Acute Inflammation Alters Brain Energy Metabolism in Mice and Humans: Role in Suppressed Spontaneous Activity, Impaired Cognition, and Delirium.

Systemic infection triggers a spectrum of metabolic and behavioral changes, collectively termed sickness behavior, which while adaptive, can affect mood and cognition. In vulnerable individuals, acute illness can also produce profound, maladaptive, cognitive dysfunction including delirium, but our understanding of delirium pathophysiology remains limited. Here, we used bacterial lipopolysaccharide (LPS) in female C57BL/6J mice and acute hip fracture in humans to address whether disrupted energy metabolism contributes to inflammation-induced behavioral and cognitive changes. LPS (250 µg/kg) induced hypoglycemia, which was mimicked by interleukin (IL)-1ß (25 µg/kg) but not prevented in IL-1RI-/- mice, nor by IL-1 receptor antagonist (IL-1RA; 10 mg/kg). LPS suppression of locomotor activity correlated with blood glucose concentrations, was mitigated by exogenous glucose (2 g/kg), and was exacerbated by 2-deoxyglucose (2-DG) glycolytic inhibition, despite preventing IL-1ß synthesis. Using the ME7 model of chronic neurodegeneration in female mice, to examine vulnerability of the diseased brain to acute stressors, we showed that LPS (100 µg/kg) produced acute cognitive dysfunction, selectively in those animals. These acute cognitive impairments were mimicked by insulin (11.5 IU/kg) and mitigated by glucose, demonstrating that acutely reduced glucose metabolism impairs cognition selectively in the vulnerable brain. To test whether these acute changes might predict altered carbohydrate metabolism during delirium, we assessed glycolytic metabolite levels in CSF in humans during inflammatory trauma-induced delirium. Hip fracture patients showed elevated CSF lactate and pyruvate during delirium, consistent with acutely altered brain energy metabolism. Collectively, the data suggest that disruption of energy metabolism drives behavioral and cognitive consequences of acute systemic inflammation.SIGNIFICANCE STATEMENT Acute systemic inflammation alters behavior and produces disproportionate effects, such as delirium, in vulnerable individuals. Delirium has serious short and long-term sequelae but mechanisms remain unclear. Here, we show that both LPS and interleukin (IL)-1ß trigger hypoglycemia, reduce CSF glucose, and suppress spontaneous activity. Exogenous glucose mitigates these outcomes. Equivalent hypoglycemia, induced by lipopolysaccharide (LPS) or insulin, was sufficient to trigger cognitive impairment selectively in animals with existing neurodegeneration and glucose also mitigated those impairments. Patient CSF from inflammatory trauma-induced delirium also shows altered brain carbohydrate metabolism. The data suggest that the degenerating brain is exquisitely sensitive to acute behavioral and cognitive consequences of disrupted energy metabolism. Thus "bioenergetic stress" drives systemic inflammation-induced dysfunction. Elucidating this may offer routes to mitigating delirium.

Multicomponent analysis using a confocal Raman microscope.

Measuring the concentration of multiple chemical components in a low-volume aqueous mixture by Raman spectroscopy has received significant interest in the literature. All of the contributions to date focus on the design of optical systems that facilitate the recording of spectra with high signal-to-noise ratio by collecting as many Raman scattered photons as possible. In this study, the confocal Raman microscope setup is investigated for multicomponent analysis. Partial least-squares regression is used to quantify physiologically relevant aqueous mixtures of glucose, lactic acid, and urea. The predicted error is 17.81 mg/dL for glucose, 10.6 mg/dL for lactic acid, and 7.6 mg/dL for urea, although this can be improved with increased acquisition times. A theoretical analysis of the method is proposed, which relates the numerical aperture and the magnification of the microscope objective, as well as the confocal pinhole size, to the performance of the technique.

Aversive prediction error signals in the amygdala.

