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Nedergaard, M.

Publications and source records attributed to Nedergaard, M..

32 records · Page 2Linked to original sources

Liver-secreted fluorescent blood plasma markers enable chronic imaging of microcirculation

Studying blood microcirculation is vital for gaining insights into vascular diseases. Acute administration of fluorescent tracers is currently used for deep tissue blood flow imaging. This is invasive, and the plasma fluorescence decreases within an hour of administration. We report a novel approach for the longitudinal study of vasculature. Using a single systemic administration of viral vectors, we express fluorescent secretory albumin-fusion proteins in the liver to label the blood in mice. All segments of the vasculature in brain and peripheral tissue are observable by two-photon microscopy within two weeks of vector administration. This approach allows for observation of circulation without the need for repeated administration for several months. We demonstrate the chronic assessment of vascular functions at micro-and mesoscopic scales. This genetic plasma labeling approach represents a versatile and cost-effective method for the chronic investigation of vasculature functions across the body in health and disease.

physiology↗

A sensitive and specific genetically encodable biosensor for potassium ions

Potassium ions (K+) play a critical role as an essential electrolyte in all biological systems. Here we report the crystal structure-guided optimization and directed evolution of an improved genetically encoded fluorescent K+ biosensor, GINKO2. GINKO2 is highly sensitive and specific for K+ and enables in vivo detection of K+ dynamics in multiple species.

bioengineering↗

A network model of glymphatic flow under different experimentally-motivated parametric scenarios

Rapidly growing evidence demonstrates that flow of cerebrospinal fluid (CSF) through perivascular spaces (PVSs) - annular channels surrounding vasculature in the brain - is a critically-important component of neurophysiology. CSF inflow contributes during physiological conditions to clearance of metabolic waste and in pathological situations to edema formation. However, brain-wide imaging methods cannot resolve PVSs, and high-resolution methods cannot access deep tissue or be applied to human subjects, so theoretical models provide essential insight. We model this CSF pathway as a network of hydraulic resistances, built from published parameters. A few parameters have very wide uncertainties, so we focus on the estimated limits of their feasible ranges by analyzing different parametric scenarios. We identify low-resistance PVSs and high-resistance parenchyma (brain tissue) as the scenario that best explains experimental observations. Our results point to the most important parameters that should be measured in future experiments. Extensions of our modeling may help predict stroke severity or lead to neurological disease treatments and drug delivery methods.

neuroscience↗

Impact of Propionic Acidemia on Brain Astrocytes

Propionic acidemia (PA) is an inborn error of metabolism (IEM) caused by mutations in the enzyme propionyl CoA carboxylase (PCC). It is characterized by the inability to break down branched chain amino acids and odd chain fatty acids, causing a buildup of toxic organic acids in blood. PA affects every organ in the body with particularly severe manifestations in the brain, like hyperammonemia, hypomyelination, seizures, cognitive impairments, optic nerve atrophy and autism spectrum disorders. Dietary management and liver transplantation have helped to ameliorate the acute expression of the disorder, but do not prevent the chronic toxicity that builds up in brain. Despite the severe brain manifestation of the disease, little is known about the mechanisms by which PA affects the nervous system. PCCA and PCCB, the two subunits required for a functional PCC enzyme, are both expressed not only in neurons but also in astrocytes. Using the two rodent genetic models of PA currently available, with mutations in PCCA, we have evaluated the involvement of astrocytes in the neuropathology of propionic acidemia. These mice exhibit cardiac pathology and hyperammonemia, similar to what is observed in patients with PA. We found that wild type (wt) astrocytes positively respond to treatment with L-Carnitine, a therapeutic approach commonly used in patients with PA, by improved survival and more efficient mitochondrial morphology. Transcriptome analysis from astrocytes derived from the wt or the mutant mice confirm that these astrocytes lack exons 3 and 4 like in the human mutations of PA. However, no other genes/exons were statistically significant with regards to differential expression between astrocytes derived from KO or from WT animals, suggesting that astrocytes in culture may be able to compensate the PCC deficiency. Histological analysis of neuronal and glial markers during brain development (TUJ1, MAP2 for neurons; nestin and Iba1 for glia) do not show significant alterations neither in distribution nor numbers of cells in the developing brain of the PCCA-/- mice. Analysis in the adult brain of mutant mice shows some variable degree of microgliosis but no indication of reactive astrocytes. No gross abnormalities were observed in cortex, hippocampus, striatum or cerebellum of adult brains of PCCA-/- mice, either. In summary, astrocytes from PCCA deficient mice show surprisingly little alterations both in vitro and in vivo. Our results evidence the need to further understand the effects of PA in brain cells to help develop potential new therapies that can preserve brain function in children affected by this devastating disease. HIGHLIGHTSO_LIAstrocytes detoxify ammonia in brain and may be affected by propionic acidemia, a disorder that causes hyperammonemia in brain. C_LIO_LIRNA seq of astrocytes in culture derived from PCCA mutant mice does not show effect in these cells. C_LIO_LIIn vivo analysis of glial and neuronal cells also shows no difference in development or adult mutant mice C_LIO_LIAstrocytes may not be an adequate target of clinical therapies in this disorder C_LI

