Microdialysis Probe Research Articles

What is the protective effect of preischemic kisspeptin-10 administration against ischemia/reperfusion injury of striatum on mice?

Kisspeptin is a neuropeptide with a primary role on the onset of puberty and has beneficial effects on ischemia/ reperfusion (I/R) injury. In this study, we aimed to investigate the effect of kisspeptin administration on striatal I/R injury in mice. Forty adult C57/BL6 mice were randomly divided into four groups: Sham, Kisspeptin, I/R, and I/R + Kisspeptin groups. The groups were administered with either physiological saline (Sham and I/R groups) or kisspeptin (Kisspeptin and I/R + Kisspeptin groups) intraperitoneally 40 min before the operation. A microdialysis probe was placed in the right striatum according to stereotaxic coordinates. During the experimental period, artificial cerebrospinal fluid was passed through the micropump. Then, transient cerebral ischemia was established by compressing both common carotid arteries with an aneurysm clip for 15 min and animals were reperfused for 2 h. Throughout the process of microdialysis (before, during and after I/R period), samples were collected to measure dopamine (DA), noradrenaline (NA), and 3,4-dihydroxyphenylglycine (DHPG) at intervals of 20 min continuously. At the end of the reperfusion period, the animals were decapitated, striatum was dissected, half of the animals were used for oxidative stress analyses (reduced glutathione, glutathione S-transferase (GST), superoxide dismutase (SOD), malondialdehyde (MDA), and the other half were used for histopathology analyses. Number of glial cells was significantly increased in kisspeptin-administered groups. DA levels in ischemic animals were decreased by kisspeptin administration (p < 0.0001). NA levels were reduced in animals administered with kisspeptin without I/R injury (p < 0.05). DHPG levels reduced during the reperfusion period in ischemic animals (p < 0.05). Kisspeptin did not exhibit a significant antioxidant activity in the ischemic animals, while GST and SOD levels were reduced in the I/R + kisspeptin group compared to the kisspeptin group (p < 0.05). Our results suggest that kisspeptin may be regulating the neurotransmitter release and metabolism, as well as inflammatory response in brain upon I/R injury.

The influence of sucrose on soil nitrogen availability – A root exudate simulation using microdialysis

Root exudates are thought to promote nitrogen (N) availability via rhizosphere interactions, but empirical evidence is difficult to obtain given the scale and temporary nature of these processes. Microdialysis has potential to simulate root exudation patterns and quantify the effects on N availability simultaneously, but this has so far not been attempted. In a conceptual root exudation study, we used sucrose as a simple C source to investigate if microdialysis could detect the effects of continuous localised C supply on soil inorganic N fluxes. Through retrodialysis we released sucrose and simultaneously monitored diffusive soil N fluxes over one week, followed by a further seven days without sucrose. Based on current understanding of rhizosphere N dynamics, we hypothesised that N fluxes are inversely related to sucrose release, and upon ceasing release, N fluxes would increase. Using a 5 mM sucrose perfusate, C releases resulted in decreased N fluxes, but contrary to our hypothesis, N fluxes did not increase after ceasing sucrose release (c.f. control soil). Diffusive sucrose efflux from microdialysis probes increased in soils amended with N-rich litter suggesting that microbial activity and associated sucrose consumption altered sucrose concentration gradients. The fluxes of sucrose breakdown products glucose and fructose were greatest in litter treatments receiving sucrose, indicative of increased invertase activity in the presence of fresh organic matter. In the short term (days), sucrose release did not prompt an increase in inorganic N availability, possibly because of stimulated microbial growth and increased N demand under C-rich conditions. Our study confirms that microdialysis allows time-sensitive insight into the dynamic interactions of carbon and N in the rhizosphere.