Prediction error signals are fundamental to learning. Here, in mice, we show that aversive prediction signals are found in the hemodynamic responses and theta oscillations recorded from the basolateral amygdala. During fear conditioning, amygdala responses evoked by footshock progressively decreased, whereas responses evoked by the auditory cue that predicted footshock concomitantly increased. Unexpected footshock evoked larger amygdala responses than expected footshock. The magnitude of the amygdala response to the footshock predicted behavioral responses the following day. The omission of expected footshock led to a decrease below baseline in the amygdala response suggesting a negative aversive prediction error signal. Thus, in mice, amygdala activity conforms to temporal difference models of aversive learning.

Differential contributions of infralimbic prefrontal cortex and nucleus accumbens during reward-based learning and extinction.

Using environmental cues for the prediction of future events is essential for survival. Such cue-outcome associations are thought to depend on mesolimbic circuitry involving the nucleus accumbens (NAc) and prefrontal cortex (PFC). Several studies have identified roles for both NAc and PFC in the expression of stable goal-directed behaviors, but much remains unknown about their roles during learning of such behaviors. To further address this question, we used in vivo oxygen amperometry, a proxy for blood oxygen level-dependent (BOLD) signal measurement in human functional magnetic resonance imaging, in rats performing a cued lever-pressing task requiring discrimination between a rewarded and nonrewarded cue. Simultaneous oxygen recordings were obtained from infralimbic PFC (IFC) and NAc throughout both acquisition and extinction of this task. Activation of NAc was specifically observed following rewarded cue onset during the entire acquisition phase and also during the first days of extinction. In contrast, IFC activated only during the earliest periods of acquisition and extinction, more specifically to the nonrewarded cue. Thus, in vivo oxygen amperometry permits a novel, stable form of longitudinal analysis of brain activity in behaving animals, allowing dissociation of the roles of different brain regions over time during learning of reward-driven instrumental action. The present results offer a unique temporal perspective on how NAc may promote actions directed toward anticipated positive outcome throughout learning, while IFC might suppress actions that no longer result in reward, but only during critical periods of learning.

A microelectrochemical biosensor for real-time in vivo monitoring of brain extracellular choline.

A first generation Pt-based polymer enzyme composite biosensor developed for real-time neurochemical monitoring was characterised in vivo for sensitive and selective detection of choline. Confirmation that the sensor responds to changes in extracellular choline was achieved using local perfusion of choline which resulted in an increase in current, and the acetylcholinesterase inhibitor neostigmine which produced a decrease. Interference by electroactive species was tested using systemic administration of sodium ascorbate which produced a rapid increase in extracellular levels before gradually returning towards baseline over several hours. There was no overall change in the response of the biosensor during the same period of monitoring. Oxygen interference was examined using pharmacological agents known to change tissue oxygenation. Chloral hydrate produced an immediate increase in O2 before gradually returning to baseline levels over 3 h. The biosensor signal displayed an initial brief decrease before increasing to a maximum after 1 h and returning to baseline within 2 h. L-NAME caused a decrease in O2 before returning to baseline levels after ca. 1.5 h. In contrast, the biosensor current increased over the same time period before slowly returning to baseline levels over several hours. Such differences in time course and direction suggest that changes in tissue O2 levels do not affect the ability of the sensor to monitor choline reliably. Although it was found to rapidly respond to behavioural activation, examination of baseline in vivo data suggests a stable viable signal for at least 14 days after implantation. Using in vitro calibration data the basal extracellular concentration of choline was estimated to be 6.3 µM.

Characterisation of a microelectrochemical biosensor for real-time detection of brain extracellular d-serine.

