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All mGlu receptor antagonists were put into the perfusate at a concentration of 100 m to yield 10 m in the tissue. Instruments, Fullerton, CA), a programmable solvent module 126 (Beckman), an KSHV ORF26 antibody analytical C18 reverse-phase column kept at 30C ]Ultrasphere ODS 5 m, 80 ? pore, 250 4.6 mm (Beckman)], and a Coulochem II electrochemical detector (ESA, Inc., Chelmsford, MA). The holding potentials were set at +350 and ?350 mV for the detection of DA, 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA). The mobile phase consisted of 80 mm sodium phosphate, 40 mm citric acid, 0.4 mm EDTA, 3 mm1-heptansulfonic acid, and 12.5% methanol, brought to pH 2.75 with phosphoric acid (run under isocratic conditions, at 1 ml/min). access to water and food, and allowed to recover for 4 d before the experiment. On the evening before the experiment, a probe was inserted into the intracerebral guide, after a dummy was removed, and mice were transferred to a plastic bowl cage with a moving arm (CMA/120 System for Freely-Moving Animals,CMA/Microdialysis), with access to water and food. Concentric vertical microdialysis probes 2 mm long and 0.24 mm in outer diameter having a cuprophane membrane with a molecular cutoff of 6000 Da (CMA/7 Microdialysis Probe, CMA/Microdialysis) were used. The probes were perfused continuously with artificial CSF at a flow rate of 1 1.5 l/min, using a microinjection pump (Bioanalytical System, West Lafayette, IN). The ACSF contained (in mm): 150 NaCl, 3 KCl, 1.7 CaCl2, and 0.9 MgCl2. This solution was not buffered, and the pH was typically 6.5. On the following morning, 30 l (20 min) of consecutive perfusate sample fractions were continuously collected by a fraction collector (CMA/142 Microfraction Collector,CMA/Microdialysis). After four sample fractions, which were used to determine the basal levels of monoamines, control mice received a single injection of MA (5 mg/kg, i.p.), and sample fractions were collected for the following 2 hr. Animals injected systemically Chlorobutanol with MPEP (20 min before MA) or locally perfused with MPEP (100 m), SIB-1757 (100 m), or CPCCOEt (100 m) received a single injection of MA (5 mg/kg, i.p.), and sample fractions were collected for the following 2 hr. In another set of animals, formation of Chlorobutanol reactive oxygen species was examined by monitoring 2,3-DHBA, a product of the reaction of salicylate (5 mm, added to artificial CSF with hydroxyl radicals. Analysis of DA and 2,3-DHBA was performed by HPLC with electrochemical detection, as described above. For the preparation of striatal synaptosomes, the brains were quickly removed and the striatum was dissected at 4C. Synaptosomal fractions were prepared as described by Gray and Whittaker (1962). Synaptosomal pellets were suspended in 0.32 mglucose (8 mg of protein/ml). The suspension was diluted 1:10 with Krebs’CRinger’s medium (130 mm NaCl, 3 mmMgSO4, 2.5 mmNa2HPO4, 1 mmascorbic acid, and 20 mm Tris buffer, pH 7.5) and preincubated for 10 min at 37C. Synaptosomes were then labeled with 0.1 mdihydroxy-phenyl-ethylamine,3,4-[7-3H] ([3H]DA) (specific activity 36 Ci/mmol; NEN-DuPont) for 10 min at 37C. After a 1:10 dilution with Krebs’CRinger’s medium, pH 7.5, containing 10 mmglucose, aliquots of Chlorobutanol the suspension were placed on Millipore filters lying at the bottom of parallel superfusion chambers thermostatically maintained at 37C (Cerrito et al., 1993). The following drugs were added to the perfusate: MA (0.1 m),d-amphetamine (0.1 m), MPEP (5 m), SIB-1757 (5 m), or SIB-1893 (5 m). The amount of [3H]DA present in each fraction and that extracted from synaptosomes at the end of the perfusion were separated from labeled metabolites on Biorex-70 columns (Smith et al., 1975), and the radioactivity present as [3H]DA was counted by scintillation spectrometry. [3H]DA release was expressed as percentage of the total [3H]DA. RESULTS mGlu5 receptor antagonists protect striatal dopaminergic terminals against MA?toxicity MA treatment (5 mg/kg, i.p.; injected three times at 2 hr intervals) led to 80% reduction in striatal DA levels, 5 d after the last injection (Fig.?(Fig.11 .