In Vivo Brain Microdialysis.

Many biological molecules in the brain interstitial fluid are involved in neuronal functions. Therefore, measuring the levels of these molecules in the extracellular fluid would provide deep insights into the physiological/pathological mechanisms underlying brain functions/disorders. In vivo microdialysis is a powerful technique used to examine the extracellular levels of various molecules in the brains of living animals. In neuroscience research, this technique has been widely used to investigate relatively small molecules including neurotransmitters and amino acids. However, recent advances in technology have made it possible to assess large molecules in the brain interstitial fluid, such as signaling peptides and proteins, using microdialysis probes with high-molecular-weight cutoff membranes. This chapter describes an in vivo microdialysis method to collect and measure the levels of large biological molecules in the extracellular fluid of the brains of freely moving mice.

The Feasibility of Semi-Continuous and Multi-Frequency Thoracic Bioimpedance Measurements by a Wearable Device during Fluid Changes in Hemodialysis Patients.

Repeated single-point measurements of thoracic bioimpedance at a single (low) frequency are strongly related to fluid changes during hemodialysis. Extension to semi-continuous measurements may provide longitudinal details in the time pattern of the bioimpedance signal, and multi-frequency measurements may add in-depth information on the distribution between intra- and extracellular fluid. This study aimed to investigate the feasibility of semi-continuous multi-frequency thoracic bioimpedance measurements by a wearable device in hemodialysis patients. Therefore, thoracic bioimpedance was recorded semi-continuously (i.e., every ten minutes) at nine frequencies (8-160 kHz) in 68 patients during two consecutive hemodialysis sessions, complemented by a single-point measurement at home in-between both sessions. On average, the resistance signals increased during both hemodialysis sessions and decreased during the interdialytic interval. The increase during dialysis was larger at 8 kHz (∆ 32.6 Ω during session 1 and ∆ 10 Ω during session 2), compared to 160 kHz (∆ 29.5 Ω during session 1 and ∆ 5.1 Ω during session 2). Whereas the resistance at 8 kHz showed a linear time pattern, the evolution of the resistance at 160 kHz was significantly different (p < 0.0001). Measuring bioimpedance semi-continuously and with a multi-frequency current is a major step forward in the understanding of fluid dynamics in hemodialysis patients. This study paves the road towards remote fluid monitoring.

A Review of Recent Advances in Microneedle-Based Sensing within the Dermal ISF That Could Transform Medical Testing.

Interstitial fluid (ISF) has attracted extensive attention in an extremely wide range of areas due to its unique advantages, such as portability, high precision, comfortable operation, and superior stability. In recent years, the microneedle (MN) technique has been considered to be an excellent tool for extracting ISF because it is painless and noninvasive. Recent reports have shown that MN has good application prospects in ISF extraction. In this review, we provide comprehensive and in-depth insight into integrated MN devices for ISF detection, covering the basic structure as well as the fabrication of integrated MN devices and various applications in ISF extraction. Challenges and prospects are highlighted, with a discussion on how to transition such MN-integrated devices toward personalized healthcare monitoring systems.

A Method to Collect Cerebrospinal Fluid from Mouse Cisterna Magna to Determine Extracellular Tau Levels.

Despite being a cytoplasmic protein abundant in neurons, tau is detectable in various extracellular fluids. In addition to being passively released from dying/degenerating neurons, tau is also actively released from living neurons in a neuronal activity-dependent mechanism. In vivo, tau released from neurons first appears in brain interstitial fluid (ISF) and subsequently drains into cerebrospinal fluid (CSF) by glymphatic system. Changes in CSF tau levels alter during the course of AD pathogenesis and are considered to predict the disease-progression of AD. A method to collect CSF from various mouse models of AD will serve as a valuable tool to investigate the dynamics of physiological/pathological tau released from neurons. In this chapter, we describe and characterize a method that reliably collects a relatively large volume of CSF from anesthetized mice.

Detection of Glymphatic Outflow of Tau from Brain to Cerebrospinal Fluid in Mice.

