G Damsma

Scope and limitations of in vivo brain dialysis: A comparison of its application to various neurotransmitter systems

Brain dialysis is rapidly becoming a routine research method with a wide range of applications. Since 1982 this sampling technique is frequently used as a method to study the in vivo release of endogenous neurotransmitters such as dopamine, noradrenaline, serotonin, acetylcholine and certain amino acids. In this review most of the studies that have appeared in this field, are evaluated. Special attention was given to the question whether the neurotransmitter content in the dialysate is related to neurotransmission. Criteria such as the presence of a high tissue/dialysate concentration ratio, the sensitivity of the transmitters to membrane active compounds and the occurrence of receptor-mediated effects, are discussed. It is concluded that dopamine, noradrenaline and acetylcholine found in the dialysate are directly derived from neurotransmission, whereas the overflow of excitatory amino acid neurotransmitters is related to neurogenic as well as to metabolic events.

THE USE OF TETRODOTOXIN FOR THE CHARACTERIZATION OF DRUG-ENHANCED DOPAMINE RELEASE IN CONSCIOUS RATS STUDIED BY BRAIN DIALYSIS

SummaryThe effect of TTX (infused during brain dialysis of the striatum and nucleus accumbens) on the in vivo release of dopamine, 3,4-dihydroxyphenylacetic acid and homovanillic acid, was investigated. In addition it was studied whether the increase in the release of dopamine, induced by various pharmacological treatments, was sensitive to TTX infusion. The following drugs were studied: haloperidol, amphetamine, haloperidol co-administered with GBR 12909, morphine and MPP+. Dialysis was carried out in the striatum, except for morphine, which was studied in the nucleus accumbens. The infusion of TTX revealed three different types of pharmacologically enhanced dopamine release in conscious rats. First, action potential dependent dopamine release (exocytosis), which was observed in untreated animals as well as in animals treated with haloperidol, haloperidol + GBR 12909, and morphine. Second, action potential independent release (carrier-mediated) was established in the case of amphetamine. Third, action potential independent DA release, probably caused by neurotoxic reactions was observed during MPP+ infusion.

MEASUREMENT OF ACETYLCHOLINE-RELEASE IN FREELY MOVING RATS BY MEANS OF AUTOMATED INTRACEREBRAL DIALYSIS

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 μM) 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.

The significance of extracellular calcium for the release of dopamine, acetylcholine and amino acids in conscious rats, evaluated by brain microdialysis

SummaryThe influence of the calcium concentration of the perfusion fluid on the release of striatal dopamine recorded by brain dialysis was investigated. The release of dopamine appeared very sensitive to the calcium concentration of the Ringer. Next we studied whether three different methods known to antagonize the effects of calcium entry, were able to affect the release of dopamine. The conditions investigated were: the use of calcium-free Ringer, infusion of magnesium, and infusion of the calcium-antagonist verapamil. Calcium-antagonism was studied on the day of implantation of the cannula as well as on several days thereafter. It appeared that magnesium infusion was the most effective condition to antagonize the effects of calcium on the release of dopamine. Magnesium infusion was also most effective in preventing drug-evoked voltage-dependent dopamine release (induced by coadministration of haloperidol and GBR 12909). In addition magnesium infusion appeared a potent antagonist of acetylcholine release. In contrast, the dialysate content of aminoacids was not influenced by magnesium infusion.

DETERMINATION OF ACETYLCHOLINE AND CHOLINE IN THE FEMTOMOLE RANGE BY MEANS OF HPLC, A POST-COLUMN ENZYME REACTOR, AND ELECTROCHEMICAL DETECTION

SummaryThe measurement of choline and acetylcholine by means of HPLC, a post-column enzyme reactor, and electrochemical detection has been simplified and optimised. The use of a cation exchanger and enzyme reactor fitted in a cartridge holder appeared to result in reproducible, sensitive, and selective measurement of endogenous choline and acetylcholine with a lower detection limit of 50 fmole.

Dopamine re-uptake inhibitors show inconsistent effects on the in vivo release of dopamine as measured by intracerebral dialysis in the rat

The effects of the dopamine (DA) re-uptake blockers amfonelic acid and GBR 12909 on the in vivo release of DA were investigated. DA release was measured by means of a fully automated brain dialysis system. GBR 12909 induced a 3-fold increase in the output of DA; amfonelic acid was without effect. These results indicate a dissociation between the behavioural effects of DA re-uptake blockers and the effects on DA release. A strong synergistic rise of DA release was observed when haloperidol was co-administered with amfonelic acid or GBR 12909. The potentiation of DA release was completely prevented by infusion of 1 microM tetrodotoxin.

