Modulation by dopamine receptors of the histamine release in the rat hypothalamus

Summary The involvement of dopaminergic neurons of the hypothalamus in the modulation of histamine release was studied by the push-pull technique. The posterior hypothalamus of the conscious, freely moving rat was superfused with artificial cerebrospinal fluid (CSF) and the release of histamine was determined radioenzymatically in the superfusate. Agonists and antagonists of dopamine D₁-, D₂- and D₃-receptors were dissolved in CSF and applied to the hypothalamus through the push-pull cannula.Hypothalamic superfusion with the D₁-, D₂- and D₃-receptor agonists dopamine or R(–)-apomorphine enhanced the release rate of histamine. (±) Apomorphine also enhanced the release of histamine, but to a lesser extent than did equimolar concentration of R(–)apomorphine. The D₃-agonist quinpirole inhibited the release of histamine, while the D₁-receptor agonist SKF 82958 [(±)-6-chloro-7,8-dihydroxy-3-allyl-l-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine] did not virtually influence the release of the neurotransmitter. On the other hand, [–]-sulpiride which predominantly blocks D₂-receptors, decreased histamine release. Hypothalamic superfusion with SKF 83566 [(±)-7-bromo-8-hydroxy-3-methyl-l-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine], which seems to be a selective antagonist of D₁-receptors, enhanced the release rate of histamine.These findings suggest that dopaminergic neurons of the hypothalamus influence the release of histamine from its neurons in a dual way. D₂-heteroreceptors stimulate the release of histamine, while D₃-heteroreceptors seem to inhibit the release of this neurotransmitter. Both types of dopamine receptors might be located presynaptically on histaminergic neurons. Alternatively, D₃- and D₂-receptors might be located on histaminergic and non-histaminergic neurons, respectively.This work was supported by the Fonds zur Förderung der wissenschaftlichen Forschung Correspondence to H. Prast at the above address     

D2 dopamine receptors and modulation of spontaneous acetylcholine (ACh) release from rat striatal synaptosomes

1. The effect of two D_3/2 dopamine receptor agonists, LY-171555 (quinpirole) and 7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT) on spontaneous [³H]-acetylcholine ([³H]-ACh) release were investigated in rat striatal synaptosomes.
2. Quinpirole and 7-OH-DPAT inhibited in a concentration-dependent manner the basal efflux of [³H]-ACh with similar E_max (maximal inhibitory effect) values (29.95±2.91% and 33.19±1.21%, respectively). Significant differences were obtained between the pEC₅₀ (−log of molar concentration) of quinpirole (7.87±0.12) and 7-OH-DPAT (7.21±0.17; P<0.01).
3. Different concentrations (0.3–10 nM) of haloperidol (D_2/3 dopamine receptor antagonist) shifted to the right the concentration-response curves elicited by quinpirole and 7-OH-DPAT, without modifications in the E_max.
4. Slopes of a Schild plot obtained with haloperidol in the presence of quinpirole and 7-OH-DPAT were not signficantly different from unity (0.85±0.05 and 1.17±0.11, respectively) and consequently haloperidol interacted with a homogeneous receptor population. The pK_B values of haloperidol obtained from Schild regression were 9.96±0.15 (in presence of quinpirole) and 9.90±0.09 (in presence of 7-OH-DPAT).
5. Specific binding of [³H]-YM-09151-2 to membranes of striatal synaptosomes and cells expressing D₂ and D₃ dopamine receptors was inhibited by haloperidol. Analysis of competition curves revealed the existence of a single population of receptors. There were no differences between the estimated pK_i (−log of molar concentration) values for synaptosomes (8.96±0.02) and cells expressing D₂ receptors (8.81±0.05), but the pK_i value from cells expressing D₃ dopamine receptors differed significantly (8.48±0.06; P<0.01).
6. In conclusion, the data obtained in the present study indicate that quinpirole and 7-OH-DPAT, two D_3/2 dopamine receptor agonists, inhibit the spontaneous [³H]-ACh efflux and this effect is competitively antagonized by haloperidol and probably mediated through dopamine D₂ receptors.

