Traumatic brain injury (TBI) is a leading cause of human death and disability with no effective therapy to fully prevent long-term neurological deficits in surviving patients. Ketone ester supplementation is protective in animal models of neurodegeneration, but its efficacy against TBI pathophysiology is unknown. Here, we assessed the neuroprotective effect of the ketone monoester, 3-hydroxybutyl-3-hydroxybutyrate, (KE) in male Sprague Dawley rats (n=32). TBI was induced using the controlled cortical impact (CCI) with Sham animals not receiving the brain impact. KE was administered daily by oral gavage (0.5 ml/kg/day) and provided ad libitum at 0.3% (v/v) in the drinking water. KE supplementation started immediately after TBI and lasted for the duration of the study. Motor and sensory deficits were assessed using the Neurobehavioral Severity Scale-Revised (NSS-R) at four weeks post-injury. The NSS-R total score in CCI\u2009+\u2009KE (1.2\u2009\u00b1\u20090.4) was significantly lower than in CCI\u2009+\u2009water (4.4\u2009\u00b1\u20090.5). Similarly, the NSS-R motor scores in CCI\u2009+\u2009KE (0.6\u2009\u00b1\u20090.7) were significantly lower than CCI\u2009+\u2009water (2.9\u2009\u00b1\u20091.5). Although the NSS-R sensory score in the CCI\u2009+\u2009KE group (0.5\u2009\u00b1\u20090.2) was significantly lower compared to CCI\u2009+\u2009water (1.8\u2009\u00b1\u20090.4), no difference was observed between CCI\u2009+\u2009water and Sham\u2009+\u2009water (1.0\u2009\u00b1\u20090.2) groups. The lesion volume was smaller in the CCI\u2009+\u2009KE (10\u2009\u00b1\u20093 mm3) compared to CCI\u2009+\u2009water (47\u2009\u00b1\u200911 mm3; p\u2009<\u20090.001). KE significantly decreased Iba1+ stained areas in the cortex and hippocampus, and GFAP+ stained areas in all brain regions analyzed - prefrontal cortex, hippocampus, cortex, amygdala (p\u2009<\u20090.01). In summary, our results indicate that KE can protect against TBI-induced morphological and functional deficits when administered immediately after an insult.
\n \n\n \n \nInflammation is a central mechanism underlying numerous diseases and incorporates multiple known and potential future therapeutic targets. However, progress in developing novel immunomodulatory therapies has been slowed by a need for improvement in noninvasive biomarkers to accurately monitor the initiation, development and resolution of immune responses as well as their response to therapies. Hyperpolarized magnetic resonance imaging (MRI) is an emerging molecular imaging technique with the potential to assess immune cell responses by exploiting characteristic metabolic reprogramming in activated immune cells to support their function. Using specific metabolic tracers, hyperpolarized MRI can be used to produce detailed images of tissues producing lactate, a key metabolic signature in activated immune cells. This method has the potential to further our understanding of inflammatory processes across different diseases in human subjects as well as in preclinical models. This review discusses the application of hyperpolarized MRI to the imaging of inflammation, as well as the progress made towards the clinical translation of this emerging technique.
\n \n\n \n \nCardiac arrhythmias are among the leading causes of mortality. They often arise from alterations in the electrophysiological properties of cardiac cells, and their underlying ionic mechanisms. It is therefore critical to further unravel the patho-physiology of the ionic basis of human cardiac electrophysiology in health and disease. In the first part of this review, current knowledge on the differences in ion channel expression and properties of the ionic processes that determine the morphology and properties of cardiac action potentials and calcium dynamics from cardiomyocytes in different regions of the heart are described. Then the cellular mechanisms promoting arrhythmias in congenital or acquired conditions of ion channel function (electrical remodelling) are discussed. The focus is human relevant findings obtained with clinical, experimental and computational studies, given that interspecies differences make the extrapolation from animal experiments to the human clinical settings difficult. Deepening the understanding of the diverse patholophysiology of human cellular electrophysiology will help developing novel and effective antiarrhythmic strategies for specific subpopulations and disease conditions.
