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An abstract of the article "Adaptive Responses of Gap Junctional Communication and Increased Myocyte Survival With Preconditioning," by R. Sundset and colleagues is presented.

Cancer immunotherapy has evolved significantly over the last decade, with therapeutics targeting the adaptive immune system showing exciting effects in clinics. Yet, the modulation of the innate immune system, particularly the tumor-associated innate immune cells which are an integral part of immune responses in cancer, remains less understood. The arginase 1 (Arg1) pathway is a pivotal metabolic pathway that tumor-associated innate immune cells exploit to create an immunosuppressive tumor microenvironment, leading to the evasion of immune surveillance. The inhibition of Arg1 presents a therapeutic opportunity to reverse this immunosuppression, and Nω‑hydroxy-l-norarginine (nor-NOHA) has emerged as a potent arginase inhibitor with promising in vivo efficacy. However, the rapid systemic clearance of nor-NOHA poses a significant challenge for its therapeutic application. This study pioneers the encapsulation of nor-NOHA in liposomes, aiming to enhance its bioavailability and prolong its inhibitory activity against Arg1. Historically, the extensive interaction between innate immune cells and nanoparticles has been one of the biggest drawbacks in nanomedicine. Here we seek to utilize this effect and deliver liposomal nor-NOHA to the arginase 1 expressing innate immune cells. We systematically investigated the effect of lipid composition, acyl chain length, manufacturing and loading methodology on the encapsulation efficiency (EE%) and release profile of nor-NOHA. Our results indicate that while the manufacturing method and lipid acyl chain length do not significantly impact EE%, they crucially influence the release kinetics of nor-NOHA, with longer acyl chains demonstrating a more sustained release of nor-NOHA from liposomes enabling continuous inhibition of Arg1. Our findings suggest that liposomal nor-NOHA retains its functional inhibitory activity and could offer improved pharmacokinetic properties, making it a compelling base for iterations for further innovative cancer immunotherapeutic strategies in preclinical and clinical evaluations., Competing Interests: Declarations of interest None., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Clinical advancements in nuclear medicine theranostics has excited a research interest in exploring novel radionuclides for medical use. The duo of the β - emitter 67 Cu and the positron emitter 64 Cu, has advantages over the well-established clinical pair 68 Ga and 177 Lu in terms of capability for high-precision therapy. Low availability has hindered the use of 67 Cu whereas 64 Cu has become established at a limited number of sites through production in low-to-medium energy biomedical cyclotrons. Via the reaction 70 Zn(p,α) 67 Cu, 67 Cu can also be cyclotron produced, although data on the cross sections of this reaction are sparse. Our aim in this study was three-fold: 1) to establish cross sections for relevant beam energies (14-16 MeV) of the 70 Zn(p,α) 67 Cu reaction; 2) determine experimentally the thick target yield for 16.5 MeV proton beam; 3) establish a routine production of 67 Cu for radiochemical and preclinical research. Additionally, our work aims to explore the feasibility of using biomedical cyclotrons for developing of novel therapeutic radionuclides. Thin layers of enriched 70 Zn were electrodeposited onto silver foils to employ the stacked foils technique for assessing the cross section at six energies. The thick target yield was measured experimentally using a pressed [ 70 Zn]ZnO target. Methods were developed for solid phase extraction separation of 67 Cu from the target material, as well as quality control of the product with regards to radionuclidic and radiochemical purity. Radiolabelling of PSMA-617 precursor was performed and the end product injected in a healthy mouse for a kinetic study. As a proof of concept for preclinical applications The animal was then SPECT imaged using the 185 keV gamma emission line. Summarizing, our data confirm that biomedical cyclotrons can contribute in developing novel radionuclides, even of low cross section, for preclinical research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: The main objective is to discuss why treatment of non-prostate cancers with [ 177 Lu]Lu-PSMA-radioligand achieved only low tumor dose in most published cases, despite high uptake on PSMA PET. We use a patient with renal cell carcinoma as an illustrative example. Furthermore, we discuss how the problem with early washout and low tumor dose might be overcome by using a radionuclide with shorter half-life, matching the target binding residence time., Case Presentation: [ 68 Ga]Ga-PSMA-11 PET/CT of a 56-year old man with metastatic renal cell carcinoma showed high lesion uptake. One dose of 6.9 GBq [ 177 Lu]Lu-PSMA-I&T was administrated. Post-therapy dosimetry was performed with SPECT/CT and whole-body planar imaging after 5, 24 and 48 h. Doses to target lesions were only 0.2-0.5 Gy. No treatment effect was achieved., Conclusion: Rapid tumor washout of [ 177 Lu]Lu-PSMA-I&T and low tumor dose despite high uptake of [ 68 Ga]Ga-PSMA-11 are most likely caused by localization of PSMA-receptors on neovasculature rather than on the tumor cells, and unlike in prostate cancer cells, the PSMA-RL / PSMA-receptor complex is not internalized. To overcome the problem with early washout, the use of a radionuclide with shorter half-life matching the target binding residence time will be needed., (© 2024. The Author(s).)