A modified development protocol and concomitant characterisation of a first generation biosensor for the detection of brain extracellular d-serine is reported. Functional parameters important for neurochemical monitoring, including sensor sensitivity, O2 interference, selectivity, shelf-life and biocompatibility were examined. Construction and development involved the enzyme d-amino acid oxidase (DAAO), utilising a dip-coating immobilisation method employing a new extended drying approach. The resultant Pt-based polymer enzyme composite sensor achieved high sensitivity to d-serine (0.76 ± 0.04 nA mm-2. µM-1) and a low µM limit of detection (0.33 ± 0.02 µM). The in-vitro response time was within the solution stirring time, suggesting potential sub-second in-vivo response characteristics. Oxygen interference studies demonstrated a 1 % reduction in current at 50 µM O2 when compared to atmospheric O2 levels (200 µM), indicating that the sensor can be used for reliable neurochemical monitoring of d-serine, free from changes in current associated with physiological O2 fluctuations. Potential interference signals generated by the principal electroactive analytes present in the brain were minimised by using a permselective layer of poly(o-phenylenediamine), and although several d-amino acids are possible substrates for DAAO, their physiologically relevant signals were small relative to that for d-serine. Additionally, changing both temperature and pH over possible in vivo ranges (34-40 °C and 7.2-7.6 respectively) resulted in no significant effect on performance. Finally, the biosensor was implanted in the striatum of freely moving rats and used to monitor physiological changes in d-serine over a two-week period.

Hyperthermia elevates brain temperature and improves behavioural signs in animal models of autism spectrum disorder.

BACKGROUND: Autism spectrum disorders (ASD) are predominantly neurodevelopmental and largely genetically determined. However, there are human data supporting the idea that fever can improve symptoms in some individuals, but those data are limited and there are almost no data to support this from animal models. We aimed to test the hypothesis that elevated body temperature would improve function in two animal models of ASD. METHODS: We used a 4 h whole-body hyperthermia (WBH) protocol and, separately, systemic inflammation induced by bacterial endotoxin (LPS) at 250 µg/kg, to dissociate temperature and inflammatory elements of fever in two ASD animal models: C58/J and Shank3B- mice. We used one- or two-way ANOVA and t-tests with normally distributed data and Kruskal-Wallis or Mann-Whitney with nonparametric data. Post hoc comparisons were made with a level of significance set at p < 0.05. For correlation analyses, data were adjusted by a linear regression model. RESULTS: Only LPS induced inflammatory signatures in the brain while only WBH produced fever-range hyperthermia. WBH reduced repetitive behaviours and improved social interaction in C58/J mice and significantly reduced compulsive grooming in Shank3B- mice. LPS significantly suppressed most activities over 5-48 h. LIMITATIONS: We show behavioural, cellular and molecular changes, but provide no specific mechanistic explanation for the observed behavioural improvements. CONCLUSIONS: The data are the first, to our knowledge, to demonstrate that elevated body temperature can improve behavioural signs in 2 distinct ASD models. Given the developmental nature of ASD, evidence that symptoms may be improved by environmental perturbations indicates possibilities for improving function in these individuals. Since experimental hyperthermia in patients would carry significant risks, it is now essential to pursue molecular mechanisms through which hyperthermia might bring about the observed benefits.

Changes in reward-related signals in the rat nucleus accumbens measured by in vivo oxygen amperometry are consistent with fMRI BOLD responses in man.

Real-time in vivo oxygen amperometry, a technique that allows measurement of regional brain tissue oxygen (O(2)) has been previously shown to bear relationship to the BOLD signal measured with functional magnetic resonance imaging (fMRI) protocols. In the present study, O(2) amperometry was applied to the study of reward processing in the rat nucleus accumbens to validate the technique with a behavioural process known to cause robust signals in human neuroimaging studies. After acquisition of a cued-lever pressing task a robust increase in O(2) tissue levels was observed in the nucleus accumbens specifically following a correct lever press to the rewarded cue. This O(2) signal was modulated by cue reversal but not lever reversal, by differences in reward magnitudes and by the motivational state of the animal consistent with previous reports of the role of the nucleus accumbens in both the anticipation and representation of reward value. Moreover, this modulation by reward value was related more to the expected incentive value rather than the hedonic value of reward, also consistent with previous reports of accumbens coding of "wanting" of reward. Altogether, these results show striking similarities to those obtained in human fMRI studies suggesting the use of oxygen amperometry as a valid surrogate for fMRI in animals performing cognitive tasks, and a powerful approach to bridge between different techniques of measurement of brain function.

