plot showed that the filling material stiffness is proportional to the stress magnitude in the endocrown, cement layer and tooth adhesive surface. In addition, the higher the pulp chamber preparation angle, the higher the stress peak in the restoration and tooth, and the lower the stress in the cement layer. Therefore, 6° and 12° pulp chamber angles showed more promising balance between the stresses of the adhesive interface structures. Under the conditions of this study, rigid filling materials were avoided to seal the orifice of root canal system when an endocrown restoration was planned as rehabilitation. In addition, the pulp chamber axial walls were prepared between 6° and 12° of divergence to balance the stress magnitude in the adhesive interface for this treatment modality.

Intravital microscopy of dynamic single-cell behavior in mouse mammary tissue.

Multiphoton intravital imaging is essential for understanding cellular behavior and function in vivo. The adipose-rich environment of the mammary gland poses a unique challenge to in vivo microscopy due to light scattering that impedes high-resolution imaging. Here we provide a protocol for high-quality, six-color 3D intravital imaging of regions across the entire mouse mammary gland and associated tissues for several hours while maintaining tissue access for microdissection and labeling. An incision at the ventral midline and along the right hind leg creates a skin flap that is then secured to a raised platform skin side down. This allows for fluorescence-guided microdissection of connective tissue to provide unimpeded imaging of mammary ducts. A sealed imaging chamber over the skin flap creates a stable environment while maintaining access to large tissue regions for imaging with an upright microscope. We provide a strategy for imaging single cells and the tissue microenvironment utilizing multicolor Confetti lineage-tracing and additional dyes using custom-designed filters and sequential excitation with dual multiphoton lasers. Furthermore, we describe a strategy for simultaneous imaging and photomanipulation of single cells using the Olympus SIM scanner and provide steps for 3D video processing, visualization and high-dimensional analysis of single-cell behavior. We then provide steps for multiplexing intravital imaging with fixation, immunostaining, tissue clearing and 3D confocal imaging to associate cell behavior with protein expression. The skin-flap surgery and chamber preparation take 1.5 h, followed by up to 12 h of imaging. Applications range from basic filming in 1 d to 5 d for multiplexing and complex analysis.

Neotropical Melyroidea group cockroaches reveal various degrees of (eu)sociality.

Eusociality in its various degrees represents an animal social system characterised by cooperative brood care, differentiation into castes and generational overlap. The fossil record indicates that eusociality is likely to have originated in hymenopterans and blattodeans during the Cretaceous. In this study, we present findings from surveys in Peruvian (Villa Carmen) and Ecuadorian (Rio Bigal, El Reventador) cloud forests revealing the first extant cockroach species living in complex, structured groups (n = 90-200 individuals, ˃20 adults). We observed and described behaviours that suggest the existence of cooperative care, nest guarding, nest chamber preparation within hardwood Casearia sp. (Salicaceae) and bamboo (Bambusoideae), multiple overlapping generations ('different stages of' instars), colony translocation, possibly a sole reproductive female (1.25 times larger white 'queen', but no potential 'king' observed), and morphologically diversified immature stages. In order to define the lineage where this type of sociality originated and occurs, the forms of Melyroidea magnifica Shelford, 1912, M. ecuadoriana sp. n., M. mimetica Shelford, 1912 and an undescribed species from Peru are also described in a separate section of this study. Blattoid morphological characteristics such as typical styli suggest categorisation within distinct Oulopterygidae (Rehn, 1951), outside Corydiidae Saussure 1864. Transitional advanced sociality or semisociality in related Aclavoidea socialis gen. et sp. n. is documented in a rotting stump (n = 80 individuals, few adults). Close phylogenetic relation between the genera, conserved morphology of numerous characters and their diverse feeding strategies generally lacking specialisation suggests a rather recent origin of a social way of life in this group. Eusociality in invertebrates and vertebrates can thus originate in various phylogenetical and ecological trajectories including predation, parasitism, care for herbs and the new one, documented through diet shift from detritivory to fungivory and algaevory. Interdisciplinary approaches reveal the low degree of knowledge of rainforest ecosystems, with fundamental groups remaining still systematically and also behaviourally undescribed.