Glymphatic system denotes a brain-wide pathway that eliminates extracellular solutes from brain. It is driven by the flow of brain interstitial fluid (ISF) and cerebrospinal fluid (CSF) via perivascular spaces. Glymphatic convective flow is driven by cerebral arterial pulsation, which is facilitated by a water channel, aquaporin-4 (AQP4) expressed in astrocytic end-foot processes. Since its discovery, the glymphatic system receives a considerable scientific attention due to its pivotal role in clearing metabolic waste as well as neurotoxic substances such as amyloid b peptide. Tau is a microtubule binding protein, however it is also physiologically released into extracellular fluids. The presence of tau in the blood stream indicates that it is eventually cleared from the brain to the periphery, however, the detailed mechanisms that eliminate extracellular tau from the central nervous system remained to be elucidated. Recently, we and others have reported that extracellular tau is eliminated from the brain to CSF by an AQP4 dependent mechanism, suggesting the involvement of the glymphatic system. In this chapter, we describe the detailed protocol of how we can assess glymphatic outflow of tau protein from brain to CSF in mice.

Cell adhesion affects the properties of interstitial fluid flow: A study using multiscale poroelastic composite modeling.

In this study, we conduct a multiscale, multiphysics modeling of the brain gray matter as a poroelastic composite. We develop a customized representative volume element based on cytoarchitectural features that encompass important microscopic components of the tissue, namely the extracellular space, the capillaries, the pericapillary space, the interstitial fluid, cell-cell and cell-capillary junctions, and neuronal and glial cell bodies. Using asymptotic homogenization and direct numerical simulation, the effective properties at the tissue level are identified based on microscopic properties. To analyze the influence of various microscopic elements on the effective/macroscopic properties and tissue response, we perform sensitivity analyses on cell junction (cluster) stiffness, cell junction diameter (dimensions), and pericapillary space width. The results of this study suggest that changes in cell adhesion can greatly affect both mechanical and hydraulic (interstitial fluid flow and porosity) features of brain tissue, consistent with the effects of neurodegenerative diseases.

Modeling Healthy and Dysbiotic Vaginal Microenvironments in a Human Vagina-on-a-Chip.

Women's health, and particularly diseases of the female reproductive tract (FRT), have not received the attention they deserve, even though an unhealthy reproductive system may lead to life-threatening diseases, infertility, or adverse outcomes during pregnancy. One barrier in the field is that there has been a dearth of preclinical, experimental models that faithfully mimic the physiology and pathophysiology of the FRT. Current in vitro and animal models do not fully recapitulate the hormonal changes, microaerobic conditions, and interactions with the vaginal microbiome. The advent of Organ-on-a-Chip (Organ Chip) microfluidic culture technology that can mimic tissue-tissue interfaces, vascular perfusion, interstitial fluid flows, and the physical microenvironment of a major subunit of human organs can potentially serve as a solution to this problem. Recently, a human Vagina Chip that supports co-culture of human vaginal microbial consortia with primary human vaginal epithelium that is also interfaced with vaginal stroma and experiences dynamic fluid flow has been developed. This chip replicates the physiological responses of the human vagina to healthy and dysbiotic microbiomes. A detailed protocol for creating human Vagina Chips has been described in this article.

Effect of interstitial fluid pressure on shear wave elastography: an experimental and computational study.

Objective. An elevated interstitial fluid pressure (IFP) can lead to strain-induced stiffening of poroelastic biological tissues. As shear wave elastography (SWE) measures functional tissue stiffness based on the propagation speed of acoustically induced shear waves, the shear wave velocity (SWV) can be used as an indirect measurement of the IFP. The underlying biomechanical principle for this stiffening behavior with pressurization is however not well understood, and we therefore studied how IFP affects SWV through SWE experiments and numerical modeling.Approach. For model set-up and verification, SWE experiments were performed while dynamically modulating IFP in a chicken breast. To identify the confounding factors of the SWV-IFP relationship, we manipulated the material model (linear poroelastic versus porohyperelastic), deformation assumptions (geometric linearity versus nonlinearity), and boundary conditions (constrained versus unconstrained) in a finite element model mimicking the SWE experiments.Main results. The experiments demonstrated a statistically significant positive correlation between the SWV and IFP. The model was able to reproduce a similar SWV-IFP relationship by considering an unconstrained porohyperelastic tissue. Material nonlinearity was identified as the primary factor contributing to this relationship, whereas geometric nonlinearity played a smaller role. The experiments also highlighted the importance of the dynamic nature of the pressurization procedure, as indicated by a different observed SWV-IFP for pressure buildup and relaxation, but its clinical relevance needs to be further investigated.Significance. The developed model provides an adaptable framework for SWE of poroelastic tissues and paves the way towards non-invasive measurements of IFP.

