Mitochondrial network remodeling of the diabetic heart: implications to ischemia related cardiac dysfunction.

Mitochondria play a central role in cellular energy metabolism, and their dysfunction is increasingly recognized as a critical factor in the pathogenesis of diabetes-related cardiac pathophysiology, including vulnerability to ischemic events that culminate in myocardial infarction on the one hand and ventricular arrhythmias on the other. In diabetes, hyperglycemia and altered metabolic substrates lead to excessive production of reactive oxygen species (ROS) by mitochondria, initiating a cascade of oxidative stress that damages mitochondrial DNA, proteins, and lipids. This mitochondrial injury compromises the efficiency of oxidative phosphorylation, leading to impaired ATP production. The resulting energy deficit and oxidative damage contribute to functional abnormalities in cardiac cells, placing the heart at an increased risk of electromechanical dysfunction and irreversible cell death in response to ischemic insults. While cardiac mitochondria are often considered to be relatively autonomous entities in their capacity to produce energy and ROS, their highly dynamic nature within an elaborate network of closely-coupled organelles that occupies 30-40% of the cardiomyocyte volume is fundamental to their ability to exert intricate regulation over global cardiac function. In this article, we review evidence linking the dynamic properties of the mitochondrial network to overall cardiac function and its response to injury. We then highlight select studies linking mitochondrial ultrastructural alterations driven by changes in mitochondrial fission, fusion and mitophagy in promoting cardiac ischemic injury to the diabetic heart.

Advanced Technologies in Radiation Research.

The U.S. Government is committed to maintaining a robust research program that supports a portfolio of scientific experts who are investigating the biological effects of radiation exposure. On August 17 and 18, 2023, the Radiation and Nuclear Countermeasures Program, within the National Institute of Allergy and Infectious Diseases, National Institutes of Health (NIH), partnered with the National Cancer Institute, NIH, the National Aeronautics and Space Administration, and the Radiation Injury Treatment Network to convene a workshop titled, Advanced Technologies in Radiation Research (ATRR), which focused on the use of advanced technologies under development or in current use to accelerate radiation research. This meeting report provides a comprehensive overview of the research presented at the workshop, which included an assembly of subject matter experts from government, industry, and academia. Topics discussed during the workshop included assessments of acute and delayed effects of radiation exposure using modalities such as clustered regularly interspaced short palindromic repeats (CRISPR) - based gene editing, tissue chips, advanced computing, artificial intelligence, and immersive imaging techniques. These approaches are being applied to develop products to diagnose and treat radiation injury to the bone marrow, skin, lung, and gastrointestinal tract, among other tissues. The overarching goal of the workshop was to provide an opportunity for the radiation research community to come together to assess the technological landscape through sharing of data, methodologies, and challenges, followed by a guided discussion with all participants. Ultimately, the organizers hope that the radiation research community will benefit from the workshop and seek solutions to scientific questions that remain unaddressed. Understanding existing research gaps and harnessing new or re-imagined tools and methods will allow for the design of studies to advance medical products along the critical path to U.S. Food and Drug Administration approval.

Mitochondrial A-kinase anchoring proteins in cardiac ventricular myocytes.

Compartmentation of cAMP signaling is a critical factor for maintaining the integrity of receptor-specific responses in cardiac myocytes. This phenomenon relies on various factors limiting cAMP diffusion. Our previous work in adult rat ventricular myocytes (ARVMs) indicates that PKA regulatory subunits anchored to the outer membrane of mitochondria play a key role in buffering the movement of cytosolic cAMP. PKA can be targeted to discrete subcellular locations through the interaction of both type I and type II regulatory subunits with A-kinase anchoring proteins (AKAPs). The purpose of this study is to identify which AKAPs and PKA regulatory subunit isoforms are associated with mitochondria in ARVMs. Quantitative PCR data demonstrate that mRNA for dual specific AKAP1 and 2 (D-AKAP1 & D-AKAP2), acyl-CoA-binding domain-containing 3 (ACBD3), optic atrophy 1 (OPA1) are most abundant, while Rab32, WAVE-1, and sphingosine kinase type 1 interacting protein (SPHKAP) were barely detectable. Biochemical and immunocytochemical analysis suggests that D-AKAP1, D-AKAP2, and ACBD3 are the predominant mitochondrial AKAPs exposed to the cytosolic compartment in these cells. Furthermore, we show that both type I and type II regulatory subunits of PKA are associated with mitochondria. Taken together, these data suggest that D-AKAP1, D-AKAP2, and ACBD3 may be responsible for tethering both type I and type II PKA regulatory subunits to the outer mitochondrial membrane in ARVMs. In addition to regulating PKA-dependent mitochondrial function, these AKAPs may play an important role by buffering the movement of cAMP necessary for compartmentation.

Compartmentation of ß2 -adrenoceptor stimulated cAMP responses by phosphodiesterase types 2 and 3 in cardiac ventricular myocytes.

