Hans Rollema

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.

IDENTIFICATION OF A POTENTIALLY NEUROTOXIC PYRIDINIUM METABOLITE OF HALOPERIDOL IN RATS

In vivo metabolic studies have revealed that haloperidol is converted to the corresponding pyridinium metabolite which has been characterized in both urine and brain tissues isolated from haloperidol treated rats. Unlike the corresponding conversion of the structurally related Parkinsonian inducing agent MPTP to the ultimate neurotoxic pyridinium metabolite MPP+, the oxidative biotransformation of haloperidol is not catalyzed by MAO-B. Microdialysis studies in the rat indicate that intrastriatal administration of this pyridinium metabolite is about 10% as effective as MPP+ in causing the irreversible depletion of striatal nerve terminal dopamine. The results point to the possibility that some of the neurological disorders observed in experimental animals and man during the course of chronic haloperidol treatment may be mediated by this pyridinium metabolite.

Partial agonists of the α3β4* neuronal nicotinic acetylcholine receptor reduce ethanol consumption and seeking in rats.

Alcohol use disorders (AUDs) impact millions of individuals and there remain few effective treatment strategies. Despite evidence that neuronal nicotinic acetylcholine receptors (nAChRs) have a role in AUDs, it has not been established which subtypes of the nAChR are involved. Recent human genetic association studies have implicated the gene cluster CHRNA3–CHRNA5–CHRNB4 encoding the α3, α5, and β4 subunits of the nAChR in susceptibility to develop nicotine and alcohol dependence; however, their role in ethanol-mediated behaviors is unknown due to the lack of suitable and selective research tools. To determine the role of the α3, and β4 subunits of the nAChR in ethanol self-administration, we developed and characterized high-affinity partial agonists at α3β4 nAChRs, CP-601932, and PF-4575180. Both CP-601932 and PF-4575180 selectively decrease ethanol but not sucrose consumption and operant self-administration following long-term exposure. We show that the functional potencies of CP-601932 and PF-4575180 at α3β4 nAChRs correlate with their unbound rat brain concentrations, suggesting that the effects on ethanol self-administration are mediated via interaction with α3β4 nAChRs. Also varenicline, an approved smoking cessation aid previously shown to decrease ethanol consumption and seeking in rats and mice, reduces ethanol intake at unbound brain concentrations that allow functional interactions with α3β4 nAChRs. Furthermore, the selective α4β2* nAChR antagonist, DHβE, did not reduce ethanol intake. Together, these data provide further support for the human genetic association studies, implicating CHRNA3 and CHRNB4 genes in ethanol-mediated behaviors. CP-601932 has been shown to be safe in humans and may represent a potential novel treatment for AUDs.

A comparison of the potencies of various dopamine receptor agonists in models for pre- and postsynaptic receptor activity.

SummarySeveral dopamine (DA) receptor agonists, notably N,N-dipropyl-2-aminotetralin analogues differing in the number and position of phenolic hydroxyl groups, were evaluated in model systems for pre- and postsynaptic dopaminergic activity. Apomorphine, piribedil and pergolide were included for comparison. All compounds inhibited the γ-butyrolactone (GBL)-induced increase in DA concentrations in the rat striatum and olfactory tubercle, although a dosedependency could not be demonstrated for one of the compounds, i.e. N,N-dipropyl-2-amino-5,6-dihydroxytetralin. In addition to the reversal of the DA-increase all compounds decreased the HVA and DOPAC levels in a dose-dependent manner, in much the same way as in normal, non GBL-pretreated rats.The potencies of the drugs to decrease HVA in normal rats and to inhibit the DA-increase and to decrease HVA in GBL-pretreated rats, both in the striatum and the olfactory tubercle were compared with each other and with the potencies to induce stereotyped behaviour. It may be concluded that (1) N,N-dipropyl-2-amino-7-hydroxytetralin shows the largest difference in activity in the biochemical and the behavioural models, suggesting a selective presynaptic activity. This was corroborated by the appearance of a marked hypomotility after low doses of this compound; (2) The potencies to decrease striatal HVA concentrations are generally somewhat different from the potencies to inhibit GBL-induced DA-increases, but appear to be comparable to the potencies to inhibit GBL-induced dihydroxyphenylalanine (DOPA)-increases; (3) There is no indication that the DA agonists in general are more potent at presynaptic receptors in the tubercle than in the striatum.

An M3-like muscarinic autoreceptor regulates the in vivo release of acetylcholine in rat striatum

Selective muscarinic antagonists were used in an attempt to characterize the muscarinic autoreceptor modulating the release of acetylcholine in the striatum of the rat. In vivo microdialysis was applied to infuse atropine, 4-DAMP (4-diphenylacetoxy-N-methylpiperidine), pirenzepine or AF-DX 116 (11-[[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5, 11-dihydro[2,3-b][1,4]benzodiazepine-6-one), leading to a dose-dependent increase in the overflow of acetylcholine, the order of potency being: atropine greater than 4-DAMP greater than pirenzepine greater than AF-DX 116. We conclude from these data that the muscarinic receptor modulating release in the striatum is of the M3 type.