Systemic oxytocin increases glutamate efflux in the nucleus accumbens core of cocaine-experienced male and female rats but only increases dopamine efflux in males

Oxytocin attenuates cocaine-seeking when administered both systemically and directly into the nucleus accumbens core. This effect is blocked by intra-accumbens antagonism of mGlu2/3 and, together with our finding that intra-accumbens oxytocin increases glutamate concentrations in this brain region, indicates that pre-synaptic regulation of glutamate release by oxytocin influences cocaine relapse. However, mGlu2/3 receptors also regulate dopamine release in the nucleus accumbens. Here we aimed to determine whether systemic oxytocin increases glutamate and dopamine concentrations in the nucleus accumbens core of cocaine-experienced and cocaine-naïve male and female rats. A subset of rats self-administered cocaine (0.5 mg/kg/infusion) and then underwent extinction training for 2–3 weeks. Rats were implanted with microdialysis probes in the accumbens core and samples were collected for a baseline period, and following saline (1 mL/kg), and oxytocin (1 mg/kg, IP) injections. Locomotion was assessed during microdialysis. In cocaine-experienced rats, oxytocin increased glutamate concentrations in the accumbens core to the same extent in males and females but only increased dopamine concentrations in male rats. Oxytocin did not alter glutamate levels in cocaine-naïve rats. Oxytocin did not produce sedation. These results extend previous findings that systemic oxytocin increases nucleus accumbens dopamine in a sex-specific manner in cocaine-experienced rats. These data are the first to find that systemic oxytocin increases nucleus accumbens glutamate after cocaine experience, providing a mechanism of action by which oxytocin attenuates the reinstatement of cocaine seeking in both male and female rats.

Methods for intratumoral microdialysis probe targeting and validation in murine brain tumor models

BackgroundMicrodialysis is a well validated sampling technique that can be used for pharmacokinetic studies of oncological drugs targeting the central nervous system. This technique has also been applied to evaluate tumor metabolism and identify pharmacodynamic biomarkers of drug activity. Despite the potential utility of microdialysis for therapeutic discovery, variability in tumor size and location hamper routine use of microdialysis as a preclinical tool. Quantitative validation of microdialysis membrane location relative to radiographically evident tumor regions could facilitate rigorous preclinical studies. However, a widely accessible standardized workflow for preclinical catheter placement and validation is needed. New methodWe provide methods for a workflow to yield tailored placement of microdialysis probes within a murine intracranial tumor and illustrate in an IDH1-mutant patient-derived xenograft (PDX) model. This detailed workflow uses a freely available on-line tool built within 3D-slicer freeware to target microdialysis probe placement within the tumor core and validate probe placement fully within the tumor. ResultsWe illustrate use of this workflow to validate microdialysis probe location relative to implanted IDH1-mutant PDXs, using the microdialysis probes to quantify levels of extracellular onco-metabolite D-2 hydroxyglutarate. Comparison with existing methodsPrevious methods have used 3D slicer to reliably measure tumor volumes. Prior microdialysis studies have targeted expected tumor locations without validation. ConclusionsThe new method offers a streamlined and freely available workflow in 3D slicer to optimize and validate microdialysis probe placement within a murine brain tumor.

Quantitative analysis of human brain microdialysate for target site pharmacokinetics of major anesthetics ketamine, midazolam and propofol

Brain microdialysis samples of intensive care patients treated with the essential anesthetics ketamine, midazolam and propofol were investigated. Importantly, despite decades of clinical use, comprehensive human cerebral pharmacokinetic data of these drugs is still missing.To encounter this apparent lack of knowledge, we combined cerebral microdialysis with leading-edge analytical instrumentation to monitor the neurochemistry of living human patients. For the quantitative analysis, high performing analytical approaches were developed that can handle minute sample volumes and possible ultralow target analyte levels. The developed methods provided detection limits below 100 ng L−1 for all target analytes and high precision (below 4% RSD intraday). Methods were linear between LODs and 100 μg L−1 for ketamine, 75 μg L−1 for midazolam and 10 μg L−1 for propofol respectively, with coefficients of determination R2≥ 0.999. Further, being aware of the error-prone and demanding translation of microdialysis levels to interstitial concentrations, in vitro approaches for recovery testing of microdialysis probes as well as internal normalization approaches were conducted.Thus, we herein report the first cerebral pharmacokinetic data of ketamine, midazolam and propofol determined in microdialysis samples of 15 neurointensive care patients. We could prove blood-brain barrier penetration of all of the investigated anesthetics and could correlate applied dosages and actual brain exposition of ketamine.However, we emphasize the need of an expanded prospective study including individual microdialysis recovery testing as well as matched serum and/or cerebrospinal fluid collection for a more comprehensive cerebral pharmacokinetic understanding.

Cocaine-Induced Changes in Tonic Dopamine Concentrations Measured Using Multiple-Cyclic Square Wave Voltammetry in vivo.