Ratiometric Single-Molecule FRET Measurements to Probe Conformational Subpopulations of Intrinsically Disordered Proteins.

Over the past few decades, numerous examples have demonstrated that intrinsic disorder in proteins lies at the heart of many vital processes, including transcriptional regulation, stress response, cellular signaling, and most recently protein liquid‐liquid phase separation. The so‐called intrinsically disordered proteins (IDPs) involved in these processes have presented a challenge to the classic protein “structure‐function paradigm,” as their functions do not necessarily involve well‐defined structures. Understanding the mechanisms of IDP function is likewise challenging because traditional structure determination methods often fail with such proteins or provide little information about the diverse array of structures that can be related to different functions of a single IDP. Single‐molecule fluorescence methods can overcome this ensemble‐average masking, allowing the resolution of subpopulations and dynamics and thus providing invaluable insights into IDPs and their function. In this protocol, we describe a ratiometric single‐molecule Förster resonance energy transfer (smFRET) routine that permits the investigation of IDP conformational subpopulations and dynamics. We note that this is a basic protocol, and we provide brief information and references for more complex analysis schemes available for in‐depth characterization. This protocol covers optical setup preparation and protein handling and provides insights into experimental design and outcomes, together with background information about theory and a brief discussion of troubleshooting. © 2020 by John Wiley & Sons, Inc. Basic Protocol: Ratiometric smFRET detection and analysis of IDPs Support Protocol 1: Fluorophore labeling of a protein through maleimide chemistry Support Protocol 2: Sample chamber preparation Support Protocol 3: Determination of direct excitation of acceptor by donor excitation and leakage of donor emission to acceptor emission channel

Endodontic treatment of maxillary second molar with two roots: A case report

&amp;lt;p&amp;gt;Introduction. The success of endodontic treatment depends on the adequate biomechanical preparation and obturation of the entire root canal system. The untreated or poorly debrided/obturated root canals are among the most frequent causes of endodontic treatment failure. The endodontic treatment of maxillary molar has the highest failure rates due to the complexity of their root canal anatomy. The aim of the study is to present the endodontic treatment of maxillary second molar with two roots and two canals. Case report. The clinical case report presents the endodontic treatment of maxillary second molar with two roots and two canals in a patient, aged 32 years. After the analysis of preoperative diagnostic radiograph, opening the pulp chamber and access cavity preparation, two canal orifices were localized. After the biomechanical preparation, root canals were obturated with the Gutta-percha points and sealer during the same visit. Conclusion. Although maxillary second molar most commonly varies in terms of the number of root canals in the buccal roots, less common variations in anatomical and morphological traits, such as the presence of two roots and two canals, should be considered during endodontic treatment.&amp;lt;/p&amp;gt;

Design of Suitable Aperture Shape for Optical Measurement Methods in Experimental Chamber

This article deals with the experimental chamber preparation, on which shockwaves, besides other measurements, will be observed using the Schlieren optic method. Because of this reason, the replaceable aperture will be used, which will be placed against the center of the observation window. The design of the optimal aperture shape was solved using mathematical – physics analysis in the Ansys Fluent system and the results will be compared with the results obtained from the experimental measurements.

Correction of the Supersonic Flow Impact on the Results of the Inserted Temperature Probe in the Differentially Pumped Chamber

This article deals with the experimental chamber preparation, on which stagnation and static temperature (besides others) will be measured. The results obtained from measurement will be compared with the results from mathematical – physics analysis of Ansys Fluent system and in this system, the temperature running and expected ratios for planned experimental measuring also will be mapped in the supersonic flow in the experimental chambers.

