Electrophysiological Effects of the Sodium-Glucose Co-Transporter-2 (SGLT2) Inhibitor Dapagliflozin on Human Cardiac Potassium Channels.

The sodium-glucose co-transporter-2 (SGLT2) inhibitor dapagliflozin is increasingly used in the treatment of diabetes and heart failure. Dapagliflozin has been associated with reduced incidence of atrial fibrillation (AF) in clinical trials. We hypothesized that the favorable antiarrhythmic outcome of dapagliflozin use may be caused in part by previously unrecognized effects on atrial repolarizing potassium (K+) channels. This study was designed to assess direct pharmacological effects of dapagliflozin on cloned ion channels Kv11.1, Kv1.5, Kv4.3, Kir2.1, K2P2.1, K2P3.1, and K2P17.1, contributing to IKur, Ito, IKr, IK1, and IK2P K+ currents. Human channels coded by KCNH2, KCNA5, KCND3, KCNJ2, KCNK2, KCNK3, and KCNK17 were heterologously expressed in Xenopus laevis oocytes, and currents were recorded using the voltage clamp technique. Dapagliflozin (100 µM) reduced Kv11.1 and Kv1.5 currents, whereas Kir2.1, K2P2.1, and K2P17.1 currents were enhanced. The drug did not significantly affect peak current amplitudes of Kv4.3 or K2P3.1 K+ channels. Biophysical characterization did not reveal significant effects of dapagliflozin on current-voltage relationships of study channels. In conclusion, dapagliflozin exhibits direct functional interactions with human atrial K+ channels underlying IKur, IKr, IK1, and IK2P currents. Substantial activation of K2P2.1 and K2P17.1 currents could contribute to the beneficial antiarrhythmic outcome associated with the drug. Indirect or chronic effects remain to be investigated in vivo.

Differential Effects of the Betablockers Carvedilol, Metoprolol and Bisoprolol on Cardiac Kv4.3 (Ito) Channel Isoforms.

Cardiac Kv4.3 channels contribute to the transient outward K+ current, Ito, during early repolarization of the cardiac action potential. Two different isoforms of Kv4.3 are present in the human ventricle and exhibit differential remodeling in heart failure (HF). Cardioselective betablockers are a cornerstone of HF with reduced ejection fraction therapy as well as ventricular arrhythmia treatment. In this study we examined pharmacological effects of betablockers on both Kv4.3 isoforms to explore their potential for isoform-specific therapy. Kv4.3 isoforms were expressed in Xenopus laevis oocytes and incubated with the respective betablockers. Dose-dependency and biophysical characteristics were examined. HEK 293T-cells were transfected with the two Kv4.3 isoforms and analyzed with Western blots. Carvedilol (100 µM) blocked Kv4.3 L by 77 ± 2% and Kv4.3 S by 67 ± 6%, respectively. Metoprolol (100 µM) was less effective with inhibition of 37 ± 3% (Kv4.3 L) and 35 ± 4% (Kv4.3 S). Bisoprolol showed no inhibitory effect. Current reduction was not caused by changes in Kv4.3 protein expression. Carvedilol inhibited Kv4.3 channels at physiologically relevant concentrations, affecting both isoforms. Metoprolol showed a weaker blocking effect and bisoprolol did not exert an effect on Kv4.3. Blockade of repolarizing Kv4.3 channels by carvedilol and metoprolol extend their pharmacological mechanism of action, potentially contributing beneficial antiarrhythmic effects in normal and failing hearts.

The rise of animal biotelemetry and genetics research data integration.

The advancement and availability of innovative animal biotelemetry and genomic technologies are improving our understanding of how the movements of individuals influence gene flow within and between populations and ultimately drive evolutionary and ecological processes. There is a growing body of work that is integrating what were once disparate fields of biology, and here, we reviewed the published literature up until January 2023 (139 papers) to better understand the drivers of this research and how it is improving our knowledge of animal biology. The review showed that the predominant drivers for this research were as follows: (1) understanding how individual-based movements affect animal populations, (2) analyzing the relationship between genetic relatedness and social structuring, and (3) studying how the landscape affects the flow of genes, and how this is impacted by environmental change. However, there was a divergence between taxa as to the most prevalent research aim and the methodologies applied. We also found that after 2010 there was an increase in studies that integrated the two data types using innovative statistical techniques instead of analyzing the data independently using traditional statistics from the respective fields. This new approach greatly improved our understanding of the link between the individual, the population, and the environment and is being used to better conserve and manage species. We discuss the challenges and limitations, as well as the potential for growth and diversification of this research approach. The paper provides a guide for researchers who wish to consider applying these disparate disciplines and advance the field.