For over 40 years, in vivo microdialysis techniques have been at the forefront in measuring the effects of illicit substances on brain tonic extracellular levels of dopamine that underlie many aspects of drug addiction. However, the size of microdialysis probes and sampling rate may limit this technique’s ability to provide an accurate assessment of drug effects in microneural environments. A novel electrochemical method known as multiple-cyclic square wave voltammetry (M-CSWV), was recently developed to measure second-to-second changes in tonic dopamine levels at microelectrodes, providing spatiotemporal resolution superior to microdialysis. Here, we utilized M-CSWV and fast-scan cyclic voltammetry (FSCV) to measure changes in tonic or phasic dopamine release in the nucleus accumbens core (NAcc) after acute cocaine administration. Carbon-fiber microelectrodes (CFM) and stimulating electrodes were implanted into the NAcc and medial forebrain bundle (MFB) of urethane anesthetized (1.5 g/kg i.p.) Sprague-Dawley rats, respectively. Using FSCV, depths of each electrode were optimized by determining maximal MFB electrical stimulation-evoked phasic dopamine release. Changes in phasic responses were measured after a single dose of intravenous saline or cocaine hydrochloride (3 mg/kg; n = 4). In a separate group, changes in tonic dopamine levels were measured using M-CSWV after intravenous saline and after cocaine hydrochloride (3 mg/kg; n = 5). Both the phasic and tonic dopamine responses in the NAcc were augmented by the injection of cocaine compared to saline control. The phasic and tonic levels changed by approximately x2.4 and x1.9, respectively. These increases were largely consistent with previous studies using FSCV and microdialysis. However, the minimal disruption/disturbance of neuronal tissue by the CFM may explain why the baseline tonic dopamine values (134 ± 32 nM) measured by M-CSWV were found to be 10-fold higher when compared to conventional microdialysis. In this study, we demonstrated phasic dopamine dynamics in the NAcc with acute cocaine administration. M-CSWV was able to record rapid changes in tonic levels of dopamine, which cannot be achieved with other current voltammetric techniques. Taken together, M-CSWV has the potential to provide an unprecedented level of physiologic insight into dopamine signaling, both in vitro and in vivo, which will significantly enhance our understanding of neurochemical mechanisms underlying psychiatric conditions.

Applicability of cerebral open flow microperfusion and microdialysis to quantify a brain-penetrating nanobody in mice

The use of biologics in the therapeutic landscape has increased exponentially since the last 3 decades. Nevertheless, patients with central nervous system (CNS) related disorders could not yet benefit from this revolution because the blood-brain barrier (BBB) severely hampers biologics from entering the brain. Considerable effort has been put into generating methods to modulate or circumvent the BBB for delivery of therapeutics to the CNS. A promising strategy is receptor-mediated transcytosis (RMT). Recently, Wouters et al. (2020) discovered a mouse anti-transferrin receptor nanobody that is able to deliver a biologically active peptide to the brain via RMT. The present study aims to sample a derivative of this brain-penetrating nanobody (Nb105) in the CNS. Therefore, we compared the applicability of cerebral open flow microperfusion (cOFM) and microdialysis as sampling techniques to directly obtain high molecular weight substances from the cerebral interstitial fluid. A custom AlphaScreen™ assay was validated to quantify nanobody concentrations in the samples. In vitro microdialysis probe (AtmosLM™, 1 MDa cut-off) recovery by gain and by loss for Nb105 was 18.3 ± 3.2% and 27.0 ± 2.5% respectively, whereas for cOFM it was 87.2 ± 4.0% and 97.3 ± 1.6%. Although a large difference in in vitro recovery is observed between cOFM and microdialysis, in vivo similar results were obtained. Immunohistochemical stainings showed an astrocytic and microglial reaction in the immediate vicinity along the implantation track for both probe types. Coronal sections showed higher fluorescein isothiocyanate-dextran and immunoglobulin G extravasation around the microdialysis probe track than after cOFM sampling experiments, however this leakage was clearly limited compared to a positive control where the BBB was disrupted. This is the first study that samples a bispecific nanobody in the brain's interstitial fluid in function of time, providing a pharmacokinetic profile of nanobodies in the CNS. Furthermore, this is the first time a cOFM study is performed in awake freely moving mice, providing data on inflammation and blood-brain barrier integrity in the mouse brain. Overall, this work demonstrates that, while taking into account the (bio)analytical considerations, both microdialysis and cOFM are suitable in vivo sampling techniques for quantification of nanobodies in the CNS.

