Regional-specific average time courses of spontaneous fluctuations in blood oxygen level dependent (BOLD) MRI contrast at 9.4T in lightly anesthetized resting rat brain are formed, and correlation coefficients between time course pairs are interpreted as measures of connectivity. A hierarchy of regional pairwise correlation coefficients (RPCCs) is observed, with the highest values found in the thalamus and cortex, both intra-and interhemisphere, and lower values between the cortex and thalamus. Independent sensory networks are distinguished by two methods: data driven, where task activation defines regions of interest (ROI), and hypothesis driven, where regions are defined by the rat histological atlas. Success in these studies is attributed in part to the use of medetomidine hydrochloride (Domitor) for anesthesia. Functional connectivity in the resting human brain using blood oxygen level dependent (BOLD) contrast is revealed by analysis of a series of MRI echo-planar images acquired over a period of several minutes with the subject at rest (1). In the present work, functional connectivity experiments are extended from human brain to rat brain and our central hypothesis is that the underlying physiology is conserved across all mammalian species.A reference time course obtained from a reference voxel (or, alternatively, an average over a cluster of voxel time courses in a region of interest [ROI]) is formed. Cross correlation of the reference time course with all voxel time courses in the slice provides a functional connectivity map. A strategy is required for selection of the reference time course, and performance of a task is commonly used to define ROIs in resting brain that can be used to form reference time courses.We have discovered that electrical stimulation using an implanted electrode on the radial nerve of the brachial plexus of a rat (2) results in activation of a network of sensorimotor brain regions, each of which is a suitable candidate for formation of a reference time course when analyzing resting-state data. Experiments not only in the sensorimotor system but also in the visual system provide further support for our central hypothesis. Functional connectivity was studied using reference waveforms obtained from areas that were found to be activated by light incident on the retina that was turned on and off in a block-trial functional MRI (fMRI) experiment.Anatomic images acquired in this experiment are of high quality, and it is possible to define anatomic regions purely by reference to the rat histological atlas (3). One can develop a reference waveform from each of these regions and test a specific hypothesis that functional connectivity to a second region is consistent with a known connectivity. We report here success in this hypothesis-driven approach to analysis of resting-state data. A total of 22 sensorimotor regions were identified, and connectivities between each of these regions and the other 21 regions were determined.Most functional connectivity studies have been in the awake human b...

The α2-adrenoreceptor agonist, medetomidine, which exhibits dose-dependent sedative effects and is gaining acceptance in small-animal functional magnetic resonance imaging (fMRI), has been studied. Rats were examined on the bench using the classic tail-pinch method with three infusion sequences: 100 μg/kg/hr, 300 μg/kg/hr, or 100 μg/kg/hr followed by 300 μg/kg/hr. Stepping the infusion rate from 100 to 300 μg/kg/hr after 2.5 hours resulted in a prolonged period of approximately level sedation that cannot be achieved by a constant infusion of either 100 or 300 μg/kg/hr. By stepping the infusion dosage, experiments as long as six hours are possible. Functional MRI experiments were carried out on rats using a frequency dependent electrical stimulation protocol—namely, forepaw stimulation at 3, 5, 7, and 10 Hz. Each rat was studied for a four-hour period, divided into two equal portions. During the first portion, rats were started at a 100 μg/kg/hr constant infusion. During the second portion, four secondary levels of infusion were used: 100, 150, 200, and 300 μg/kg/hr. The fMRI response to stimulation frequency was used as an indirect measure of modulation of neuronal activity through pharmacological manipulation. The frequency response to stimulus was attenuated at the lower secondary infusion dosages 100 or 150 μg/kg/hr but not at the higher secondary infusion dosages 200 or 300 μg/kg/hr. Parallel experiments with the animal at rest were carried out using both electroencephalogram (EEG) and functional connectivity MRI (fcMRI) methods with consistent results. In the secondary infusion period using 300 μg/kg/hr, resting-state functional connectivity is enhanced.

Rationale: A hallmark of chronic inflammatory disorders is persistence of pro-inflammatory macrophages in diseased tissues. In atherosclerosis this is associated with dyslipidemia and oxidative stress, but mechanisms linking these phenomena to macrophage activation remain incompletely understood. Objective: To investigate mechanisms linking dyslipidemia, oxidative stress and macrophage activation through modulation of immunometabolism, and to explore therapeutic potential targeting specific metabolic pathways. Methods and Results: Using a combination of biochemical, immunological, and ex vivo cell metabolic studies, we report that CD36 mediates a mitochondrial metabolic switch from oxidative phosphorylation to superoxide production in response to its ligand, oxLDL. Mitochondrial-specific inhibition of superoxide inhibited oxLDL-induced NF-κB activation and inflammatory cytokine generation. RNAseq, flow cytometry, 3H-labeled palmitic acid uptake, lipidomic analysis, confocal and EM imaging, and functional energetics revealed that oxLDL upregulated effectors of long-chain fatty acid (FA) uptake and mitochondrial import, while downregulating FA oxidation and inhibiting ATP5A, an electron transport chain (ETC) component. The combined effect is long-chain FA accumulation, alteration of mitochondrial structure and function, repurposing of the ETC to superoxide production, and NF-κB activation. Apoe null mice challenged with high fat diet showed similar metabolic changes in circulating Ly6C+ monocytes and peritoneal macrophages, along with increased CD36 expression. Moreover, mitochondrial ROS was positively correlated with CD36 expression in aortic lesional macrophages. Conclusions: These findings reveal that oxLDL/CD36 signaling in macrophages links dys-regulated FA metabolism to oxidative stress from the mitochondria, which drives chronic inflammation. Thus, targeting to CD36 and its downstream effectors may serve as potential new strategies against chronic inflammatory diseases such as atherosclerosis.

man. 20-HETE contributes to the acute fall in cerebral blood flow after subarachnoid hemorrhage in the rat. Am J Physiol Heart Circ Physiol 282: H1556-H1565, 2002. First published December 6, 2001 10.1152/ajpheart.00924.2001.-This study examined the effects of blocking the formation of 20-hydroxyeicosatetraenoic acid (20-HETE) on the acute fall in cerebral blood flow after subarachnoid hemorrhage (SAH) in the rat. In vehicle-treated rats, regional cerebral blood flow (rCBF) measured with laser-Doppler flowmetry fell by 30% 10 min after the injection of 0.3 ml of arterial blood into the cisterna magna, and it remained at this level for 2 h. Pretreatment with inhibitors of the formation of 20-HETE, 17-octadecynoic acid (17-ODYA; 1.5 nmol intrathecally) and N-hydroxy-NЈ-(4-butyl-2-methylphenyl)formamidine (HET0016; 10 mg/kg iv), reduced the initial fall in rCBF by 40%, and rCBF fully recovered 1 h after induction of SAH. The concentration of 20-HETE in the cerebrospinal fluid rose from 12 Ϯ 2 to 199 Ϯ 17 ng/ml after SAH in vehicle-treated rats. 20-HETE levels averaged only 15 Ϯ 11 and 39 Ϯ 13 ng/ml in rats pretreated with 17-ODYA or HET0016, respectively. HET0016 selectively inhibited the formation of 20-HETE in rat renal microsomes with an IC50 of Ͻ15 nM and human recombinant CYP4A11, CYP4F2, and CYP4F3 enzymes with an IC50 of 42, 125, and 100 nM, respectively. These results indicate that 20-HETE contributes to the acute fall in rCBF after SAH in rats.

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