Hypothermia applied after hypoxia offers neuroprotection in neonatal animals, but the mechanisms involved remain unknown. Hypoxia was induced in newborn piglets and changes in excitatory amino acids (EAAs) and the citrulline:arginine ratio (CAR) were followed by microdialysis for 5 h. After the 45 min hypoxic insult, the animals were randomized to receive normothermia (39 degrees C; n=7) or hypothermia (35 degrees C; n = 7). After reoxygenation, extracellular glutamate, aspartate and the excitotoxic index were significantly lower in the cerebral cortex of hypothermic animals than in normothermic animals. A progressive rise of the CAR occurred during reoxygenation in the normothermic group whereas the ratio tended to decrease in the hypothermic group. In conclusion, post-hypoxic hypothermia attenuated NO production and overflow of EAAs.

We have studied the ability of the vigilance-promoting drug modafinil to counteract the ischemic lesion produced by a unilateral microinjection of endothelin-1 (ET-1) in the neostriatum of the rat using a combined morphometrical, biochemical, cardiovascular and behavioral analysis. ET-1 was injected unilaterally into the neostriatum. The ET-1-induced lesion volume, which was determined by a computer-assisted morphometrical analysis, was reduced by the 7-day modafinil treatment (10, 30, and 100 mg/kg i.p.) in a dose-related way. Modafinil also produced a dose-related counteraction of the ET-1-induced increase of perfusate lactate levels, as determined by intrastriatal microdialysis without affec ting the ET-1 induced reduction of striatal blood flow, as determined by laser-Doppler flowmetry. The ipsilateral rotational behavior induced by apomorphine in the ET-1-lesioned rats was reduced dose-dependently by modafinil treatment. Thus, morphological, neurochemical, and behavioral evidence that the putative ischemic striatal injury induced by microinjection of ET-1 in the rat neostriatum is counteracted in a dose-dependent way by modafinil treatment has been obtained. The mechanism does not appear to involve an increase in striatal blood flow. It is instead speculated that its powerful preventive action in striatal ischemic injury may be related to a reduced anaerobic metabolism.

The present study examines the possibility that lesions induced by intrastriatal injections of endothelin-1 (ET-1, 0.43 nmol/0.5 µl) are ischemic in nature due to a vasoconstriction of the cerebral microvessels. In time course and dose-response experiments with ET-1 and in comparisons with ET-3, the volume of the lesions has been determined based mainly on the disappearance of striatal nerve cells, using a computer assisted morphometrical analysis. The blood flow in the neostriatum close to the site of injection of ET-1 was determined acutely by Laser-Doppler flowmetry. The acute metabolic effects of ET-1 were also studied on striatal superfusate levels of lactate, pyruvate, dopamine and its metabolites DOPAC (3,4-dihydroxyphenylacetic acid) and homovanilic acid (HVA) using an instrastriatal microdialysis probe. Dose related striatal lesions were observed with ET-1 (0.043–0.43 nmol) with a peak lesion volume after 24–48 h and the possible existence of a penumbra area. ET-3 showed a reduced potency to produce striatal lesions compared to ET-1. The lesions induced by ET-1 were prevented by coinjection with dihydralazine, a vasodilator drug. Acutely ET-1 (0.43 nmol/0.5 µl) produced a prolonged reduction of the cerebral blood flow down to 40% of control values and temporary increases of striatal lactate and DA efflux, the latter change being very marked. Also a significant reduction of DOPAC and HVA was observed. These neurochemical changes were all prevented by treatment with dihydralazine. These results indicate that ET-1 injected in the neostriatum may produce lesions by causing local ischemia, related to its vasoconstrictor activity and possibly also to an activation of ET-1 receptors in the astroglial-endothelial complex. Based on the present results it seems possible that ET-1 may participate in the multifactorial pathogenesis of cerebral ischemia.

Recently a histological study has demonstrated that intrastriatally injected endothelin-1 (ET-1) produced ischemia-like lesions in the neostriatum. The present study was undertaken to investigate whether intrastriatally injected ET-1 produces ischemic responses such as a decrease in the striatal blood flow and increases in extracellular lactate and dopamine levels in the neostriatum as seen in other models of ischemia. A small needle (for injection of ET-1), a microdialysis probe (for collecting extracellular substances), and a probe of a laser Doppler flowmeter (for measuring local cerebral blood flow) were implanted with their tips close to each other in the neostriatum of halothane-anesthetized rats. Focal administration of ET-1 (430 pmol) into the neostriatum resulted in a marked decrease in striatal blood flow without any change in systemic blood pressure. It also markedly increased extracellular lactate and dopamine levels, whereas it decreased pyruvate and dopamine metabolite levels. These changes agreed well with those known to be produced by ischemia. Intracerebral injection of ET-1 will therefore provide a new model for production of local ischemia in experimental animals.