Gas phase microdialysis and chemiluminescence detection: A small, fast, selective, and sensitive method to monitor aqueous nitric oxide

A method using a gas-phase microdialysis probe interfaced with a modified commercially available nitric oxide (NO) detector is shown to selectively measure aqueous NO at low μM levels with high selectivity. The detector measures chemiluminescence resulting from the gas-phase reaction of NO with ozone. The microdialysis probe is small enough (3 mm × 200 μm) to be used in vivo. Because the processes of extraction across the microdialysis membrane and transport from the probe to the detector are both very fast, the response time is shorter than 5 s. The method was verified using two different quantifiable sources of NO: nitrite and methylamine hexamethylene methylamine (MAHMA) NONOates. To demonstrate ruggedness and to show the impact of matrix on NO generation, the method was used to measure NO in a cell culture matrix. The continuous extraction, fast response time, and rugged nature make the method useful for monitoring NO in biological applications. Our results also show that predicting NO concentration for in vitro experiments based on NONOate concentration may be a poor assumption due to the pH dependence of NO formation and the rapid decline in NO concentration.

Penetration and pharmacokinetics of ferulic acid after dermal administration

Purpose: To study the in vitro penetration and in vivo pharmacokinetics of ferulic acid (FA), and the correlation between them after dermal administration.&#x0D; Methods: Franz diffusion cell was used to study in vitro penetration of FA. The concentration of FA in the Franz receiver solution was assessed by high performance liquid chromatography (HPLC). Prior to in vivo pharmacokinetics experiments, probe recovery was validated with respect to influencing factors such as flow rate, FA concentration, within-day stability and reproducibility of the probes. In in vivo pharmacokinetic experiment, six male CD-1 hairless mice were used. The micro-dialysis (MD) probe was implanted in the dermis of the rat skin, and dialysates from probe outlet were quantified directly by HPLC. In in vivo studies, deconvolution methods were used to determine the relationship between in vitro and in vivo data, and the correlation coefficient of linear equations.&#x0D; Results: There was significant effect of pH (5 ~ 8) on the penetration of FA. Increase in pH caused commensurate decrease in permeability. The Cmax of FA was 300.74 ± 31.86 ng/mL while Tmax was 138.00 ± 22.80 min after dermal administration of 1 mg/mL FA dissolved in phosphate buffered saline (PBS). The correlation coefficient (r) between in vitro and in vivo data was 0.9905.&#x0D; Conclusion: Both in vivo and in vitro experiments demonstrate that FA permeates the stratum corneum of skin rapidly. The unionized form of FA shows better penetration than the ionic form. In addition, results from correlation analysis indicate that the in vitro penetration characteristics of FA can be applied to predict its in vivo pharmacokinetics.

Miniaturized probe on polymer SU-8 with array of individually addressable microelectrodes for electrochemical analysis in neural and other biological tissues.

An SU-8 probe with an array of nine, individually addressable gold microband electrodes (100μm long, 4μm wide, separated by 4-μm gaps) was photolithographically fabricated and characterized for detection of low concentrations of chemicals in confined spaces and in vivo studies of biological tissues. The probe's shank (6mm long, 100μm wide, 100μm thick) is flexible, but exhibits sufficient sharpness and rigidity to be inserted into soft tissue. Laser micromachining was used to define probe geometry by spatially revealing the underlying sacrificial aluminum layer, which was thenetched to free the probes from a silicon wafer. Perfusion with fluorescent nanobeads showed that, like a carbon fiber electrode, the probe produced no noticeable damage when inserted into rat brain, in contrast to damage from an inserted microdialysis probe. The individual addressability of the electrodes allows single and multiple electrode activation. Redox cycling is possible, where adjacent electrodes serve as generators (that oxidize or reduce molecules) and collectors (that do the opposite) to amplify signals of small concentrations without background subtraction. Information about electrochemical mechanisms and kinetics may also be obtained. Detection limits for potassium ferricyanide in potassium chloride electrolyte of 2.19, 1.25, and 2.08μM and for dopamine in artificial cerebral spinal fluid of 1.94, 1.08, and 5.66μM for generators alone and for generators and collectors during redox cycling, respectively, were obtained.

