Long-wavelength traveling waves of vasomotion modulate the perfusion of cortex.

Brain arterioles are active, multicellular complexes whose diameters oscillate at ∼ 0.1 Hz. We assess the physiological impact and spatiotemporal dynamics of vaso-oscillations in the awake mouse. First, vaso-oscillations in penetrating arterioles, which source blood from pial arterioles to the capillary bed, profoundly impact perfusion throughout neocortex. The modulation in flux during resting-state activity exceeds that of stimulus-induced activity. Second, the change in perfusion through arterioles relative to the change in their diameter is weak. This implies that the capillary bed dominates the hydrodynamic resistance of brain vasculature. Lastly, the phase of vaso-oscillations evolves slowly along arterioles, with a wavelength that exceeds the span of the cortical mantle and sufficient variability to establish functional cortical areas as parcels of uniform phase. The phase-gradient supports traveling waves in either direction along both pial and penetrating arterioles. This implies that waves along penetrating arterioles can mix, but not directionally transport, interstitial fluids.

A lone spike in blood glucose can enhance the thrombo-inflammatory response in cortical venules.

How transient hyperglycemia contributes to cerebro-vascular disease has been a challenge to study under controlled physiological conditions. We use amplified, ultrashort laser-pulses to physically disrupt brain-venule endothelium at targeted locations. This vessel disruption is performed in conjunction with transient hyperglycemia from a single injection of metabolically active D-glucose into healthy mice. The observed real-time responses to laser-induced disruption include rapid serum extravasation, platelet aggregation, and neutrophil recruitment. Thrombo-inflammation is pharmacologically ameliorated by a platelet inhibitor, by a scavenger of reactive oxygen species, and by a nitric oxide donor. As a control, vessel thrombo-inflammation is significantly reduced in mice injected with metabolically inert L-glucose. Venules in mice with diabetes show a similar response to laser-induced disruption and damage is reduced by restoration of normo-glycemia. Our approach provides a controlled method to probe synergies between transient metabolic and physical vascular perturbations and can reveal new aspects of brain pathophysiology.

Baseline oxygen consumption decreases with cortical depth.

The cerebral cortex is organized in cortical layers that differ in their cellular density, composition, and wiring. Cortical laminar architecture is also readily revealed by staining for cytochrome oxidase-the last enzyme in the respiratory electron transport chain located in the inner mitochondrial membrane. It has been hypothesized that a high-density band of cytochrome oxidase in cortical layer IV reflects higher oxygen consumption under baseline (unstimulated) conditions. Here, we tested the above hypothesis using direct measurements of the partial pressure of O2 (pO2) in cortical tissue by means of 2-photon phosphorescence lifetime microscopy (2PLM). We revisited our previously developed method for extraction of the cerebral metabolic rate of O2 (CMRO2) based on 2-photon pO2 measurements around diving arterioles and applied this method to estimate baseline CMRO2 in awake mice across cortical layers. To our surprise, our results revealed a decrease in baseline CMRO2 from layer I to layer IV. This decrease of CMRO2 with cortical depth was paralleled by an increase in tissue oxygenation. Higher baseline oxygenation and cytochrome density in layer IV may serve as an O2 reserve during surges of neuronal activity or certain metabolically active brain states rather than reflecting baseline energy needs. Our study provides to our knowledge the first quantification of microscopically resolved CMRO2 across cortical layers as a step towards better understanding of brain energy metabolism.

Neurophotonic tools for microscopic measurements and manipulation: status report.

Neurophotonics was launched in 2014 coinciding with the launch of the BRAIN Initiative focused on development of technologies for advancement of neuroscience. For the last seven years, Neurophotonics' agenda has been well aligned with this focus on neurotechnologies featuring new optical methods and tools applicable to brain studies. While the BRAIN Initiative 2.0 is pivoting towards applications of these novel tools in the quest to understand the brain, this status report reviews an extensive and diverse toolkit of novel methods to explore brain function that have emerged from the BRAIN Initiative and related large-scale efforts for measurement and manipulation of brain structure and function. Here, we focus on neurophotonic tools mostly applicable to animal studies. A companion report, scheduled to appear later this year, will cover diffuse optical imaging methods applicable to noninvasive human studies. For each domain, we outline the current state-of-the-art of the respective technologies, identify the areas where innovation is needed, and provide an outlook for the future directions.

A Suite of Neurophotonic Tools to Underpin the Contribution of Internal Brain States in fMRI.

