Exosome-derived CIRP: An amplifier of inflammatory diseases.

Cold-inducible RNA-binding protein (CIRP) is an intracellular stress-response protein and a type of damage-associated molecular pattern (DAMP) that responds to various stress stimulus by altering its expression and mRNA stability. Upon exposure to ultraviolet (UV) light or low temperature, CIRP get translocated from the nucleus to the cytoplasm through methylation modification and stored in stress granules (SG). During exosome biogenesis, which involves formation of endosomes from the cell membrane through endocytosis, CIRP also gets packaged within the endosomes along with DNA, and RNA and other proteins. Subsequently, intraluminal vesicles (ILVs) are formed following the inward budding of the endosomal membrane, turning the endosomes into multi-vesicle bodies (MVBs). Finally, the MVBs fuse with the cell membrane to form exosomes. As a result, CIRP can also be secreted out of cells through the lysosomal pathway as Extracellular CIRP (eCIRP). Extracellular CIRP (eCIRP) is implicated in various conditions, including sepsis, ischemia-reperfusion damage, lung injury, and neuroinflammation, through the release of exosomes. In addition, CIRP interacts with TLR4, TREM-1, and IL-6R, and therefore are involved in triggering immune and inflammatory responses. Accordingly, eCIRP has been studied as potential novel targets for disease therapy. C23 and M3, polypeptides that oppose eCIRP binding to its receptors, are beneficial in numerous inflammatory illnesses. Some natural molecules such as Luteolin and Emodin can also antagonize CIRP, which play roles similar to C23 in inflammatory responses and inhibit macrophage-mediated inflammation. This review aims to provide a better understanding on CIRP translocation and secretion from the nucleus to the extracellular space and the mechanisms and inhibitory roles of eCIRP in diverse inflammatory illnesses.

Chloride ions evoke taste sensations by binding to the extracellular ligand-binding domain of sweet/umami taste receptors.

Salt taste sensation is multifaceted: NaCl at low or high concentrations is preferably or aversively perceived through distinct pathways. Cl- is thought to participate in taste sensation through an unknown mechanism. Here, we describe Cl- ion binding and the response of taste receptor type 1 (T1r), a receptor family composing sweet/umami receptors. The T1r2a/T1r3 heterodimer from the medaka fish, currently the sole T1r amenable to structural analyses, exhibited a specific Cl- binding in the vicinity of the amino-acid-binding site in the ligand-binding domain (LBD) of T1r3, which is likely conserved across species, including human T1r3. The Cl- binding induced a conformational change in T1r2a/T1r3LBD at sub- to low-mM concentrations, similar to canonical taste substances. Furthermore, oral Cl- application to mice increased impulse frequencies of taste nerves connected to T1r-expressing taste cells and promoted their behavioral preferences attenuated by a T1r-specific blocker or T1r3 knock-out. These results suggest that the Cl- evokes taste sensations by binding to T1r, thereby serving as another preferred salt taste pathway at a low concentration.