\n \n\n \n \nBackground: Kinase oxidation is a critical signalling mechanism through which changes in the intracellular redox state alter cardiac function. In the myocardium, type-1 protein kinase A (PKARI\u03b1) can be reversibly oxidized, forming interprotein disulfide bonds within the holoenzyme complex. However, the effect of PKARI\u03b1 disulfide formation on downstream signaling in the heart, particularly under states of oxidative stress such as ischemia and reperfusion (I/R), remains unexplored. Methods: Atrial tissue obtained from patients before and after cardiopulmonary bypass and reperfusion and left ventricular (LV) tissue from mice subjected to I/R or sham surgery were used to assess PKARI\u03b1 disulfide formation by immunoblot. To determine the impact of disulfide formation on PKARI\u03b1 catalytic activity and sub-cellular localization, live-cell fluorescence imaging and stimulated emission depletion super-resolution microscopy were performed in prkar1 knock-out mouse embryonic fibroblasts, neonatal myocytes or adult LV myocytes isolated from 'redox dead' (Cys17Ser) PKARI\u03b1 knock-in mice and their wild-type littermates. Comparison of intracellular calcium dynamics between genotypes was assessed in fura2-loaded LV myocytes whereas I/R-injury was assessed ex vivo. Results: In both humans and mice, myocardial PKARI\u03b1 disulfide formation was found to be significantly increased (2-fold in humans, p=0.023; 2.4-fold in mice, p<0.001) in response to I/R in vivo. In mouse LV cardiomyocytes, disulfide-containing PKARI\u03b1 was not found to impact catalytic activity, but instead led to enhanced A-kinase-anchoring protein (AKAP) binding with preferential localization of the holoenzyme to the lysosome. Redox-dependent regulation of lysosomal two pore channels (TPC) by PKARI\u03b1 was sufficient to prevent global calcium release from the sarcoplasmic reticulum in LV myocytes, without affecting intrinsic ryanodine receptor leak or phosphorylation. Absence of I/R-induced PKARI\u03b1 disulfide formation in \"redox dead\" knock-in mouse hearts resulted in larger infarcts (2-fold, p<0.001) and a concomitant reduction in LV contractile recovery (1.6-fold, p<0.001), which was prevented by administering the lysosomal TPC inhibitor Ned-19 at the time of reperfusion. Conclusions: Disulfide-modification targets PKARI\u03b1 to the lysosome where it acts as a gatekeeper for TPC-mediated triggering of global calcium release. In the post-ischemic heart, this regulatory mechanism is critical for protecting from extensive injury and offers a novel target for the design of cardioprotective therapeutics.
\n \n\n \n \nNeurohumoral activation is an early hallmark of cardiovascular disease and contributes to the etiology of the pathophysiology. Stellectomy has reemerged as a positive therapeutic intervention to modify the progression of dysautonomia, although the biophysical properties underpinning abnormal activity of this ganglia are not fully understood in the initial stages of the disease. We investigated whether stellate ganglia neurons from prehypertensive SHRs (spontaneously hypertensive rats) are hyperactive and describe their electrophysiological phenotype guided by single-cell RNA sequencing, molecular biology, and perforated patch clamp to uncover the mechanism of abnormal excitability. We demonstrate the contribution of a plethora of ion channels, in particular inhibition of M current to stellate ganglia neuronal firing, and confirm the conservation of expression of key ion channel transcripts in human stellate ganglia. We show that hyperexcitability was curbed by M-current activators, nonselective sodium current blockers, or inhibition of Nav1.1-1.3, Nav1.6, or INaP. We conclude that reduced activity of M current contributes significantly to abnormal firing of stellate neurons, which, in part, contributes to the hyperexcitability from rats that have a predisposition to hypertension. Targeting these channels could provide a therapeutic opportunity to minimize the consequences of excessive sympathetic activation.