Recent developments in optical microscopy, applicable for large-scale and longitudinal imaging of cortical activity in behaving animals, open unprecedented opportunities to gain a deeper understanding of neurovascular and neurometabolic coupling during different brain states. Future studies will leverage these tools to deliver foundational knowledge about brain state-dependent regulation of cerebral blood flow and metabolism as well as regulation as a function of brain maturation and aging. This knowledge is of critical importance to interpret hemodynamic signals observed with functional magnetic resonance imaging (fMRI).

Diurnal T2-changes of the intervertebral discs of the entire spine and the influence of weightlifting.

The purpose was to study if (1) diurnal changes occur in the entire spine and if (2) intervertebral discs (IVDs) of weightlifters (WL) have decreased baseline T2-values in the morning as well as (3) increased diurnal changes throughout the day. This prospective cohort study investigated healthy volunteers between 2015 and 2017. WL were required to have participated in weightlifting ≥ 4×/week for ≥ 5 years, while non-weightlifters (NWL) were limited to < 2×/week for ≥ 5 years. Both groups underwent magnetic resonance imaging (MRI) of the entire spine in the morning and evening. WL were requested to perform weightlifting in-between imaging. IVD regions of interest (nucleus pulposus) were defined and T2-maps were measured. Analysis consisted of unpaired t-test, paired t-test, propensity-score matching (adjusting for age and sex), and Pearson correlation. Twenty-five individuals (15 [60.0%] males) with a mean age of 29.6 (standard deviation [SD 6.9]) years were analyzed. Both groups (WL: n = 12 versus [vs.] NWL: n = 13) did not differ demographic characteristics. Mean IVD T2-values of all participants significantly decreased throughout the day (95.7 [SD 15.7] vs. 86.4 [SD 13.9] milliseconds [ms]) in IVDs of the cervical (71.8 [SD 13.4] vs. 64.4 [SD 14.1] ms), thoracic (98.8 [SD 19.9] vs. 88.6 [SD 16.3] ms), and lumbar (117.0 [SD 23.7] vs. 107.5 [SD 21.6] ms) spine (P < 0.001 each). There were no differences between both groups in the morning (P = 0.635) and throughout the day (P = 0.681), even after adjusting for confounders. It can be concluded that diurnal changes of the IVDs occurred in the entire (including cervical and thoracic) spine. WL and NWL showed similar morning baseline T2-values and diurnal changes. Weightlifting may not negatively affect IVDs chronically or acutely.

Reference values of physiological 18F-FET uptake: Implications for brain tumor discrimination.

PURPOSE: The aim of this study was to derive reference values of 18F-fluoro-ethyl-L-tyrosine positron emission tomography (18F-FET-PET) uptake in normal brain and head structures to allow for differentiation from tumor tissue. MATERIALS AND METHODS: We examined the datasets of 70 patients (median age 53 years, range 15-79), whose dynamic 18F-FET-PET was acquired between January 2016 and October 2017. Maximum standardized uptake value (SUVmax), target-to-background standardized uptake value ratio (TBR), and time activity curve (TAC) of the 18F-FET-PET were assessed in tumor tissue and in eight normal anatomic structures and compared using the t-test and Mann-Whitney U-test. Correlation analyses were performed using Pearson or Spearman coefficients, and comparisons between several variables with Pearson's chi-squared tests and Kruskal-Wallis tests as well as the Benjamini-Hochberg correction. RESULTS: All analyzed structures showed an 18F-FET uptake higher than background (threshold: TBR > 1.5). The venous sinuses and cranial muscles exhibited a TBR of 2.03±0.46 (confidence interval (CI) 1.92-2.14), higher than the uptake of caudate nucleus, pineal gland, putamen, and thalamus (TBR 1.42±0.17, CI 1.38-1.47). SUVmax, TBR, and TAC showed no difference in the analyzed structures between subjects with high-grade gliomas and subjects with low-grade gliomas, except the SUVmax of the pineal gland (t-tests of the pineal gland: SUVmax: p = 0.022; TBR: p = 0.411). No significant differences were found for gender and age. CONCLUSION: Normal brain tissue demonstrates increased 18F-FET uptake compared to background tissue. Two distinct clusters have been identified, comprising venous structures and gray matter with a reference uptake of up to SUVmax of 2.99 and 2.33, respectively.