Background: This study aimed to develop a novel positron emission tomography (PET) tracer, [ 68 Ga]Ga-TD-01, for CXCR4 imaging. To achieve this goal, the molecular scaffold of TIQ15 was tuned by conjugation with the DOTA chelator to make it suitable for 68 Ga radiolabeling., Methods: A bifunctional chelator was prepared by conjugating the amine group of TIQ15 with p-NCS-Bz-DOTA, yielding TD-01, with a high yield (68.92%). TD-01 was then radiolabeled with 68 Ga using 0.1 M ammonium acetate at 60 °C for 10 min. A 1-h dynamic small animal PET/MRI study of the labeled compound in GL261-luc2 tumor-bearing mice was performed, and brain tumor uptake was assessed. Blocking studies involved pre-administration of TIQ15 (10 mg/kg) 10 min before the PET procedure started., Results: [ 68 Ga]Ga-TD-01 exhibited a radiochemical yield (RCY) of 36.33 ± 1.50% (EOS), with a radiochemical purity > 99% and a molar activity of 55.79 ± 1.96 GBq/µmol (EOS). The radiotracer showed in vitro stability in PBS and human plasma for over 4 h. Biodistribution studies in healthy animals revealed favorable kinetics for subsequent PET pharmacokinetic modeling with low uptake in the brain and moderate uptake in lungs, intestines and spleen. Elimination could be assigned to a renal-hepatic pathway as showed by high uptake in kidneys, liver, and urinary bladder. Importantly, [ 68 Ga]Ga-TD-01 uptake in glioblastoma (GBM)-bearing mice significantly decreased upon competition with TIQ15, with a baseline tumor-to-background ratios > 2.5 (20 min p.i.), indicating high specificity., Conclusion: The newly developed CXCR4 PET tracer, [ 68 Ga]Ga-TD-01, exhibited a high binding inhibition for CXCR4, excellent in vitro stability, and favorable pharmacokinetics, suggesting that the compound is a promising candidate for full in vivo characterization of CXCR4 expression in GBM, with potential for further development as a tool in cancer diagnosis., (© 2024. The Author(s).)

Background: Treatment of castration-resistant metastatic prostate cancer with [¹⁷⁷Lu]PSMA radioligand., Case Presentation: A man in his seventies with metastatic prostate cancer received castration therapy for four years, developing castration-resistant disease. PET/CT with [⁶⁸Ga]PSMA-11 showed high uptake in metastatic lymph nodes. The patient received 7.4 GBq [¹⁷⁷Lu]PSMA-I&T (Curium, Finland) as five treatments at five-week intervals. Five weeks after the first treatment, p-PSA dropped from 154 to 53 µg/L. Five weeks after the fifth treatment, p-PSA was 1.8 µg/L. [⁶⁸Ga]PSMA-11 PET/CT showed significant reduction in the size of metastases, with the largest decreasing in diameter from 10 to 4 mm. Seven months after the fifth treatment, p-PSA increased to 14.3 µg/L, and [⁶⁸Ga]PSMA-11 PET/ CT revealed additional skeletal metastases, while the lymph node metastases remained unchanged. Thus, the treatment had a good but temporary effect on the metastases., Interpretation: Treatment with [¹⁷⁷Lu]PSMA radioligand resulted in a temporary regression of the metastases.

Dynamic positron emission tomography and kinetic modeling play a critical role in tracer development research using small animals. Kinetic modeling from dynamic PET imaging requires accurate knowledge of an input function, ideally determined through arterial blood sampling. Arterial cannulation in mice, however, requires complex, time-consuming and terminal surgery, meaning that longitudinal studies are impossible. The aim of the current work was to develop and evaluate a non-invasive, deep-learning-based prediction model (DLIF) that directly takes the PET data as input to predict a usable input function. We first trained and evaluated the DLIF model on 68 [ 18 F]Fluorodeoxyglucose mouse scans with image-derived targets using cross validation. Subsequently, we evaluated the performance of a trained DLIF model on an external dataset consisting of 8 mouse scans where the input function was measured by continuous arterial blood sampling. The results showed that the predicted DLIF and image-derived targets were similar, and the net influx rate constants following from Patlak modeling using DLIF as input function were strongly correlated to the corresponding values obtained using the image-derived input function. There were somewhat larger discrepancies when evaluating the model on the external dataset, which could be attributed to systematic differences in the experimental setup between the two datasets. In conclusion, our non-invasive DLIF prediction method may be a viable alternative to arterial blood sampling in small animal [ 18 F]FDG imaging. With further validation, DLIF could overcome the need for arterial cannulation and allow fully quantitative and longitudinal experiments in PET imaging studies of mice., Competing Interests: RL is a co-founder and acting Chief Scientific Officer of Imaging Research & Technology Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024 Kuttner, Luppino, Convert, Sarrhini, Lecomte, Kampffmeyer, Sundset and Jenssen.)