Ability of a Blue Hemoglobin-Based Liquid as a Novel Technology to Stain Initial Enamel Demineralization: A Proof-of-Concept in vitro Study.

During orthodontic treatment, enamel demineralization can occur. Its early detection is the basis for efficient preventive measures to arrest or remineralize lesions. In the present study, the application of a novel blue hemoglobin-based liquid (BlueCheck) was evaluated as proof of concept for detection of artificially demineralized smooth surfaces. 60 samples from extracted human posterior teeth were randomly assigned to four groups (15 per group). In 30 of these samples (groups A and B), superficial enamel was removed to create a ground surface. On the surface of other 30 samples (group C and D), orthodontic metal brackets were bonded. On each surface, BC liquid was applied and rinsed with water after 3 min (baseline). All surfaces were checked by two independent observers for presence of blue areas. On each sample, one side was covered by nail varnish to protect this enamel part from demineralization. The samples were demineralized with lactic acid (pH 4.6) for 7 days (group A and C) and 14 days (group B and D), respectively. Mineral loss was determined using quantitative light-induced fluorescence after demineralization. BlueCheck dye was again applied on the samples and evaluated for presence of stained areas. Histological sections were prepared from randomly selected samples and lesion depth was measured. Kruskal-Wallis test was used for group comparison (α = 0.05). After demineralization, median ΔF value for all samples was -8.25% indicating the presence of an initial demineralization. The difference of ΔF values was not statistically significant between samples at 7 or 14 days of demineralization, nor for samples with and without orthodontic brackets (p = 0.13). At baseline, none of the sample surfaces showed discoloration, whereas a distinctive blue color was visible after demineralization in all samples exposed to acid-exposed areas, corresponding to 100% sensitivity. The internal control surfaces (without demineralization) did not show any staining, corresponding to 100% specificity. Histologically measured lesion depths ranged between 200 and 254 µm. In this in vitro study, staining of demineralized enamel surface areas were shown to be reliable. Based on our results, this easily applicable product seems useful to be an adjuvant method to clinical examination to monitor oral health during an orthodontic treatment on tooth surfaces after removal of dental biofilm.

Differential regulation of KCa 2.1 (KCNN1) K+ channel expression by histone deacetylases in atrial fibrillation with concomitant heart failure.

Atrial fibrillation (AF) with concomitant heart failure (HF) poses a significant therapeutic challenge. Mechanism-based approaches may optimize AF therapy. Small-conductance, calcium-activated K+ (KCa , KCNN) channels contribute to cardiac action potential repolarization. KCNN1 exhibits predominant atrial expression and is downregulated in chronic AF patients with preserved cardiac function. Epigenetic regulation is suggested by AF suppression following histone deacetylase (HDAC) inhibition. We hypothesized that HDAC-dependent KCNN1 remodeling contributes to arrhythmogenesis in AF complicated by HF. The aim of this study was to assess KCNN1 and HDAC1-7 and 9 transcript levels in AF/HF patients and in a pig model of atrial tachypacing-induced AF with reduced left ventricular function. In HL-1 atrial myocytes, tachypacing and anti-Hdac siRNAs were employed to investigate effects on Kcnn1 mRNA levels. KCNN1 expression displayed side-specific remodeling in AF/HF patients with upregulation in left and suppression in right atrium. In pigs, KCNN1 remodeling showed intermediate phenotypes. HDAC levels were differentially altered in humans and pigs, reflecting highly variable epigenetic regulation. Tachypacing recapitulated downregulation of Hdacs 1, 3, 4, 6, and 7 with a tendency towards reduced Kcnn1 levels in vitro, indicating that atrial high rates induce remodeling. Finally, Kcnn1 expression was decreased by knockdown of Hdacs 2, 3, 6, and 7 and enhanced by genetic Hdac9 inactivation, while anti-Hdac 1, 4, and 5 siRNAs did not affect Kcnn1 transcript levels. In conclusion, KCNN1 and HDAC expression is differentially remodeled in AF complicated by HF. Direct regulation of KCNN1 by HDACs in atrial myocytes provides a basis for mechanism-based antiarrhythmic therapy.