ELIMU-MDx: a web-based, open-source platform for storage, management and analysis of diagnostic qPCR data.

The Electronic Laboratory Information and Management Utensil for Molecular Diagnostics (ELIMU-MDx) is a user-friendly platform designed and built to accelerate the turnaround time of diagnostic qPCR assays. ELIMU-MDx is compliant with Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines and has extensive data-import capabilities for all major qPCR instruments by using the RDML data standard. This platform was designed as an open-source software tool and can be accessed through the web browser on all major operating systems.

In Vivo Evidence for a Lactate Gradient from Astrocytes to Neurons.

Investigating lactate dynamics in brain tissue is challenging, partly because in vivo data at cellular resolution are not available. We monitored lactate in cortical astrocytes and neurons of mice using the genetically encoded FRET sensor Laconic in combination with two-photon microscopy. An intravenous lactate injection rapidly increased the Laconic signal in both astrocytes and neurons, demonstrating high lactate permeability across tissue. The signal increase was significantly smaller in astrocytes, pointing to higher basal lactate levels in these cells, confirmed by a one-point calibration protocol. Trans-acceleration of the monocarboxylate transporter with pyruvate was able to reduce intracellular lactate in astrocytes but not in neurons. Collectively, these data provide in vivo evidence for a lactate gradient from astrocytes to neurons. This gradient is a prerequisite for a carrier-mediated lactate flux from astrocytes to neurons and thus supports the astrocyte-neuron lactate shuttle model, in which astrocyte-derived lactate acts as an energy substrate for neurons.

NH4(+) triggers the release of astrocytic lactate via mitochondrial pyruvate shunting.

Neural activity is accompanied by a transient mismatch between local glucose and oxygen metabolism, a phenomenon of physiological and pathophysiological importance termed aerobic glycolysis. Previous studies have proposed glutamate and K(+) as the neuronal signals that trigger aerobic glycolysis in astrocytes. Here we used a panel of genetically encoded FRET sensors in vitro and in vivo to investigate the participation of NH4(+), a by-product of catabolism that is also released by active neurons. Astrocytes in mixed cortical cultures responded to physiological levels of NH4(+) with an acute rise in cytosolic lactate followed by lactate release into the extracellular space, as detected by a lactate-sniffer. An acute increase in astrocytic lactate was also observed in acute hippocampal slices exposed to NH4(+) and in the somatosensory cortex of anesthetized mice in response to i.v. NH4(+). Unexpectedly, NH4(+) had no effect on astrocytic glucose consumption. Parallel measurements showed simultaneous cytosolic pyruvate accumulation and NADH depletion, suggesting the involvement of mitochondria. An inhibitor-stop technique confirmed a strong inhibition of mitochondrial pyruvate uptake that can be explained by mitochondrial matrix acidification. These results show that physiological NH4(+) diverts the flux of pyruvate from mitochondria to lactate production and release. Considering that NH4(+) is produced stoichiometrically with glutamate during excitatory neurotransmission, we propose that NH4(+) behaves as an intercellular signal and that pyruvate shunting contributes to aerobic lactate production by astrocytes.

Channel-mediated lactate release by K⁺-stimulated astrocytes.

Excitatory synaptic transmission is accompanied by a local surge in interstitial lactate that occurs despite adequate oxygen availability, a puzzling phenomenon termed aerobic glycolysis. In addition to its role as an energy substrate, recent studies have shown that lactate modulates neuronal excitability acting through various targets, including NMDA receptors and G-protein-coupled receptors specific for lactate, but little is known about the cellular and molecular mechanisms responsible for the increase in interstitial lactate. Using a panel of genetically encoded fluorescence nanosensors for energy metabolites, we show here that mouse astrocytes in culture, in cortical slices, and in vivo maintain a steady-state reservoir of lactate. The reservoir was released to the extracellular space immediately after exposure of astrocytes to a physiological rise in extracellular K(+) or cell depolarization. Cell-attached patch-clamp analysis of cultured astrocytes revealed a 37 pS lactate-permeable ion channel activated by cell depolarization. The channel was modulated by lactate itself, resulting in a positive feedback loop for lactate release. A rapid fall in intracellular lactate levels was also observed in cortical astrocytes of anesthetized mice in response to local field stimulation. The existence of an astrocytic lactate reservoir and its quick mobilization via an ion channel in response to a neuronal cue provides fresh support to lactate roles in neuronal fueling and in gliotransmission.