Experimental and numerical investigation of microdialysis probes for ethanol metabolism studies.

Microdialysis is an important technique for in vivo sampling of tissue's biochemical composition. Understanding the factors that affect the performance of the microdialysis probes and developing methods for sample analysis are crucial for obtaining reliable results. In this work, we used experimental and numerical procedures to study the performance of microdialysis probes having different configurations, membrane materials and dimensions. For alcohol research, it is important to understand the dynamics of ethanol metabolism, particularly in the brain and in other organs, and to simultaneously measure the concentrations of ethanol and its metabolites - acetaldehyde and acetate. Our work provides a comprehensive characterization of three microdialysis probes, in terms of recovery rates and backpressure, allowing for interpretation and optimization of experimental procedures. In vivo experiments were performed to measure the time course concentration of ethanol, acetaldehyde, and acetate in the rat brain dialysate. Additionally, the combination of in vitro experimental results with numerical simulations enabled us to calculate diffusion coefficients of molecules in the microdialysis membranes and study the extent of the depletion effect caused by continuous microdialysis sampling, thus providing additional insights for probe selection and data interpretation.

Fluorocitrate inhibition of astrocytes reduces nicotine self-administration and alters extracellular levels of glutamate and dopamine within the nucleus accumbens in male wistar rats.

Emerging evidence suggests an important role of astrocytes in mediating behavioral and molecular effects of commonly misused drugs. Passive exposure to nicotine alters molecular, morphological, and functional properties of astrocytes. However, a potential involvement of astrocytes in nicotine reinforcement remains largely unexplored. The overall hypothesis tested in the current study is that astrocytes play a critical role in nicotine reinforcement. Protein levels of the astrocyte marker glial fibrillary acidic protein (GFAP) were examined in key mesocorticolimbic regions following chronic nicotine intravenous self-administration. Fluorocitrate, a metabolic inhibitor of astrocytes, was tested for its effects on behaviors related to nicotine reinforcement and relapse. Effects of fluorocitrate on extracellular neurotransmitter levels, including glutamate, GABA, and dopamine, were determined with microdialysis. Chronic nicotine intravenous self-administration increased GFAP expression in the nucleus accumbens core (NACcr), but not other key mesocorticolimbic regions, compared to saline intravenous self-administration. Both intra-ventricular and intra-NACcr microinjection of fluorocitrate decreased nicotine self-administration. Intra-NACcr fluorocitrate microinjection also inhibited cue-induced reinstatement of nicotine seeking. Local perfusion of fluorocitrate decreased extracellular glutamate levels, elevated extracellular dopamine levels, but did not alter extracellular GABA levels in the NACcr. Fluorocitrate did not alter basal locomotor activity. These results indicate that nicotine reinforcement upregulates the astrocyte marker GFAP expression in the NACcr, metabolic inhibition of astrocytes attenuates nicotine reinforcement and relapse, and metabolic inhibition of astrocytes disrupts extracellular dopamine and glutamate transmission. Overall, these findings suggest that astrocytes play an important role in nicotine reinforcement and relapse, potentially through regulation of extracellular glutamate and dopamine neurotransmission.

Repetitive mild traumatic brain injury impairs norepinephrine system function and psychostimulant responsivity.

Traumatic brain injury (TBI) is a complex pathophysiological process that results in a variety of neurotransmitter, behavioral, and cognitive deficits. The locus coeruleus-norepinephrine (LC-NE) system is a critical regulator of arousal levels and higher executive processes affected by TBI including attention, working memory, and decision making. LC-NE axon injury and impaired signaling within the prefrontal cortex (PFC) is a potential contributor to the neuropsychiatric symptoms after single, moderate to severe TBI. The majority of TBIs are mild, yet long-term cognitive deficits and increased susceptibility for further injury can accumulate after each repetitive mild TBI. As a potential treatment for restoring cognitive function and daytime sleepiness after injury psychostimulants, including methylphenidate (MPH) that increase levels of NE within the PFC, are being prescribed "off-label". The impact of mild and repetitive mild TBI on the LC-NE system remains limited. Therefore, we determined the extent of LC-NE and arousal dysfunction and response to therapeutic doses of MPH in rats following experimentally induced single and repetitive mild TBI. Microdialysis measures of basal NE efflux from the medial PFC and arousal measures were significantly lower after repetitive mild TBI. Females showed higher baseline PFC-NE efflux than males following single and repetitive mild TBI. In response to MPH challenge, males exhibited a blunted PFC-NE response and persistent arousal levels following repetitive mild TBI. These results provide critical insight into the role of catecholamine system dysfunction associated with cognitive deficits following repeated injury, outcome differences between sex/gender, and lack of success of MPH as an adjunctive therapy to improve cognitive function following injury.

