The enzyme glucose oxidase was immobilized on the surface of carbon fiber microelectrodes (CFMEs) either by cross-linking in glutaraldehyde vapor or by enzyme entrapment in electropolymerized films of m-phenylenediamine or resorcinol. The cross-linked enzymatic layer was, in the given conditions, covered with an additional membrane of Nafion or cellulose acetate. The prepared glucose sensors were tested using differential normal pulse voltammetry (DNPV, in which the scan comprises successive double pulses ("prepulse and pulse"), the prepulses are of increasing amplitude, and the current measured is the differential of the current existing between each prepulse and pulse). With properly chosen DNPV parameters, the response to glucose presented a peak at a potential of about 1 V versus an Ag/AgC1-reference, owing to the oxidation of enzymatically produced hydrogen peroxide. The calibration curves obtained (peak height/glucose concentration) were linear from 0.3-0.5 up to 1.5-6.5 mM and showed a sensitivity ranging from 1.4 up to 34.5 mA M-1 cm-2, depending on the sensor type. The DNPV response to glucose exhibited an essential insensitivity toward easily oxidizable interfering substances such as ascorbic acid and acetaminophen present at physiological concentrations. Peptides, the interfering species typical of the cerebral medium, were effectively retained by the above additional membranes. Concentration values of glucose in plasma and cerebrospinal fluid, determined in vitro from the DNPV peak height, agreed well with those measured by standard procedures. In the anesthetized rat, extracellular brain concentration of glucose was also monitored during administration of either insulin or glucagon. Under such pharmacological conditions, the changes observed in the peak height were in perfect agreement with the known effects induced by both substances.

Cortical lactate was monitored voltammetrically in freely moving rats equipped with polygraphic electrodes. Differential normal pulse voltammetric measurements were carried out using a lactate biosensor coated with lactate oxidase and cellulose acetate. Changes occurring in lactate level were in keeping with sleep-wake states. During slow wave sleep (SWS), the lactate level decreased significantly (-16.2%) vs. the spontaneous waking state (W) referenced to as 100%. During paradoxical sleep (PS), and still vs. W, it remained low (-9.0%) but this variation was not statistically significant. However, when this PS change was compared to the SWS variation, a significant increase in lactate level was then revealed (+8.5%). Finally, during the active waking (aW) triggered by a water puff stress, lactate level rose significantly in accordance with the animal activity (+53% compared to W). Long-term monitoring also allowed the determination of a circadian component in lactate production, the lowest and highest values being monitored during light and dark periods, respectively. The acrophasis of the circadian change occurred during the dark period, about 3 h after the light-off (+89%). It is suggested that during wakefulness astrocyte metabolism allows the transformation of the blood-borne glucose into lactate. The increase in this substrate observed during PS may fulfil the oxidative phosphorylation in order to supply the important ATP need of PS.

In the present study, cortical extracellular levels of glucose were monitored for the first time throughout the sleep-wake states of the freely moving rat. For this purpose, polygraphic recordings (electroencephalogram of the fronto-occipital cortices and electromyogram of the neck muscles) were achieved in combination with differential normal pulse voltammetry (DNPV) using a specific glucose sensor. Data obtained reveal that the basal extracellular glucose concentration in the conscious rat is 0.59 +/- 0.3 m M while under chloral hydrate anaesthesia (0.4 g/kg, i.p.) it increases up to 180% of its basal concentration. Regarding the sleep-wake cycle, the existence of spontaneous significant variations in the mean glucose level during slow-wave sleep (SWS = +13%) and paradoxical sleep (PS = -11%) compared with the waking state (100%) is also reported. It is to be noticed that during long periods of active waking, glucose level tends towards a decrease that becomes significant after 15 min (active waking = -32%). On the contrary, during long episodes of slow-wave sleep, it tends towards an increase which becomes significant after 12 min (SWS = +28%). It is suggested that voltammetric techniques using enzymatic biosensors are useful tools allowing direct glucose measurements in the freely moving animal. On the whole, paradoxical sleep is pointed out as a state highly dependent on the availability of energy and slow-wave sleep as a period of energy saving.