BACKGROUND AND PURPOSE: In cardiac myocytes, cyclic AMP (cAMP) produced by both ß1 - and ß2 -adrenoceptors increases L-type Ca2+ channel activity and myocyte contraction. However, only cAMP produced by ß1 -adrenoceptors enhances myocyte relaxation through phospholamban-dependent regulation of the sarco/endoplasmic reticulum Ca2+ ATPase 2 (SERCA2). Here we have tested the hypothesis that stimulation of ß2 -adrenoceptors produces a cAMP signal that is unable to reach SERCA2 and determine what role, if any, phosphodiesterase (PDE) activity plays in this compartmentation. EXPERIMENTAL APPROACH: The cAMP responses produced by ß1 -and ß2 -adrenoceptor stimulation were studied in adult rat ventricular myocytes using two different fluorescence resonance energy transfer (FRET)-based biosensors, the Epac2-camps, which is expressed uniformly throughout the cytoplasm of the entire cell and the Epac2-αKAP, which is targeted to the SERCA2 signalling complex. KEY RESULTS: Selective activation of ß1 - or ß2 -adrenoceptors produced cAMP responses detected by Epac2-camps. However, only stimulation of ß1 -adrenoceptors produced a cAMP response detected by Epac2-αKAP. Yet, stimulation of ß2 -adrenoceptors was able to produce a cAMP signal detected by Epac2-αKAP in the presence of selective inhibitors of PDE2 or PDE3, but not PDE4. CONCLUSION AND IMPLICATIONS: These results support the conclusion that cAMP produced by ß2 -adrenoceptor stimulation was not able to reach subcellular locations where the SERCA2 pump is located. Furthermore, this compartmentalized response is due at least in part to PDE2 and PDE3 activity. This discovery could lead to novel PDE-based therapeutic treatments aimed at correcting cardiac relaxation defects associated with certain forms of heart failure.

Reveal LINQ Versus Reveal XT Implantable Loop Recorders: Intra- and Post-Procedural Comparison.

OBJECTIVES: To compare the procedure, recovery, hospitalization times, and costs along with patient/parent satisfaction after newer-generation cardiac implantable loop recorder (Reveal LINQ; Medtronic Inc, Minneapolis, Minnesota) and previous-generation implantable loop recorder (Reveal XT; Medtronic Inc). STUDY DESIGN: A prospective study of patients undergoing LINQ implantations between April 2014 and October 2015 was performed. Retrospective chart review of patients undergoing XT implantations was performed for comparison. RESULTS: Thirty-one patients received LINQ and 15 patients received XT. Indications included syncope/palpitations (28/46, 61%), history of arrhythmias (9/46, 20%), arrhythmia burden in congenital heart disease (5/46, 10%), and monitoring in channelopathies (4/46, 9%). The LINQ group underwent more conscious sedation procedures than the XT group (8/31 vs 0/15, P = .04) with shorter procedural time (9 vs 34 minutes, P <.001), room occupation time (38 vs 81 minutes, P <.001), recovery time (21 vs 67 minutes, P <.001), and total hospital time (214 vs 264 minutes, P = .046). The LINQ group also had shorter return to activity time (2 vs 5 days, P = 1). Three device erosions in the LINQ group required reintervention. The LINQ group had fewer body image issues than the XT group (1/26 vs 5/14, P = .01) with both groups scoring 5/5 overall patient/parent satisfaction score at follow-up. Both groups had comparable total direct hospital costs (US $5905 vs $5438, P = .8). CONCLUSIONS: LINQ offers better procedural and recovery time compared with XT. LINQ implantations under conscious sedation reduce total hospitalization time.

The Xenopus oocyte cut-open vaseline gap voltage-clamp technique with fluorometry.

The cut-open oocyte Vaseline gap (COVG) voltage clamp technique allows for analysis of electrophysiological and kinetic properties of heterologous ion channels in oocytes. Recordings from the cut-open setup are particularly useful for resolving low magnitude gating currents, rapid ionic current activation, and deactivation. The main benefits over the two-electrode voltage clamp (TEVC) technique include increased clamp speed, improved signal-to-noise ratio, and the ability to modulate the intracellular and extracellular milieu. Here, we employ the human cardiac sodium channel (hNaV1.5), expressed in Xenopus oocytes, to demonstrate the cut-open setup and protocol as well as modifications that are required to add voltage clamp fluorometry capability. The properties of fast activating ion channels, such as hNaV1.5, cannot be fully resolved near room temperature using TEVC, in which the entirety of the oocyte membrane is clamped, making voltage control difficult. However, in the cut-open technique, isolation of only a small portion of the cell membrane allows for the rapid clamping required to accurately record fast kinetics while preventing channel run-down associated with patch clamp techniques. In conjunction with the COVG technique, ion channel kinetics and electrophysiological properties can be further assayed by using voltage clamp fluorometry, where protein motion is tracked via cysteine conjugation of extracellularly applied fluorophores, insertion of genetically encoded fluorescent proteins, or the incorporation of unnatural amino acids into the region of interest(1). This additional data yields kinetic information about voltage-dependent conformational rearrangements of the protein via changes in the microenvironment surrounding the fluorescent molecule.