neuroscience↗

Bulk flow of cerebrospinal fluid observed in periarterial spaces is not an artifact of injection

Cerebrospinal fluid (CSF) flowing through periarterial spaces is integral to the brains mechanism for clearing metabolic waste products. Experiments that track tracer particles injected into the cisterna magna of mouse brains have shown evidence of pulsatile CSF flow in pial periarterial spaces, with a bulk flow in the same direction as blood flow. However, the driving mechanism remains elusive. Several studies have suggested that the bulk flow might be an artifact, driven by the injection itself. Here, we address this hypothesis with new in vivo experiments where tracer particles are injected into the cisterna magna using a dual-syringe system, with simultaneous injection and withdrawal of equal amounts of fluid. This method produces no net increase in CSF volume and no significant increase in intracranial pressure. Yet, particle-tracking reveals flows in the pial periarterial spaces that are completely consistent with the flows observed in earlier experiments with single-syringe injection.

neuroscience↗

Sleep-specific changes in physiological brain pulsations

Sleep is known to increase the convection of interstitial brain metabolites along with cerebrospinal fluid (CSF). We used ultrafast magnetic resonance encephalography (MREGBOLD) to quantify the effect of sleep on physiological (vasomotor, respiratory and cardiac) brain pulsations driving the CSF convection in humans. Transition to electroencephalography verified sleep occurred in conjunction with power increase and reduced spectral entropy (SE) of physiological brain pulsations. During sleep, the greatest increase in spectral power was in very-low frequency (VLF < 0.1 Hz) waves, followed by respiratory and cardiac brain pulsations. SE reduction coincided with decreased vigilance in awake state and could robustly (ROC 0.88, p < 0.001) differentiate between sleep vs. awake states, indicating the sensitivity of SE of the MREGBOLD signal as a marker for sleep level. In conclusion, the three physiological brain pulsation contribute to the sleep-associated increase in glymphatic CSF convective flow in an inverse frequency order. HighlightsO_LIBrain tissue contains almost no connective tissue, this enabling pressure waves to initiate long-distance brain pulsations C_LIO_LIBrain pulsations are induced by vasomotion, respiration, and the cardiac cycle C_LIO_LISleep strikingly increases spectral power and decreases spectral entropy of brain pulsations, especially for the very low frequency vasomotor waves C_LIO_LISpectral entropy of brain pulsations detected by MREG is a sensitive measure of vigilance, resembling the corresponding entropy changes detected by scalp EEG C_LI

neuroscience↗

Dynamic fluctuations of the locus coeruleus-norepinephrine system underlie sleep state transitions

We normally regard sleep and wake as two distinct opposing brain states, where sleep requires silence of wake-promoting structures such as the locus coeruleus (LC)-norepinephrine (NE) system. We set out to investigate how cortical NE dynamics and NE-related astrocytic activity relates to LC population activity during sleep states. We show that LC displays regular phasic activity bouts during NREM sleep leading to a slow oscillatory pattern of prefrontal NE levels of which the majority of NE increases does not lead to awakening. NE troughs link to sleep spindles and continued NE decline transitions into REM sleep. Last, we show that prefrontal astrocytes have reduced sensitivity towards NE during sleep. Our results suggest that dynamic changes in the activity of wake-promoting systems during sleep create alternation between crucial sleep processes and broadening of sensitivity towards incoming sensory input. HighlightsO_LIExtracellular levels of norepinephrine display dynamic changes during NREM and REM sleep C_LIO_LIPhasic activity of locus coeruleus neurons during NREM underlies slow norepinephrine oscillations C_LIO_LISpindles occur at norepinephrine troughs and are abolished by norepinephrine increases C_LIO_LIIncreased spindles prior to REM reflect the beginning of a long-lasting norepinephrine decline C_LIO_LIREM episodes are characterized by a sub-threshold continuous norepinephrine decline C_LIO_LIThe responsiveness of astrocytic Ca2+ to norepinephrine is reduced during sleep C_LI

neuroscience↗

Perivascular pumping in the mouse brain: Realistic boundary conditions reconcile theory, simulation, and experiment