The Multiplicity of the D1 Dopamine Receptor

Dopaminergic neurotransmission is known to modulate a variety of behaviors, including ambulation (Ungerstedt and Arbuthnott, 1970; Pijnenburg et al., 1976), stereotyped behaviors (Creese and Iversen, 1973), self-stimulation (Phillips and Fibiger, 1973), conditioned avoidance responding (Seiden and Carlsson, 1963), stimulus control (Ho and Huang, 1975), and feeding and drinking (Ungerstedt, 1971; Fitzsimons and Setler, 1975). It is not surprising, therefore, that drugs which are believed to act primarily as dopamine receptor agonists or antagonists have important clinical utility. Our work has sought to address two questions of some neuropharmacological importance. First, what is the nature of mechanisms by which dopamine initiates many of these psychopharmacological effects, and second, is it possible to design highly specific drugs targeted only at a selected subpopulation of dopamine receptors?

Effect of non-catecholic 2-aminotetralin derivatives on dopamine metabolism in the rat striatum

SummaryThe concentrations of dopamine (DA), dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were measured in the striatum of rats after i.p. injection of dipropyl-2-aminotetralin and the four positional isomers of monohydroxy-dipropyl-2-aminotetralin. All compounds except 8-OH dipropylaminotetralin caused a decrease in DOPAC-and HVA-concentrations. In addition, 5-OH-dipropylaminotetralin produced a small elevation in DA concentrations. In contrast, 7-OH dipropylaminotetralin, in doses of 100 μmol/kg and more, decreased DA to 50% and initially increased HVA and DOPAC to about 200%, after which the concentrations of the metabolites fell to 30% or less. The 5-OH isomer was found to be the most potent compound, decreasing HVA concentrations to 70% at a dose of 0.14 μmol/kg. The potencies are compared to those of catechol-group containing DA-agonists such as apomorphine and N,N-dipropyl-5,6-dihydroxy-2-aminotetralin. In addition, a comparison is made with reported behavioural data. It is suggested that the more active N-alkylated 2-aminotetralins have a conformation which corresponds to that of the α rotamer of dopamine.

Effect of dihydroxy-2-aminotetralin derivatives on dopamine metabolism in the rat striatum

SummaryConcentrations of dopamine (DA), dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were measured in the striatum of rats after i.p. injection of apomorphine, N,N-dipropyldopamine and a series of alkylated and/or esterified dopamine analogues of the dihydroxyaminotetralin type.All compounds tested caused a decrease in DOPAC- and HVA-concentrations. The N-alkylated derivatives had a rapid onset of action, showing a maximal HVA decrease after 15–45 min, after which time the metabolite concentrations slowly returned to control values. In addition, the dihydroxyaminotetralins, especially N,N-dipropylamino-5,6-dihydroxytetrahydronaphthalene (DiPr-5,6-ADTN), produced a rapid, short lasting elevation of DA concentrations. The esterified primary amines, dibenzoyl-5,6-and dibenzoyl-6,7-dihydroxyaminotetralin, had a delayed effect, causing a maximal HVA decrease after 4–6 h.DiPr-5,6-ADTN was found to be the most potent compound, with a maximal effect at a dose of 0.33 μmol/kg, it being 30 times more potent than apomorphine and DiPr-6,7-ADTN. The results corroborate reported behavioural data, and the relative potencies of the alkylated derivatives in this test system for dopaminergic activity are in agreement with those based on stereotyped behaviour.

Dopaminergic prodrugs: Brain concentrations and neurochemical effects of 5,6- and 6,7-ADTN after administration as dibenzoyl esters

Abstract Regional levels of the dopamine agonists 5,6- and 6,7-ADTN in rat brain were studied after administration of the parent compound and/or the dibenzoyl-ester derivative (a prodrug). A method for measuring 5,6- and 6,7-ADTN in the picomole range is described. The method is based on high performance liquid chromatography in conjunction with an electrochemical detector. ADTN levels in brain after administration of the dibenzoyl prodrug were about 5 times higher than when non-esterified ADTN was applied. The concentrations of the 5,6-isomer were 5–7 times higher than those of the 6,7-isomer. Constant brain levels of 5,6-ADTN (for at least 10 h) indicated that this isomer leaves the brain only with great difficulty. The susceptibility to metabolic degradation appears to play a crucial role in the in vivo activity of ADTN derivatives. An accumulation of 5,6-ADTN in DA-rich areas of the brain was observed after administration of the dibenzoyl derivative. By comparing the striatal levels of both ADTN isomers (after administration as dibenzoyl derivatives) with the data on their potency to decrease homovanillic acid concentrations in this brain area, we were able to quantitatively determine the in vivo activity of these isomers. 6,7-ADTN appeared to be about 12–14 times more potent than the 5,6-derivative, at the receptor site. These results strongly support the hypothesis that the active conformation of dopamine is closer to the β-rotamer than to the α-rotamer.