Astrocytes have a higher antioxidant potential in comparison to neurons. Pathways associated with this selective advantage include the transcriptional regulation of antioxidant enzymes via the action of the Cap'n'Collar transcription factor Nrf2 at the antioxidant response element (ARE). Here we show that Nrf2 overexpression can reengineer neurons to express this glial pathway and enhance antioxidant gene expression. However, Nrf2-mediated protection from oxidative stress is conferred primarily by glia in mixed cultures. The antioxidant properties of Nrf2-overexpressing glia are more pronounced than those of neurons, and a relatively small number of these glia (Ͻ 1% of total cell number added) could protect fully cocultured naive neurons from oxidative glutamate toxicity associated with glutathione (GSH) depletion. Microarray and biochemical analyses indicate a coordinated upregulation of enzymes involved in GSH biosynthesis (xCT cystine antiporter, ␥-glutamylcysteine synthetase, and GSH synthase), use (glutathione S-transferase and glutathione reductase), and export (multidrug resistance protein 1) with Nrf2 overexpression, leading to an increase in both media and intracellular GSH. Selective inhibition of glial GSH synthesis and the supplementation of media GSH indicated that an Nrf2-dependent increase in glial GSH synthesis was both necessary and sufficient for the protection of neurons, respectively. Neuroprotection was not limited to overexpression of Nrf2, because activation of endogenous glial Nrf2 by the small molecule ARE inducer, tert-butylhydroquinone, also protected against oxidative glutamate toxicity.
Neural activity in the brain is followed by localized changes in blood flow and volume. We address the relative change in volume for arteriole vs. venous blood within primary vibrissa cortex of awake, head-fixed mice. Two-photon laser-scanning microscopy was used to measure spontaneous and sensory evoked changes in flow and volume at the level of single vessels. We find that arterioles exhibit slow (<1 Hz) spontaneous increases in their diameter, as well as pronounced dilation in response to both punctate and prolonged stimulation of the contralateral vibrissae. In contrast, venules dilate only in response to prolonged stimulation. We conclude that stimulation that occurs on the time scale of natural stimuli leads to a net increase in the reservoir of arteriole blood. Thus, a "bagpipe" model that highlights arteriole dilation should augment the current "balloon" model of venous distension in the interpretation of fMRI images.ocalized changes in the flow and volume of oxygenated blood in the brain are commonly used as a correlate of heightened neural activity. For two important imaging modalities, blood oxygen level-dependent functional magnetic resonance imaging (BOLD fMRI) (1) and intrinsic optical signal imaging (IOS) (2), the signals are generated by a complex interplay of the rate of oxidative metabolism, the flux of blood in the underlying angioarchitecture, and changes in vascular volume (3). The locus for the increase in vascular volume that follows sensory stimulation (i.e., arterioles or venules) is an enduring controversy that bears directly on interpreting and quantifying fMRI signals (4-6). To resolve this question, we used in vivo two-photon laserscanning microscopy to image spontaneous and sensory evoked vascular dynamics in the vibrissa area of parietal cortex of awake, head-fixed mice. All data were collected through a reinforced thin-skull window (7) (Fig. 1A); this method obviates potential complications from inflammation or changes in cranial pressure that may occur with a craniotomy (8). ResultsImages of the pial surface and measurements of the diameter of the lumen of surface arterioles and venules were performed while the mouse sat passively (Fig. 1B). Dilations greater than 5% of the baseline diameter occurred with frequencies of 0.07 ± 0.05 Hz (mean ± SD, n = 118 arteries in six mice). The diameters of short segments of arterioles (red in Fig. 1B) exhibited relatively large spontaneous increases in the spectral range between 0.1 Hz and 1 Hz ( Fig. 1 C and D). The peak amplitude of these fluctuations was 23% ± 10% of the initial vessel diameter across all arterioles, with instances of a 50% increases in diameter. Further, these lowfrequency oscillations were strongly coherent and synchronous over a distance of several hundred micrometers across cortex (198 pairs of vessels, in six mice, that were separated by 20-315 μm; slope of coherence = 0.001 μm −1 [nonsignificant (NS)] and slope of phase shift = 0.0009 rad/μm
We present a method to form an optical window in the mouse skull that spans millimeters and is stable for months without inflammation of the brain. This enabled us to repeatedly image blood flow in cortical capillaries of awake animals and determine long-range correlations in speed. We further demonstrate repeated cortical imaging of dendritic spines, microglia, and angioarchitecture, as well as illumination to drive motor output via optogenetics and induce microstrokes via photosensitizers.
The cerebral vascular system services the constant demand for energy during neuronal activity in the brain. Attempts to delineate the logic of neurovascular coupling have been greatly aided by the advent of two-photon laser scanning microscopy to image both blood flow and the activity of individual cells below the surface of the brain. Here we provide a technical guide to imaging cerebral blood flow in rodents. We describe in detail the surgical procedures required to generate cranial windows for optical access to the cortex of both rats and mice and the use of two-photon microscopy to accurately measure blood flow in individual cortical vessels concurrent with local cellular activity. We further provide examples on how these techniques can be applied to the study of local blood flow regulation and vascular pathologies such as small-scale stroke.