Nanomedicine holds immense potential for therapeutic manipulation of phagocytic immune cells. However, in vitro studies often fail to accurately translate to the complex in vivo environment. To address this gap, we employed an ex vivo human whole-blood assay to evaluate liposome interactions with immune cells. We systematically varied liposome size, PEG-surface densities and sphingomyelin and ganglioside content. We observed differential uptake patterns of the assessed liposomes by neutrophils and monocytes, emphasizing the importance of liposome design. Interestingly, our results aligned closely with published in vivo observations in mice and patients. Moreover, liposome exposure induced changes in cytokine release and cellular responses, highlighting the potential modulation of immune system. Our study highlights the utility of human whole-blood models in assessing nanoparticle-immune cell interactions and provides insights into liposome design for modulating immune responses., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: Glioblastoma multiforme (GBM) is the most common glioma and standard therapies can only slightly prolong the survival. Neo-vascularization is a potential target to image tumor microenvironment, as it defines its brain invasion. We investigate [ 18 F]rhPSMA-7.3 with PET/MRI for quantitative imaging of neo-vascularization in GBM bearing mice and human tumor tissue and compare it to [ 18 F]FET and [ 18 F]fluciclovine using PET pharmacokinetic modeling (PKM)., Methods: [ 18 F]rhPSMA-7.3, [ 18 F]FET, and [ 18 F]fluciclovine were i.v. injected with 10.5 ± 3.1 MBq, 8.0 ± 2.2 MBq, 11.5 ± 1.9 MBq (n = 28, GL261-luc2) and up to 90 min PET/MR imaged 21/28 days after surgery. Regions of interest were delineated on T2-weighted MRI for (i) tumor, (ii) brain, and (iii) the inferior vena cava. Time-activity curves were expressed as SUV mean, SUVR and PKM performed using 1-/2-tissue-compartment models (1TCM, 2TCM), Patlak and Logan analysis (LA). Immunofluorescent staining (IFS), western blotting, and autoradiography of tumor tissue were performed for result validation., Results: [ 18 F]rhPSMA-7.3 showed a tumor uptake with a tumor-to-background-ratio (TBR) = 2.1-2.5, in 15-60 min. PKM (2TCM) confirmed higher K1 (0.34/0.08, p = 0.0012) and volume of distribution V T (0.24/0.1, p = 0.0017) in the tumor region compared to the brain. Linearity in LA and similar k3 = 0.6 and k4 = 0.47 (2TCM, tumor, p = ns) indicated reversible binding. K1, an indicator for vascularization, increased (0.1/0.34, 21 to 28 days, p < 0.005). IFS confirmed co-expression of PSMA and tumor vascularization. [ 18 F]fluciclovine showed higher TBR (2.5/1.8, p < 0.001, 60 min) and V S (1.3/0.7, p < 0.05, tumor) compared to [ 18 F]FET and LA indicated reversible binding. V T increased (p < 0.001, tumor, 21 to 28 days) for [ 18 F]FET (0.5-1.4) and [ 18 F]fluciclovine (0.84-1.5)., Conclusion: [ 18 F]rhPSMA-7.3 showed to be a potential candidate to investigate the tumor microenvironment of GBM. Following PKM, this uptake was associated with tumor vascularization. In contrast to what is known from PSMA-PET in prostate cancer, reversible binding was found for [ 18 F]rhPSMA-7.3 in GBM, contradicting cellular trapping. Finally, [ 18 F]fluciclovine was superior to [ 18 F]FET rendering it more suitable for PET imaging of GBM., (© 2022. The Author(s).)