Preliminary Data Monitoring In Vivo Sources of Reactive Oxygen Species Production in Human Obesity

Introduction Excessive reactive oxygen species (ROS) production is a prominent feature of obesity-related conditions including impairments in endothelial function. NADPH oxidases (Nox) and mitochondria are major sources of ROS production within the vasculature. However, studies involving direct measurement of in vivo ROS production are limited in humans. Thus, the relative contributions of Nox- and mitochondrial-derived ROS in human obesity remain to be fully characterized. The objective of this study was to determine the predominant sources of in vivo ROS production in the skeletal muscle microvasculature of individuals with obesity. We hypothesize that both Nox- and mitochondria-dependent ROS production are upregulated in human obesity. Methods Two sedentary men with Class II and above obesity (body mass index ≥ 35 kg/m2) and the Metabolic Syndrome were studied. Microdialysis was used for measurements of in vivo skeletal muscle ROS production. Microdialysis probes were perfused with saline containing Amplex Ultrared, horseradish peroxidase and superoxide dismutase to measure local hydrogen peroxide and superoxide concentrations. To assess ROS production from distinct sources, microdialysis probes were perfused with the following: apocynin (non-specific Nox inhibitor), MitoTEMPO (mitochondrial-ROS inhibitor), and GKT 137831 (specific Nox 4 inhibitor). Results The concentrations of hydrogen peroxide and superoxide decreased upon addition of apocynin to microdialysis probes (mean ± SEM; hydrogen peroxide: 1.18 ± 0.58 to 0.76 ± 0.01 μM; superoxide: 1.47 ± 0.46 to 1.10 ± 0.62 μM). MitoTEMPO perfusion elicited an increase in hydrogen peroxide (1.12 ± 0.76 to 1.51 ± 1.08 μM) and decrease in superoxide (1.47 ± 0.46 to 0.78 ± 0.10 μM). The combination of apocynin plus MitoTEMPO resulted in concentrations of hydrogen peroxide (0.22 ± 0.10 μM) and superoxide (0.23 ± 0.06 μM) that were 69.7% and 79.1% lower than apocynin alone. In addition, the hydrogen peroxide (0.56 ± 0.25 μM) and superoxide (0.43 ± 0.18 μM) concentrations with co-perfusion of apocynin and GKT were 26.3% and 60.9% lower than with apocynin alone. Conclusions Our preliminary data suggest that mitochondria and Nox together are significant sources of ROS production within the skeletal muscle microvasculature of obese individuals. The local inhibition of Nox- and mitochondrial-derived ROS resulted in marked decreases in extracellular ROS levels within the skeletal muscle microvasculature. Although the small sample size currently limits our interpretation of the results, this study is anticipated to become the first to report on distinct sources of in vivo ROS production in human obesity.

Investigations on the Oral and Brain Uptake Mechanisms of D‐ and L‐eflornithine in the Rat

Eflornithine is a recommended treatment of the otherwise fatal disease late stage Gambian human African trypanosomiasis (HAT). Whereas racemic eflornithine is today administered intravenously, an oral eflornithine treatment option would increase accessibility for patients in affected areas in sub-Saharan Africa. Oral absorption of eflornithine is stereoselective when dosed as a racemic mixture with the more potent L-enantiomer disfavoured [1, 2]. In late stage Gambian HAT, the brain is the primary site of pharmacological action. Knowledge on whether stereoselective passage also occurs across the blood-brain barrier is limited. This study aimed to investigate the rate, extent and potential stereoselectivity of brain uptake of D- and L-eflornithine using in vivo microdialysis in the rat. Microdialysis probes were inserted in the third brain ventricle with simultaneous catheterizing of the jugular vein for blood sampling in Sprague-Dawley rats (n=16). Racemic eflornithine was dosed (1200 mg/kg) by oral gavage. Plasma and microdialysis samples were collected up to nine hours after dosing. Stereoselective eflornithine concentrations were determined by high performance liquid chromatography [3]. Plasma data was complemented with data from a prior study [1]. Areas under the curve (AUC) were determined using pharmacokinetic modelling with naïve pooled data analysis as implemented in Phoenix (Version 8.3, Certara, Princeton, NJ, USA). Pharmacokinetics of D- and L-eflornithine were best described by a two-compartment drug disposition model. Transit compartments were used to model the delay in oral absorption. Observed AUCs in plasma and brain dialysate for D-eflornithine were 160 min•mM and 1.0 min•mM, respectively. For L-eflornithine, AUCs were 60 min•mM and 0.38 min•mM in plasma and brain dialysate, respectively. Oral uptake of D- and L-eflornithine was, as expected, stereoselective whereas the uptake to the third ventricle in the rat brain was not stereoselective since the dialysate/plasma AUC ratio was similar for the enantiomers. Overall, results suggested a brain efflux of both D- and L-eflornithine to a similar extent. The findings will be used in future translational investigations to model the feasibility of an oral eflornithine based therapy against late stage Gambian HAT.