Cerebrospinal fluid (CSF) flows through the perivascular spaces (PVSs) surrounding cerebral arteries. Revealing the mechanisms driving that flow could bring improved understanding of brain waste transport and insights for disorders including Alzheimers disease and stroke. In vivo velocity measurements of CSF in surface PVSs in mice have been used to argue that flow is driven primarily by the pulsatile motion of artery walls -- perivascular pumping. However, fluid dynamics theory and simulation have predicted that perivascular pumping produces flows differing from in vivo observations starkly, particularly in the phase and relative amplitude of flow oscillation. Here we show that coupling theoretical and simulated flows to realistic end boundary conditions, using resistance and compliance values measured in mice, results in velocities that match observations closely in phase, relative amplitude of oscillation, and mean flow speed. This new, quantitative agreement among theory, simulation, and in vivo measurement further supports the idea that perivascular pumping is a primary CSF driver in physiological conditions.

neuroscience↗

The Ocular Glymphatic Clearance System

Despite high metabolic activity, the retina and optic nerve head lack traditional lymphatic drainage. We here identified a novel ocular glymphatic clearance route for fluid and wastes via the proximal optic nerve. Amyloid-{beta} (A{beta}) was cleared from the vitreous via a pathway driven by the ocular-cranial pressure difference. After traversing the lamina barrier, intra-axonal A{beta} was cleared via the perivenous space and subsequently drained to lymphatic vessels. Light-induced pupil constriction enhanced, while atropine or raising intracranial pressure blocked efflux. In two distinct murine models of glaucoma, A{beta} leaked from the eye via defects in the lamina barrier instead of directional axonal efflux. The discovery of a novel pathway for removal of fluid and metabolites from the intraocular space prompts a reevaluation of the core principles governing eye physiology and provides a framework for new therapeutic approaches to treat common eye diseases, including glaucoma. One Sentence SummaryGlymphatic pathway clears ocular amyloid-{beta} via optic nerve and fails in glaucoma.

physiology↗

Haplotype of the astrocytic water channel AQP4 modulates slow wave energy in human NREM sleep

Cerebrospinal fluid (CSF) flow through the brain parenchyma is facilitated by the astrocytic water channel aquaporin 4 (AQP4). Homeostatically regulated electroencephalographic (EEG) slow waves are a hallmark of deep non-rapid-eye-movement (NREM) sleep and have been implicated in the regulation of parenchymal CSF flow and brain clearance. The human AQP4 gene harbors several single nucleotide polymorphisms (SNPs) associated with AQP4 expression, brain-water homeostasis and neurodegenerative diseases. To date, their role in sleep-wake regulation is unknown. To investigate whether functional variants in AQP4 modulate human sleep, nocturnal EEG-recordings and cognitive performance were investigated in 123 healthy participants genotyped for a common eight-SNP AQP4-haplotype. We show that this AQP4-haplotype is associated with distinct modulations of NREM slow wave energy, strongest in early sleep and mirrored by changes in sleepiness and reaction times during extended wakefulness. The study provides the first human evidence for a link between AQP4, deep NREM sleep and cognitive consequences of prolonged wakefulness.

neuroscience↗

Glymphatic Optimal Mass Transport with Lagrangian Workflow Reveals Advective and Diffusion Driven Solute Transport

The presence of advection in neuropil is contested and solute transport is claimed to occur by diffusion only. To address this controversy, we implemented a regularized version of the optimal mass transport (rOMT) problem, wherein the advection/diffusion equation is the only a priori assumption required. rOMT analysis with a Lagrangian perspective of glymphatic system (GS) transport revealed that solute speed was faster in cerebrospinal fluid (CSF) compared to grey and white matter. rOMT analysis also demonstrated 2-fold differences in regional particle speed within the brain parenchyma. Collectively, these results imply that advective transport dominates in CSF while diffusion and advection both contribute to transport in parenchyma. In rats with chronic hypertension, solute transport in perivascular spaces (PVS) and PVS-to-tissue transfer was slower compared to normotension. Thus, the analytical framework of rOMT provides novel insights in local variation and dynamics of GS transport that may have implications for neurodegenerative diseases.