Cerebral blood flow (CBF) can be measured with dynamic positron emission tomography (PET) of 15 O-labeled water by using tracer kinetic modelling. However, for quantification of regional CBF, an arterial input function (AIF), obtained from arterial blood sampling, is required. In this work we evaluated a novel, non-invasive approach for input function prediction based on machine learning (MLIF), against AIF for CBF PET measurements in human subjects.Twenty-five subjects underwent two 10 min dynamic 15 O-water brain PET scans with continuous arterial blood sampling, before (baseline) and following acetazolamide medication. Three different image-derived time-activity curves were automatically segmented from the carotid arteries and used as input into a Gaussian process-based AIF prediction model, considering both baseline and acetazolamide scans as training data. The MLIF approach was evaluated by comparing AIF and MLIF curves, as well as whole-brain grey matter CBF values estimated by kinetic modelling derived with either AIF or MLIF.The results showed that AIF and MLIF curves were similar and that corresponding CBF values were highly correlated and successfully differentiated before and after acetazolamide medication. In conclusion, our non-invasive MLIF method shows potential to replace the AIF obtained from blood sampling for CBF measurements using 15 O-water PET and kinetic modelling.

We report the synthesis, radiosynthesis and biological characterisation of two gonadotropin-releasing hormone receptor (GnRH-R) antagonists with nanomolar binding affinity. A small library of GnRH-R antagonists was synthesised in 20-67 % overall yield with the aim of identifying a high-affinity antagonist capable of crossing the blood-brain barrier. Binding affinity to rat GnRH-R was determined by autoradiography in competitive-binding studies against [ 125 I]buserelin, and inhibition constants were calculated by using the Cheng-Prusoff equation. The radioligands were obtained in 46-79 % radiochemical yield and >95 % purity and with a molar activity of 19-38 MBq/nmol by direct nucleophilic radiofluorination. Positron emission tomography imaging in rat under baseline conditions in comparison to pretreatment with a receptor-saturating dose of GnRH antagonist revealed saturable uptake (0.1 %ID/mL) into the brain., (© 2020 The Authors. Published by Wiley-VCH GmbH.)

This audit describes ionizing and non-ionizing diagnostic imaging at a regional trauma centre. All 144 patients (males 79.2%, median age 31 years) met with trauma team activation from 1 January 2015 to 31 December 2015 were included. We used data from electronic health records to identify all diagnostic imaging and report radiation exposure as dose area product (DAP) for conventional radiography (X-ray) and dose length product (DLP) and effective dose for CT. During hospitalization, 134 (93.1%) underwent X-ray, 122 (84.7%) CT, 92 (63.9%) focused assessment with sonography for trauma (FAST), 14 (9.7%) ultrasound (FAST excluded) and 32 (22.2%) magnetic resonance imaging. One hundred and sixteen (80.5%) underwent CT examinations during trauma admissions, and 73 of 144 (50.7%) standardized whole body CT (SWBCT). DAP values were below national reference levels. Median DLP and effective dose were 2396 mGycm and 20.42 mSv for all CT examinations, and 2461 mGycm (national diagnostic reference level 2400) and 22.29 mSv for a SWBCT., (© The Author(s) 2020. Published by Oxford University Press.)

The increased expression of gonadotropin releasing hormone receptor (GnRH-R) in brain has been strongly linked to Alzheimer disease. Therefore, the development of radiolabeled imaging agents for GnRH-R is relevant for early diagnosis of Alzheimer disease. We have recently disclosed the discovery of two promising compounds displaying nanomolar-range affinity for the GnRH-R. In the present study, a preclinical evaluation of the compound properties was performed to evaluate their potential as single photon emission computed tomography (SPECT) radiotracers for imaging the GnRH-receptor. The compounds were assessed in vitro by performing serum stability analysis by human and rat serum, metabolic profiling by human liver microsomes, and exploratory rat brain autoradiography. The investigated compounds displayed satisfactory stability against human, rat serum, and liver microsomal metabolism, which favors their potential as SPECT-imaging agents. Additionally, we identified and quantified the formation rate of the metabolites by fragmentation of up to five mass spectrometric stages. The GnRH-R rat brain specificity of these compounds was tested in competition with a known ligand for the receptor and the in vitro autoradiography confirmed that compounds 3 and 4 binds to rat GnRH-R in different rat brain regions., (© 2019 The Authors. Journal of Labelled Compounds and Radiopharmaceuticals published by John Wiley & Sons Ltd.)