Cerebral Microdialysis Probe Location as an Indicator of Metabolic Distress in Traumatic Brain Injury. (4758)

Cerebral Microdialysis Probe Location as an Indicator of Metabolic Distress in Traumatic Brain Injury. (4758)

Neural dysfunctions following experimental permanent occlusions of bilateral common carotid arteries cause an increase of rat voluntary alcohol drinking behavior.

We have previously reported that ischemic animal models treated with a respiratory inhibitor, rotenon, show an increased voluntary alcohol intake. Although it is clear that ischemic brain, as a result of reduced-blood flow, shows pathological events and/or neuro-degenerations apparently, little is known of causal relationship between the mechanism of neural dysfunction and voluntary alcohol consumption. Authors have investigated effects of permanent two-vessel occlusion (p2VO) on rat voluntary alcohol drinking behavior. In first experiment the p2VO-treated rats showed an increase of voluntary alcohol drinking behavior, as compared with sham controls. Using brain microdialysis technique, increases of only nucleus accumbens (ACC) dopamine (DA) releases were suppressed in the p2VO-treated rats significantly, following the high K+ (40mM) perfusion through the microdialysis probe membrane. Alcohol (200mM) perfusion-induced DA and serotonin (5-HT) releases in the ACC of the p2VO-treated rats were suppressed significantly in the second experiment, as compared with the sham-treated rats. In third experiment p2VO-treated rats showed significant decreases of the contents of DA, not 5-HT, in the ACC, caudate-putamen (C/P), ventral tegmental area-substantia nigra (VT/SN) and lateral hypothalamus (LH). Dopaminergic neurons in the ACC showed more functional vulnerability against the p2VO treatments, as compared with the serotonergic neurons. An increase of alcohol intake in the p2VO-treated rats means the compensation for the neural degeneration of the dopaminergic system in the ACC consisted brain rewarding system. It was likely suggested that neural disturbance of higher functions involved with incomplete global brain ischemia leads the risk of an abnormal alcohol drinking in human.

Space and time-resolved monitoring of phosphorus release from a fertilizer pellet and its mobility in soil using microdialysis and X-ray computed tomography.

Phosphorus is an essential nutrient for crops. Precise spatiotemporal application of P fertilizer can improve plant P acquisition and reduce run‐off losses of P. Optimizing application would benefit from understanding the dynamics of P release from a fertilizer pellet into bulk soil, which requires space‐ and time‐resolved measurements of P concentration in soil solutions. In this study, we combined microdialysis and X‐ray computed tomography to investigate P transport in soil. Microdialysis probes enabled repeated solute sampling from one location with minimal physical disturbance, and their small dimensions permitted spatially resolved monitoring. We observed a rapid initial release of P from the source, producing high dissolved P concentrations within the first 24 h, followed by a decrease in dissolved P over time compatible with adsorption onto soil particles. Soils with greater bulk density (i.e., reduced soil porosity) impeded the P pulse movement, which resulted in a less homogeneous distribution of total P in the soil column at the end of the experiment. The model fit to the data showed that the observed phenomena can be explained by diffusion and adsorption. The results showed that compared with conventional measurement techniques (e.g., suction cups), microdialysis measurements present a less invasive alternative. The time‐resolved measurements ultimately highlighted rapid P dynamics that require more attention for improving P use efficiency.

