In vitro effects of wound-dressings on key wound healing properties of dermal fibroblasts.

Healing of complex wounds requires dressings that must, at least, not hinder and should ideally promote the activity of key healing cells, in particular fibroblasts. This in vitro study assessed the effects of three wound-dressings (a pure Ca2+ alginate: Algostéril®, a Ca2+ alginate + carboxymethylcellulose: Biatain alginate® and a polyacrylate impregnated with lipido-colloid matrix: UrgoClean®) on dermal fibroblast activity. The results showed the pure calcium alginate to be non-cytotoxic, whereas the other wound-dressings showed moderate to strong cytotoxicity. The two alginates stimulated fibroblast migration and proliferation, whereas the polyacrylate altered migration and had no effect on proliferation. The pure Ca2+ alginate significantly increased the TGF-ß-induced fibroblast activation, which is essential to healing. This activation was confirmed by a significant increase in Vascular endothelial growth factor (VEGF) secretion and a higher collagen production. The other dressings reduced these fibroblast activities. The pure Ca2+ alginate was also able to counteract the inhibitory effect of NK cell supernatants on fibroblast migration. These in vitro results demonstrate that tested wound-dressings are not equivalent for fibroblast activation. Only Algostéril was found to promote all the fibroblast activities tested, which could contribute to its healing efficacy demonstrated in the clinic.

High-performance serial block-face SEM of nonconductive biological samples enabled by focal gas injection-based charge compensation.

A longstanding limitation of imaging with serial block-face scanning electron microscopy is specimen surface charging. This charging is largely due to the difficulties in making biological specimens and the resins in which they are embedded sufficiently conductive. Local accumulation of charge on the specimen surface can result in poor image quality and distortions. Even minor charging can lead to misalignments between sequential images of the block-face due to image jitter. Typically, variable-pressure SEM is used to reduce specimen charging, but this results in a significant reduction to spatial resolution, signal-to-noise ratio and overall image quality. Here we show the development and application of a simple system that effectively mitigates specimen charging by using focal gas injection of nitrogen over the sample block-face during imaging. A standard gas injection valve is paired with a precisely positioned but retractable application nozzle, which is mechanically coupled to the reciprocating action of the serial block-face ultramicrotome. This system enables the application of nitrogen gas precisely over the block-face during imaging while allowing the specimen chamber to be maintained under high vacuum to maximise achievable SEM image resolution. The action of the ultramicrotome drives the nozzle retraction, automatically moving it away from the specimen area during the cutting cycle of the knife. The device described was added to a Gatan 3View system with minimal modifications, allowing high-resolution block-face imaging of even the most charge prone of epoxy-embedded biological samples.

Altered fMRI resting-state connectivity in individuals with fibromyalgia on acute pain stimulation.

BACKGROUND: Fibromyalgia is a chronic widespread pain condition, with patients commonly reporting other symptoms such as sleep difficulties, memory complaints and fatigue. The use of magnetic resonance imaging (MRI) in fibromyalgia has allowed for the detection of neural abnormalities, with alterations in brain activation elicited by experimental pain and alterations in resting state connectivity related to clinical pain. METHODS: In this study, we sought to monitor state changes in resting brain connectivity following experimental pressure pain in fibromyalgia patients and healthy controls. Twelve fibromyalgia patients and 15 healthy controls were studied by applying discrete pressure stimuli to the thumbnail bed during MRI. Resting-state functional MRI scanning was performed before and immediately following experimental pressure pain. We investigated changes in functional connectivity to the thalamus and the insular cortex. RESULTS: Acute pressure pain increased insula connectivity to the anterior cingulate and the hippocampus. Additionally, we observed increased thalamic connectivity to the precuneus/posterior cingulate cortex, a known part of the default mode network, in patients but not in controls. This connectivity was correlated with changes in clinical pain. CONCLUSIONS: These data reporting changes in resting-state brain activity following a noxious stimulus suggest that the acute painful stimuli may contribute to the alteration of the neural signature of chronic pain. WHAT DOES THIS STUDY/ADD?: In this study acute pain application shows an echo in functional connectivity and clinical pain changes in chronic pain.