Tracer kinetic modelling, based on dynamic 18 F-fluorodeoxyglucose (FDG) positron emission tomography (PET) is used to quantify glucose metabolism in humans and animals. Knowledge of the arterial input-function (AIF) is required for such measurements. Our aim was to explore two non-invasive machine learning-based models, for AIF prediction in a small-animal dynamic FDG PET study. 7 tissue regions were delineated in images from 68 FDG PET/computed tomography mouse scans. Two machine learning-based models were trained for AIF prediction, based on Gaussian processes (GP) and a long short-term memory (LSTM) recurrent neural network, respectively. Because blood data were unavailable, a reference AIF was formed by fitting an established AIF model to vena cava and left ventricle image data. The predicted and reference AIFs were compared by the area under curve (AUC) and root mean square error (RMSE). Net-influx rate constants, K i , were calculated with a two-tissue compartment model, using both predicted and reference AIFs for three tissue regions in each mouse scan, and compared by means of error, ratio, correlation coefficient, P value and Bland-Altman analysis. The impact of different tissue regions on AIF prediction was evaluated by training a GP and an LSTM model on subsets of tissue regions, and calculating the RMSE between the reference and the predicted AIF curve. Both models generated AIFs with AUCs similar to reference. The LSTM models resulted in lower AIF RMSE, compared to GP. K i from both models agreed well with reference values, with no significant differences. Myocardium was highlighted as important for AIF prediction, but AIFs with similar RMSE were obtained also without myocardium in the input data. Machine learning can be used for accurate and non-invasive prediction of an image-derived reference AIF in FDG studies of mice. We recommend the LSTM approach, as this model predicts AIFs with lower errors, compared to GP.

Background: Hospitals must improve patient safety and quality continuously. Clinical quality registries can drive such improvement. Trauma registries code injuries according to the Abbreviated Injury Scale (AIS) and benchmark outcomes based on the Injury Severity Score (ISS) and New ISS (NISS). The primary aim of this study was to validate the injury codes and severities registered in a national trauma registry. Secondarily, we aimed to examine causes for missing and discordant codes, to guide improvement of registry data quality., Methods: We conducted an audit and established an expert coder group injury reference standard for patients met with trauma team activation in 2015 in a Level 1 trauma centre. Injuries were coded according to the AIS. The audit included review of all data in the electronic health records (EHR), and new interpretation of all images in the picture archiving system. Validated injury codes were compared with the codes registered in the registry. The expert coder group's interpretations of reasons for discrepancies were categorised and registered. Inter-rater agreement between registry data and the reference standard was tested with Bland-Altman analysis., Results: We validated injury data from 144 patients (male sex 79.2%) with median age 31 (inter quartile range 19-49) years. The total number of registered AIS codes was 582 in the registry and 766 in the reference standard. All injuries were concordantly coded in 62 (43.1%) patients. Most non-registered codes (n = 166 in 71 (49.3%) patients) were AIS 1, and information in the EHR overlooked by registrars was the dominating cause. Discordant coding of head injuries and extremity fractures were the most common causes for 157 discordant AIS codes in 74 (51.4%) patients. Median ISS (9) and NISS (12) for the total population did not differ between the registry and the reference standard., Conclusions: Concordance between the codes registered in the trauma registry and the reference standard was moderate, influencing individual patients' injury codes validity and ISS/NISS reliability. Nevertheless, aggregated median group ISS/NISS reliability was acceptable.

Objectives: In vivo evaluations of a gonadotropin releasing hormone-receptor single photon emission computed tomography radiotracer for non-invasive detection of gonadotropin releasing homone-receptors in brain., Results: We have used a simple, robust and high-yielding procedure to radiolabel an alpha-halogenated bioactive compound with high radiochemical yield. Literature findings showed similar alpha-halogenated compounds suitable for in vivo evaluations. The compound was found to possess nano molar affinity for the gonadotropin releasing hormone-receptor in a competition dependent inhibition study. Furthermore, liquid chromatography-mass spectrometry analysis in saline, human and rat serum resulted in 46%, 52% and 44% stability after incubation for 1 h respectively. In addition, rat brain single photon emission computed tomography and biodistribution studies gave further insight into the nature of the compound as a radiotracer.

In this communication, we report the synthesis and characterization of a library of small molecule antagonists of the human gonadotropin releasing hormone receptor based upon the 2-(4- tert -butylphenyl)-4-piperazinyl-benzimidazole scaffold via Cu-catalysed azide alkyne cycloaddition. Our main purpose was to find a more soluble compound based on the WAY207024 lead with nanomolar potency to inhibit the GnRH receptor. A late stage diversification by the use of click chemistry was, furthermore developed to allow for expansion of the library in future optimisations. All compounds were tested in a functional assay to determine the individual potency of inhibiting stimulation of the receptor by the endogenous agonist GnRH. In conclusion, we found that compound 8a showed improved solubility compared to WAY207024 and nanomolar affinity to GnRH receptor.