Trigger-Specific Remodeling of KCa2 Potassium Channels in Models of Atrial Fibrillation.

AIM: Effective antiarrhythmic treatment of atrial fibrillation (AF) constitutes a major challenge, in particular, when concomitant heart failure (HF) is present. HF-associated atrial arrhythmogenesis is distinctly characterized by prolonged atrial refractoriness. Small-conductance, calcium-activated K+ (KCa, SK, KCNN) channels contribute to cardiac action potential repolarization and are implicated in AF susceptibility and therapy. The mechanistic impact of AF/HF-related triggers on atrial KCa channels is not known. We hypothesized that tachycardia, stretch, ß-adrenergic stimulation, and hypoxia differentially determine KCa2.1-2.3 channel remodeling in atrial cells. METHODS: KCNN1-3 transcript levels were assessed in AF/HF patients and in a pig model of atrial tachypacing-induced AF with reduced left ventricular function. HL-1 atrial myocytes were subjected to proarrhythmic triggers to investigate the effects on Kcnn mRNA and KCa channel protein. RESULTS: Atrial KCNN1-3 expression was reduced in AF/HF patients. KCNN2 and KCNN3 suppression was recapitulated in the corresponding pig model. In contrast to human AF, KCNN1 remained unchanged in pigs. Channel- and stressor-specific remodeling was revealed in vitro. Lower expression levels of KCNN1/KCa2.1 were linked to stretch and ß-adrenergic stimulation. Furthermore, KCNN3/KCa2.3 expression was suppressed upon tachypacing and hypoxia. Finally, KCNN2/KCa2.2 abundance was specifically enhanced by hypoxia. CONCLUSION: Reduction of KCa2.1-2.3 channel expression might contribute to the action potential prolongation in AF complicated by HF. Subtype-specific KCa2 channel remodeling induced by tachypacing, stretch, ß-adrenergic stimulation, or hypoxia is expected to differentially determine atrial remodeling, depending on patient-specific activation of each triggering factor. Stressor-dependent KCa2 regulation in atrial myocytes provides a starting point for mechanism-based antiarrhythmic therapy.

HDAC2-dependent remodeling of KCa2.2 (KCNN2) and KCa2.3 (KCNN3) K+ channels in atrial fibrillation with concomitant heart failure.

AIMS: Atrial fibrillation (AF) with concomitant heart failure (HF) is associated with prolonged atrial refractoriness. Small-conductance, calcium-activated K+ (KCa, KCNN) channels promote action potential (AP) repolarization. KCNN2 and KCNN3 variants are associated with AF risk. In addition, histone deacetylase (HDAC)-related epigenetic mechanisms have been implicated in AP regulation. We hypothesized that HDAC2-dependent remodeling of KCNN2 and KCNN3 expression contributes to atrial arrhythmogenesis in AF complicated by HF. The objectives were to assess HDAC2 and KCNN2/3 transcript levels in AF/HF patients and in a pig model, and to investigate cellular epigenetic effects of HDAC2 inactivation on KCNN expression. MATERIALS AND METHODS: HDAC2 and KCNN2/3 transcript levels were quantified in patients with AF and HF, and in a porcine model of atrial tachypacing-induced AF and reduced left ventricular function. Tachypacing and anti-Hdac2 siRNA treatment were employed in HL-1 atrial myocytes to study effects on KCNN2/3 mRNA and KCa protein abundance. KEY FINDINGS: Atrial KCNN2 and KCNN3 expression was reduced in AF/HF patients and in a corresponding pig model. HDAC2 displayed significant downregulation in humans and a tendency towards reduced expression in right atrial tissue of pigs. Tachypacing recapitulated downregulation of Kcnn2/KCa2.2, Kcnn3/KCa2.3 and Hdac2/HDAC2, indicating that high atrial rates trigger epigenetic remodeling mechanisms. Finally, knock-down of Hdac2 in vitro reduced Kcnn3/KCa2.3 expression. SIGNIFICANCE: KCNN2/3 and HDAC2 expression is suppressed in AF complicated by HF. Hdac2 directly regulates Kcnn3 mRNA levels in atrial cells. The mechanistic and therapeutic significance of epigenetic electrophysiological effects in AF requires further validation.