Engineering Lymphangiogenesis-On-Chip: The Independent and Cooperative Regulation by Biochemical Factors, Gradients, and Interstitial Fluid Flow.

Despite the crucial role of lymphangiogenesis during development and in several diseases with implications for tissue regeneration, immunity, and cancer, there are significantly fewer tools to understand this process relative to angiogenesis. While there has been a major surge in modeling angiogenesis with microphysiological systems, they have not been rigorously optimized or standardized to enable the recreation of the dynamics of lymphangiogenesis. Here, a Lymphangiogenesis-Chip (L-Chip) is engineered, within which new sprouts form and mature depending upon the imposition of interstitial flow, growth factor gradients, and pre-conditioning of endothelial cells with growth factors. The L-Chip reveals the independent and combinatorial effects of these mechanical and biochemical determinants of lymphangiogenesis, thus ultimately resulting in sprouts emerging from a parent vessel and maturing into tubular structures up to 1 mm in length within 4 days, exceeding prior art. Further, when the constitution of the pre-conditioning cocktail and the growth factor cocktail used to initiate and promote lymphangiogenesis are dissected, it is found that endocan (ESM-1) results in more dominant lymphangiogenesis relative to angiogenesis. Therefore, The L-Chip provides a foundation for standardizing the microfluidics assays specific to lymphangiogenesis and for accelerating its basic and translational science at par with angiogenesis.

Multisensory gamma stimulation promotes glymphatic clearance of amyloid.

The glymphatic movement of fluid through the brain removes metabolic waste1-4. Noninvasive 40 Hz stimulation promotes 40 Hz neural activity in multiple brain regions and attenuates pathology in mouse models of Alzheimer's disease5-8. Here we show that multisensory gamma stimulation promotes the influx of cerebrospinal fluid and the efflux of interstitial fluid in the cortex of the 5XFAD mouse model of Alzheimer's disease. Influx of cerebrospinal fluid was associated with increased aquaporin-4 polarization along astrocytic endfeet and dilated meningeal lymphatic vessels. Inhibiting glymphatic clearance abolished the removal of amyloid by multisensory 40 Hz stimulation. Using chemogenetic manipulation and a genetically encoded sensor for neuropeptide signalling, we found that vasoactive intestinal peptide interneurons facilitate glymphatic clearance by regulating arterial pulsatility. Our findings establish novel mechanisms that recruit the glymphatic system to remove brain amyloid.

Neuronal dynamics direct cerebrospinal fluid perfusion and brain clearance.

The accumulation of metabolic waste is a leading cause of numerous neurological disorders, yet we still have only limited knowledge of how the brain performs self-cleansing. Here we demonstrate that neural networks synchronize individual action potentials to create large-amplitude, rhythmic and self-perpetuating ionic waves in the interstitial fluid of the brain. These waves are a plausible mechanism to explain the correlated potentiation of the glymphatic flow1,2 through the brain parenchyma. Chemogenetic flattening of these high-energy ionic waves largely impeded cerebrospinal fluid infiltration into and clearance of molecules from the brain parenchyma. Notably, synthesized waves generated through transcranial optogenetic stimulation substantially potentiated cerebrospinal fluid-to-interstitial fluid perfusion. Our study demonstrates that neurons serve as master organizers for brain clearance. This fundamental principle introduces a new theoretical framework for the functioning of macroscopic brain waves.

TriFNet: A triple-branch feature fusion network for pH determination by surface-enhanced Raman spectroscopy.