Background: It is challenging to obtain a similar access to positron emission tomography/computed tomography (PET-CT) within the whole region served. In the subarctic and arctic region of Norway, significant distances, weather conditions and seasonable darkness have been challenging when the health care provider has aimed for a high quality PET-CT service with similar availability to all inhabitants., Methods: The PET-CT service at the University Hospital of North Norway (UNN) was established in May 2010. The glucose analogue tracer fluorine-18 fluorodeoxyglucose (FDG) was delivered from Helsinki, Finland. An ambulatory PET-CT scanner was initially employed and a permanent local one was introduced in October 2011. In March 2014, we analysed retrospectively all data on the PET-CT exams performed at the Section of Nuclear Medicine, Department of Radiology during a 32 months time period 2010-13. The following patient data were recorded: gender, age, diagnosis, residence and distance of travelling. There were in total 796 exams in 706 patients., Results: Four hundred sixty-one PET-CT exams per million inhabitants were, on average, performed per year. Lung cancer (32.7%), malignant melanoma (11.3%), colorectal cancer (10.9%) and lymphoma (9.7%) constituted two-thirds of all exams. Three-fourths were males and the median age was 63.5 years (range 15.2-91.4 years). The access to PET-CT exam varied within the region. The southern county (Nordland) experienced a significantly less access (p < 0.0001) to the regional service. Except for malignant melanoma, this finding was observed in all major cancer subgroups. In colorectal cancer and lymphoma a lower consumption of PET-CT was also observed in the northeastern county (Finnmark). Patients' mean distance of travelling by car (one way) was 373 km (median 313 km, range 5-936 km)., Conclusion: PET-CT was not similarly available within the region. Especially, inhabitants in the southern county experienced less access to the regional service. National and regional standards of care, new scanners and improved collaboration between hospital trusts may alter this situation.

Aim: Gap junction intercellular communication (GJIC) and hemichannel permeability may have important roles during an ischemic insult. Our aim was to evaluate the effect of ischemia on gap junction channels and hemichannels., Methods: We used neonatal rat heart myofibroblasts and simulated ischemia with a HEPES buffer with high potassium, low pH, absence of glucose, and oxygen tension was reduced by dithionite. Microinjection, western blot, immunofluorescence, cell viability and dye uptake were used to evaluate the effects induced by dithionite. Isolated perfused rat hearts were used to analyse infarct size., Results: Short period with simulated ischemia reduced the ability to transfer a dye between neighbouring cells, which indicated reduced GJIC. Prolonged exposure to simulated ischemia caused opening of hemichannels, and cell death was apparent while gap junction channels remained closed. Connexin 43 became partially dephosphorylated and the total amount decreased during simulated ischemia. We were not able to detect the alternative hemichannel-forming protein, Pannexin 1, in these cells. The potential importance of Connexin 43 or Pannexin 1 hemichannels in ischemia-induced infarct in the intact heart was studied by perfusion of the heart in the presence of peptides that block one or the other type of hemichannels. The connexin-derived peptide, Gap26, significantly reduced the infract/risk zone ratio (control 48.7±4.2% and Gap26 19.4±4.1%, p<0.001), while the pannexin-derived peptide, (10)Panx1, did not change infarct/risk ratio., Conclusion: Connexin 43 is most likely responsible for both closure of gap junction channels and opening of hemichannels during simulated ischemia in neonatal rat heart myofibroblasts. Opening of connexin 43 hemichannels during ischemia-reperfusion seems to be an important mechanism for ischemia-reperfusion injury in the heart. By preventing the opening of these channels during early ischemia-reperfusion the infarct size becomes significantly reduced., (Copyright © 2011 S. Karger AG, Basel.)

Background: Recently, cardiac resynchronization therapy, by using biventricular pacemakers, has become implemented in the treatment of patients with severe heart failure. However, using the classical inclusion criteria, 30 % of patients treated with resynchronization do not improve symptoms or activity level. Phase analysis of radionuclide ventriculography gives information about pattern of ventricular contraction and may detect dyssynchrony. The method may therefore be used to select patients with dyssynctrony to resynchronization therapy. In this study we have investigated the amount of dyssynchrony, by using phase analysis of radionuclide ventriculography, in men and women as a function of left ventricular ejection fraction., Material and Methods: The study is based on 1 266 radionuclide ventriculographies performed at Section of Nuclear Medicine, University Hospital North-Norway, during 1998 - 2006. The relationship between left ventricular ejection fraction and number of patients with ventricular dyssynchrony was investigated. 90 patients with no known heart problems were considered as reference values for synchrony data., Results: The phase analysis showed that 35 % of the women and 34 % of the men with left ventricular ejection fraction below 35 % had both inter- and intra-ventricular dyssynchrony., Interpretation: Today's criteria for including patients to resynchonizing therapy are not good enough. Phase analysis of radionuclide ventriculography of patients with left ventricular ejection fraction below 35 % shows that one third of the patients have both inter- and intra- ventricular dyssynchrony. These patients might be responders to resynchronization therapy.