     

Effects of morphine on release of acetylcholine in the rat striatum: an in vivo microdialysis study

Summary We examined the effect of morphine on the release of acetylcholine (ACh) in the striatum of freely moving rats using the in vivo microdialysis method. The basal level of ACh was 3.01 ± 0.51 pmol/30 l/15 min in the presence of neostigmine (10 M). Tetrodotoxin (1 M), a selective blocker of voltage-dependent Na⁺ channels, rapidly decreased the release of ACh in the striatal perfusates. Morphine at a dose of 10 mg/kg (i.p.) caused a reduction of ACh release in the striatum at 90–150 min. However, a lower dose of morphine (5 mg/kg, i.p.) did not affect ACh release in the striatum. The reduction following intraperitoneal administration of morphine was abolished by naloxone (1.0 mg/kg).After microinjection of the neurotoxin 6-hydroxydopamine (6 g/3 l, 7 days before) in the substantia nigra, the morphine (10 mg/kg)-induced decrease of ACh was attenuated, and a similar result occurred following reserpine (2 mg/kg, i.p.) 24 h before combined with -methyl-p-tyrosine (300 mg/kg, i. p.) 2.5 h before.These findings indicate that morphine exerts an inhibitory influence on striatal ACh release in freely moving rats and that this inhibitory effect is mediated by the nigro-striatal dopaminergic system.Correspondence to K. Taguchi at the above address     

Functional crosstalk and heteromerization of serotonin 5-HT2A and dopamine D2 receptors

The serotonin 5-HT_2A receptor (5-HT_2AR) and dopamine D₂ receptor (D₂R) are high-affinity G protein-coupled receptor targets for two different classes of antipsychotic drugs used to treat schizophrenia. Interestingly, the antipsychotic effects are not based on the regulation of same signaling mediators since activation of the 5-HT_2AR and of the D₂R regulate G_q/11 protein and G_i/o protein, respectively. Here we use radioligand binding and second messenger production assays to provide evidence for a functional crosstalk between 5-HT_2AR and D₂R in brain and in HEK293 cells. D₂R activation increases the hallucinogenic agonist affinity for 5-HT_2AR and decreases the 5-HT_2AR induced inositol phosphate production. In vivo, 5-HT_2AR expression is necessary for the full effects of D₂R antagonist on MK-801-induced locomotor activity. Co-immunoprecipitation studies show that the two receptors can physically interact in HEK293 cells and raise the possibility that a receptor heterocomplex mediates the crosstalk observed. The existence of this 5-HT_2AR-D₂R heteromer and crosstalk may have implications for diseases involving alterations of serotonin and dopamine systems and for the development of new classes of therapeutic drugs.     

Activation of A1 adenosine receptors decreases the release of serotonin in the rabbit hippocampus,but not in the caudate nucleus

Summary The effects of A1 adenosine receptor ligands on the evoked release of serotonin (5-HT) were studied in slices of the hippocampus and the caudate nucleus of the rabbit, preincubated with ³H-5-HT. In hippocampal tissue electrical stimulation elicited a release which was inhibited by the adenosine receptor agonist N⁶-cyclohexyladenosine (CHA) and enhanced by the selective A₁ receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). The concentration-response curve of CHA was shifted to the right by DPCPX. The shift corresponded to a pA₂ value of 9.4 for DPCPX. CHA, R-N⁶-phenylisopropyladenosine (R-PIA) and DPCPX were ineffective in caudate nucleus tissue. When instead of electrical pulses high K⁺ was used to induce 5-HT release in the presence of the Na⁺ channel blocker tetrodotoxin (TTX), which was present in order to exclude effects mediated by interneurones, CHA was equally effective in the hippocampus but again failed to modify 5-HT release in the caudate nucleus. The disinhibition by DPCPX of the evoked 5-HT release was used to calculate the extracellular concentration of endogenous adenosine at the A₁ receptor. The calculation greatly depended on the dissociation constant of adenosine at the A1 receptor.It is concluded that A1 adenosine receptors, activated by the endogenous agonist at a concentration of about 0.7 mol/l, are located on serotonergic nerve endings in the hippocampus, but not in the caudate nucleus. The estimated extracellular concentration of endogenous adenosine is in reasonable agreement with actually measured concentrations reported in the literature. Send offprint requests to T. J. Feuerstein at the above address     

Discriminative stimulus properties of cocaine: modulation by dopamine D1 receptors in the nucleus accumbens

Dopamine (DA) D₁ and D₂ receptors are involved in mediating the behavioral effects of cocaine, including its discriminative stimulus properties. The purpose of the present study was to investigate the role of the nucleus accumbens and, in particular, accum bens DA D₁ receptors in modulating the stimulus effects of cocaine. Thus, rats were trained to discriminate cocaine (10 mg/kg, IP) from saline using a two-lever, water-reinforced FR 20 drug discrimination task. In substitution tests, systemic (IP) administration of cocaine (0.625–20 mg/kg) produced a dose-related increase in cocaine-appropriate responding. Microinjections of cocaine (2.5–40 µg) into the nucleus accumbens also engendered dose-dependent and complete substitutions (> 80% drug-lever responding) for the systemic training dose of cocaine, whereas intra-accumbens artificial cerebrospinal fluid (1 µl/side) produced primarily saline-appropriate responding. In antagonism tests, pretreatment with the DA D₁ antagonist SCH 23390 (3–12 µg/kg) completely antagonized (<20% drug-lever responding) a dose of cocaine (5 mg/kg) that produced greater than 90% cocaine-lever responding when given alone. Additionally, intra-accumbens injections of SCH 23390 (0.025–0.4 µg) prior to systemic cocaine (5 mg/kg) also significantly blocked the cocaine stimulus. The present results confirm the importance of the nucleus accumbens in mediating the discriminative stimulus properties of cocaine and suggest a primary role of accumbens DA D₁ receptors in modulating this behavior.Some of these data were presented at the annual FASEB Experimental Biology meeting in New Orleans (1993)     