Metabolome analyses of skin dialysate: Insights into skin interstitial fluid biomarkers.

BACKGROUND: Metabolites in biofluids can serve as biomarkers for diagnosing diseases and monitoring body conditions. Among the available biofluids, interstitial fluid (ISF) in the skin has garnered considerable attention owing to its advantages, which include inability to clot, easy access to the skin, and possibility of incorporating wearable devices. However, the scientific understanding of skin ISF composition is limited. OBJECTIVE: In this study, we aimed to compare metabolites between skin dialysate containing metabolites from the skin ISF and venous blood (plasma) samples, both collected under resting states. METHODS: We collected forearm skin dialysate using intradermal microdialysis alongside venous blood (plasma) samples from 12 healthy young adults. We analyzed these samples using capillary electrophoresis-fourier transform mass spectrometry-based metabolomics (CE-FTMS). RESULTS: Significant positive correlations were observed in 39 metabolites between the skin dialysate and plasma, including creatine (a mitochondrial disease biomarker), 1-methyladenosine (an early detection of cancer biomarker), and trimethylamine N-oxide (a posterior predictor of heart failure biomarker). Based on the Human Metabolome Technologies database, we identified 12 metabolites unique to forearm skin dialysate including nucleic acids, benzoate acids, fatty acids, amino acids, ascorbic acid, 3-methoxy-4-hydroxyphenylethyleneglycol (an Alzheimer's disease biomarker), and cysteic acid (an acute myocardial infarction biomarker). CONCLUSION: We show that some venous blood biomarkers may be predicted from skin dialysate or skin ISF, and that these fluids may serve as diagnostic and monitoring tools for health and clinical conditions.

(+)-Borneol Protects Dopaminergic Neuronal Loss in Methyl-4-phenyl-1,2,3,6-tetrahydropyridine-Induced Parkinson's Disease Mice: A Study of Dopamine Level using In Vivo Brain Microdialysis.

Considerable research efforts have been directed toward the symptom relief of Parkinson's disease (PD) by attenuating dopamine (DA) depletion. One common feature of these existing therapies is their unavailability of preventing the neurodegenerative process of dopaminergic neurons. (+)-Borneol, a natural highly lipid-soluble bicyclic monoterpene, has been reported to regulate the levels of monoamine neurotransmitters in the central nervous system and exhibit neuroprotective effects. However, the effect of (+)-borneol on the dopaminergic neuronal loss of methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice is not defined. Herein, we first report that 30 mg/kg (+)-borneol significantly attenuated the motor deficits of PD mice, which benefits from markedly increasing the level of DA and decreasing the metabolic rate of DA in the striatum of conscious and freely moving mouse detected by ultraperformance liquid chromatography tandem mass spectrometry online combined with in vivo brain microdialysis sampling. It is worth noting that the enhanced level of DA by (+)-borneol was enabled by the reduction in loss of tyrosine hydroxylase-immunoreactive dopaminergic neurons in the substantia nigra and striatum and promotion of reserpine- or nomifensine-induced DA release in PD mice. Interestingly, (+)-borneol evidently inhibited the decreased expression levels of DA transporter (DAT) and vesicular monoamine transporter 2 (VMAT2) on the MPTP mouse model of PD. Moreover, (+)-borneol suppressed the neuroinflammation by inhibiting the production of IL-1ß, IL-6, and TNF-α and attenuated oxidative stress by decreasing the level of MDA and increasing the activities of SOD and GSH-px in PD mice. These findings demonstrate that (+)-borneol protects DA neurons by inhibiting neuroinflammation and oxidative stress. Further research work for the neuroprotection mechanism of (+)-borneol will focus on reactive oxygen species-mediated apoptosis. Therefore, (+)-borneol is a potential therapeutic candidate for retarding the neurodegenerative process of PD.