Humans perceive taste when proteins called taste receptors on the surface of the tongue are activated by molecules of food. These receptors turn on nerve cells that send signals the brain can read as sweet, sour, salty, bitter, or umami, depending on which receptor was activated. Most animals with backbones share the same five types of taste receptors. In food, salty flavors are usually the result of adding table salt, which has two components: a sodium ion and chloride ion. The main taste receptors that signal to the brain that a food is salty become activated when they bind to the sodium ion. However, some studies have shown that salt is also perceived as sweet when eaten in minuscule amounts. It is poorly understood why this happens, but it is possible that the chloride half of salt drives the sweet taste. In 2017, scientists worked out the structure of a taste receptor from a fish, that is equivalent to the sweet receptor in humans. Curiously, one part of this receptor, known as T1r2a/T1r3LBD, was bound to a chloride ion. This prompted Atsumi, Yasumatsu et al. to think about the 'sweet' taste of salt, leading them to take a closer look at T1r2a/T1r3LBD and whether chloride could indeed activate it. Atsumi, Yasumatsu et al. used structural biology techniques to examine T1r2a/T1r3LBD and found evidence that the receptor might be binding chloride. Further biophysical experiments confirmed that chloride does indeed bind to the receptor, and that it also causes it to change shape. Usually, changes in shape are hallmarks of receptor activation, suggesting that chloride may activate T1r2a/T1r3LBD. Next, Atsumi, Yasumatsu et al. checked whether chloride could stimulate the neurons that signal when food tastes sweet, by using an approach known as electrophysiology to measure the activity of these neurons in mice. The results showed that the neurons became active when a solution containing small amounts of chloride was placed on the mouse's tongue. This activity went away when a compound that can block the receptor's activity was delivered alongside the chloride. Additionally, when mice were given a choice of plain water or water containing chloride, they seemed to prefer the latter. This confirmed that mice recognized the sweetness of chloride via the activation of sweet taste receptors and neurons. Based on these findings, Atsumi, Yasumatsu et al. propose that small amounts of salt may taste sweet because the chloride ions in the salt activate sweet taste receptors and their linked neurons. Their results also suggest that animals sense salt in many ways, likely because balanced salt levels are essential for the body to work properly. Future experiments on human taste receptors may reveal how these pathways help assess salt levels in humans.

Success probability of high-affinity DNA aptamer generation by genetic alphabet expansion.

Nucleic acid aptamers as antibody alternatives bind specifically to target molecules. These aptamers are generated by isolating candidates from libraries with random sequence fragments, through an evolutionary engineering system. We recently reported a high-affinity DNA aptamer generation method that introduces unnatural bases (UBs) as a fifth letter into the library, by genetic alphabet expansion. By incorporating hydrophobic UBs, the affinities of DNA aptamers to target proteins are increased over 100-fold, as compared with those of conventional aptamers with only the natural four letters. However, there is still plenty of room for improvement of the methods for routinely generating high-affinity UB-containing DNA (UB-DNA) aptamers. The success probabilities of the high-affinity aptamer generation depend on the existence of the aptamer candidate sequences in the initial library. We estimated the success probabilities by analysing several UB-DNA aptamers that we generated, as examples. In addition, we investigated the possible improvement of conventional aptamer affinities by introducing one UB at specific positions. Our data revealed that UB-DNA aptamers adopt specific tertiary structures, in which many bases including UBs interact with target proteins for high affinity, suggesting the importance of the UB-DNA library design. This article is part of the theme issue 'Reactivity and mechanism in chemical and synthetic biology'.

A novel interaction between extracellular vimentin and fibrinogen in fibrin formation.

INTRODUCTION: Thrombosis is frequently manifested in critically ill patients with systemic inflammation, including sepsis and COVID-19. The coagulopathy in systemic inflammation is often associated with increased levels of fibrinogen and D-dimer. Because elevated levels of vimentin have been detected in sepsis, we sought to investigate the relationship between vimentin and the increased fibrin formation potential observed in these patients. MATERIALS AND METHODS: This hypothesis was examined by using recombinant human vimentin, anti-vimentin antibodies, plasma derived from healthy and critically ill patients, confocal microscopy, co-immunoprecipitation assays, and size exclusion chromatography. RESULTS: The level of vimentin in plasma derived from critically ill subjects with systemic inflammation was on average two-fold higher than that of healthy volunteers. We determined that vimentin directly interacts with fibrinogen and enhances fibrin formation. Anti-vimentin antibody effectively blocked fibrin formation ex vivo and caused changes in the fibrin structure in plasma. Additionally, confocal imaging demonstrated plasma vimentin enmeshed in the fibrin fibrils. Size exclusion chromatography column and co-immunoprecipitation assays demonstrated a direct interaction between extracellular vimentin and fibrinogen in plasma from critically ill patients but not in healthy plasma. CONCLUSIONS: The results describe that extracellular vimentin engages fibrinogen in fibrin formation. In addition, the data suggest that elevated levels of an apparent aberrant extracellular vimentin potentiate fibrin clot formation in critically ill patients with systemic inflammation; consistent with the notion that plasma vimentin contributes to the pathogenesis of thrombosis.

hERG1 channel subunit composition mediates proton inhibition of rapid delayed rectifier potassium current (IKr) in cardiomyocytes derived from hiPSCs.