Dissociation between macro- and microvascular parameters in the early phase of hemorrhagic shock

Hemorrhagic shock (HS) therapy is based on macrohemodynamic improvement, but it is not clear if this therapy correlates directly with increases in tissue perfusion. Aiming to clarify this point, we compared norepinephrine (NE, a vasoconstrictor), sodium nitroprusside (NP, a vasodilator) and levosimendan (LEV, an inodilator) treatments on macro and microvascular parameters using the hamster dorsal skinfold chamber preparation. One hour after HS, animals received Ringer's lactate (RL) solution within 10 min, then animals received RL, NP, NE and LEV during 90 min via jugular vein. Macrovascular variables: mean arterial pressure (MAP), heart rate (HR), maximal ventricle pressure (MVP), change in ventricular pressure over time (dP/dt) and microvascular variables: arteriolar and venular diameters, functional capillary density (FCD) and red blood cell velocity (RBCV) were evaluated at baseline, 60 min after HS, 60 and 90 min after treatments. Lactate blood concentrations were evaluated at baseline, 60 min after HS and 90 min after treatments. Hematocrit (Hct), cardiac output (CO), stroke volume (SV) and number of rolling and adhered leukocytes were assessed at 90 min after treatments. Data were considered significant when p < 0.05. NE increased significantly all macrohemodynamic variables compared to baseline (except MAP), and it was the only treatment that increased Hct, CO and SV significantly. NE decreased significantly all microvascular variables in comparison to baseline. NP increased HR, FCD and RBCV and reduced MVP and dP/dt significantly. LEV decreased MVP and dP/dt, arteriolar diameter and FCD and augmented RBCV significantly in comparison to baseline. Blood concentration of lactate increased significantly 60 min after HS. Leukocyte rolling and adhesion were not different between groups. We concluded that, early, during hemorrhagic shock, norepinephrine associated to fluid therapy improved macrohemodynamic parameters but failed to improved microvascular flow. Conversely, sodium nitroprusside association had the opposite effect. Despite its inodilator properties, levosimendan did not improve macro or microhemodynamic parameters when combined to fluid therapy.

Modulation of Exposure to Static Magnetic Field Affects Targeted Therapy of Solid Tumors In Vivo.

Static magnetic fields (SMF) exhibit antitumoral activity and enhance the efficacy of chemotherapy by opening the tumor-blood barrier. This study aimed to analyze different SMF-exposure protocols on epidermal growth factor receptor (EGFR)-overexpressing tumors, as well as their combination with cetuximab. Experiments were performed in skinfold chamber preparations of C57Bl/6-and CD-1nu/nu mice bearing LLC-1 tumors. Animals were exposed to 587 mT magnetic field following different exposure protocols. A subgroup received additional cetuximab injections. Using in vivo-fluorescence microscopy and planimetry, tumor angiogenesis, growth and microcirculation were repeatedly analyzed for 13 days. In contrast to daily short SMF exposure, three-fold SMF exposure for 2 h led to a significant 46% reduction of tumor growth. Adding cetuximab to SMF exposure did not yield any benefit, although cetuximab monotherapy was highly effective (53% reduction of tumor growth), indicating a potential interference of SMF and EGFR signaling. No effects on microcirculation, angiogenesis or leukocyte-endothelium interactions were documented. The use of SMF is promising in the treatment of solid tumors; however, it appears to interfere with EGFR-targeted therapy.

In vivo functional and morphological characterization of bone and striated muscle microcirculation in NSG mice.

Organ-specific microcirculation plays a central role in tumor growth, tumor cell homing, tissue engineering, and wound healing. Mouse models are widely used to study these processes; however, these mouse strains often possess unique microhemodynamic parameters, making it difficult to directly compare experiments. The full functional characterization of bone and striated muscle microcirculatory parameters in non-obese diabetic-severe combined immunodeficiency/y-chain; NOD-Prkds IL2rg (NSG) mice has not yet been reported. Here, we established either a dorsal skinfold chamber or femur window in NSG mice (n = 23), allowing direct analysis of microcirculatory parameters in vivo by intravital fluorescence microscopy at 7, 14, 21, and 28 days after chamber preparation. Organ-specific differences were observed. Bone had a significantly lower vessel density but a higher vessel diameter than striated muscle. Bone also showed higher effective vascular permeability than striated muscle. The centerline velocity values were similar in the femur window and dorsal skinfold chamber, with a higher volumetric blood flow in bone. Interestingly, bone and striated muscle showed similar tissue perfusion rates. Knowledge of physiological microhemodynamic values of bone and striated muscle in NSG mice makes it possible to analyze pathophysiological processes at these anatomic sites, such as tumor growth, tumor metastasis, and tumor microcirculation, as well as the response to therapeutic agents.