Inhibition of cardiac Kv4.3 (Ito) channel isoforms by class I antiarrhythmic drugs lidocaine and mexiletine.

Transient outward K+ current, Ito, contributes to cardiac action potential generation and is primarily carried by Kv4.3 (KCND3) channels. Two Kv4.3 isoforms are expressed in human ventricle and show differential remodeling in heart failure (HF). Lidocaine and mexiletine may be applied in selected patients to suppress ventricular arrhythmias, without effects on sudden cardiac death or mortality. Isoform-dependent effects of antiarrhythmic drugs on Kv4.3 channels and potential implications for remodeling-based antiarrhythmic management have not been assessed to date. We sought to test the hypotheses that Kv4.3 channels are targeted by lidocaine and mexiletine, and that drug sensitivity is determined in isoform-specific manner. Expression of KCND3 isoforms was quantified using qRT-PCR in left ventricular samples of patients with HF due to either ischemic or dilated cardiomyopathies (ICM or DCM). Long (Kv4.3-L) and short (Kv4.3-S) isoforms were heterologously expressed in Xenopus laevis oocytes to study drug sensitivity and effects on biophysical characteristics activation, deactivation, inactivation, and recovery from inactivation. In the present HF patient cohort KCND3 isoform expression did not differ between ICM and DCM. In vitro, lidocaine (IC50-Kv4.3-L: 0.8 mM; IC50-Kv4.3-S: 1.2 mM) and mexiletine (IC50-Kv4.3-L: 146 µM; IC50-Kv4.3-S: 160 µM) inhibited Kv4.3 with different sensitivity. Biophysical analyses identified accelerated and enhanced inactivation combined with delayed recovery from inactivation as primary biophysical mechanisms underlying Kv4.3 current reduction. In conclusion, differential effects on Kv4.3 isoforms extend the electropharmacological profile of lidocaine and mexiletine. Patient-specific remodeling of Kv4.3 isoforms may determine individual drug responses and requires consideration during clinical application of compounds targeting Kv4.3.

Optimal light conditions for Daphnia filtration.

Daphnia populations are present in lakes and ponds. They are known to experience diurnal vertical migrations according to their feeding needs. During the day they migrate downwards to avoid predation in light-receiving layers and at night they migrate upwards, searching for food in the shallow productive layers. The light photoperiod and light intensity vary depending on the latitude and, therefore, the precise location of lakes and ponds will be an additional and crucial parameter in determining the development of Daphnia. Here we will focus on a population of Daphnia magna (a genus of the Cladocera order). The effect of both light intensity and photoperiod on Daphnia filtration was studied in laboratory experiments. An increase in the light intensity resulted in two D. magna responses depending on the exposure time of individuals to light. Short time exposures to a decrease in the light intensity of less than one day produced an increase in the D. magna filtration. However, exposures of longer than one day resulted in a decrease in the D. magna filtration along with a decrease in the light intensity. Photoperiod exposures of 8, 12 and 16 h produced greater D. magna filtrations than photoperiods of 0, 4 and 24 h. In this study, regulation of the light intensity and the period of exposure were used in laboratory experiments to establish D. magna development thresholds by latitudinal variation in the photoperiod.

Mediated food and hydrodynamics on the ingestion of microplastics by Daphnia magna.

There is consensus on the need to study the potential impact microplastics (MP) have on freshwater planktonic organisms. It is not yet fully understood how MP enter the aquatic food web or the effect they have on all the trophic levels. As a result of the potential for MP to accumulate throughout food webs, there is increasing interest in evaluating their fate in a variety of environmental conditions. This study investigated the variability in the ingestion of MP to food ratios and the exposed time of MP to Daphnia magna in non-sheared and sheared conditions. The sheared environment provided Daphnia magna with the conditions for optimal filtering capacity. Regardless of the ratios of MP concentration to food concentration (MP:Food), the filtration capacity of the Daphnia magna was enhanced in the sheared experiments. In both the sheared and non-sheared experiments, filtration capacity decreased when the ratios of MP to food concentration and the exposure times to MP were increased. Mortality was mainly enhanced in the non-sheared conditions at higher MP concentrations and exposure times to MP. No mortality was found in the sheared conditions for the exposure times studied. Therefore, in aquatic systems that undergo constant low sheared conditions, Daphnia magna can survive longer when exposed to MP than in calm conditions, provided food concentrations do not limit their capacity to filter.