Design optimisation and characterisation of an amperometric glutamate oxidase-based composite biosensor for neurotransmitter l-glutamic acid.

A polymer/enzyme composite biosensor for monitoring neurochemical glutamate was performance optimised in vitro for sensitivity, selectivity and stability. This first generation Pt/glutamate oxidase-based sensor displayed appropriate sensitivity (90.4 ± 2.0 nA cm-2 µM-1). It also has ideal stability/biocompatibility with no significant decrease in response observed for repeated calibrations, exposure to electron beam sterilisation, or following storage at 4 °C either dry (28 days) or in ex-vivo rodent brain tissue (14 days). Potential non-glutamate contributing signals, generated by extracellular levels of the principal endogenous electroactive interferents, were typically <5% of the basal (10 µM) glutamate response. Changes in molecular oxygen (the natural enzyme mediator) over the normal brain tissue range of 40-80 µM had minimal effect on the glutamate signal for concentrations of 10 and 100 µM (Mean KMO2 = 1.86 ± 0.74 µM, [O2]90% = ca. 15 µM). Additionally, a low µM calculated limit of detection (0.44 ± 0.05) and rapid response time (ca. 1.67 ± 0.06 s), combined with no effect of pH and temperature changes over physiologically relevant ranges (7.2-7.6 and 34-40 °C respectively), collectively suggest that this composite biosensor should reliably detect l-glutamate when used for neurochemical monitoring. Preliminary experiments involving implantation in the striatum of freely moving rats demonstrated stable recording over several weeks, and reliable detection of physiological changes in glutamate in response to behavioural/neuronal activation (locomotor activity and restraint stress).

Brain tissue oxygen amperometry in behaving rats demonstrates functional dissociation of dorsal and ventral hippocampus during spatial processing and anxiety.

Traditionally, the function of the hippocampus (HPC) has been viewed in unitary terms, but there is growing evidence that the HPC is functionally differentiated along its septotemporal axis. Lesion studies in rodents and functional brain imaging in humans suggest a preferential role for the septal HPC in spatial learning and a preferential role for the temporal HPC in anxiety. To better enable cross-species comparison, we present an in vivo amperometric technique that measures changes in brain tissue oxygen at high temporal resolution in freely-moving rats. We recorded simultaneously from the dorsal (septal; dHPC) and ventral (temporal; vHPC) HPC during two anxiety tasks and two spatial tasks on the radial maze. We found a double-dissociation of function in the HPC, with increased vHPC signals during anxiety and increased dHPC signals during spatial processing. In addition, dHPC signals were modulated by spatial memory demands. These results add a new dimension to the growing consensus for a differentiation of HPC function, and highlight tissue oxygen amperometry as a valuable tool to aid translation between animal and human research.

Close temporal coupling of neuronal activity and tissue oxygen responses in rodent whisker barrel cortex.

Neuronal activity elicits metabolic and vascular responses, during which oxygen is first consumed and then supplied to the tissue via an increase in cerebral blood flow. Understanding the spatial and temporal dynamics of blood and tissue oxygen (To2) responses following neuronal activity is crucial for understanding the physiological basis of functional neuroimaging signals. However, our knowledge is limited because previous To2 measurements have been made at low temporal resolution (>100 ms). Here we recorded To2 at high temporal resolution (1 ms), simultaneously with co-localized field potentials, at several cortical depths from the whisker region of the somatosensory cortex in anaesthetized rats and mice. Stimulation of the whiskers produced rapid, laminar-specific changes in To2. Positive To2 responses (i.e. increases) were observed in the superficial layers within 50 ms of stimulus onset, faster than previously reported. Negative To2 responses (i.e. decreases) were observed in the deeper layers, with maximal amplitude in layer IV, within 40 ms of stimulus onset. The amplitude of the negative, but not the positive, To2 response correlated with local field potential amplitude. Disruption of neurovascular coupling, via nitric oxide synthase inhibition, abolished positive To2 responses to whisker stimulation in the superficial layers and increased negative To2 responses in all layers. Our data show that To2 responses occur rapidly following neuronal activity and are laminar dependent.