Due to the acidic tumor microenvironment caused by metabolic changes in tumor cells, the accurate pH detection of extracellular fluid is helpful for doctors in precise tumor resection. The combination of Raman spectroscopy and deep learning provides a solution for pH detection. However, most existing studies use one-dimensional convolutional neural networks (1D-CNNs) for spectral analysis, which limits the performance due to insufficient feature extraction. In this work, we propose a 2D triple-branch feature fusion network (TriFNet) for accurate pH determination using surface-enhanced Raman spectra (SERS). Specifically, we design a triple-branch network structure by converting Raman spectra into three types of images to extensively extract complex patterns in spectra. In addition, an attention fusion module, which leverages the complementarity among features in both space and channel, is designed to obtain the valuable information, achieving further accurate pH determination. On our Raman spectral dataset containing 14,137 samples, we achieved mean absolute error (MAE) of 0.059, standard deviation of the absolute error (SD) of 0.07, root mean squared error (RMSE) of 0.092, and coefficient of determination (R2) of 0.991 on the test set. Compared with other published methods, the four metrics showed an average improvement of 47%, 39%, 43%, and 6%, respectively. In addition, visualization validates the diagnostic capability of our model to correlate with biomolecular signatures. Meanwhile, our model has robustness to different SERS chips. These results prove the potential of our method to develop an effective technology based on Raman spectroscopy for accurate pH determination to guide surgery.

Novel method for collecting hippocampal interstitial fluid extracellular vesicles (EVISF ) reveals sex-dependent changes in microglial EV proteome in response to Aß pathology.

Brain-derived extracellular vesicles (EVs) play an active role in Alzheimer's disease (AD), relaying important physiological information about their host tissues. The internal cargo of EVs is protected from degradation, making EVs attractive AD biomarkers. However, it is unclear how circulating EVs relate to EVs isolated from disease-vulnerable brain regions. We developed a novel method for collecting EVs from the hippocampal interstitial fluid (ISF) of live mice. EVs (EVISF ) were isolated via ultracentrifugation and characterized by nanoparticle tracking analysis, immunogold labelling, and flow cytometry. Mass spectrometry and proteomic analyses were performed on EVISF cargo. EVISF were 40-150 nm in size and expressed CD63, CD9, and CD81. Using a model of cerebral amyloidosis (e.g., APPswe, PSEN1dE9 mice), we found protein concentration increased but protein diversity decreased with Aß deposition. Genotype, age, and Aß deposition modulated proteostasis- and immunometabolic-related pathways. Changes in the microglial EVISF proteome were sexually dimorphic and associated with a differential response of plaque associated microglia. We found that female APP/PS1 mice have more amyloid plaques, less plaque associated microglia, and a less robust- and diverse- EVISF microglial proteome. Thus, in vivo microdialysis is a novel technique for collecting EVISF and offers a unique opportunity to explore the role of EVs in AD.

Reduced graphene oxide-functionalized polymer microneedle for continuous and wide-range monitoring of lactate in interstitial fluid.

Despite substantial developments in minimally invasive lactate monitoring microneedle electrodes, most such electrode developments have focused on either sensitivity or invasiveness while ignoring a wide range of detection, which is the most important factor in measuring the normal range of lactate in interstitial fluid (ISF). Herein, we present a polymer-based planar microneedle electrode fabrication using microelectromechanical and femtosecond laser technology for the continuous monitoring of lactate in ISF. The microneedle is functionalized with two-dimensional reduced graphene oxide (rGO) and electrochemically synthesized platinum nanoparticles (PtNPs). A particular quantity of Nafion (1.25 wt%) is applied on top of the lactate enzyme to create a diffusion-controlled membrane. Due to the combined effects of the planar structure of the microneedle, rGO, and membrane, the biosensor exhibited excellent linearity up to 10 mM lactate with a limit of detection of 2.04 µM, high sensitivity of 43.96 µA mM-1cm-2, a reaction time of 8 s and outstanding stability, selectivity, and repeatability. The feasibility of the microneedle is evaluated by using it to measure lactate concentrations in artificial ISF and human serum. The results demonstrate that the microneedle described here has great potential for use in real-time lactate monitoring for use in sports medicine and treatment.

Demonstrating the Analytical Potential of a Wearable Microneedle-Based Device for Intradermal CO2 Detection.