Uninterrupted in vivo cerebral microdialysis measures of the acute neurochemical response to a single or repeated concussion in a rat model combining force and rotation.

Altered extracellular amino acid concentrations following concussion or mild traumatic brain injury can result in delayed neuronal damage through overactivation of NMDA glutamatergic receptors. However, the consequences of repeated concussions prior to complete recovery are not well understood. In this study, we utilized in vivo cerebral microdialysis and a weight-drop model to investigate the acute neurochemical response to single and repeated concussions in adult rats that were fully conscious. A microdialysis probe was inserted into the hippocampus and remained in place during impact. Primary outcomes included concentrations of glutamate, GABA, taurine, glycine, glutamine, and serine, while secondary outcomes were righting times and excitotoxic indices. Compared to sham injury, the first concussion resulted in significant increases in glutamate, GABA, taurine, and glycine levels, longer righting times, and higher excitotoxic indices. Following the second concussion, righting times were significantly longer, suggesting cumulative effects of repeated concussion while only partial increases were observed in glutamate and taurine levels. GABA and glycine levels, and excitotoxic indices were comparable to sham injury. These findings suggest that single and repeated concussions may induce acute increases in several amino acids, while repeated concussions could exacerbate neurological symptoms despite less pronounced neurochemical changes.

UHPLC-MS/MS combined with microdialysis for simultaneous determination of nicotine and neurotransmitter metabolites in the rat hippocampal brain region: application to pharmacokinetic and pharmacodynamic study.

Nicotine crosses the blood-brain barrier and interacts with nicotinic acetylcholine receptors, initiating a cascade of neurotransmitter effects with potential therapeutic implications for neurodegenerative conditions such as Alzheimer's and Parkinson's disease. The hippocampus, pivotal for cognitive processes, plays a crucial role in nicotine-mediated cognitive enhancement due to its abundant expression of nicotinic acetylcholine receptors, particularly the α7 subtype, which is heavily implicated in hippocampus-related behavioral functions and dysfunctions. However, the intricate process of nicotine metabolism within the hippocampus remains poorly understood, impeding our comprehension of how nicotine and its metabolites modulate neurotransmitter dynamics. To address this gap, we have developed and validated a novel methodology combining microdialysis with UHPLC-MS/MS, enabling simultaneous detection of 12 neurotransmitters, nicotine, and its seven metabolites within the rat hippocampus. The linearity range of the targeted compounds is satisfactory (R2 > 0.9970), with intra-day and inter-day precision not exceeding 12.7%, and accuracy ranging from -12.4% to 13.7%. Our findings reveal differential pharmacokinetics of nicotine and its metabolites in the α7KO group compared to the control group, characterized by heightened nicotine absorption and slower elimination and distribution in the former. Notably, the pharmacokinetic parameters of cotinine exhibit similarity across both groups. Studies investigating the impact of nicotine on monoamine neurotransmitters have elucidated its capacity to augment the release of dopamine, serotonin, norepinephrine, glutamate, and acetylcholine in the rat hippocampus. This integrated approach facilitates a comprehensive analysis of neurotransmitter alterations within the hippocampal region following nicotine administration, thereby providing robust technical support and scientific rationale for understanding the neurochemical effects of nicotine and its metabolites. Further exploration into the pharmacokinetics and pharmacodynamics of nicotine holds promise for uncovering novel therapeutic avenues in the management of neurodegenerative diseases such as Alzheimer's.

Friedman's Gradient-Boosting Algorithm Predicts Lactate-Pyruvate Ratio Trends in Cases of Intracerebral Hemorrhages.

OBJECTIVE: The local effects of an intracerebral hemorrhage (ICH) on surrounding brain tissue can be detected bedside using multimodal brain monitoring techniques. The aim of this study is to design a gradient boosting regression model using the R package boostmtree with the ability to predict lactate-pyruvate ratio measurements in ICH. METHODS: We performed a retrospective analysis of 6 spontaneous ICH and 6 traumatic ICH patients who underwent surgical removal of the clot with microdialysis catheters placed in the perihematomal zone. Predictors of glucose, lactate, pyruvate, age, sex, diagnosis, and operation status were used to design our model. RESULTS: In a holdout analysis, the model forecasted lactate-pyruvate ratio trends in a representative in-sample testing set. We anticipate that boostmtree could be applied to designs of similar regression models to analyze trends in other multimodal monitoring features across other types of acute brain injury. CONCLUSIONS: The model successfully predicted hourly lactate-pyruvate ratios in spontaneous ICH and traumatic ICH cases after the hemorrhage evacuation and displayed significantly better performance than linear models. Our results suggest that boostmtree may be a powerful tool in developing more advanced mathematical models to assess other multimodal monitoring parameters for cases in which the perihematomal environment is monitored.