The voltage-gated channel, hERG1, conducts the rapid delayed rectifier potassium current (IKr) and is critical for human cardiac repolarization. Reduced IKr causes long QT syndrome and increases the risk for cardiac arrhythmia and sudden death. At least two subunits form functional hERG1 channels, hERG1a and hERG1b. Changes in hERG1a/1b abundance modulate IKr kinetics, magnitude, and drug sensitivity. Studies from native cardiac tissue suggest that hERG1 subunit abundance is dynamically regulated, but the impact of altered subunit abundance on IKr and its response to external stressors is not well understood. Here, we used a substrate-driven human-induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) maturation model to investigate how changes in relative hERG1a/1b subunit abundance impact the response of native IKr to extracellular acidosis, a known component of ischemic heart disease and sudden infant death syndrome. IKr recorded from immatured hiPSC-CMs displays a 2-fold greater inhibition by extracellular acidosis (pH 6.3) compared with matured hiPSC-CMs. Quantitative RT-PCR and immunocytochemistry demonstrated that hERG1a subunit mRNA and protein were upregulated and hERG1b subunit mRNA and protein were downregulated in matured hiPSC-CMs compared with immatured hiPSC-CMs. The shift in subunit abundance in matured hiPSC-CMs was accompanied by increased IKr. Silencing hERG1b's impact on native IKr kinetics by overexpressing a polypeptide identical to the hERG1a N-terminal Per-Arnt-Sim domain reduced the magnitude of IKr proton inhibition in immatured hiPSC-CMs to levels comparable to those observed in matured hiPSC-CMs. These data demonstrate that hERG1 subunit abundance is dynamically regulated and determines IKr proton sensitivity in hiPSC-CMs.

Local diffusion in the extracellular space of the brain.

The brain extracellular space (ECS) is a vast interstitial reticulum of extreme morphological complexity, composed of narrow gaps separated by local expansions, enabling interconnected highways between neural cells. Constituting on average 20% of brain volume, the ECS is key for intercellular communication, and understanding its diffusional properties is of paramount importance for understanding the brain. Within the ECS, neuroactive substances travel predominantly by diffusion, spreading through the interstitial fluid and the extracellular matrix scaffold after being focally released. The nanoscale dimensions of the ECS render it unresolvable by conventional live tissue compatible imaging methods, and historically diffusion of tracers has been used to indirectly infer its structure. Novel nanoscopic imaging techniques now show that the ECS is a highly dynamic compartment, and that diffusivity in the ECS is more heterogeneous than anticipated, with great variability across brain regions and physiological states. Diffusion is defined primarily by the local ECS geometry, and secondarily by the viscosity of the interstitial fluid, including the obstructive and binding properties of the extracellular matrix. ECS volume fraction and tortuosity both strongly determine diffusivity, and each can be independently regulated e.g. through alterations in glial morphology and the extracellular matrix composition. Here we aim to provide an overview of our current understanding of the ECS and its diffusional properties. We highlight emerging technological advances to respectively interrogate and model diffusion through the ECS, and point out how these may contribute in resolving the remaining enigmas of the ECS.

Capture of Electrophilic Quinones in the Extracellular Space: Evidence for a Phase Zero Reaction.