Electrophysiological remodeling involving gap junctions has been demonstrated in failing hearts and may contribute to intercellular uncoupling, delayed conduction, enhanced arrhythmias, and vulnerability to sudden death in patients with heart failure. Recently, we showed that failing human hearts exhibit marked increases in connexin45 (Cx45) expression in addition to previously documented decreases in connexin43 (Cx43) expression. Each of these changes results in reduced gap junction coupling. The objective of the present study was to examine functional consequences of increased Cx45 in cardiac gap junctions. Transgenic mice with cardiac-selective overexpression of the developmentally downregulated cardiac connexin, connexin45 (Cx45OE mice) were subjected to in vivo electrophysiology studies in which an intracardiac catheter was used to induce ventricular arrhythmias in anesthetized mice, and in which ambulatory ECG monitoring was used to detect spontaneous arrhythmias in unanesthetized mice. Hearts were analyzed by TaqMan RT-PCR, immunostaining, immunoblotting, and echocardiography. Lucifer yellow and neurobiotin dye transfer was used to assess coupling in transgenic and control myocyte cultures. Cx45 mRNA was two orders of magnitude greater in Cx45OE mice. Cx45-immunoreactive signal at gap junctions increased twofold and total Cx45 protein by immunoblotting increased 25% in Cx45OE mice compared with nontransgenic littermate controls. Functionally, Cx45OE mice exhibited more inducible ventricular tachycardia than controls but did not exhibit any other functional or structural derangements as assessed by echocardiography. Ventricular myocytes isolated from Cx45OE mice exhibited diminished intercellular transfer of Lucifer yellow dye and increased transfer of neurobiotin, consistent with altered cell-to-cell communication. Thus increased myocardial expression of Cx45 results in remodeling of intercellular coupling and greater susceptibility to ventricular arrhythmias in vivo.

Ischemic preconditioning increases the heart's tolerance to a subsequent longer ischemic period. The purpose of this study was to investigate the role of gap junction communication in simulated preconditioning in cultured neonatal rat cardiac myofibroblasts. Gap junctional intercellular communication was assessed by Lucifer yellow dye transfer. Preconditioning preserved intercellular coupling after prolonged ischemia. An initial reduction in coupling in response to the preconditioning stimulus was also observed. This may protect neighboring cells from damaging substances produced during subsequent regional ischemia in vivo, and may preserve gap junctional communication required for enhanced functional recovery during subsequent reperfusion.

Introduction: Heart failure is associated with reduced expression of the major gap junction protein connexin43 (Cx43), which may contribute to arrhythmias and sudden cardiac death in this patient population. Other cardiac connexins may be altered as well. Because connexin45 (Cx45) has been shown to colocalize with Cx43, we determined whether the number, size, or distribution of Cx45 gap junctions is altered in the failing heart., Methods and Results: Cx45 expression levels were measured by immunoblotting and quantitative immunostaining in failing and control human left ventricles. Total Cx45 protein was significantly (P = 0.021) up-regulated 1.8-fold in failing hearts. Cx45 immunohistochemical signal was increased by 80% (P = 0.005) due to a 3.5-fold increase in the number of gap junctions containing Cx45. Cx45 mRNA was not altered in failing hearts, suggesting reduced degradation of Cx45 protein in the failing heart. Cx43 signal, on the other hand, was reduced by 49% in failing hearts. Double-label experiments demonstrated colocalization of Cx45 and Cx43 in the same gap junctions., Conclusion: Cx45 is markedly enhanced in the failing heart. Up-regulation of Cx45 in conjunction with down-regulation of Cx43 could result in abnormal impulse propagation and generation of ventricular arrhythmias, thereby predisposing patients in heart failure to sudden cardiac death.