Functional responses of Daphnia magna to zero-mean flow turbulence.

Daphnia are important to understanding the biogeochemistry of aquatic ecosystems, mainly because of their ability to filter bacteria, algae and inorganic particles as well. Although there are many studies on the general effects that biotic and abiotic stressors, increased temperature and hypoxia, salinity, metals, pharmaceuticals, pesticides, etc., have on Daphnia populations, little is known about the impact elevated turbulence has. Here, we show that turbulence affects Daphnia magna survival, swimming behaviour and filtering capacity. Our data demonstrate that altering their habitat by induced mixing from turbulence, induces an increased filtering capacity of the Daphnia magna individuals, provided the level of background turbulence (defined by the dissipation of turbulent kinetic energy) is lower than ε = 0.04 cm2 s-3. The filtering capacity reduced exponentially with increasing ε, and at ε > 1 cm2 s-3 both mobility and filtration were suppressed and eventually led to the death of all the Daphnia magna individuals.

Temperature-driven response reversibility and short-term quasi-acclimation of Daphnia magna.

Analysing the effect water temperature has on Daphnia magna is essential in anticipating the impact climate change will have on this freshwater zooplanktonic keystone species. While many authors have followed this line of research, few have covered an extensive temperature range or complex temperature change scenarios. Global warming is mostly associated with increased extreme temperature events, such as heat waves, as well as earlier and more intense thermal stratification. Both of these events may directly influence D. magna fitness, especially in those populations performing diel vertical migration (DVM). We analysed the effect water temperatures, ranging from 11 to 29°C, have on the filtration capacity (FC) of D. magna, to anticipate the effects of acclimation, temperature change rate (TCR) and potential reversibility of responses to such conditions. Results show that sudden temperature changes have an immediate negative impact on the FC of D. magna and is more severe at higher temperatures and higher TCRs. However, D. magna individuals have shown themselves to be capable of quasi-acclimating to temperatures ranging from 11 to 25°C in around a week and achieving much higher FCs; albeit never reaching the optimal FC achieved at 20°C. That said, 29°C is lethal for D. magna individuals within approximately five days. Finally, non-optimal temperature acclimated individuals can recover maximal FC within 2-4 days of the optimal long-term acclimation temperature (20°C) being re-established, thus proving temperature responses to be reversible.

Cardiovascular pharmacology of K2P17.1 (TASK-4, TALK-2) two-pore-domain K+ channels.

K2P17.1 (TASK-4, TALK-2) potassium channels are expressed in the heart and represent potential targets for pharmacological management of atrial and ventricular arrhythmias. Reduced K2P17.1 expression was found in atria and ventricles of heart failure (HF) patients. Modulation of K2P17.1 currents by antiarrhythmic compounds has not been comprehensively studied to date. The objective of this study was to investigate acute effects of clinically relevant antiarrhythmic drugs on human K2P17.1 channels to provide a more complete picture of K2P17.1 electropharmacology. Whole-cell patch clamp and two-electrode voltage clamp electrophysiology was employed to study human K2P17.1 channel pharmacology. K2P17.1 channels expressed in Xenopus laevis oocytes were screened for sensitivity to antiarrhythmic drugs, revealing significant activation by propafenone (+ 296%; 100 µM), quinidine (+ 58%; 100 µM), mexiletine (+ 21%; 100 µM), propranolol (+ 139%; 100 µM), and metoprolol (+ 17%; 100 µM) within 60 min. In addition, the currents were inhibited by amiodarone (- 13%; 100 µM), sotalol (- 10%; 100 µM), verapamil (- 21%; 100 µM), and ranolazine (- 8%; 100 µM). K2P17.1 channels were not significantly affected by ajmaline and carvedilol. Concentration-dependent K2P17.1 activation by propafenone was characterized in more detail. The onset of activation was fast, and current-voltage relationships were not modulated by propafenone. K2P17.1 activation was confirmed in mammalian Chinese hamster ovary cells, revealing 7.8-fold current increase by 100 µM propafenone. Human K2P17.1 channels were sensitive to multiple antiarrhythmic drugs. Differential pharmacological regulation of repolarizing K2P17.1 background K+ channels may be employed for personalized antiarrhythmic therapy.