An in vitro characterisation comparing carbon paste and Pt microelectrodes for real-time detection of brain tissue oxygen.

In vitro characterisation results for O(2) reduction at Pt-based microelectrodes are presented and compared with those for carbon-paste electrodes (CPEs). Cyclic voltammetry indicates a potential of -650 mV vs. SCE is required for cathodic reduction at both electrode types, and calibration experiments at this potential revealed a significantly higher sensitivity for Pt (-0.091 ± 0.006 µAmm(-2)µM(-1) vs. -0.048 ± 0.002 µAmm(-2)µM(-1) for CPEs). Since Pt electrodes are readily poisoned through contact with biological samples selected surface coated polymers (polyphenylenediamine (PPD), polymethyl methacrylate (PMMA) and Rhoplex(®)) were examined in biocompatibility studies performed in protein, lipid and brain tissue solutions. While small and comparable decreases in sensitivity were observed for bare Pt, Pt-Rhoplex and PMMA there was minimal change at the Pt-PPD modified electrode for each 24h treatment, including an extended 3 day exposure to brain tissue. The polymers themselves had no effect on the O(2) response characteristics. Further characterisation studies at the Pt-based microelectrodes confirmed interference free signals, no effect of pH and ion changes, and a comparable detection limit (0.08 ± 0.01 µM) and response time (<1 s) to CPEs. Although a significant temperature effect (ca. 3% change in signal for each 1 °C) was observed it is predicted that this will not be important for in vivo brain tissue O(2) measurements due to brain temperature homeostasis. These results suggest that amperometric Pt electrodes have the potential to be used reliably as an alternative to CPEs to monitor brain tissue O(2) over extended periods in freely-moving animals.

Real-time electrochemical monitoring of brain tissue oxygen: a surrogate for functional magnetic resonance imaging in rodents.

Long-term in-vivo electrochemistry (LIVE) enables real-time monitoring and measurement of brain metabolites. In this study we have simultaneously obtained blood oxygenation level dependent (BOLD) fMRI and amperometric tissue O(2) data from rat cerebral cortex, during both increases and decreases in inspired O(2) content. BOLD and tissue O(2) measurements demonstrated close correlation (r=0.7898) during complete (0%) O(2) removal, with marked negative responses occurring ca. 30s after the onset of O(2) removal. Conversely, when the inspired O(2) was increased (50, 70 and 100% O(2) for 1min) similar positive rapid changes (ca. 15s) in both the BOLD and tissue O(2) signals were observed. These findings demonstrate, for the first time, the practical feasibility of obtaining real-time metabolite information during fMRI acquisition, and that tissue O(2) concentration monitored using an O(2) sensor can serve as an index of changes in the magnitude of the BOLD response. As LIVE O(2) sensors can be used in awake animals performing specific behavioural tasks the technique provides a viable animal surrogate of human fMRI experimentation.

Development and validation of a real-time microelectrochemical sensor for clinical monitoring of tissue oxygenation/perfusion.

Oxygen is of critical importance to tissue viability and there is increasing demand for its reliable real-time clinical monitoring in order to prevent, diagnose, and treat several pathological disorders, including hypoxia, stroke and reperfusion injury. Herein we report the development and characterisation of a prototype clinical O2 sensor, and its validation in vivo, including proof-of-concept monitoring in patients undergoing surgery for carpal tunnel release. An integrated platinum-based microelectrochemical device was custom designed and controlled using a miniaturised telemetry-operated single channel clinical potentiostat. The in vitro performance of different sensor configurations is presented, with the best sensor design (S2) displaying appropriate linearity (R2 = 0.994) and sensitivity (0.569 ± 0.022 nA µM-1). Pre-clinical validation of S2 was performed in the hind limb muscle of anaesthetised rats; tourniquet application resulted in a significant rapid decrease in signal (90 ± 27%, [ΔO2] ca. 140 ± 18 µM), with a return to baseline within a period of ca. 3 min following tourniquet release. Similar trends were observed in the clinical study; an immediate decrease in signal (39 ± 3%, [ΔO2] ca. 30 ± 20 µM), with basal levels re-established within 2 min of tourniquet release. These results confirm that continuous real-time monitoring of dynamic changes in tissue O2 can serve as an indicator of reperfusion status in patients undergoing carpal tunnel surgery, and suggests the potential usefulness of the developed microelectrochemical sensor for other medical conditions where clinical monitoring of O2 and perfusion is important.