Microinfarctions are present in the aged and injured human brain. Their clinical significance is controversial, with postulated sequelae ranging from cognitive sparing to vascular dementia. To address the consequences of microinfarcts, we used controlled optical methods to create occlusions of individual penetrating arterioles or venules within rat cortex. Single microinfarcts, targeted to encompass all or part of a cortical column, impaired performance in a macrovibrissa-based behavioral task. Further, multiple targeted vessels caused tissue damage that coalesced across cortex, even though the intervening penetrating vessels were acutely patent. Post-occlusion administration of Memantine, a glutamate receptor antagonist that reduces cognitive decline in Alzheimer’s disease, ameliorated tissue damage and perceptual deficits. Collectively, these data imply that microinfarcts are likely contributors to cognitive decline. Strategies that have received limited success in the treatment of ischemic injury, which include therapeutics against excitotoxicity, may be successful against the progressive nature of vascular dementia.
The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) coordinates expression of genes required for free radical scavenging, detoxification of xenobiotics, and maintenance of redox potential. Previously, activation of this pleiotropic response was neuroprotective in cell culture models that simulate components of stroke damage. However, the role of Nrf2 in limiting stroke damage in vivo remained unclear. We report that Nrf2 activation protects the brain from cerebral ischemia in vivo. Acute (1-3 d) intracerebroventricular or intraperitoneal pretreatment with tert-butylhydroquinone (tBHQ), an Nrf2 activity inducer, reduced cortical damage and sensorimotor deficit at 24 h and even 1 month after ischemia-reperfusion in rats. Cortical glutathione levels robustly increased with tBHQ administration to rats and Nrf2-expressing mice, but not Nrf2 Ϫ/Ϫ mice. Basal and inducible activities of antioxidant/detoxification enzymes in Nrf2Ϫ/Ϫ mice were reduced when compared with Nrf2 ϩ/ϩ controls. Interestingly, larger infarcts were observed in Nrf2 Ϫ/Ϫ mice at 7 d after stroke, but not at 24 h, suggesting that Nrf2 may play a role in shaping the penumbra well after the onset of ischemia. Neuronal death caused by a "penumbral" model of stroke, using intracortical endothelin-1 microinjection, was attenuated by tBHQ administration to Nrf2 ϩ/ϩ , but not to Nrf2 Ϫ/Ϫ mice, confirming the Nrf2-specific action of tBHQ in vivo. We conclude that Nrf2 plays a role in modulating ischemic injury in vivo. Accordingly, Nrf2 activation by small molecule inducers may be a practical preventative treatment for stroke-prone patients.
Pericytes are essential for normal brain function, but many aspects of their physiology remain enigmatic due to a lack of tools to genetically target this cell population. Here, we characterize brain pericytes using two existing Cre-recombinase driver mouse lines that can serve distinct purposes in cerebrovascular research. One line expresses an inducible version of Cre under the NG2 proteoglycan promoter, which provides the sparse labeling necessary to define the morphology of single cells. These mice reveal structural differences between pericytes adjacent to arterioles versus those broadly distributed in the capillary bed that may underlie differential roles in control of vessel caliber. A second line expresses Cre constitutively under the platelet-derived growth factor receptor β promoter and provides continuous, highly specific and near-complete labeling of pericytes and myocytes along the entire cerebrovasculature. This line provides a three-dimensional view of pericyte distribution along the cortical angioarchitecture following optical clearing of brain tissue. In combination with recent reporter lines for expression of optogenetic actuators and activity-sensitive probes, these mice may be key tools for studying pericyte biology in the intact brain.
Cerebral microinfarcts are small lesions that are presumed to be ischaemic. Despite the small size of these lesions, affected individuals can have hundreds to thousands of cerebral microinfarcts, which cause measurable disruption to structural brain connections, and are associated with dementia that is independent of Alzheimer’s disease pathology or larger infarcts (ie, lacunar infarcts, and large cortical and non-lacunar subcortical infarcts). Substantial progress has been made with regard to understanding risk factors and functional consequences of cerebral microinfarcts, partly driven by new in-vivo detection methods and the development of animal models that closely mimic multiple aspects of cerebral microinfarcts in human beings. Evidence from these advances suggests that cerebral microinfarcts can be manifestations of both small vessel and large vessel disease, that cerebral microinfarcts are independently associated with cognitive impairment, and that these lesions are likely to cause damage to brain structure and function that extends beyond their actual lesion boundaries. Criteria for the identification of cerebral microinfarcts with in-vivo MRI are provided to support further studies of the association between these lesions and cerebrovascular disease and dementia.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
334 Leonard St
Brooklyn, NY 11211
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.