The effects of cerebral pressure autoregulation status and CPP levels on cerebral metabolism in pediatric traumatic brain injury.

BACKGROUND: Cerebral perfusion pressure (CPP) management in the developing child with traumatic brain injury (TBI) is challenging. The pressure reactivity index (PRx) may serve as marker of cerebral pressure autoregulation (CPA) and optimal CPP (CPPopt) may be assessed by identifying the CPP level with best (lowest) PRx. To evaluate the potential of CPPopt guided management in children with severe TBI, cerebral microdialysis (CMD) monitoring levels of lactate and the lactate/pyruvate ratio (LPR) (indicators of ischemia) were related to actual CPP levels, autoregulatory state (PRx) and deviations from CPPopt (ΔCPPopt). METHODS: Retrospective study of 21 children ≤ 17 years with severe TBI who had both ICP and CMD monitoring were included. CPP, PRx, CPPopt and ΔCPPopt where calculated, dichotomized and compared with CMD lactate and lactate-pyruvate ratio. RESULTS: Median age was 16 years (range 8-17) and median Glasgow coma scale motor score 5 (range 2-5). Both lactate (p = 0.010) and LPR (p = < 0.001) were higher when CPP ≥ 70 mmHg than when CPP < 70. When PRx ≥ 0.1 both lactate and LPR were higher than when PRx < 0.1 (p = < 0.001). LPR was lower (p = 0.012) when CPPopt ≥ 70 mmHg than when CPPopt < 70, but there were no differences in lactate levels. When ΔCPPopt > 10 both lactate (p = 0.026) and LPR (p = 0.002) were higher than when ΔCPPopt < -10. CONCLUSIONS: Increased levels of CMD lactate and LPR in children with severe TBI appears to be related to disturbed CPA (PRx). Increased lactate and LPR also seems to be associated with actual CPP levels ≥ 70 mmHg. However, higher lactate and LPR values were also seen when actual CPP was above CPPopt. Higher CPP appears harmful when CPP is above the upper limit of pressure autoregulation. The findings indicate that CPPopt guided CPP management may have potential in pediatric TBI.

Pharmacokinetic analysis of antiviral drug ritonavir across the blood-brain barrier and its interaction with Scutellaria baicalensis using multisite microdialysis in rats.

Ritonavir, an excellent inhibitor of CYP3A4, has recently been combined with nirmatrelvir to form Paxlovid for the treatment of severe acute respiratory syndrome coronavirus 2 infections. The root of Scutellaria baicalensis Georgi (S. baicalensis), a traditional Chinese medicinal (TCM) herb commonly used to treat heat/inflammation in the lung and digestive tracts, which are major organs targeted by viral infections, contains flavones that can influence the CYP3A metabolism pathway. To investigate the ability of ritonavir to cross the bloodbrain barrier (BBB) and its potential herb-drug interactions with an equivalent TCM clinical dose of S. baicalensis, multisite microdialysis coupled with an LCMS/MS system was developed using rat model. Pretreatment with S. baicalensis extract for 5 days, which contains less flavones than those used in previous studies, had a significant influence on ritonavir, resulting in a 2-fold increase in the total concentration of flavones in the blood and brain. Treatment also boosted the maximum blood concentration of flavones by 1.5-fold and the maximum brain concentration of flavones by 2-fold, all the while exerting no noticeable influence on the transfer ratio across the bloodbrain barrier. These experimental results demonstrated that the use of a typical traditional Chinese medicinal dose of S. baicalensis is sufficient to influence the metabolic pathway and synergistically increase the concentration of ritonavir in rats.

Continuous evaluation of single-dose moxifloxacin concentrations in brain extracellular fluid, cerebrospinal fluid, and plasma: a novel porcine model.