The sodium-hydrogen exchanger (NHE) helps the cell to recover from intracellular acidosis. In this study, we have investigated the effect of HOE 642 (a specific NHE1 blocker) on papillary muscles from rats and guinea pigs during transient acidosis and PKC activation by recording developed force (DF), action potential characteristics, and electrical conductance (stimulus-response interval). Two protocols were used, with or without HOE 642 (10(-5) mol/L): papillary muscle was exposed (i) for 15 min to a glucose-free, nonoxygenated HEPES buffer containing lactate (20 mmol/L) (pH 6.8) followed by 15 min recovery or (ii) to a PKC activator (phorbolmyristate acetate (PMA) (10(-9) mol/L)) for 30 min. The DF after acidification remained significantly decreased in the NHE-blocked papillary muscles. During recovery from acidosis, papillary muscles exposed to HOE 642 remained at a higher electrical resistance. The present study shows that post-acidotic continued depression of DF and change in tissue electrophysiological properties might occur as a result of blocking the NHE. During infarct development, the tissue-protecting effect of NHE blockade has been well documented. When acidosis or reduced contractile function is present, however, blocking NHE by HOE 642 might not improve the situation.

Unlabelled: Brief ischemia or hypoxia has been found to protect the heart against subsequent long-lasting ischemia and to improve contractile dysfunction as well to reduce cell necrosis and the incidence of lethal arrhythmias. This phenomenon, termed preconditioning (PC) has been demonstrated in different species. However, little is known about PC in guinea pigs. Moreover, electrophysiological changes underlying protection have not been studied so far in conjuntion with force recovery in a setting of PC. The aim of the study was to study PC in a guinea pig papillary muscle, using recovery of contractility after long hypoxic challenge as the main end-point of protection, and to investigate concomitant electrophysiological alterations. In guinea pig papillary muscle preparations contracting isometrically (paced at 2 Hz), transmembrane action potentials (AP) and developed force (DF) were recorded by conventional microelectrode technique and a force transducer. In addition, effective refractory periods (ERP) were determined. Hypoxia was induced by superfusion with 100% N2 (pO2 < 5 kPa) and pacing at 3,3 Hz. In the control group, long hypoxia lasted for 45 min and was followed by 30 min reoxygenation. In the PC group, muscles were subjected to 5 min hypoxia followed by 10 min recovery prior to sustained hypoxia/reoxygenation., Results: Long hypoxia induced a similar depression of DF in both, PC and control groups. However, a loss of contractile activity occured earlier in the PC group. AP duration and ERP decreased faster and were significantly shorter after PC. Upon reoxygenation, preconditioned muscles showed significantly better recovery of function (DF 86% of prehypoxic value vs. 36% in controls; p < 0,05). AP and ERP were completely restored in both, PC and control groups. Guinea pig papillary muscle can be preconditioned with a brief hypoxic challenge against contractile dysfunction upon long-lasting hypoxia/reoxygenation. Shortening of AP and loss of contractility occured more quickly during hypoxia and may participate in the protective effect of preconditioning. Possible mechanisms might involve facilitated opening of K(ATP)-dependent channels.

Because glycolysis is thought to be important for maintenance of cellular ion homeostasis, the aim of the present study was to examine the role of glycolysis in the control of cytosolic calcium ([Ca2+]i) and cell shortening during conditions of increased calcium influx. Thus, [Ca2+]i and unloaded cell shortening were measured in fura-2/AM loaded rat ventricular myocytes. All cells were superfused with Tyrode's solution containing glucose and pyruvate (to preserve oxidative metabolism), and glycolysis was inhibited by iodoacetate (IAA, 100 microM). Calcium influx was increased, secondary to an increase in intracellular sodium, by addition of veratrine (1 microgram/ml), or directly by either elevating [Ca2+]o from 2 to 5 mM or by exposing the cells to isoproterenol (1 to 100 nm). Veratrine exposure caused a time-dependent increase in both diastolic and systolic [Ca2+]i that resulted in cellular calcium overload and hypercontraction. The rate of increase in [Ca2+]i was more rapid in IAA-treated than in untreated myocytes, leading to a 13+/-3 v 5+/-2% increase (P<0.05) in diastolic [Ca2+]i after 5 min of exposure. The corresponding increases in systolic [Ca2+]i were 43+/-6 and 24+/-5% (P<0.05). Elevated [Ca2+]o resulted in increased [Ca2+]i transient amplitudes and cell shortening. These responses were each attenuated by inhibiting glycolysis, so that the increase was 38+/-5 v 68+/-9% ([Ca2+]i transient amplitude, P<0.05) and 41+/-11 v 91+/-18% (cell shortening, P<0.05). Inhibition of glycolysis did not, however, affect the increase in calcium transient or cell shortening during addition of isoproterenol. We conclude that glycolysis plays an essential role in the maintenance of intracellular calcium homeostasis during severe calcium overload. Glycolysis was also essential for signalling the inotropic effect that accompanied elevation in extracellular calcium, while the changes in intracellular calcium following administration of isoproterenol were not influenced by glycolysis in the present model., (Copyright 1998 Academic Press.)