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Atrial granules as acidic calcium stores in cardiomyocytes.
Acidic calcium stores significantly influence basal calcium transient amplitude and β-adrenergic responses in cardiomyocytes. Atrial myocytes contain atrial granules (AGs), small acidic organelles that store and secrete atrial natriuretic peptide (ANP) and are absent in healthy ventricular myocytes. AGs are known to be acidic and calcium-rich, but their number and location relative to other signalling sites remain unexplored. Labelling of acidic organelles in adult guinea pig cardiomyocytes showed the presence of acidic puncta throughout the cytosol. Atrial myocytes exhibited an increased concentration of acidic organelles at the nuclear poles. Live cell fluorescent studies using 4-phenyl-3-butenoic acid (PBA) to inhibit peptidylglycine α-amidating monooxygenase, a crucial component of AGs membranes, effectively eliminated staining at the nuclear poles and most acidic puncta in atrial cells, but not in ventricular cells. Our immunofluorescent labelling also emphasizes the differences in acidic punctae between atrial and ventricular myocytes by showing minimal co-localization between AG-specific ANP and lysosomal-associated membrane protein. Electron microscopy studies on goat atrial fibrillation (AF) and sham control tissue allowed visualization of AGs. Quantitative analysis revealed that AGs were positioned significantly further away from the nearest sarcoplasmic reticulum and were closer to mitochondria in AF compared to sinus rhythm control tissue. We raise the question whether the positioning of AGs is strategic for communication with other calcium-containing organelles.
Lesion level and severity acutely influence metabolomic profiles in spinal cord injury.
Changes in the peripheral metabolome, particularly in the blood, may provide biomarkers for assessing lesion severity and predicting outcomes after spinal cord injury (SCI). Using principal component analysis (PCA) and Orthogonal Partial Least Squares Discriminatory Analysis (OPLS-DA), we sought to discover how SCI severity and location acutely affect the nuclear magnetic resonance-acquired metabolome of the blood, spinal cord, and liver at 6 h post-SCI in mice. Unsupervised PCA of the spinal cord metabolome separated mild (30 kdyne) and severe (70 kdyne) contusion injury groups but did not distinguish between lesion level. However, OPLS-DA could discriminate thoracic level T2 from T9 lesions in both blood plasma (accuracy 86 ± 6%) and liver (accuracy 89 ± 5%) samples. These differences were dependent on alterations in energy metabolites (lactate and glucose), lipoproteins, and lipids. Lactate was the most discriminatory between mild and severe injury at T2, whereas overlapping valine/proline resonances were most discriminatory between injury severities at T9. Plasma lactate correlated with blood-spinal cord barrier breakdown and plasma glucose with microglial density. We propose that peripheral biofluid metabolites can serve as biomarkers of SCI severity and associated pathology at the lesion site; their predictive value is most accurate when the injury level is also considered.
Role of B vitamins in modulating homocysteine and metabolic pathways linked to brain atrophy: Metabolomics insights from the VITACOG trial.
INTRODUCTION: Elevated total homocysteine (tHcy) is a major predictor of brain atrophy, cognitive decline, and Alzheimer's disease (AD) progression. The VITACOG trial, a randomized, placebo-controlled study in mild cognitive impairment (MCI), previously showed that B vitamin supplementation lowered tHcy, slowing brain atrophy and cognitive decline; however, the underlying mechanisms remained unclear. METHODS: We used untargeted, multi-platform metabolomics, with nuclear magnetic resonance and liquid chromatography-mass spectrometry to analyze serum samples from 89 B vitamin-treated and 84 placebo-treated MCI participants over a 2 year follow-up period. RESULTS: Multivariate modeling distinguished treated from placebo groups with 91.2 ± 1.8% accuracy. B vitamin supplementation induced significant metabolic reprogramming, lowering quinolinic acid, α-ketoglutarate, α-ketobutyrate, glucose, and glutamate. DISCUSSION: These findings reveal that B vitamins influence metabolic pathways beyond tHcy reduction, particularly the tricarboxylic acid cycle and glutamine-glutamate cycling, critical for brain energy homeostasis and neurotransmission. This metabolic signature supports B vitamin supplementation as a strategy for slowing MCI progression. HIGHLIGHTS: Nuclear magnetic resonance and multi-platform liquid chromatography tandem mass spectrometry metabolomics were performed on serum samples from 89 B vitamin-treated and 84 placebo participants in the VITACOG trial. Multi-platform metabolomics revealed B vitamin-driven metabolic reprogramming, achieving 91% classification accuracy. B vitamin supplementation modulates key neuroprotective metabolic pathways. Regulation of energy metabolism and neurotransmission by B vitamins contributes to brain health in elderly individuals. B vitamins demonstrate potential as an adjunct therapy in mild cognitive impairment, potentially mitigating progression to Alzheimer's disease.