BACKGROUND: Knowledge regarding CNS pharmacokinetics of moxifloxacin is limited, with unknown consequences for patients with meningitis caused by bacteria resistant to beta-lactams or caused by TB. OBJECTIVE: (i) To develop a novel porcine model for continuous investigation of moxifloxacin concentrations within brain extracellular fluid (ECF), CSF and plasma using microdialysis, and (ii) to compare these findings to the pharmacokinetic/pharmacodynamic (PK/PD) target against TB. METHODS: Six female pigs received an intravenous single dose of moxifloxacin (6 mg/kg) similar to the current oral treatment against TB. Subsequently, moxifloxacin concentrations were determined by microdialysis within five compartments: brain ECF (cortical and subcortical) and CSF (ventricular, cisternal and lumbar) for the following 8 hours. Data were compared to simultaneously obtained plasma samples. Chemical analysis was performed by high pressure liquid chromatography with mass spectrometry. The applied PK/PD target was defined as a maximum drug concentration (Cmax):MIC ratio >8. RESULTS: We present a novel porcine model for continuous in vivo CNS pharmacokinetics for moxifloxacin. Cmax and AUC0-8h within brain ECF were significantly lower compared to plasma and lumbar CSF, but insignificantly different compared to ventricular and cisternal CSF. Unbound Cmax:MIC ratio across all investigated compartments ranged from 1.9 to 4.3. CONCLUSION: A single dose of weight-adjusted moxifloxacin administered intravenously did not achieve adequate target site concentrations within the uninflamed porcine brain ECF and CSF to reach the applied TB CNS target.

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.

Negligible In Vitro Recovery of Macromolecules from Microdialysis Using 100 kDa Probes and Dextran in Perfusion Fluid.

Microdialysis is applied in neurointensive care to monitor cerebral glucose metabolism. If recoverable, macromolecules may also serve as biomarkers in brain disease and provide clues to their passage across the blood-brain barrier. Our study aimed to investigate the in vitro recovery of human micro- and macromolecules using microdialysis catheters and perfusion fluids approved for clinical use. In vitro microdialysis of a bulk solution containing physiological or supraphysiological concentrations of glucose, lactate, pyruvate, human IgG, serum albumin, and hemoglobin was performed using two different catheters and perfusion fluids. One had a membrane cut-off of 20 kDa and was used with a standard CNS perfusion fluid, and the other had a membrane cut-off of 100 kDa and was perfused with the same solution supplemented with dextran. The flow rate was 0.3 µl/min. We used both push and push-pull methods. Dialysate samples were collected at 2-h intervals for 6 h and analyzed for relative recovery of each substance. The mean relative recovery of glucose, pyruvate, and lactate was > 90% in all but two sets of experiments. In contrast, the relative recovery of human IgG, serum albumin, and hemoglobin from both bulk solutions was below the lower limit of quantification (LLOQ). Using a push-pull method, recovery of human IgG, serum albumin, and hemoglobin from a bulk solution with supraphysiological concentrations were above LLOQ but with low relative recovery (range 0.9%-1.6%). In summary, exchanging the microdialysis setup from a 20 kDa catheter with a standard perfusion fluid for a 100 kDa catheter with a perfusion solution containing dextran did not affect the relative recovery of glucose and its metabolites. However, it did not result in any useful recovery of the investigated macromolecules at physiological levels, either with or without a push-pull pump system.

Characterization of the Neurochemical and Behavioral Effects of the Phenethylamine 2-Cl-4,5-MDMA in Adolescent and Adult Male Rats.