Pharmacological aspects of R-(+)-7-OH-DPAT, a putative dopamine D3 receptor ligand.

The R-(+)-isomer of 7-hydroxy-2-(N,N-di-n-propylamino)tetralin (7-OH-DPAT) bound with a more than 200-fold higher affinity to cloned human dopamine D3 receptors (Ki = 0.57 nM) than to dopamine D2 receptors; the corresponding S-(-)-enantiomer had considerably less affinity for both dopamine receptor subtypes, indicating that the known enantiomer selectivity of 7-OH-DPAT for the classical dopamine D2 receptor subtype extends to the recently discovered dopamine D3 receptor subtype. In rats R-(+)-7-OH-DPAT dose dependently (10-1000 nmol/kg) decreased dopamine release and induced yawning, while sniffing behaviour occurred at the highest dose tested (1000 nmol/kg). The possibility that the inhibition of dopamine release and the elicitation of yawning are mediated by dopamine D3 receptors is considered.

Basal acetylcholine release in freely moving rats detected by on-line trans-striatal dialysis: Pharmacological aspects

The basal release of acetylcholine (without the use of an esterase inhibitor) from brain tissue was quantified by means of transversal striatal dialysis in freely moving rats, coupled on-line to an HPLC analysis system. Basal release of acetylcholine was shown to be fully calcium dependent and tetrodotoxin sensitive. A comparison between a U-shaped and a transversally localized dialysis probe was made and some important differences were noticed. The use of a transversal probe resulted in a 20 times higher recovery when compared with the U-shaped cannula. The effect of the cholinomimetic oxotremorine and the anticholinergic atropine on the basal acetylcholine output was determined. Atropine increased the output of acetylcholine, whereas oxotremorine induced a decrease in the output of the transmitter. Application of various degrees of esterase inhibition (by addition of neostigmine to the perfusion fluid) did not qualitatively interfere with the effects of oxotremorine or atropine on the release of acetylcholine.

Effect of Ouabain Applied by Intrastriatal Microdialysis on the In Vivo Release of Dopamine, Acetylcholine, and Amino Acids in the Brain of Conscious Rats

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 Mg2+.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.

The effect of intrastriatal application of directly and indirectly acting dopamine agonists and antagonists on the in vivo release of acetylcholine measured by brain microdialysis

SummaryThe effect of intrastriatal application of D-1, D-2 and indirect dopaminergic drugs on the release of striatal acetylcholine as a function of the post-implantation intervals was studied using in vivo microdialysis. The dopamine, D-2 agonists LY 171555 and (−)N0437 inhibited the release of striatal acetylcholine to 40% of control values 16–24 h after implantation of the dialysis cannula. When LY 171555 was infused 40–48 h after implantation of the dialysis cannula, the response was attenuated to 20% of control values. Meanwhile, the effectiveness of infusions of the antagonists (−)sulpiride and haloperidol was augmented from a non significant effect at 16–24 h to a 150% increase 40–48 h after implantation of the cannula.Infusions of the dopamine releasing agent amphetamine or the dopamine uptake inhibitor nomifensine resulted in a dose-dependent increase in the overflow of dopamine. Not until a sevenfold increase in the level of dopamine was seen, the release of acetylcholine was significantly affected. This hyporesponsiveness of the striatal cholinergic interneurons to endogenous dopamine could not be attributed to dopamine D-1 receptor activation, since no effects on striatal acetylcholine release were found by intrastriatal infusions of the selective D-1 agonist CY 208-243 or the selective D-1 antagonist SCH 23390.The results indicate that dopamine D-2 receptors are involved in the regulation of striatal acetylcholine release and that these receptors are tonically occupied by endogenous dopamine under the present experimental conditions 40–48 h after probe implantation. The fact that cholinergic responses to intrastriatally applied dopaminergic agents are dynamic with respect to the time between implantation surgery of the dialysis tube and the experimental measurements suggests that the striatal neuronal system is perturbated by the implantation of a dialysis probe for a considerable length of time.