Resting state connectivity correlates with drug and placebo response in fibromyalgia patients.

Fibromyalgia is a chronic pain syndrome characterized by widespread pain, fatigue, and memory and mood disturbances. Despite advances in our understanding of the underlying pathophysiology, treatment is often challenging. New research indicates that changes in functional connectivity between brain regions, as can be measured by magnetic resonance imaging (fcMRI) of the resting state, may underlie the pathogenesis of this and other chronic pain states. As such, this parameter may be able to be used to monitor changes in brain function associated with pharmacological treatment, and might also be able to predict treatment response. We performed a resting state fcMRI trial using a randomized, placebo-controlled, cross-over design to investigate mechanisms of action of milnacipran (MLN), a selective serotonin and norepinephrine reuptake inhibitor (SNRI), in fibromyalgia patients. Our aim was to identify functional connectivity patterns at baseline that would differentially predict treatment response to MLN as compared to placebo. Since preclinical studies of MLN suggest that this medication works by augmenting antinociceptive processes, we specifically investigated brain regions known to be involved in pain inhibition. 15 fibromyalgia patients completed the study, consisting of 6 weeks of drug and placebo intake (order counterbalanced) with an interspersed 2 week wash out period. As a main finding we report that reductions in clinical pain scores during MLN were associated with decreased functional connectivity between pro-nociceptive regions and antinociceptive pain regions at baseline, specifically between the rostral part of the anterior cingulate cortex (ACC) and the insular cortex (IC), as well as between the periaqueductal gray (PAG) and the IC: patients with lower preexisting functional connectivity had the greatest reduction in clinical pain. This pattern was not observed for the placebo period. However a more robust placebo response was associated with lower baseline functional connectivity between the ACC and the dorsolateral prefrontal cortex. This study indicates that ACC-IC connectivity might play a role in the mechanism of action of MLN, and perhaps more importantly fcMRI might be a useful tool to predict pharmacological treatment response.

The autism brain imaging data exchange: towards a large-scale evaluation of the intrinsic brain architecture in autism.

Autism spectrum disorders (ASDs) represent a formidable challenge for psychiatry and neuroscience because of their high prevalence, lifelong nature, complexity and substantial heterogeneity. Facing these obstacles requires large-scale multidisciplinary efforts. Although the field of genetics has pioneered data sharing for these reasons, neuroimaging had not kept pace. In response, we introduce the Autism Brain Imaging Data Exchange (ABIDE)-a grassroots consortium aggregating and openly sharing 1112 existing resting-state functional magnetic resonance imaging (R-fMRI) data sets with corresponding structural MRI and phenotypic information from 539 individuals with ASDs and 573 age-matched typical controls (TCs; 7-64 years) (http://fcon_1000.projects.nitrc.org/indi/abide/). Here, we present this resource and demonstrate its suitability for advancing knowledge of ASD neurobiology based on analyses of 360 male subjects with ASDs and 403 male age-matched TCs. We focused on whole-brain intrinsic functional connectivity and also survey a range of voxel-wise measures of intrinsic functional brain architecture. Whole-brain analyses reconciled seemingly disparate themes of both hypo- and hyperconnectivity in the ASD literature; both were detected, although hypoconnectivity dominated, particularly for corticocortical and interhemispheric functional connectivity. Exploratory analyses using an array of regional metrics of intrinsic brain function converged on common loci of dysfunction in ASDs (mid- and posterior insula and posterior cingulate cortex), and highlighted less commonly explored regions such as the thalamus. The survey of the ABIDE R-fMRI data sets provides unprecedented demonstrations of both replication and novel discovery. By pooling multiple international data sets, ABIDE is expected to accelerate the pace of discovery setting the stage for the next generation of ASD studies.

Functional connectivity of human chewing: an fcMRI study.