BACKGROUND: The proliferation of novel psychoactive substances (NPS) in the drug market raises concerns about uncertainty on their pharmacological profile and the health hazard linked to their use. Within the category of synthetic stimulant NPS, the phenethylamine 2-Cl-4,5-methylenedioxymethamphetamine (2-Cl-4,5-MDMA) has been linked to severe intoxication requiring hospitalization. Thereby, the characterization of its pharmacological profile is urgently warranted. METHODS: By in vivo brain microdialysis in adolescent and adult male rats we investigated the effects of 2-Cl-4,5-MDMA on dopamine (DA) and serotonin (5-HT) neurotransmission in two brain areas critical for the motivational and rewarding properties of drugs, the nucleus accumbens (NAc) shell and the medial prefrontal cortex (mPFC). Moreover, we evaluated the locomotor and stereotyped activity induced by 2-Cl-4,5-MDMA and the emission of 50-kHz ultrasonic vocalizations (USVs) to characterize its affective properties. RESULTS: 2-Cl-4,5-MDMA increased dialysate DA and 5-HT in a dose-, brain area-, and age-dependent manner. Notably, 2-Cl-4,5-MDMA more markedly increased dialysate DA in the NAc shell and mPFC of adult than adolescent rats, while the opposite was observed on dialysate 5-HT in the NAc shell, with adolescent rats being more responsive. Furthermore, 2-Cl-4,5-MDMA stimulated locomotion and stereotyped activity in both adolescent and adult rats, although to a greater extent in adolescents. Finally, 2-Cl-4,5-MDMA did not stimulate the emission of 50-kHz USVs. CONCLUSIONS: This is the first pharmacological characterization of 2-Cl-4,5-MDMA demonstrating that its neurochemical and behavioral effects may differ between adolescence and adulthood. These preclinical data could help understanding the central effects of 2-Cl-4,5-MDMA by increasing awareness on possible health damage in users.

Brain Regional Energy Metabolism in Patients with Traumatic Brain Injury: A Cerebral Microdialysis Guided Study.

BACKGROUND: In traumatic brain injuries (TBI), cerebral microdialysis (CMD)-derived parameters, especially the lactate to pyruvate ratio (LP ratio), have been utilized for cerebral perfusion optimization. The objectives were to identify cerebral ischemia as measured by CMD in TBI patients requiring decompressive craniectomy and to observe the correlation between cerebral perfusion pressure (CPP), intracranial pressure (ICP), and CMD variables in these patients. Our secondary aim was to observe the effect of CPP augmentation on ischemia biomarkers. METHODS: After the Institute Ethics Committee approvals, seven adult patients requiring decompressive craniectomy following TBI were enrolled and CMD data were obtained prospectively for 72 h. CPP was augmented by 20% with noradrenaline infusion if LP ratio >40. Correlations were done with bootstrapping (n = 500) to obtain the confidence intervals (CI) due to the small sample size. RESULTS: One patient had cerebral ischemia (median LP ratio of 265.5 and median pyruvate of 38 µmol/L), while another patient had non-ischemic mitochondrial dysfunction (median LP ratio 40.7 and median pyruvate 278.5). The coefficients of correlation between the LP ratio with CPP and ICP were r = -0.05 (CI = -0.14-0.03) and r = 0.09 (CI = -0.03-0.24), respectively. The coefficient of correlation between cerebral and blood glucose was r = 0.38, (CI - 0.35-0.14). Only two patients needed CPP augmentation, however, postaugmentation cerebral biochemistry did not change appreciably. CONCLUSION: CMD can identify cerebral ischemia, however, no correlations were observed between the LP ratio and CPP or ICP. CPP augmentation did not improve cerebral biochemistry. More studies are required to understand and treat cerebral metabolism in TBI.

Brain metabolism response to intrahospital transfers in neurocritical ill patients and the impact of microdialysis probe location.

Intrahospital transfer (IHT), a routine in the management of neurocritical patients requiring imaging or interventions, might affect brain metabolism. Studies about IHT effects using microdialysis (MD) have produced conflicting results. In these studies, only the most damaged hemisphere was monitored, and those may not reflect the impact of IHT on overall brain metabolism, nor do they address differences between the hemispheres. Herein we aimed to quantify the effect of IHT on brain metabolism by monitoring both hemispheres with bilateral MD. In this study, 27 patients with severe brain injury (10 traumatic brain injury and 17 subarachnoid hemorrhage patients) were included, with a total of 67 IHT. Glucose, glycerol, pyruvate and lactate were measured by MD in both hemispheres for 10 h pre- and post-IHT. Alterations in metabolite levels after IHT were observed on both hemispheres; although these changes were more marked in hemisphere A (most damaged) than B (less damaged). Our results suggest that brain metabolism is altered after an IHT of neurocritical ill patients particularly but not limited to the damaged hemisphere. Bilateral monitorization may be more sensitive than unilateral monitorization for detecting metabolic disturbances not directly related to the course of the disease.

Microdialysis coupled with droplet microfluidics and mass spectrometry for determination of neurotransmitters in vivo with high temporal resolution.

