Abstract: In vivo microdialysis was employed in order to characterize the steady‐state kinetics of the turnover of specific dopamine and serotonin metabolites in the rat striaturn 48 h after surgery. Inhibitors of monoamine oxidase (MAO; pargyline) and catechol‐O‐methyltransferase (COMT; Ro 40‐7592) were administered, either separately or in conjunction, at doses sufficient to block these enzymes in the CNS. In some experiments, the acid metabolite carrier was blocked with probenecid. Temporal changes were then observed in the efflux of interstitial dopamine, 3‐methoxytyramine (3‐MT), 3,4‐dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 5‐hydroxyindoleacetic acid (5‐HIAA). The fractional rate constants for the accumulation or disappearance of the metabolites could be determined after pharmacological blockade of catabolic enzymes or the acid metabolite carrier. Interstitial 5‐HIAA was found to be cleared with a half‐life of approximately 2 h. After blockade of either MAO or COMT, HVA disappeared with a half‐life of 17 min. Experiments employing probenecid suggested that some of the interstitial HVA was cleared by the acid metabolite carrier, the remainder being cleared by a probenecid‐insensitive process, possibly conjugation. After MAO inhibition, DOPAC disappeared with an apparent half‐life of 11.3 min. The rate of 3‐MT accumulation after pargyline indicated that the majority of interstitial HVA (>95%) is formed from DOPAC rather than 3‐MT. The formation of 3‐MT from interstitial dopamine, calculated from the accumulation rate of 3‐MT after pargyline, appeared to follow first‐order kinetics (k = 0.1 min −1 ).
Abstract: In the present study we have applied a brain microdialysis technique to investigate the effects of ouabain infusion on the release of dopamine, acetylcholine, and amino acids from striatal neurons in freely moving rats. Ouabain caused an increase in the dialysate levels of dopamine; its metabolite 3,4‐dihydroxyphenylacetic acid (DOPAC); and the amino acids glutamate, aspartate, taurine, glycine, alanine, serine, asparagine, and threonine. The ouabain‐induced increase in dopamine was dose dependent and explosive (100‐ fold at an infusion concentration of 1 mmol/L) and contrasted strongly with the small effect of the glycoside on the output of DOPAC. We investigated the nature of ouabain‐induced transmitter release by determining its sensitivity to coinfusion with tetrodotoxin or the calcium antagonist Mg 2+ .In the case of dopamine two mechanisms of ouabain‐induced release could be established. At lower infusion concentrations ouabain induced an exocytotic type of release whereas at higher concentrations the release was probably camer mediated. In the case of amino acids we noticed a calcium‐independent release which was nerve impulse flow dependent in the case of glutamate and aspartate and impulse flow independent in the case of alanine, serine, glycine, threonine, and asparagine. Ouabain induced a decrease in the release of acetylcholine and glutamine.
Abstract: The present study demonstrates the feasibility of measuring acetylcholine in perfusion samples collected by means of in vivo brain dialysis in the striata of freely moving rats. The output of the dialysis device was directly connected to an automated sample valve of a HPLC‐assay system that comprises a cation exchanger, a post‐column enzyme reactor, and an electrochemical detector. The presence of an acetylcholinesterase inhibitor (neostigmine) in the perfusion fluid was required for the detection of acetylcholine in the perfusate. Increasing concentrations of neostigmine induced increasing amounts of acetylcholine. Continuous perfusion with a fixed concentration (2 μ M ) of neostigmineresultedingraduallyincreasingamounts of collected acetylcholine over time although a considerable variation between successive samples exists. The brain dialysis technique was further validated by studying the effect of various drugs. Systemically administered atropine increased the output of acetylcholine, whereas the addition of tetrodotoxin to the perfusion fluid resulted in a complete disappearance of the neurotransmitter.
Abstract In vivo brain microdialysis was used to characterize the effects of some dopamine uptake inhibitors on the extracellular concentrations of dopamine (DA) and its metabolitesdihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the striata of awake, freely moving rats. d‐Amphetamine, GBR 12909, cocaine, nomifensine, methylphenidate, bupropion, and benztropine were administered directly to the striatum via the perfusion fluid in increasing concentrations (1–1,000 μM). All drugs increased extracellular DA in a dose‐dependent manner; however, only d‐amphetamine produced dose‐dependent decreases in DOPAC and HVAconcentrations. The shapes of the dose‐reponse functions differed considerably between the drugs. At 100 and 1000 μM d‐amphetamine had biphasic effects (an increase followed by a decrease) on dialysate DA concentration. GBR 12909, methylphenidate, and benztropine also had biphasic effects when applied at the 1,000 μM concentration. In contrast, cocaine, nomifensine, and bupropion produced relatively monophasic increases in extracellular DA. Tetrodotoxin (TTX), which prevents action potentials by blocking voltage‐dependent Na + channels, did not prevent d‐amphetamine induced increases in extracellular DA, but blocked completely the effects of cocaine, nomifensine, bupropion, and methylphenidate. While low doses (10 μM) of GBR 12909 and benztropine were highly sensitive to TTX, the toxin was only partially effective against higher doses of the compounds. The rank order of potency of the drugs as determined by the increases in extracellular DA produced by 10 or 100 μM (following correction for dialysis efficiency of the test compounds in vitro) was GBR 12909< benztropine< amphetamine= nomifensine= methylphenidate< cocaine< bupropion. The in vivo characterization of changes in extracellular DA following direct, local application of DA uptake inhibitors can be used to provide useful information aboout the mechanisms of action and potency of these compounds.