Mastication is one of the most important orofacial functions. The neurobiological mechanisms of masticatory control have been investigated in animal models, but less so in humans. This project used functional connectivity magnetic resonance imaging (fcMRI) to assess the positive temporal correlations among activated brain areas during a gum-chewing task. Twenty-nine healthy young-adults underwent an fcMRI scanning protocol while they chewed gum. Seed-based fcMRI analyses were performed with the motor cortex and cerebellum as regions of interest. Both left and right motor cortices were reciprocally functionally connected and functionally connected with the post-central gyrus, cerebellum, cingulate cortex, and precuneus. The cerebellar seeds showed functional connections with the contralateral cerebellar hemispheres, bilateral sensorimotor cortices, left superior temporal gyrus, and left cingulate cortex. These results are the first to identify functional central networks engaged during mastication.

Vitamin D metabolism, functions and needs: from science to health claims.

BACKGROUND: Vitamin D is a nutrient long considered as essential for skeletal health but is now attracting interest from medical and nutritional communities as knowledge emerges of its biological function and its association with decreased risk of many chronic diseases. RESULTS: A question emerges: How much more vitamin D do we need for these new functions of vitamin D? This review discusses vitamin D physiology and hypovitaminosis D and presents two vitamin D dietary policies: that according to regulatory authorities and that of nutrition scientists. Scientific evidence suggests that 25(OH)D serum levels should be over 75 nmol/L; otherwise, there is no beneficial effect of vitamin D on long-latency diseases. Current regulatory authority recommendations are insufficient to reach this level of adequacy. Observational and some prospective data show that vitamin D has a role in the prevention of cancer as well as immunity, diabetes and cardiovascular and muscle disorders, which supports the actions of 1α,25(OH)2D at cellular and molecular levels. The recent assessments done by the European Food Safety Authority should lead to new health claims. CONCLUSIONS: Vitamin D, through food fortification and supplementation, is a promising new health strategy and thus provides opportunities for food industry and nutrition researchers to work together towards determining how to achieve this potential health benefit.

L-DOPA changes spontaneous low-frequency BOLD signal oscillations in Parkinson's disease: a resting state fMRI study.

Analysis of the amplitude of low frequency BOLD signal fluctuations (ALFF) in the resting state has recently been used to study the dynamics of intrinsic neural activity. Several studies have also suggested its potential as a biomarker for neuropsychiatric disease. In the current study, we quantified ALFF to determine changes in intrinsic neural oscillations in patients with Parkinson's disease (PD) on and off L-DOPA. Twenty-four PD patients and 24 healthy age-matched controls participated in the study. PD patients underwent two resting state fMRI sessions, either ON a controlled dose of L-DOPA or following a placebo pill (OFF). Control participants underwent one test session. We found that there was increased amplitude of low frequency BOLD signal oscillations for PD patients OFF L-DOPA in the primary and secondary motor areas, and in the middle and medial prefrontal cortices. L-DOPA significantly reduced the amplitude of low frequency oscillations within these regions. The degree of ALFF in the premotor cortex predicted patients' motor performance as measured by the Grooved Pegboard task, such that greater ALFF was associated with poorer performance. These results are in line with the pathophysiology of PD, which shows changes in neural oscillations. Thus, frequency domain analyses of resting state BOLD fMRI signals may provide a useful means to study the pathophysiology of PD and the physiology of the brain's dopaminergic pathways.

An increase in the redox state during reperfusion contributes to the cardioprotective effect of GIK solution.