Monitoring the concentration fluctuations of neurotransmitters in vivo is valuable for elucidating the chemical signals that underlie brain functions. Microdialysis sampling is a widely used tool for monitoring neurochemicals in vivo. The volume requirements of most techniques that have been coupled to microdialysis, such as HPLC, result in fraction collection times of minutes, thus limiting the temporal resolution possible. Further the time of analysis can become long for cases where many fractions are collected. Previously we have used direct analysis of dialysate by low-flow electrospray ionization-tandem mass spectrometry (ESI-MS/MS) on a triple quadrupole mass spectrometer to monitor acetylcholine, glutamate, and γ-amino-butyric acid to achieve multiplexed in vivo monitoring with temporal resolution of seconds. Here, we have expanded this approach to adenosine, dopamine, and serotonin. The method achieved limits of detection down to 2 nM, enabling basal concentrations of all these compounds, except serotonin, to be measured in vivo. Comparative analysis with LC-MS/MS showed accurate results for all compounds except for glutamate, possibly due to interference for this compound in vivo. Pairing this analysis with droplet microfluidics yields 11 s temporal resolution and can generate dialysate fractions down to 3 nL at rates up to 3 fractions per s from a microdialysis probe. The system is applied to multiplexed monitoring of neurotransmitter dynamics in response to stimulation by 100 mM K+ and amphetamine. These applications demonstrate the suitability of the droplet ESI-MS/MS method for monitoring short-term dynamics of up to six neurotransmitters simultaneously.

Dual DAT and sigma receptor inhibitors attenuate cocaine effects on nucleus accumbens dopamine dynamics in rats.

Psychostimulant use disorders (PSUD) are prevalent; however, no FDA-approved medications have been made available for treatment. Previous studies have shown that dual inhibitors of the dopamine transporter (DAT) and sigma receptors significantly reduce the behavioral/reinforcing effects of cocaine, which have been associated with stimulation of extracellular dopamine (DA) levels resulting from DAT inhibition. Here, we employ microdialysis and fast scan cyclic voltammetry (FSCV) procedures to investigate the effects of dual inhibitors of DAT and sigma receptors in combination with cocaine on nucleus accumbens shell (NAS) DA dynamics in naïve male Sprague Dawley rats. In microdialysis studies, administration of rimcazole (3, 10 mg/kg; i.p.) or its structural analog SH 3-24 (1, 3 mg/kg; i.p.), compounds that are dual inhibitors of DAT and sigma receptors, significantly reduced NAS DA efflux stimulated by increasing doses of cocaine (0.1, 0.3, 1.0 mg/kg; i.v.). Using the same experimental conditions, in FSCV tests, we show that rimcazole pretreatments attenuated cocaine-induced stimulation of evoked NAS DA release but produced no additional effect on DA clearance rate. Under the same conditions, JJC8-091, a modafinil analog and dual inhibitor of DAT and sigma receptors, similarly attenuated cocaine-induced stimulation of evoked NAS DA release but produced no additional effect on DA clearance rate. Our results provide the neurochemical groundwork towards understanding actions of dual inhibitors of DAT and sigma receptors on DA dynamics that likely mediate the behavioral effects of psychostimulants like cocaine.

Intraparenchymal near-infrared spectroscopy for detection of delayed cerebral ischemia in poor-grade aneurysmal subarachnoid hemorrhage.