Real-time monitoring of brain tissue oxygen using a miniaturized biotelemetric device implanted in freely moving rats.

A miniaturized biotelemetric device for the amperometric detection of brain tissue oxygen is presented. The new system, derived from a previous design, has been coupled with a carbon microsensor for the real-time detection of dissolved O(2) in the striatum of freely moving rats. The implantable device consists of a single-supply sensor driver, a current-to-voltage converter, a microcontroller, and a miniaturized data transmitter. The oxygen current is converted to a digital value by means of an analog-to-digital converter integrated in a peripheral interface controller (PIC). The digital data is sent to a personal computer using a six-byte packet protocol by means of a miniaturized 434 MHz amplitude modulation (AM) transmitter. The receiver unit is connected to a personal computer (PC) via a universal serial bus. Custom developed software allows the PC to store and plot received data. The electronics were calibrated and tested in vitro under different experimental conditions and exhibited high stability, low power consumption, and good linear response in the nanoampere current range. The in vivo results confirmed previously published observations on oxygen dynamics in the striatum of freely moving rats. The system serves as a rapid and reliable model for studying the effects of different drugs on brain oxygen and brain blood flow and it is suited to work with direct-reduction sensors or O(2)-consuming biosensors.

Contributions by a novel edge effect to the permselectivity of an electrosynthesized polymer for microbiosensor applications.

Pt electrodes of different sizes (2 x 10(-5)-2 x 10(-2) cm(2)) and geometries (disks and cylinders) were coated with the ultrathin non-conducting form of poly(o-phenylenediamine), PPD, using amperometric electrosynthesis. Analysis of the ascorbic acid (AA) and H(2)O(2) apparent permeabilities for these Pt/PPD sensors revealed that the PPD deposited near the electrode insulation (Teflon or glass edge) was not as effective as the bulk surface PPD for blocking AA access to the Pt substrate. This discovery impacts on the design of implantable biosensors where electrodeposited polymers, such as PPD, are commonly used as the permselective barrier to block electroactive interference by reducing agents present in the target medium. The undesirable "edge effect" was particularly marked for small disk electrodes which have a high edge density (ratio of PPD-insulation edge length to electrode area), but was essentially absent for cylinder electrodes with a length of >0.2 mm. Sample biosensors, with a configuration based on these findings (25 microm diameter Pt fiber cylinders) and designed for brain neurotransmitter L-glutamate, behaved well in vitro in terms of Glu sensitivity and AA blocking.

An integrative dynamic model of brain energy metabolism using in vivo neurochemical measurements.

An integrative, systems approach to the modelling of brain energy metabolism is presented. Mechanisms such as glutamate cycling between neurons and astrocytes and glycogen storage in astrocytes have been implemented. A unique feature of the model is its calibration using in vivo data of brain glucose and lactate from freely moving rats under various stimuli. The model has been used to perform simulated perturbation experiments that show that glycogen breakdown in astrocytes is significantly activated during sensory (tail pinch) stimulation. This mechanism provides an additional input of energy substrate during high consumption phases. By way of validation, data from the perfusion of 50 microM propranolol in the rat brain was compared with the model outputs. Propranolol affects the glucose dynamics during stimulation, and this was accurately reproduced in the model by a reduction in the glycogen breakdown in astrocytes. The model's predictive capacity was verified by using data from a sensory stimulation (restraint) that was not used for model calibration. Finally, a sensitivity analysis was conducted on the model parameters, this showed that the control of energy metabolism and transport processes are critical in the metabolic behaviour of cerebral tissue.