\n \n\n \n \nPurpose: Phosphorus saturation-transfer experiments can quantify metabolic\nfluxes non-invasively. Typically, the forward flux through the creatine-kinase\nreaction is investigated by observing the decrease in phosphocreatine (PCr)\nafter saturation of $\\gamma$-ATP. The quantification of total ATP utilisation\nis currently under-explored, as it requires simultaneous saturation of\ninorganic phosphate (Pi) and PCr. This is challenging, as currently available\nsaturation pulses reduce the already-low $\\gamma$-ATP signal present.\n Methods: Using a hybrid optimal-control and Shinnar-Le-Roux method, a\nquasi-adiabatic RF pulse was designed for the dual-saturation of PCr and Pi to\nenable determination of total ATP utilisation. The pulses were evaluated in\nBloch equation simulations, compared with a conventional hard-cosine DANTE\nsaturation sequence, before application to perfused rat hearts at 11.7 Tesla.\n Results: The quasi-adiabatic pulse was insensitive to a $>2.5$-fold variation\nin $B_1$, producing equivalent saturation with a 53% reduction in delivered\npulse power and a 33-fold reduction in spillover at the minimum effective\n$B_1$. This enabled the complete quantification of the synthesis and\ndegradation fluxes for ATP in 30-45 minutes in the perfused rat heart. While\nthe net synthesis flux ($3.3\\pm0.4$ mM/s, SEM) was not significantly different\nfrom degradation flux ($8\\pm2$ mM/s) and both measures are consistent with\nprior work, nonlinear error analysis highlights uncertainties in the Pi-to-ATP\nmeasurement that may explain the possible imbalance.\n Conclusion: This work demonstrates a novel quasi-adiabatic dual-saturation RF\npulse with significantly improved performance that can be used to measure ATP\nturnover in the heart in vivo.
\n \n\n \n \nResearch into potential targets for cardiac repair encompasses recognition of tissue-resident cells with intrinsic regenerative properties. The adult vertebrate heart is covered by mesothelium, named the epicardium, which becomes active in response to injury and contributes to repair, albeit suboptimally. Motivation to manipulate the epicardium for treatment of myocardial infarction is deeply rooted in its central role in cardiac formation and vasculogenesis during development. Moreover, the epicardium is vital to cardiac muscle regeneration in lower vertebrate and neonatal mammalian-injured hearts. In this review, we discuss our current understanding of the biology of the mammalian epicardium in development and injury. Considering present challenges in the field, we further contemplate prospects for reinstating full embryonic potential in the adult epicardium to facilitate cardiac regeneration.
\n \n\n \n \nProcessing in the sensory periphery involves various mechanisms that enable the detection and discrimination of sensory information. Despite their biological complexity, could these processing steps sub-serve a relatively simple transformation of sensory inputs, which are then transmitted to the CNS? Here we explored both biologically-detailed and very simple models of the auditory periphery to find the appropriate input to a phenomenological model of auditory cortical responses to natural sounds. We examined a range of cochlear models, from those involving detailed biophysical characteristics of the cochlea and auditory nerve to very pared-down spectrogram-like approximations of the information processing in these structures. We tested the capacity of these models to predict the time-course of single-unit neural responses recorded in the ferret primary auditory cortex, when combined with a linear non-linear encoding model. We show that a simple model based on a log-spaced, log-scaled power spectrogram with Hill-function compression performs as well as biophysically-detailed models of the cochlea and the auditory nerve. These findings emphasize the value of using appropriate simple models of the periphery when building encoding models of sensory processing in the brain, and imply that the complex properties of the auditory periphery may together result in a simpler than expected functional transformation of the inputs.
\n \n\n \n \n<jats:p>The sequential activation of neurons has been observed in various areas of the brain, but in no case is the underlying network structure well understood. Here we examined the circuit anatomy of zebra finch HVC, a cortical region that generates sequences underlying the temporal progression of the song. We combined serial block-face electron microscopy with light microscopy to determine the cell types targeted by HVC(RA) neurons, which control song timing. Close to their soma, axons almost exclusively targeted inhibitory interneurons, consistent with what had been found with electrical recordings from pairs of cells. Conversely, far from the soma the targets were mostly other excitatory neurons, about half of these being other HVC(RA) cells. Both observations are consistent with the notion that the neural sequences that pace the song are generated by global synaptic chains in HVC embedded within local inhibitory networks.</jats:p>
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