physiology↗

Hydraulic resistance of perivascular spaces in the brain

Perivascular spaces (PVSs) are annular channels that surround arteries and carry cerebrospinal fluid into the brain, sweeping away metabolic waste. In vivo observations reveal that they are not concentric, circular annuli, however: the outer boundaries are often oblate, and the arteries that form the inner boundaries are often offset from the central axis. Here we show that the observed shapes of PVSs have lower hydraulic resistance than concentric, circular annuli of the same size, and therefore allow faster, more efficient flow of cerebrospinal fluid. To calculate hydraulic resistance, we parameterize PVS cross-sections as circles surrounded by ellipses, varying the radii of the circles, major and minor axes of the ellipses, and two-dimensional eccentricities of the circles with respect to the ellipses. For each shape, we solve the governing Navier-Stokes equation to determine the velocity profile for steady laminar flow and the corresponding hydraulic resistance. We find that the minimum hydraulic resistance (and therefore maximum flow rate) for a given PVS cross-sectional area occurs when the ellipse is elongated and intersects the circle, dividing the PVS into two lobes, as is common around pial arteries. We also find that if both the inner and outer boundaries are circular, the minimum hydraulic resistance occurs when the eccentricity is large, as is common around penetrating arteries. The circular, concentric annuli assumed in recent studies maximize hydraulic resistance and therefore minimize flow. Our parameterization can incorporate more realistic resistances into hydraulic network models of flow of cerebrospinal fluid in the brain.\n\nSignificance StatementRecent experiments show that there is bulk flow of cerebrospinal fluid in brain perivascular spaces (PVSs) in the direction of the blood flow, driven by pulsations of arterial walls due to the heartbeat. Published hydraulic network models have argued against such a flow due to the purported high hydraulic resistance of the system. Here we show that these high resistances are due to misrepresentations of the size and shape of PVSs. We present an adjustable model PVS that can match closely the actual shapes observed in vivo and show that these shapes are often nearly optimal, in the sense of offering the least hydraulic resistance. This optimization may well represent an evolutionary adaptation that maximizes clearance of metabolic waste from the brain.

neuroscience↗

Panoptic vDISCO imaging reveals neuronal connectivity, remote trauma effects and meningeal vessels in intact transparent mice

Analysis of entire transparent rodent bodies could provide holistic information on biological systems in health and disease. However, it has been challenging to reliably image and quantify signal from endogenously expressed fluorescent proteins in large cleared mouse bodies due to the low signal contrast. Here, we devised a pressure driven, nanobody based whole-body immunolabeling technology to enhance the signal of fluorescent proteins by up to two orders of magnitude. This allowed us to image subcellular details in transparent mouse bodies through bones and highly autofluorescent tissues, and perform quantifications. We visualized for the first-time whole-body neuronal connectivity of an entire adult mouse and discovered that brain trauma induces degeneration of peripheral axons. We also imaged meningeal lymphatic vessels and immune cells through the intact skull and vertebra in naive animals and trauma models. Thus, our new approach can provide an unbiased holistic view of biological events affecting the nervous system and the rest of the body.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC=\"FIGDIR/small/374785_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (62K):\norg.highwire.dtl.DTLVardef@8fad72org.highwire.dtl.DTLVardef@c41ed5org.highwire.dtl.DTLVardef@1c470eaorg.highwire.dtl.DTLVardef@1d144cb_HPS_FORMAT_FIGEXP M_FIG C_FIG Note: Manuscript videos are available at Supplementary material section of BioRxiv and at the following link http://vdisco.isd-muc.de/

neuroscience↗

Aquaporin-4 dependent glymphatic solute transport in rodent brain

The glymphatic system is a brain-wide metabolite clearance pathway, impairment of which in post-traumatic and ischemic brain or healthy aging is proposed to contribute to intracerebral accumulation of amyloid-{beta} and tau proteins. Glymphatic perivascular influx of cerebrospinal fluid (CSF) depends upon the expression and perivascular localization of the astroglial water channel aquaporin-4 (AQP4). Prompted by a recent publication that failed to find an effect of Aqp4 knockout on perivascular CSF tracer influx and interstitial fluid (ISF) tracer dispersion, four independent research groups have herein re-examined the importance of Aqp4 in glymphatic fluid transport. We concur in finding that CSF tracer influx, as well as fluorescently-tagged amyloid-{beta} efflux, are significantly faster in wild-type mice than in three different transgenic lines featuring disruption of the Aqp4 gene and one line in which AQP4 expression lacks the critical perivascular localization (Snta1 knockout). These data validate the role of AQP4 in supporting fluid and solute transport and efflux in brain in accordance with the glymphatic system model.

neuroscience↗