The Cumulative Amount of Exuded Citrate Controls Its Efficiency to Mobilize Soil Phosphorus

Root exudation of citrate is discussed as mechanism to mobilize P from the soils' solid phase. Microbial processes can mitigate the mobilization efficiency of citrate. Due to higher microbial activity in topsoils compared to subsoils, we hypothesized a lower mobilization efficiency of exuded citrate in topsoils than in the subsoils. As a model system we used microdialysis (MD) probes and we followed diffusive fluxes of citrate from the perfusate into the soil and of phosphate from the soil into the dialysate in three soil horizons (Oa, Ah, Bw) of a Fagus sylvatica L. stand Cambisol. Three different MD perfusates with a KCl background concentration have been used: control, 1, and 3 mmol L−1 citric acid. Fluxes have been measured after 24, 48, and 144 h. The high-citrate perfusate increased the cumulative 144 h P-influx by a factor of 8, 13, and 113 in the Oa, Ah, and Bw horizon, respectively. With the high-citrate treatment, P mobilization efficiency decreased over time, whereas for the low citrate, P mobilization efficiency had a maximum at day 2. Minimum P mobilization efficiency of citrate was 1:25,000 mol phosphate per mol citrate in the Oa horizon between days 2 and 6, and maximum was 1:286 in the Bw-horizon during day 2. An increasing citrate efflux over time indicated an increasing sink term for citrate in the soil due to microbial decay or immobilization processes. Cumulative phosphate influx could be fitted to cumulative citrate efflux and soil horizon in a logarithmic model explaining 87% of the variability. For the first time, we could follow the localized P-uptake with citrate exudation over several days. Cumulative citrate efflux as the main control of P-mobilization has been barely discussed yet, however, it could explain some gaps in the role of carboxylates in the rhizosphere. Batch experiments are not capable to elucidate microscale dynamic competition for phosphate and carboxylates. MD is a promising tool for spatially explicit investigation of phosphate–citrate exchange, since such detailed insights in are not possible with batch experiments. In combination with the analysis of microbial properties, this technique has a huge potential to identify mobilization processes in soils as induced by citrate.

Dexamethasone-Enhanced Microdialysis and Penetration Injury.

Microdialysis probes, electrochemical microsensors, and neural prosthetics are often used for in vivo monitoring, but these are invasive devices that are implanted directly into brain tissue. Although the selectivity, sensitivity, and temporal resolution of these devices have been characterized in detail, less attention has been paid to the impact of the trauma they inflict on the tissue or the effect of any such trauma on the outcome of the measurements they are used to perform. Factors affecting brain tissue reaction to the implanted devices include: the mechanical trauma during insertion, the foreign body response, implantation method, and physical properties of the device (size, shape, and surface characteristics. Modulation of the immune response is an important step toward making these devices with reliable long-term performance. Local release of anti-inflammatory agents such as dexamethasone (DEX) are often used to mitigate the foreign body response. In this article microdialysis is used to locally deliver DEX to the surrounding brain tissue. This work discusses the immune response resulting from microdialysis probe implantation. We briefly review the principles of microdialysis and the applications of DEX with microdialysis in (i) neuronal devices, (ii) dopamine and fast scan cyclic voltammetry, (iii) the attenuation of microglial cells, (iv) macrophage polarization states, and (v) spreading depolarizations. The difficulties and complexities in these applications are herein discussed.

Effects of Ice Massage Prior to an Iontophoresis Treatment Using Dexamethasone Sodium Phosphate.

Low current intensity iontophoresis treatments have increased skin perfusion over 700% from baseline potentially altering drug clearance from or diffusion to the targeted area. To determine the effects of a preceding 10-minute ice massage on subcutaneous dexamethasone sodium phosphate (Dex-P) concentration and skin perfusion during and after a 4-mA iontophoresis treatment. Controlled laboratory study. Research laboratory. Twenty-four participants (male = 12, female = 12; age = 25.6 [4.5]y, height = 173.9 [8.51]cm, mass = 76.11 [16.84]kg). Participants were randomly assigned into 2 groups: (1)pretreatment 10-minute ice massage and (2)no pretreatment ice massage. Treatment consisted of an 80-mA·minute (4mA, 20min) Dex-P iontophoresis treatment. Microdialysis probes (3 mm deep in the forearm) were used to assess Dex-P, dexamethasone (Dex), and its metabolite (Dex-Met) concentrations. Skin perfusion was measured using laser Doppler flowmetry. Microdialysis samples were collected at baseline, at conclusion of treatment, and every 20 minutes posttreatment for 60minutes. Samples were analyzed to determine Dex-Total (Dex-Total = Dex-P + Dex + Dex-Met). Skin perfusion was calculated as a percentage change from baseline. A mixed-design analysis of variance was used to determine Dex-Total and skin perfusion difference between groups overtime. There was no difference between groups (P = .476), but [Dex-Total] significantly increased over the course of the iontophoresis and posttreatment time (P < .001). Dex-P was measured in 18 of 24 participants with a mean concentration of 0.67 (1.09)μg/mL. Skin perfusion was significantly greater in the no ice treatment group (P = .002). Peak skin perfusion reached 27.74% (47.49%) and 117.39% (103.45%) from baseline for the ice and no ice groups, respectively. Ice massage prior to iontophoresis does not alter the tissue [Dex-Total] even with less skin perfusion.