Comparative Neuroplasticity in Frontal- and Lateral-Eyed Mammals With Induced-Binocular Vision Dysfunction: Insights From Monocular Deprivation Models.
Visual cortical plasticity during early postnatal life is profoundly shaped by species-specific ocular anatomy and ecological demands. This review synthesizes comparative evidence on how monocular deprivation (MD)-a classical model of amblyopia-affects visual system development in frontal- versus lateral-eyed mammals. Frontal-eyed species, including cats and primates, exhibit extensive binocular field overlaps and columnar architecture in the primary visual cortex (V1), making them highly susceptible to MD-induced shifts in ocular dominance and synaptic remodeling. In contrast, lateral-eyed species such as rodents and ungulates possess limited binocular overlaps and lack well-defined ocular dominance columns yet still demonstrate significant MD-induced plasticity involving callosal reorganization, glial activation, and extracellular matrix remodeling. We examine shared and divergent cellular mechanisms underpinning these responses, including the role of parvalbumin-expressing interneurons, perineuronal nets, and neuromodulators like BDNF and NRG1. Rodent models support the notion that even in the absence of classical columnar organization, lateral-eyed species can undergo region-specific structural remodeling in V1 following MD. These distinctions underscore how binocular integration circuits are fine-tuned through extended critical periods in frontal-eyed species, whereas plasticity in lateral-eyed species is more diffusely distributed. The integration of cross-species data revealed conserved principles of visual cortical plasticity and identified mechanisms potentially targetable for amblyopia therapy. Understanding the ecological and anatomical context of plasticity allows for a more accurate interpretation of animal models and supports the development of precision strategies for visual rehabilitation. This comparative framework expands the scope of amblyopia research and offers new avenues for translational interventions.
Elevated Acetylation of MFN2 is Accompanied by the Disruption of Mitochondrial Energy Metabolism and Inflammation in a Mouse Model of Depression.
Mitofusin-2 (MFN2) is recognized as an important regulator of mitochondrial function. The activity of MFN2 is increased by deacetylation, but while MFN2 levels have been reported to be increased in major depressive disorder, the relationship between acetylation status of MFN2, mitochondrial energy production, and inflammation in depression-like disease in rodents has not been studied. Here, we induced a depression-like syndrome in mice with a 14-day-long chronic restraint stress (CRS) model, and the levels of acetylated MFN2 and SIRT1 activity were measured. The interaction of MFN2 with complex I was identified by immunoprecipitation, and the levels of mitochondrial metabolites were measured by GC-MS. MFN2 levels were unaltered by CRS, but SIRT1 expression and activity were reduced in the CRS-exposed mice, and levels of acetylated MFN2 were significantly increased. CRS affected mitochondrial energy metabolism by reducing the expression and activity of complexes I-V, decreasing levels of NAD+ and ATP synthase, and diminishing ATP production. Thus, while the expression of Mfn2 was unchanged by CRS, the inhibition of MFN2 deacetylation, via loss of SIRT1 activity, was associated with impaired mitochondrial oxidative phosphorylation, increased oxidative stress markers, and increased levels of inflammatory markers under the control of the SIRT1 target NFκB. The results presented here highlight the profound influence of acetylation/deacetylation-mediated control associated with depression-like behaviors.
Thio-NHS esters are non-innocent protein acylating reagents.
N-Hydroxysuccinimide (NHS)-ester derivatives are widely used reagents in biological chemistry and chemical biology. Their efficacy relies critically on the exclusive chemoselectivity of activated acyl over that of the imidic acyl moieties in the succinimide. Here, through systematic structural variation that modulates acyl reactivity, coupled with a statistically controlled ultra-rapid screen for unknown modifications in tandem mass spectra as well as lysine profiling across complex lysine environments, including those within proteomes containing many thousands of proteins, we reveal that ring-opening to afford N-succinamide derivatives is a present, sometimes dominant, side-reaction. The extent of side-reaction is shown to be site-dependent, with side-reaction and desired reaction occurring within the same protein substrate. The resulting formation of bioconjugates with unintended, unstable linkages and modifications suggests the re-evaluation of: (i) known commercial reagents; and (ii) functional conclusions previously drawn using NHS esters in areas as diverse as antibody-drug biotherapy, vaccination and cross-link-enabled structural analyses.
Circadian rhythms in metabolism and mental health: a reciprocal regulatory network with implications for metabolic and neuropsychiatric disorders
Circadian rhythms orchestrate metabolism and brain function, aligning internal physiological processes with the 24-hour day–night cycle. Growing evidence highlights a reciprocal relationship between circadian regulation, metabolism, and neurobiological processes. Circadian disruption impairs glucose and lipid homeostasis, alters neurotransmitter and endocrine signalling, and triggers stress response, forming a feedback loop that impacts metabolism and brain function. These disturbances are implicated in many conditions, such as obesity, diabetes, depression, and bipolar disorder. This review examines recent advances in the interplay between circadian regulation, metabolism, and mental health, emphasising shared molecular mechanisms and their role in disease progression. Understanding these connections may ultimately inform therapeutic strategies that integrate circadian-based approaches to improve treatments for metabolic and psychiatric disorders.