This study aimed at determining whether glucose-insulin-potassium (GIK) solutions modify the NADH/NAD(+) ratio during postischemic reperfusion and whether their cardioprotective effect can be attributed to this change in part through reduction of the mitochondrial reactive oxygen species (ROS) production. The hearts of 72 rats were perfused with a buffer containing glucose (5.5 mM) and hexanoate (0.5 mM). They were maintained in normoxia for 30 min and then subjected to low-flow ischemia (0.5% of the preischemic coronary flow for 20 min) followed by reperfusion (45 min). From the beginning of ischemia, the perfusate was subjected to various changes: enrichment with GIK solution, enrichment with lactate (2 mM), enrichment with pyruvate (2 mM), enrichment with pyruvate (2 mM) plus ethanol (2 mM), or no change for the control group. Left ventricular developed pressure, heart rate, coronary flow, and oxygen consumption were monitored throughout. The lactate/pyruvate ratio of the coronary effluent, known to reflect the cytosolic NADH/NAD(+) ratio and the fructose-6-phosphate/dihydroxyacetone-phosphate (F6P/DHAP) ratio of the reperfused myocardium, were evaluated. Mitochondrial ROS production was also estimated. The GIK solution improved the recovery of mechanical function during reperfusion. This was associated with an enhanced cytosolic NADH/NAD(+) ratio and reduced mitochondrial ROS production. The cardioprotection was also observed when the hearts were perfused with fluids known to increase the cytosolic NADH/NAD(+) ratio (lactate, pyruvate plus ethanol) compared with the other fluids (control and pyruvate groups). The hearts with a high mechanical recovery also displayed a low F6P/DHAP ratio, suggesting that an accelerated glycolysis rate may be responsible for increased cytosolic NADH production. In conclusion, the cardioprotection induced by GIK solutions could occur through an increase in the cytosolic NADH/NAD(+) ratio, leading to a decrease in mitochondrial ROS production.

Age differences in symbolic representations of motor sequence learning.

We recently reported that young adults (YA) preferentially recruit cerebellar lobule HVI for symbolic motor sequence learning [3]. Learning magnitude in the symbolic condition was correlated with activation level in lobule HVI. Here, we evaluated age differences in the symbolic representation of motor sequence learning. Fourteen YA and 14 older adults (OA) performed the alternating serial reaction time task (ASRT) under conditions in which the spatial processing component was selectively eliminated from stimulus presentation (spatial versus symbolic), response execution (manual versus vocal), or both. Results showed that OA had reduced learning magnitudes relative to YA. Using the cerebellum lobule HVI as a region-of-interest, we found that OA had significantly lower activation in this region than YA during the symbolic learning conditions (FWE, P<0.05). Similar to YA, OA also showed a significant correlation between learning magnitude and cerebellar activation in the symbolic conditions. These results suggest that although YA and OA recruit similar neural networks during implicit learning, OA under-recruit relevant brain areas which may partially explain their implicit sequence learning deficits.

Symbolic representations in motor sequence learning.

It has been shown that varying the spatial versus symbolic nature of stimulus presentation and response production, which affects stimulus-response (S-R) mapping requirements, influences the magnitude of implicit sequence learning (Koch and Hoffman, 2000). Here, we evaluated how spatial and symbolic stimuli and responses affect the neural bases of sequence learning. We selectively eliminated the spatial component of stimulus presentation (spatial vs. symbolic), response execution (manual vs. vocal), or both. Fourteen participants performed the alternating serial reaction time task under these conditions in an MRI scanner, with interleaved acquisition to allow for recording of vocal response reaction times. Nine regions of interest (ROIs) were selected to test the hypothesis that the dorsolateral prefrontal cortex (DLPFC) was preferentially engaged for spatially cued conditions and cerebellum lobule HVI, crus I and II were associated with symbolically cued learning. We found that the left cerebellum lobule HVI was selectively recruited for symbolic learning and the percent signal change in this region was correlated with learning magnitude under the symbolic conditions. In contrast, the DLPFC did not exhibit selective activation for learning under spatial conditions. The inferior parietal lobule exhibited increased activation during learning regardless of the condition, supporting its role in forming an abstract representation of learned sequences. These findings reveal different brain networks that are flexibly engaged depending on the conditions of sequence learning.

The OptIPuter microscopy demonstrator: enabling science through a transatlantic lightpath.

The OptIPuter microscopy demonstrator project has been designed to enable concurrent and remote usage of world-class electron microscopes located in Oxford and San Diego. The project has constructed a network consisting of microscopes and computational and data resources that are all connected by a dedicated network infrastructure using the UK Lightpath and US Starlight systems. Key science drivers include examples from both materials and biological science. The resulting system is now a permanent link between the Oxford and San Diego microscopy centres. This will form the basis of further projects between the sites and expansion of the types of systems that can be remotely controlled, including optical, as well as electron, microscopy. Other improvements will include the updating of the Microsoft cluster software to the high performance computing (HPC) server 2008, which includes the HPC basic profile implementation that will enable the development of interoperable clients.