Electrophilic quinones are produced during the combustion of gasoline in the atmosphere. Although these reactive species covalently bind to protein-based nucleophiles in cells, resulting in the formation of protein adducts involved in the modulation of redox signaling pathways and cytotoxicity, the extracellular regulation of quinones is not understood. In this study, incubation of 1,2-naphthoquinone (1,2-NQ) with the low-molecular-weight fraction of mouse plasma resulted in the consumption of cysteine (CysSH) in the plasma in a concentration-dependent manner. Covalent modification of albumin was markedly repressed by the addition of either the low-molecular-weight fraction of mouse plasma or CysSH, suggesting that CysSH protects by forming a conjugate with 1,2-NQ. Similar phenomena also occurred for other atmospheric quinones 1,4-NQ and 1,4-benzoquinone (1,4-BQ). The addition of cystine to a culture medium without amino acids enhanced the release of CysSH from A431 cells and blocked 1,2-NQ-mediated arylation of intracellular proteins, suggesting that 1,2-NQ interacts with extracellular CysSH. Liquid chromatography-tandem mass spectrometry analysis revealed that 1,2-NQ and 1,4-BQ undergoes nucleophilic attack by CysSH, yielding a 1,2-NQH2-SCys adduct and 1,4-BQH2-SCys adduct, respectively. Unlike 1,2-NQ and 1,4-BQ, the authentic 1,2-NQH2-SCys adduct and 1,4-BQH2-SCys adduct had little effect on the covalent modification of cellular proteins and viability of A431 cells. These results suggest that electrophilic quinones are readily trapped by CysSH released from A431 cells, forming less-toxic CysSH adducts and thereby repressing covalent modification of cellular proteins. These findings provide evidence for the existence of a "phase zero" reaction of electrophiles prior to their uptake by cells.

Is There a Role for Immunoregulatory and Antiviral Oligonucleotides Acting in the Extracellular Space? A Review and Hypothesis.

Here, we link approved and emerging nucleic acid-based therapies with the expanding universe of small non-coding RNAs (sncRNAs) and the innate immune responses that sense oligonucleotides taken up into endosomes. The Toll-like receptors (TLRs) 3, 7, 8, and 9 are located in endosomes and can detect nucleic acids taken up through endocytic routes. These receptors are key triggers in the defense against viruses and/or bacterial infections, yet they also constitute an Achilles heel towards the discrimination between self- and pathogenic nucleic acids. The compartmentalization of nucleic acids and the activity of nucleases are key components in avoiding autoimmune reactions against nucleic acids, but we still lack knowledge on the plethora of nucleic acids that might be released into the extracellular space upon infections, inflammation, and other stress responses involving increased cell death. We review recent findings that a set of single-stranded oligonucleotides (length of 25-40 nucleotides (nt)) can temporarily block ligands destined for endosomes expressing TLRs in human monocyte-derived dendritic cells. We discuss knowledge gaps and highlight the existence of a pool of RNA with an approximate length of 30-40 nt that may still have unappreciated regulatory functions in physiology and in the defense against viruses as gatekeepers of endosomal uptake through certain routes.

Cancer-derived extracellular succinate: a driver of cancer metastasis.

Succinate is a tricarboxylic acid (TCA) cycle intermediate normally confined to the mitochondrial matrix. It is a substrate of succinate dehydrogenase (SDH). Mutation of SDH subunits (SDHD and SDHB) in hereditary tumors such as paraganglioma or reduction of SDHB expression in cancer results in matrix succinate accumulation which is transported to cytoplasma and secreted into the extracellular milieu. Excessive cytosolic succinate is known to stabilize hypoxia inducible factor-1α (HIF-1α) by inhibiting prolyl hydroxylase. Recent reports indicate that cancer-secreted succinate enhances cancer cell migration and promotes cancer metastasis by activating succinate receptor-1 (SUCNR-1)-mediated signaling and transcription pathways. Cancer-derived extracellular succinate enhances cancer cell and macrophage migration through SUCNR-1 â†’ PI-3 K â†’ HIF-1α pathway. Extracellular succinate induces tumor angiogenesis through SUCNR-1-mediated ERK1/2 and STAT3 activation resulting in upregulation of vascular endothelial growth factor (VEGF) expression. Succinate increases SUCNR-1 expression in cancer cells which is considered as a target for developing new anti-metastasis drugs. Furthermore, serum succinate which is elevated in cancer patients may be a theranostic biomarker for selecting patients for SUCNR-1 antagonist therapy.