Influence of striatal blood flow changes on recovery of extracellular lactate in vivo through a microdialysis probe implanted into the neostriatum was investigated in halothane anesthetized rats. Relative radioactivity loss of [14C]lactate from perfusate medium through a microdialysis probe was continuously measured in vivo as an indicator of relative recovery of extracellular lactate through the probe because both the relative recovery of lactate and the relative hot loss of [14C]lactate through a microdialysis probe were similar to each other in vitro. The relative hot loss of [14C]lactate decreased in parallel with decreases in the striatal blood flow, while it did not significantly change in response to increases in the blood flow up to 200% of control. These results demonstrate that recovery of extracellular lactate in vivo through the microdialysis probe is not directly influenced by changes in the cerebral blood flow.

Endothelin-1 and its receptors are widely distributed in the brain of rodents and humans. In view of its potent and long-lasting vasoconstrictor activity, a role of endothelin-1 has been proposed in brain ischemia. In the present paper, the local injection of endothelin-1 was utilized to induce ischemia in rat striatum. An evaluation of the rostrocaudal extension of the lesion is reported. By using intracerebral microdialysis, a marked increase of lactate and dopamine, but not glutamate, was observed in this region upon endothelin-1 administration. Moreover, preliminary data reported show a protective effect of ganglioside treatment on endothelin-1 lesion of rat striatum. The characteristics of the present model of brain ischemia are discussed in comparison with well characterized models, such as the Pulsinelli's four vessel occlusion and the middle cerebral artery occlusion.

The effect of general anaesthesia on extracellular levels of acetylcholine (ACh) in the caudate-putamen of freely moving rats was studied by microdialysis. ACh concentrations were determined in the same perfusate samples by radioenyzymatic and HPLC/electrochemical procedures in order to compare the assays. The concentration of ACh in perfusate samples was estimated to be 0.30 μM in conscious unrestrained rats. However, when these rats were administered chloral hydrate (400 mg/kg i.p.), the level of ACh was decreased immediately by 50%, attaining a value of 0.06 μM within 20–40 min following the injection. Upon recovery of the righting reflex, ACh levels were once again re-elevated. The levels of choline (Ch), the precursor of ACh, were unaffected by anaesthesia. It was apparent that the level of consciousness (i.e. awake vs. anaesthetized) is an important factor determining ACh overflow. Radioenzymatic and HPLC assays proved to give identical results for the analysis of ACh and Ch.

Cerebral blood flow (CBF) and extracellular lactate in the parietal cortex were simultaneously measured in halothane-anesthetized rats. Focal electrical stimulation of the magnocellular nucleus of the basal forebrain (nucleus basalis of Meynert; NBM) for 10 min produced a significant increase in cortical CBF without any significant changes in extracellular lactate in the parietal cortex or in systemic arterial blood pressure (BP). Cutaneous pinching of a hindpaw for 10 min increased cortical CBF and BP, but did not influence extracellular cortical lactate. Systemic hypoxia for 10 min reducing the end-tidal O2 concentration from 18% to 6–8% produced a remarkable increase in extracellular cortical lactate. It was suggested that the increased cortical CBF following either NBM stimulation or pinching of a hindpaw was not due to metabolic changes in the cortical neurons of anesthetized rats.

An isocratic high-performance liquid chromatographic (HPLC) method was developed for the simultaneous determination of lactate, pyruvate, and ascorbate in tissue and blood, sampled in vivo using the microdialysis technique. The compounds were separated on a silica-based, reversed-phase column or on a resin-based, cation exchange column. The acids were monitored by ultraviolet detection.

Extracellular (EC) ascorbate concentrations were measured in microdialysates from the striatum bilaterally in rats subjected to unilateral middle cerebral artery occlusion (MCAO). The focal cerebral ischemia induced a dramatic increase in ascorbate on the ipsilateral (operated) side while the levels remained at the preocclusion level in the striatum of the contralateral (control) hemisphere. The possibility that ascorbate may aggravate ischemic neuronal damage by its proposed neuromodulatory properties and/or by its ability to induce lipid peroxidation is discussed.

Microdialysis is a new technique to monitor levels of chemical compounds in the extracellular space over time. It involves the implantation of a microdialysis probe into the brain tissue. The probe is similar to a push-pull probe but the perfusate is contained inside a semi-permeable membrane located at the tip of the probe. Substances in the extracellular fluid will diffuse into the perfusate while substances included in the perfusate will diffuse into the tissue. This principle opens up a wealth of possibilities to monitor chemical events within the brain and to study the working mechanism of various drugs. The perfusate may be analysed by a number of different techniques. In this paper we give a short summary of various HPLC techniques that have proven particularly useful.