Nitric oxide monitoring in brain extracellular fluid: characterisation of Nafion-modified Pt electrodes in vitro and in vivo.

A Nafion(5 pre-coats/2 dip-coats)-modified Pt sensor developed for real-time neurochemical monitoring has now been characterised in vitro for the sensitive and selective detection of nitric oxide (NO). A potentiodynamic profile at bare Pt established +0.9 V (vs. SCE) to be the most appropriate applied potential for NO oxidation. The latter was confirmed using oxyhaemoglobin and N(2), both of which reduced the NO signal to baseline levels. Results indicated enhanced NO sensitivity at the Nafion(5/2) sensor (1.67 +/- 0.08 nA microM(-1)) compared to bare Pt (1.08 +/- 0.20 nA microM(-1)) and negligible interference from a wide range of endogenous electroactive interferents such as ascorbic acid, dopamine and its metabolites, NO(2)(-) and H(2)O(2). The response time of 33.7 +/- 2.7 s was found to improve (19.0 +/- 3.4 s) when the number of Nafion layers was reduced to 2/1 and an insulating outer layer of poly(o-phenylenediamine) added. When tested under physiological conditions of 37 degrees C the response time of the Nafion(5/2) sensor improved to 14.00 +/- 2.52 s. In addition, the NO response was not affected by physiological concentrations of O(2) despite the high reactivity of the two species for each other. The limit of detection (LOD) was estimated to be 5 nM while stability tests in lipid (phosphatidylethanolamine; PEA) and protein (bovine serum albumin; BSA) solutions (10%) found an initial ca. 38% drop in sensitivity in the first 24 h which remained constant thereafter. Preliminary in vivo experiments involving systemic administration of NO and L-arginine produced increases in the signals recorded at the Nafion(5/2) sensor implanted in the striatum of freely-moving rats, thus supporting reliable in vivo recording of NO.

Development of a voltammetric technique for monitoring brain dopamine metabolism: compensation for interference caused by DOPAC electrogenerated during homovanillic acid detection.

The established stability of carbon-paste electrodes (CPEs) in brain extracellular fluid was exploited to develop a voltammetric technique to monitor the dopamine metabolite, homovanillic acid (HVA), at 10 s intervals. At the scan rates needed for this time resolution, 3,4-dihydroxyphenylacetic acid (DOPAC), electrogenerated as a result of HVA oxidation, was observed in the cyclic staircase voltammograms, and this interfered with the straightforward reliable quantification of HVA. However, correction of the HVA signal, recorded in mixtures, with currents from the DOPAC and ascorbate regions of the voltammogram allowed the reproducible construction of well behaved HVA calibration plots. These showed good linearity, LOD values, selectivity and stability during six days of continuous CPE exposure to a lipid medium, which served as an in-vitro model of CPE implantation in brain tissue for future applications.

Biotelemetric monitoring of brain neurochemistry in conscious rats using microsensors and biosensors.

In this study we present the real-time monitoring of three key brain neurochemical species in conscious rats using implantable amperometric electrodes interfaced to a biotelemetric device. The new system, derived from a previous design, was coupled with carbon-based microsensors and a platinum-based biosensor for the detection of ascorbic acid (AA), O(2) and glucose in the striatum of untethered, freely-moving rats. The miniaturized device consisted of a single-supply sensor driver, a current-to-voltage converter, a microcontroller and a miniaturized data transmitter. The redox currents were digitized to digital values by means of an analog-to-digital converter integrated in a peripheral interface controller (PIC), and sent to a personal computer by means of a miniaturized AM transmitter. The electronics were calibrated and tested in vitro under different experimental conditions and exhibited high stability, low power consumption and good linear response in the nanoampere current range. The in-vivo results confirmed previously published observations on striatal AA, oxygen and glucose dynamics recorded in tethered rats. This approach, based on simple and inexpensive components, could be used as a rapid and reliable model for studying the effects of different drugs on brain neurochemical systems.