Rapid reduction of liver steatosis in omega3-depleted rats injected with a novel lipid emulsion.

The bolus intravenous administration of a novel medium-chain triglyceride: fish oil emulsion (MCT:FO) to normal subjects was recently found to increase within 60 min the amount of long-chain polyunsaturated omega3 fatty acids ( omega3) in platelet and leukocyte phospholipids and, hence, was proposed as a tool to prevent such pathological events as cardiac arrhythmia in selected patients who have to undergo urgent anesthesia and/or surgery. This study investigates whether other cells located outside the vascular bed may also benefit from this procedure for replenishing phospholipids with omega3. For such a purpose, the MCT:FO emulsion (1.0 ml) was injected into normal or omega3-depleted rats examined, one hour later, for the content and fatty acid pattern of liver triglycerides and phospholipids. Control experiments included the administration of saline or a medium-chain triglyceride:olive oil emulsion. The results reveal that the bolus intravenous injection of MCT:FO to the omega3-depleted rats resulted in the enrichment of liver phospholipids in omega3 and a marked reduction in hepatic steatosis. In conclusion, it is proposed that such a procedure may indeed allow a rapid supply of omega3 not only to circulating and vascular endothelial cells but also to extravascular cells, with a resulting correction of the biochemical and biophysical defects linked to a deficiency in these fatty acids.

Cardioprotective effect of spontaneous activity.

In the perspective of giving a better understanding of the cardioprotective effects attributable to the tandem low caloric intake and training, Lou/C rats would be an interesting model since these animals exhibit spontaneously these two characteristics for months, without any dietary manipulations or stressor stimuli. No information was so far available on their cardiac function. Therefore, the aim of this pilot study was (i) to document cardiac function before and after ischemia in this strain, and (ii) to investigate whether spontaneous wheel-running activity can improve the ability of cardiac muscle to recover its function after an ischemic period. Cardiac mechanical and metabolic functions were measured in isolated Langendorff hearts from Wistar sedentary, Lou/C sedentary, and Lou/C wheel-running male rats submitted to a 20-min low-flow ischemia and 20-min reperfusion. In Lou/C sedentary rats, rate-pressure product, an index of cardiac work, was decreased before ischemia as compared to Wistar sedentary animals (- 24 %, p < 0.05). After ischemia, cardiac mechanical function recovery did not significantly differ between these two groups. Nevertheless, flux of non-oxidative glycolysis was lower before and after ischemia in Lou/C sedentary animals than in Wistar sedentary rats. In Lou/C rats, during normoxic perfusion, wheel-running activity significantly decreased heart rate (- 15 %), oxygen consumption (- 2.2 %) and cardiac efficiency (- 37 %), whereas coronary flow and flux of non-oxidative glycolysis were significantly increased (+ 15 % and + 263 %, respectively). After ischemia, recovery of cardiac mechanical function and cardiac efficiency were improved in Lou/C wheel-running rats versus Lou/C sedentary animals (p < 0.05). In conclusion, the impact of ischemia-reperfusion is similar between Lou/C- and Wistar sedentary rats. Spontaneous wheel-running activity decreases cardiac efficiency before ischemia and confers a protection against ischemia- and reperfusion-induced injury in isolated Lou/C rat hearts.

Accelerated clearance of plasma triglycerides and free fatty acids in omega3-depleted rats.

The present study aims mainly at exploring the effects of a severe depletion in polyunsaturated long-chain omega3 fatty acids upon the fate of circulating lipids. The plasma concentration and fatty acid pattern of triglycerides, diglycerides, free fatty acids, and phospholipids were measured in omega3-depleted and control rats injected intravenously one hour before sacrifice with either saline, a control medium-chain triglyceride:olive oil emulsion or a medium-chain triglyceride:fish oil emulsion recently found to rapidly increase the phospholipid content of C20:5omega3 and C22:6omega3 in different cell types. The estimated fractional removal rate of the injected triglycerides and the clearance of free fatty acids from circulation were both higher in omega3-depleted rats than in control animals. The injection of the lipid emulsions apparently inhibited intracellular lipolysis, this being least pronounced in omega3-depleted rats. The increased clearance of circulating triglycerides and unesterified fatty acids in omega3-depleted rats may favor the cellular accumulation of lipids. In turn, such an accumulation and the lesser regulatory inhibition of tissular lipolysis may match the increased clearance of circulating unesterified fatty acids and, hence, account for the lack of any significant difference in plasma unesterified fatty acid concentration between these and control animals.