Monitoring of carbon dioxide (CO2) body levels is crucial under several clinical conditions (e.g., human intensive care and acid-base disorders). To date, painful and risky arterial blood punctures have been performed to obtain discrete CO2 measurements needed in clinical setups. Although noninvasive alternatives have been proposed to assess CO2, these are currently limited to benchtop devices, requiring trained personnel, being tedious, and providing punctual information, among other disadvantages. To the best of our knowledge, the literature and market lack a wearable device for real-time, on-body monitoring of CO2. Accordingly, we have developed a microneedle (MN)-based sensor array, labeled as CO2-MN, comprising a combination of potentiometric pH- and carbonate (CO32-)-selective electrodes together with the reference electrode. The CO2-MN is built on an epidermal patch that allows it to reach the stratum corneum of the skin, measuring pH and CO32- concentrations directly into the interstitial fluid (ISF). The levels for the pH-CO32- tandem are then used to estimate the PCO2 in the ISF. Assessing the response of each individual MN, we found adequate response time (t95 < 5s), sensitivity (50.4 and -24.6 mV dec-1 for pH and CO32-, respectively), and stability (1.6 mV h-1 for pH and 2.1 mV h-1 for CO32-). We validated the intradermal measurements of CO2 at the ex vivo level, using pieces of rat skin, and then, with in vivo assays in anesthetized rats, showing the suitability of the CO2-MN wearable device for on-body measurements. A good correlation between ISF and blood CO2 concentrations was observed, demonstrating the high potential of the developed MN sensing technology as an alternative to blood-based analysis in the near future. Moreover, these results open new horizons in the noninvasive, real-time monitoring of CO2 as well as other clinically relevant gases.

Harvesting and manipulating sweat and interstitial fluid in microfluidic devices.

Microfluidic devices began to be used to facilitate sweat and interstitial fluid (ISF) sensing in the mid-2010s. Since then, numerous prototypes involving microfluidics have been developed in different form factors for sensing biomarkers found in these fluids under in vitro, ex vivo, and in vivo (on-body) settings. These devices transport and manipulate biofluids using microfluidic channels composed of silicone, polymer, paper, or fiber. Fluid flow transport and sample management can be achieved by controlling the flow rate, surface morphology of the channel, and rate of fluid evaporation. Although many devices have been developed for estimating sweat rate, electrolyte, and metabolite levels, only a handful have been able to proceed beyond laboratory testing and reach the stage of clinical trials and commercialization. To further this technology, this review reports on the utilization of microfluidics towards sweat and ISF management and transport. The review is distinguished from other recent reviews by focusing on microfluidic principles of sweat and ISF generation, transport, extraction, and management. Challenges and prospects are highlighted, with a discussion on how to transition such prototypes towards personalized healthcare monitoring systems.

Wearable Microneedle Patch for Transdermal Electrochemical Monitoring of Urea in Interstitial Fluid.

Microneedle-based wearable electrochemical biosensors are the new frontier in personalized health monitoring and disease diagnostic devices that provide an alternative tool to traditional blood-based invasive techniques. Advancements in micro- and nanofabrication technologies enabled the fabrication of microneedles using different biomaterials and morphological features with the aim of overcoming existing challenges and enhancing sensing performance. In this work, we report a microneedle array featuring conductive recessed microcavities for monitoring urea levels in the interstitial fluid of the skin. Microcavities are small pockets on the tip of each microneedle that can accommodate the sensing layer, provide protection from delamination during skin insertion or removal, and position the sensing layer in a deep layer of the skin to reach the interstitial fluid. The wearable urea patch has shown to be highly sensitive and selective in monitoring urea, with a sensitivity of 2.5 mV mM-1 and a linear range of 3 to 18 mM making it suitable for monitoring urea levels in healthy individuals and patients. Our ex vivo experiments have shown that recessed microcavities can protect the sensing layer from delamination during skin insertion and monitor changing urea levels in interstitial fluid. This biocompatible platform provides alternative solutions to the critical issue of maintaining the performance of the biosensor upon skin insertion and holds great potential for advancing transdermal sensor technology.

Current Understanding of the Anatomy, Physiology, and Magnetic Resonance Imaging of Neurofluids: Update From the 2022 "ISMRM Imaging Neurofluids Study group" Workshop in Rome.