Electroosmotic Perfusion-Microdialysis Probe Created by Direct Laser Writing for Quantitative Assessment of Leucine Enkephalin Hydrolysis by Insulin-Regulated Aminopeptidase in Vivo.

There are many processes that actively alter the concentrations of solutes in the extracellular space. Enzymatic reactions, either by soluble enzymes or membrane-bound ectoenzymes, and uptake or clearance are two such processes. Investigations of ectoenzymatic reactions in vivo is challenging, particularly in the brain. Studies using microdialysis have revealed some qualitative information about what enzymes may be present, but microdialysis is a sampling technique so it is not designed to control conditions such as a substrate concentration outside the probe. Micropush-pull perfusion has been used to determine which nitric oxide synthase enzymes are active in discrete regions of the rat retina. Ectopeptidases are a particularly important class of ectoenzymes. As far as it is known, the extracellular activity of active peptides in the brain is controlled by ectopeptidases. To understand ectopeptidase activity, we developed a physical probe and an accompanying method. The probe has a two-channel source that supplies substrate or substrate plus inhibitor using electroosmotic perfusion (EOP). It also has a microdialysis probe to collect products and unreacted substrate. The method provides quantitative estimates of substrate-to-product conversion and the influence of inhibitors on this process. The quantitative estimates are made possible by including a d-amino acid-containing peptide analog of the substrate in the substrate-containing solution infused. Quantitative analysis of substrate, substrate analog, and products is carried out by quantitative, online capillary liquid chromatography-tandem mass spectrometry. The electroosmotic perfusion-microdialysis probe and associated method were used to determine the effect of the selective inhibitor HFI-419 on insulin-regulated aminopeptidase (EC 3.4.11.3) in the rat neocortex.

Contribution of afferent pathway to vagal nerve stimulation-induced myocardial interstitial acetylcholine release in rats.

Vagal nerve stimulation (VNS) has been explored as a potential therapy for chronic heart failure. The contribution of the afferent pathway to myocardial interstitial acetylcholine (ACh) release during VNS has yet to be clarified. In seven anesthetized Wistar-Kyoto rats, we implanted microdialysis probes in the left ventricular free wall and measured the myocardial interstitial ACh release during right VNS with the following combinations of stimulation frequency (F in Hz) and voltage readout (V in volts): F0V0 (no stimulation), F5V3, F20V3, F5V10, and F20V10. F5V3 did not affect the ACh level. F20V3, F5V10, and F20V10 increased the ACh level to 2.83 ± 0.47 (P < 0.01), 4.31 ± 1.09 (P < 0.001), and 4.33 ± 0.82 (P < 0.001) nM, respectively, compared with F0V0 (1.76 ± 0.22 nM). After right vagal afferent transection (rVAX), F20V3 and F20V10 increased the ACh level to 2.90 ± 0.53 (P < 0.001) and 3.48 ± 0.63 (P < 0.001) nM, respectively, compared with F0V0 (1.61 ± 0.19 nM), but F5V10 did not (2.11 ± 0.24 nM). The ratio of the ACh levels after rVAX relative to before was significantly <100% in F5V10 (59.4 ± 8.7%) but not in F20V3 (102.0 ± 8.7%). These results suggest that high-frequency and low-voltage stimulation (F20V3) evoked the ACh release mainly via direct activation of the vagal efferent pathway. By contrast, low-frequency and high-voltage stimulation (F5V10) evoked the ACh release in a manner dependent on the vagal afferent pathway.