Transport in the Brain Extracellular Space: Diffusion, but Which Kind?

The mechanisms of transport of substances in the brain parenchyma have been a hot topic in scientific discussion in the past decade. This discussion was triggered by the proposed glymphatic hypothesis, which assumes a directed flow of cerebral fluid within the parenchyma, in contrast to the previous notion that diffusion is the main mechanism. However, when discussing the issue of "diffusion or non-diffusion", much less attention was given to the question that diffusion itself can have a different character. In our opinion, some of the recently published results do not fit into the traditional understanding of diffusion. In this regard, we outline the relevant new theoretical approaches on transport processes in complex random media such as concepts of diffusive diffusivity and time-dependent homogenization, which expands the understanding of the forms of transport of substances based on diffusion.

Intracellular-to-extracellular localization switch of acidic lipase in Enterobacter cloacae through multi-objective medium optimization: aqueous two-phase purification and activity kinetics.

As physiological impairments that require replacement therapy continue to increase, so also does the need for improved production of acidic lipase from new microbial sources. Enterobacter cloacae strain UCCM 00116 produced a novel acidic lipase in kernel oil-processing waste-basal broth with 0.023:1 extracellular: intracellular localization ratio. This research re-directed enzyme localization to the extracellular milieu to reduce recovery cost using multi-objective response surface optimization of medium parameters. Results revealed a 1:0.32 extracellular:intracellular lipase ratio. Product formation kinetics, modeled by the Luedeking-Piret function, showed a significant switch from a completely growth-associated intracellular production to a predominantly non-growth-associated extracellular localization through medium optimization. Aqueous two-phase system purification conditions extracted 95.22% lipase with 72.36 purity, a Vmax of 370.37 µmolmin-1, and a Km of 0.63 mmol. Enzyme activity was enhanced by K+ and Ca2+ ions, stable in many organic solvents, except acetone, and had pH and temperature optima at 2.5-3.5 and 50 °C, respectively.

Possible involvement of extracellular polymeric substrates of Antarctic cyanobacterium Nostoc sp. strain SO-36 in adaptation to harsh environments.

Cyanobacteria are some of the primary producers in extremely cold biospheres such as the Arctic, Antarctic, and vast ice sheets. Many genera of cyanobacteria are identified from these harsh environments, but their specific mechanisms for cold adaptation are not fully understood. Nostoc sp. strain SO-36 is a cyanobacterium isolated in Antarctica more than 30 years ago and regarded as a psychrotolelant species. To determine whether the strain is psychrotolelant or psychrophilic, it was first grown at 30 °C and 10 °C. The cells grew exponentially at 30 °C, but their growth stopped at 10 °C, indicating that the strain is only psychrotolerant. Microscopic analysis revealed that the morphology of the cells grown at 30 °C was filamentous and differentiated heterocysts, which are specialized cells for gaseous nitrogen fixation under nitrogen-deprived conditions, indicating that the strain can grow diazotrophically. The cells grown at 10 °C have a smaller size, shortened filament length and decreased chlorophyll content per cell. At 10 °C, the cells are aggregated with extracellular polymeric substrates (EPSs), which is a common mechanism to protect cells from ultraviolet light. These results imply that segmentation into short filaments was induced by photodamage at low temperatures. To fully understand the adaptation mechanisms of Nostoc sp. strain SO-36 for low-temperature conditions, next-generation sequencing analyses were conducted. Complete genome sequence of the strain revealed that it has one main chromosome of approximately 6.8 Mbp with 4 plasmids, including 6855 coding sequences, 48 tRNA genes, 4 copies of rRNA operons, and 5 CRISPR regions. Putative genes for EPS biosynthesis were found to be conserved in Nostocaceae regardless of their habitat. These results provide basic information to understand the adaptation mechanisms at low temperatures, and the strain can be a model organism to analyze adaptation to extreme environments.