High dietary sucrose triggers hyperinsulinemia, increases myocardial beta-oxidation, reduces glycolytic flux and delays post-ischemic contractile recovery.

Although the causal relationship between insulin resistance (IR) and hypertension is not fully resolved, the importance of IR in cardiovascular dysfunction is recognized. As IR may follow excess sucrose or fructose diet, the aim of this study was to test whether dietary starch substitution with sucrose results in myocardial dysfunction in energy substrate utilization and contractility during normoxic and post-ischemic conditions. Forty-eight male Wistar rats were randomly allocated to three diets, differing only in their starch to sucrose (S) ratio (13, 2 and 0 for the Low S, Middle S and High S groups, respectively), for 3 weeks. Developed pressure and rate x pressure product (RPP) were determined in Langendorff mode-perfused hearts. After 30 min stabilization, hearts were subjected to 25 min of total normothermic global ischemia, followed by 45-min reperfusion. Oxygen consumption, beta-oxidation rate (using 1-13C hexanoate and Isotopic Ratio Mass Spectrometry of CO2 produced in the coronary effluent) and flux of non-oxidative glycolysis were also evaluated. Although fasting plasma glucose levels were not affected by increased dietary sucrose, high sucrose intake resulted in increased plasma insulin levels, without significant rise in plasma triglyceride and free fatty acid concentrations. Sucrose-rich diet reduced pre-ischemic baseline measures of heart rate, RPP and non-oxidative glycolysis. During reperfusion, post-ischemic recovery of RPP was impaired in the Middle S and High S groups, as compared to Low S, mainly due to delayed recovery of developed pressure, which by 45 min of reperfusion eventually resumed levels matching Low S. At the start of reperfusion, delayed post-ischemic recovery of contractile function was accompanied by: (i) reduced lactate production; (ii) decreased lactate to pyruvate ratio; (iii) increased beta-oxidation; and (iv) depressed metabolic efficiency. In conclusion, sucrose rich-diet increased plasma insulin levels, in intact rat, and increased cardiac beta-oxidation and coronary flow-rate, but reduced glycolytic flux and contractility during normoxic baseline function of isolated perfused hearts. Sucrose rich-diet impaired early post-ischemic recovery of isolated heart cardiac mechanical function and further augmented cardiac beta-oxidation but reduced glycolytic and lactate flux.

Visual cortex activation in kinesthetic guidance of reaching.

The purpose of this research was to determine the cortical circuit involved in encoding and controlling kinesthetically guided reaching movements. We used (15)O-butanol positron emission tomography in ten blindfolded able-bodied volunteers in a factorial experiment in which arm (left/right) used to encode target location and to reach back to the remembered location and hemispace of target location (left/right side of midsagittal plane) varied systematically. During encoding of a target the experimenter guided the hand to touch the index fingertip to an external target and then returned the hand to the start location. After a short delay the subject voluntarily moved the same hand back to the remembered target location. SPM99 analysis of the PET data contrasting left versus right hand reaching showed increased (P < 0.05, corrected) neural activity in the sensorimotor cortex, premotor cortex and posterior parietal lobule (PPL) contralateral to the moving hand. Additional neural activation was observed in prefrontal cortex and visual association areas of occipital and parietal lobes contralateral and ipsilateral to the reaching hand. There was no statistically significant effect of target location in left versus right hemispace nor was there an interaction of hand and hemispace effects. Structural equation modeling showed that parietal lobe visual association areas contributed to kinesthetic processing by both hands but occipital lobe visual areas contributed only during dominant hand kinesthetic processing. This visual processing may also involve visualization of kinesthetically guided target location and use of the same network employed to guide reaches to visual targets when reaching to kinesthetic targets. The present work clearly demonstrates a network for kinesthetic processing that includes higher visual processing areas in the PPL for both upper limbs and processing in occipital lobe visual areas for the dominant limb.