A hyperspectral vessel image registration method for blood oxygenation mapping.

Blood oxygenation mapping by the means of optical oximetry is of significant importance in clinical trials. This paper uses hyperspectral imaging technology to obtain in vivo images for blood oxygenation detection. The experiment involves dorsal skin fold window chamber preparation which was built on adult (8–10 weeks of age) female BALB/c nu/nu mice and in vivo image acquisition which was performed by hyperspectral imaging system. To get the accurate spatial and spectral information of targets, an automatic registration scheme is proposed. An adaptive feature detection method which combines the local threshold method and the level-set filter is presented to extract target vessels. A reliable feature matching algorithm with the correlative information inherent in hyperspectral images is used to kick out the outliers. Then, the registration images are used for blood oxygenation mapping. Registration evaluation results show that most of the false matches are removed and the smooth and concentrated spectra are obtained. This intensity invariant feature detection with outliers-removing feature matching proves to be effective in hyperspectral vessel image registration. Therefore, in vivo hyperspectral imaging system by the assistance of the proposed registration scheme provides a technique for blood oxygenation research.

A portable somatic cell counter based on a multi-functional counting chamber and a miniaturized fluorescence microscope

A somatic cell count is the concentration or density of somatic cells in milk, and is an important indicator for monitoring mastitis incidence and milk quality in the dairy industry. Managing and controlling mastitis based on somatic cell counts can help ensure high milk quality and yield. A major challenge when translating existing cell counting methods to such application is that they require off-chip sample preparation and complicated sample and reagent delivery steps that cannot be easily performed in resource-limited settings such as dairy farms. Here, we describe an integrated cell counting platform that enables automatic sample delivery into a cell counting chamber and on-chip sample preparation without requiring any off-chip processes, and that simultaneously provides a miniaturized, hand-held fluorescence device for the identification of fluorescently-labelled somatic cells. Our platform thus allows simple, rapid and accurate enumeration of somatic cells in milk. We successfully demonstrated its capability of counting somatic cells in milk, which can be easily performed even by non-experts without additional instrumentation. The platform represents a promising tool for everyday milk-quality tracking and for controlling mastitis occurrence.

Ivabradine Attenuates the Microcirculatory Derangements Evoked by Experimental Sepsis.

Experimental data suggest that ivabradine, an inhibitor of the pacemaker current in sinoatrial node, exerts beneficial effects on endothelial cell function, but it is unclear if this drug could prevent microcirculatory dysfunction in septic subjects, improving tissue perfusion and reducing organ failure. Therefore, this study was designed to characterize the microcirculatory effects of ivabradine on a murine model of abdominal sepsis using intravital videomicroscopy. Twenty-eight golden Syrian hamsters were allocated in four groups: sham-operated animals, nontreated septic animals, septic animals treated with saline, and septic animals treated with ivabradine (2.0 mg/kg intravenous bolus + 0.5 mg · kg · h). The primary endpoint was the effect of ivabradine on the microcirculation of skinfold chamber preparations, assessed by changes in microvascular reactivity and rheologic variables, and the secondary endpoint was its effects on organ function, evaluated by differences in arterial blood pressure, motor activity score, arterial blood gases, and hematologic and biochemical parameters among groups. Compared with septic animals treated with saline, those treated with ivabradine had greater functional capillary density (90 ± 4% of baseline values vs. 71 ± 16%; P < 0.001), erythrocyte velocity in capillaries (87 ± 11% of baseline values vs. 62 ± 14%; P < 0.001), and arteriolar diameter (99 ± 4% of baseline values vs. 91 ± 7%; P = 0.041) at the end of the experiment. Besides that, ivabradine-treated animals had less renal, hepatic, and neurologic dysfunction. Ivabradine was effective in reducing microvascular derangements evoked by experimental sepsis, which was accompanied by less organ dysfunction. These results suggest that ivabradine yields beneficial effects on the microcirculation of septic animals.