Near-Infrared Carbon Nanotube Tracking Reveals the Nanoscale Extracellular Space around Synapses.

We provide evidence of a local synaptic nanoenvironment in the brain extracellular space (ECS) lying within 500 nm of postsynaptic densities. To reveal this brain compartment, we developed a correlative imaging approach dedicated to thick brain tissue based on single-particle tracking of individual fluorescent single wall carbon nanotubes (SWCNTs) in living samples and on speckle-based HiLo microscopy of synaptic labels. We show that the extracellular space around synapses bears specific properties in terms of morphology at the nanoscale and inner diffusivity. We finally show that the ECS juxta-synaptic region changes its diffusion parameters in response to neuronal activity, indicating that this nanoenvironment might play a role in the regulation of brain activity.

Thiol Modifications in the Extracellular Space-Key Proteins in Inflammation and Viral Infection.

Posttranslational modifications (PTMs) allow to control molecular and cellular functions in response to specific signals and changes in the microenvironment of cells. They regulate structure, localization, stability, and function of proteins in a spatial and temporal manner. Among them, specific thiol modifications of cysteine (Cys) residues facilitate rapid signal transduction. In fact, Cys is unique because it contains the highly reactive thiol group that can undergo different reversible and irreversible modifications. Upon inflammation and changes in the cellular microenvironment, many extracellular soluble and membrane proteins undergo thiol modifications, particularly dithiol-disulfide exchange, S-glutathionylation, and S-nitrosylation. Among others, these thiol switches are essential for inflammatory signaling, regulation of gene expression, cytokine release, immunoglobulin function and isoform variation, and antigen presentation. Interestingly, also the redox state of bacterial and viral proteins depends on host cell-mediated redox reactions that are critical for invasion and infection. Here, we highlight mechanistic thiol switches in inflammatory pathways and infections including cholera, diphtheria, hepatitis, human immunodeficiency virus (HIV), influenza, and coronavirus disease 2019 (COVID-19).

Cellular mechanisms mediating activity-dependent extracellular space shrinkage in the retina.

Volume transmission plays an essential role in CNS function, with neurotransmitters released from synapses diffusing through the extracellular space (ECS) to distant sites. Changes in the ECS volume fraction (α) will influence the diffusion and the concentration of transmitters within the ECS. We have recently shown that neuronal activity evoked by physiological photic stimuli results in rapid decreases in ECS α as large as 10% in the retina. We now characterize the cellular mechanisms responsible for this ECS shrinkage. We find that block of inwardly rectifying K+ channels with Ba2+ , inhibition of the Na+ /K+ /2Cl- cotransporter with bumetanide, or block of AQP4 water channels with TGN-020 do not diminish the light-evoked ECS decrease. Inhibition of the Na+ /HCO3 - cotransporter by removing HCO3 - from the superfusate, in contrast, reduces the light-evoked ECS decrease by 95.6%. Inhibition of the monocarboxylate transporter with alpha-cyano-4-hydroxycinnamate (4-CIN) also reduces the ECS shrinkage, but only by 32.5%. We tested whether the swelling of Müller cells, the principal glial cells of the retina, is responsible for the light-evoked ECS shrinkage. Light stimulation evoked a 6.3% increase in the volume of the fine processes of Müller cells. This volume increase was reduced by 97.1% when HCO3 - was removed from the superfusate. We conclude that a large fraction of the activity-dependent decrease in ECS α is generated by the activation of the Na+ /HCO3 - cotransporter in Müller cells. The monocarboxylate transporter may also contribute to the response.

Unassembled cell wall proteins form aggregates in the extracellular space of Chlamydomonas reinhardtii strain UVM4.