Acute in vivo administration of a fish oil-containing emulsion improves post-ischemic cardiac function in n-3-depleted rats.

A novel i.v. lipid preparation (MCT:FO) containing 80% medium chain-triacylglycerols and 20% fish oil was recently developed to rapidly replenish cell membrane phospholipids with omega 3 (n-3) polyunsaturated fatty acids (PUFA). In regard of this property, we investigated the effect of a single i.v. administration of MCT:FO on the recovery of cardiac function after ischemia in control and n-3-depleted rats. Results were compared with those obtained either with a control preparation, where FO was replaced by triolein (MCT:OO), or with saline. Saline (1 ml) or lipid preparation (also 1 ml) was injected as a bolus via the left saphenous vein. After 60 min the heart was removed and perfused for 20 min in normoxic conditions according to Langendorff. Thereafter, the heart was subjected to a 20 min zero-flow normothermic ischemia, followed by 40 min reperfusion. Cardiac mechanical and metabolic functions were monitored. In control rats, the previous administration of a lipid preparation (MCT:FO or MCT:OO) versus saline improved cardiac function during aerobic reperfusion post-ischemia. N-3-depleted rats showed decreased basal cardiac function and impaired recovery following ischemia. However, the bolus injection of MCT:FO opposed the deleterious effect of long-term n-3-deficiency and, in this respect, was superior to MCT:OO over the first 20 min of reperfusion. This novel approach to rapidly correct n-3 PUFA-deficiency might be clinically relevant and offer interesting perspectives in the management of acute ischemic accidents.

Automated microscopy system for mosaic acquisition and processing.

An automatic mosaic acquisition and processing system for a multiphoton microscope is described for imaging large expanses of biological specimens at or near the resolution limit of light microscopy. In a mosaic, a larger image is created from a series of smaller images individually acquired systematically across a specimen. Mosaics allow wide-field views of biological specimens to be acquired without sacrificing resolution, providing detailed views of biological specimens within context. The system is composed of a fast-scanning, multiphoton, confocal microscope fitted with a motorized, high-precision stage and custom-developed software programs for automatic image acquisition, image normalization, image alignment and stitching. Our current capabilities allow us to acquire data sets comprised of thousands to tens of thousands of individual images per mosaic. The large number of individual images involved in creating a single mosaic necessitated software development to automate both the mosaic acquisition and processing steps. In this report, we describe the methods and challenges involved in the routine creation of very large scale mosaics from brain tissue labelled with multiple fluorescent probes.

Effect of exogenous adenosine and monensin on glycolytic flux in isolated perfused normoxic rat hearts: role of pyruvate kinase.

We studied the effect of exogenous adenosine in isolated perfused normoxic rat hearts on glycolytic flux through pyruvate kinase (PK). We compared its effect with that of myxothiazol, an inhibitor of mitochondrial ATP production. Moreover, we tested whether an increase of membrane ionic flux with monensin is linked to a stimulation of glycolytic flux through PK. After a 20-min stabilization period adenosine, myxothiazol or monensin were administrated to the perfusate continuously at various concentrations during 10 min. The contraction was monitored and the lactate production in coronary effluents evaluated. The amount of adenine nucleotides and phosphoenolpyruvate was measured in the frozen hearts. Myxothiazol induced a decrease of the left ventricular developed pressure (LVDP : -40%) together with a stimulation of glycolytic flux secondary to PK activation. In contrast, adenosine primarily reduced heart rate (HR: -30%) with only marginal effects on LVDP. This was associated with an inhibition of glycolysis at the level of PK. The Na+ ionophore monensin affected HR (+14%) and LVDP (+25%). This effect was associated with a stimulation of glycolysis secondary to the stimulation of PK. These results provide new information of action of adenosine in the heart and support the concept of a direct coupling between glycolysis and process regulating sarcolemmal ionic fluxes.