Neurofluids is a term introduced to define all fluids in the brain and spine such as blood, cerebrospinal fluid, and interstitial fluid. Neuroscientists in the past millennium have steadily identified the several different fluid environments in the brain and spine that interact in a synchronized harmonious manner to assure a healthy microenvironment required for optimal neuroglial function. Neuroanatomists and biochemists have provided an incredible wealth of evidence revealing the anatomy of perivascular spaces, meninges and glia and their role in drainage of neuronal waste products. Human studies have been limited due to the restricted availability of noninvasive imaging modalities that can provide a high spatiotemporal depiction of the brain neurofluids. Therefore, animal studies have been key in advancing our knowledge of the temporal and spatial dynamics of fluids, for example, by injecting tracers with different molecular weights. Such studies have sparked interest to identify possible disruptions to neurofluids dynamics in human diseases such as small vessel disease, cerebral amyloid angiopathy, and dementia. However, key differences between rodent and human physiology should be considered when extrapolating these findings to understand the human brain. An increasing armamentarium of noninvasive MRI techniques is being built to identify markers of altered drainage pathways. During the three-day workshop organized by the International Society of Magnetic Resonance in Medicine that was held in Rome in September 2022, several of these concepts were discussed by a distinguished international faculty to lay the basis of what is known and where we still lack evidence. We envision that in the next decade, MRI will allow imaging of the physiology of neurofluid dynamics and drainage pathways in the human brain to identify true pathological processes underlying disease and to discover new avenues for early diagnoses and treatments including drug delivery. Evidence level: 1 Technical Efficacy: Stage 3.

Is increased subjective thirst associated with greater interdialytic weight gains, extracellular fluid and dietary sodium intake?

BACKGROUND: Some previous studies have reported an effect of increasing subjective thirst and interdialytic weight gains (IDWG), and that this may be influenced by nonadherence to dietary sodium restrictions, whereas others reported no such association. As such we wished to review the effect of self-reported thirst on IDWGs and dietary sodium intake. METHODS: Dialysis patients were asked to complete visual analogues thirst, distress thermometer (DT) scores and complete a sodium food frequency questionnaire (SFFQ). IDWG and pre and post dialysis volumes were measured with multifrequency bioelectrical impedance. RESULTS: One hundred and eleven patients completed the questionnaires and had bioimpedance measurements: 63% male, mean age 63.8 ± 16.1 years, 33% diabetic with a median thirst score 3 (0-5) and SFFQ 52.0 ± 18, and IDWG 2.1 ± 1.3%. Thirst was associated with DT (r = 0.28, p = 0.004) and negatively with age (r = -0.31, p < 0.001), but not SFFQ, IDWG, extracellular water, or dialysate sodium, or dialysate to plasma gradient. Patients with higher thirst scores were younger (58.0 ± 15.2 vs. 69.4 ± 15.0 years, p < 0.001) with higher DT scores (5 [2-7] vs. 2 [0-5], p < 0.001). On multivariate logistic analysis, only age was associated with self-reported thirst (odds ratio 0.95, 95% confidence limits 0.92-0.98, p < 0.001). CONCLUSION: We found that subjective thirst was greater for younger patients and those who reported higher levels of distress, but no association with IDWGs, dietary sodium intake, or dialysate sodium. However, most of our patients followed the dietary advice, as evidenced by the low SFFQ scores and % IDWGs. Whether thirst increases distress or distress increases subjective thirst remains to be determined.

Clearance of interstitial fluid (ISF) and CSF (CLIC) group-part of Vascular Professional Interest Area (PIA), updates in 2022-2023. Cerebrovascular disease and the failure of elimination of Amyloid-ß from the brain and retina with age and Alzheimer's disease: Opportunities for therapy.

This editorial summarizes advances from the Clearance of Interstitial Fluid and Cerebrospinal Fluid (CLIC) group, within the Vascular Professional Interest Area (PIA) of the Alzheimer's Association International Society to Advance Alzheimer's Research and Treatment (ISTAART). The overarching objectives of the CLIC group are to: (1) understand the age-related physiology changes that underlie impaired clearance of interstitial fluid (ISF) and cerebrospinal fluid (CSF) (CLIC); (2) understand the cellular and molecular mechanisms underlying intramural periarterial drainage (IPAD) in the brain; (3) establish novel diagnostic tests for Alzheimer's disease (AD), cerebral amyloid angiopathy (CAA), retinal amyloid vasculopathy, amyloid-related imaging abnormalities (ARIA) of spontaneous and iatrogenic CAA-related inflammation (CAA-ri), and vasomotion; and (4) establish novel therapies that facilitate IPAD to eliminate amyloid ß (Aß) from the aging brain and retina, to prevent or reduce AD and CAA pathology and ARIA side events associated with AD immunotherapy.