Evidence that 5-HT2A receptor signalling efficacy and not biased agonism differentiates serotonergic psychedelic from non-psychedelic drugs.
BACKGROUND AND PURPOSE: Serotonergic psychedelic drugs are under investigation as therapies for various psychiatric disorders, including major depression. Although serotonergic psychedelic drugs are 5-HT2A receptor agonists, some such agonists are not psychedelic, potentially due to differences in 5-HT2A receptor ligand bias or signalling efficacy. Here, we investigated 5-HT2A receptor signalling properties of selected psychedelic and non-psychedelic drugs. EXPERIMENTAL APPROACH: Gq-coupled (Ca2+ and IP1) and β-arrestin2 signalling effects of six psychedelic drugs (psilocin, 5-MeO-DMT, LSD, mescaline, 25B-NBOMe and DOI) and three non-psychedelic drugs (lisuride, TBG and IHCH-7079) were characterised using SH-SY5Y cells expressing human 5-HT2A receptors. Ligand bias and signalling efficacy were measured using concentration-responses curves, compared with 5-HT. The generality of findings was tested using rat C6 cells which express endogenous 5-HT2A receptors. KEY RESULTS: In SH-SY5Y cells, all psychedelic drugs were partial agonists at both 5-HT2A receptor signalling pathways and none showed significant ligand bias. In comparison, the non-psychedelic drugs were not distinguishable from psychedelic drugs in terms of ligand bias properties but exhibited the lowest 5-HT2A receptor signalling efficacy of all drugs tested. The latter result was confirmed in C6 cells. CONCLUSION AND IMPLICATIONS: In summary, all psychedelic drugs tested were unbiased, partial 5-HT2A receptor agonists. Importantly, the non-psychedelic drugs lisuride, TBG and IHCH-7079 were discriminated from psychedelic drugs, not through ligand bias but rather by low efficacy. Therefore, low 5-HT2A receptor signalling efficacy may explain why some 5-HT2A receptor agonists are not psychedelic, although a larger panel of drugs should be tested to confirm this idea.
P21-Activated Kinase 2 as a Novel Target for Ventricular Tachyarrhythmias Associated with Cardiac Adrenergic Stress and Hypertrophy.
Ventricular arrhythmias associated with cardiac adrenergic stress and hypertrophy pose a significant clinical challenge. We explored ventricular anti-arrhythmic effects of P21-activated kinase 2 (Pak2), comparing in vivo and ex vivo cardiomyocyte-specific Pak2 knockout (Pak2cko) or overexpression (Pak2ctg) murine models, under conditions of acute adrenergic stress, and hypertrophy following chronic transverse aortic constriction (TAC). Pak2 was downregulated 5 weeks following the latter TAC challenge. Cellular physiological, optical action potential and Ca2+ transient, measurements, demonstrated increased incidences of triggered ventricular arrhythmias, and prolonged action potential durations (APD) and altered Ca2+ transients with increases in their beat-to beat variations, in Pak2cko hearts. Electron microscopic, proteomic, and molecular biological methods revealed a mitochondrial localization of stress-related proteins on proteomic and phosphoproteomic analyses, particularly in TAC stressed Pak2cko mice. They further yielded accompanying evidence for mitochondrial oxidative stress, increased reactive oxygen species (ROS) biosynthesis, reduced mitochondrial complexes I-V, diminished ATP synthesis and elevated NADPH oxidase 4 (NOX4) levels. Pak2 overexpression and the novel Pak2 activator JB2019A ameliorated these effects, enhanced cardiac function and decreased the frequencies of triggered ventricular arrhythmias. Pak2 activation thus protects against ventricular arrhythmia associated with cardiac stress and hypertrophy, through unique mechanisms offering potential novel therapeutic anti-arrhythmic targets.
p21-Activated Kinases Present a New Drug Target for Hypertrophic Cardiomyopathy
Hypertrophic cardiomyopathy (HCM), primarily involving mutations in sarcomeric proteins, is the most common form of inherited heart disease and a leading cause of sudden death in young adults and athletes. HCM patients present with cardiac hypertrophy, fibrosis, and diastolic dysfunction often in a progressive manner. Despite significant progress made in understanding the molecular genetic basis of HCM, there remains a lack of effective and specific treatment for preventing disease progression in HCM. This article first provides an overview of recent progress in understanding the pathogenic basis of disease progression in HCM, in particular dysfunctional calcium handling, mitochondrial impairment, and endoplasmic reticulum stress. This article then analyses the evidence for critical roles of the multifunctional enzymes P21-activated kinase-1 and 2 (Pak1/2) in the heart and our opinion on their therapeutic value as a promising druggable target in pathological hypertrophy and associated ventricular arrhythmias.