The green microalga Chlamydomonas reinhardtii is emerging as a promising cell biofactory for secreted recombinant protein (RP) production. In recent years, the generation of the broadly used cell wall-deficient mutant strain UVM4 has allowed for a drastic increase in secreted RP yields. However, purification of secreted RPs from the extracellular space of C. reinhardtii strain UVM4 is challenging. Previous studies suggest that secreted RPs are trapped in a matrix of cell wall protein aggregates populating the secretome of strain UVM4, making it difficult to isolate and purify the RPs. To better understand the nature and behaviour of these extracellular protein aggregates, we analysed and compared the extracellular proteome of the strain UVM4 to its cell-walled ancestor, C. reinhardtii strain 137c. When grown under the same conditions, strain UVM4 produced a unique extracellular proteomic profile, including a higher abundance of secreted cell wall glycoproteins. Further characterization of high molecular weight extracellular protein aggregates in strain UVM4 revealed that they are largely comprised of pherophorins, a specific class of cell wall glycoproteins. Our results offer important new insights into the extracellular space of strain UVM4, including strain-specific secreted cell wall proteins and the composition of the aggregates possibly related to impaired RP purification. The discovery of pherophorins as a major component of extracellular protein aggregates will inform future strategies to remove or prevent aggregate formation, enhance purification of secreted RPs, and improve yields of recombinant biopharmaceuticals in this emerging cell biofactory. KEY POINTS: • Extracellular protein aggregates hinder purification of recombinant proteins in C. reinhardtii • Unassembled cell wall pherophorins are major components of extracellular protein aggregates • Known aggregate composition informs future strategies for recombinant protein purification.

Honey, I shrunk the extracellular space: Measurements and mechanisms of astrocyte swelling.

Astrocyte volume fluctuation is a physiological phenomenon tied closely to the activation of neural circuits. Identification of underlying mechanisms has been challenging due in part to use of a wide range of experimental approaches that vary between research groups. Here, we first review the many methods that have been used to measure astrocyte volume changes directly or indirectly. While the field has recently shifted towards volume analysis using fluorescence microscopy to record cell volume changes directly, established metrics corresponding to extracellular space dynamics have also yielded valuable insights. We then turn to analysis of mechanisms of astrocyte swelling derived from many studies, with a focus on volume changes tied to increases in extracellular potassium concentration ([K+ ]o ). The diverse methods that have been utilized to generate the external [K+ ]o environment highlight multiple scenarios of astrocyte swelling mediated by different mechanisms. Classical potassium buffering theories are tempered by many recent studies that point to different swelling pathways optimized at particular [K+ ]o and that depend on local/transient versus more sustained increases in [K+ ]o .

Arterial vasodilation drives convective fluid flow in the brain: a poroelastic model.

The movement of fluid into, through, and out of the brain plays an important role in clearing metabolic waste. However, there is controversy regarding the mechanisms driving fluid movement in the fluid-filled paravascular spaces (PVS), and whether the movement of metabolic waste in the brain extracellular space (ECS) is primarily driven by diffusion or convection. The dilation of penetrating arterioles in the brain in response to increases in neural activity (neurovascular coupling) is an attractive candidate for driving fluid circulation, as it drives deformation of the brain tissue and of the PVS around arteries, resulting in fluid movement. We simulated the effects of vasodilation on fluid movement into and out of the brain ECS using a novel poroelastic model of brain tissue. We found that arteriolar dilations could drive convective flow through the ECS radially outward from the arteriole, and that this flow is sensitive to the dynamics of the dilation. Simulations of sleep-like conditions, with larger vasodilations and increased extracellular volume in the brain showed enhanced movement of fluid from the PVS into the ECS. Our simulations suggest that both sensory-evoked and sleep-related arteriolar dilations can drive convective flow of cerebrospinal fluid not just in the PVS, but also into the ECS through the PVS around arterioles.