OBJECTIVE: Detection of delayed cerebral ischemia (DCI) is challenging in comatose patients with poor-grade aneurysmal subarachnoid hemorrhage (aSAH). Brain tissue oxygen pressure (PbtO2) monitoring may allow early detection of its occurrence. Recently, a probe for combined measurement of intracranial pressure (ICP) and intraparenchymal near-infrared spectroscopy (NIRS) has become available. In this pilot study, the parameters PbtO2, Hboxy, Hbdeoxy, Hbtotal and rSO2 were measured in parallel and evaluated for their potential to detect perfusion deficits or cerebral infarction. METHODS: In patients undergoing multimodal neuromonitoring due to poor neurological condition after aSAH, Clark oxygen probes, microdialysis and NIRS-ICP probes were applied. DCI was suspected when the measured parameters in neuromonitoring deteriorated. Thus, perfusion CT scan was performed as follow up, and DCI was confirmed as perfusion deficit. Median values for PbtO2, Hboxy, Hbdeoxy, Hbtotal and rSO2 in patients with perfusion deficit (Tmax > 6 s in at least 1 vascular territory) and/or already demarked infarcts were compared in 24- and 48-hour time frames before imaging. RESULTS: Data from 19 patients (14 University Hospital Zurich, 5 Charité Universitätsmedizin Berlin) were prospectively collected and analyzed. In patients with perfusion deficits, the median values for Hbtotal and Hboxy in both time frames were significantly lower. With perfusion deficits, the median values for Hboxy and Hbtotal in the 24 h time frame were 46,3 [39.6, 51.8] µmol/l (no perfusion deficits 53 [45.9, 55.4] µmol/l, p = 0.019) and 69,3 [61.9, 73.6] µmol/l (no perfusion deficits 74,6 [70.1, 79.6] µmol/l, p = 0.010), in the 48 h time frame 45,9 [39.4, 51.5] µmol/l (no perfusion deficits 52,9 [48.1, 55.1] µmol/l, p = 0.011) and 69,5 [62.4, 74.3] µmol/l (no perfusion deficits 75 [70,80] µmol/l, p = 0.008), respectively. In patients with perfusion deficits, PbtO2 showed no differences in both time frames. PbtO2 was significantly lower in patients with infarctions in both time frames. The median PbtO2 was 17,3 [8,25] mmHg (with no infarctions 29 [22.5, 36] mmHg, p = 0.006) in the 24 h time frame and 21,6 [11.1, 26.4] mmHg (with no infarctions 31 [22,35] mmHg, p = 0.042) in the 48 h time frame. In patients with infarctions, the median values of parameters measured by NIRS showed no significant differences. CONCLUSIONS: The combined NIRS-ICP probe may be useful for early detection of cerebral perfusion deficits and impending DCI. Validation in larger patient collectives is needed.

Understanding the nebulisation of antibiotics: the key role of lung microdialysis studies.

BACKGROUND: Nebulisation of antibiotics is a promising treatment for ventilator-associated pneumonia (VAP) caused by multidrug-resistant organisms. Ensuring effective antibiotic concentrations at the site of infection in the interstitial space fluid is crucial for clinical outcomes. Current assessment methods, such as epithelial lining fluid and tissue homogenates, have limitations in providing longitudinal pharmacokinetic data. MAIN BODY: Lung microdialysis, an invasive research technique predominantly used in animals, involves inserting probes into lung parenchyma to measure antibiotic concentrations in interstitial space fluid. Lung microdialysis offers unique advantages, such as continuous sampling, regional assessment of antibiotic lung concentrations and avoidance of bronchial contamination. However, it also has inherent limitations including the cost of probes and assay development, the need for probe calibration and limited applicability to certain antibiotics. As a research tool in VAP, lung microdialysis necessitates specialist techniques and resource-intensive experimental designs involving large animals undergoing prolonged mechanical ventilation. However, its potential impact on advancing our understanding of nebulised antibiotics for VAP is substantial. The technique may enable the investigation of various factors influencing antibiotic lung pharmacokinetics, including drug types, delivery devices, ventilator settings, interfaces and disease conditions. Combining in vivo pharmacokinetics with in vitro pharmacodynamic simulations can become feasible, providing insights to inform nebulised antibiotic dose optimisation regimens. Specifically, it may aid in understanding and optimising the nebulisation of polymyxins, effective against multidrug-resistant Gram-negative bacteria. Furthermore, lung microdialysis holds promise in exploring novel nebulisation therapies, including repurposed antibiotic formulations, bacteriophages and immunomodulators. The technique's potential to monitor dynamic biochemical changes in pneumonia, such as cytokines, metabolites and inflammation/infection markers, opens avenues for developing theranostic tools tailored to critically ill patients with VAP. CONCLUSION: In summary, lung microdialysis can be a potential transformative tool, offering real-time insights into nebulised antibiotic pharmacokinetics. Its potential to inform optimal dosing regimen development based on precise target site concentrations and contribute to development of theranostic tools positions it as key player in advancing treatment strategies for VAP caused by multidrug-resistant organisms. The establishment of international research networks, exemplified by LUMINA (lung microdialysis applied to nebulised antibiotics), signifies a proactive step towards addressing complexities and promoting multicentre experimental studies in the future.