In vivo occupancy of dopamine D2 receptors by antipsychotic drugs and novel compounds in the mouse striatum and olfactory tubercles

Interaction with dopamine D₂-like receptors plays a major role in the therapeutic effects of antipsychotic drugs. We examined in vivo dopamine D₂ receptor occupancy of various established and potential antipsychotics in mouse striatum and olfactory tubercles 1 h after administration of the compound, using [³H]nemonapride as a ligand. All the compounds reduced in vivo binding of [³H]nemonapride in the striatum. When administered systemically, conventional antipsychotics, D₂ antagonists, nemonapride (ID₅₀: 0.034 mg/kg), eticlopride (0.047), haloperidol (0.11) and raclopride (0.11) potently inhibited [³H]nemonapride binding. The ‘atypical’ antipsychotics, risperidone (0.18), ziprasidone (0.38), aripiprazole (1.6), olanzapine (0.99), and clozapine (11.1) were less potent for occupying D₂-like receptors. New compounds, displaying marked agonism at 5-HT_1A receptors in addition to D₂ receptor affinity, exhibited varying D₂ receptor occupancy: bifeprunox (0.25), SLV313 (0.78), SSR181507 (1.6) and sarizotan (6.7). ID₅₀ values for inhibition of [³H]nemonapride binding in the striatum correlated with those in the olfactory tubercles (r=0.95, P<0.0001). These values also correlated with previously-reported in vitro affinity of the compounds at rat D₂ receptors (r=0.85, P=0.0001) and with inhibition of apomorphine-induced climbing in mice (r=0.79 P=0.0005). In contrast, there was no significant correlation between ID₅₀ values herein and previously-reported ED₅₀ values for catalepsy in mice. These data indicate that: (1) there is no difference in D₂ receptor occupancy in limbic versus striatal regions between most classical and atypical or potential antipsychotics; and (2) high occupancy of D₂ receptors can be dissociated from catalepsy, if the drugs also activate 5-HT_1A receptors. Taken together, these data support the strategy of simultaneously targeting D₂ receptor blockade and 5-HT_1A receptor activation for new antipsychotics.     

Norepinephrine release in amygdala of rats during chronic nicotine self-administration: an in vivo microdialysis study

The essential role of the amygdala in learning and memory, including cue-associated learning, is influenced by local release of norepinephrine (NE). The current study investigated changes in amygdaloid NE secretion in rats learning to self-administer nicotine in an unlimited access model (23 h/day). In vivo microdialysis of NE was performed for 9 h intervals during three phases of nicotine self-administration: acquisition (day 1); early maintenance, when self-administration rates first stabilized (day 8.4+/-0.7); and later, during fully stable maintenance (day 17.6+/-1.0). On day 1, a greater number of self-administration episodes (SAEs) were associated with elevated NE levels in rats bar-pressing for nicotine (88% vs. 39% with saline). By early maintenance, such episodes increased threefold and overall NE levels were greater. During later maintenance, however, bar-pressing behavior was similar and NE was elevated by the first SAE of the day, but total daily NE levels were no longer elevated. In all the three phases, the enhanced NE release during the first daily SAE did not occur in the last SAE 9 h later. Thus, in an animal model of unlimited nicotine self-administration that approximates the human pattern of nicotine consumption via smoking, the amygdaloid NE response to nicotine diminishes over each day and with the stabilization of self-administration. The decline of amygdaloid NE secretion after long-term nicotine self-administration likely reflects desensitization to the pharmacological effects of nicotine. In addition, amygdaloid NE release, which enhances the consolidation of amygdala-dependent memory, may no longer be necessary once self-administration behavior has been established.     

Activation of dopamine D1 receptors does not affect D2 receptor-mediated inhibition of acetylcholine release in rabbit striatum

Summary The possible involvement of dopamine D₁ receptors in the regulation of acetylcholine release in the rabbit caudate nucleus was investigated. Caudate slices, preincubated with [³H]choline, were superfused continuously and subjected to electrical field stimulation with only a single pulse. In agreement with the view that the release of acetylcholine evoked by a single electrical pulse is not influenced by endogenous transmitters, atropine and domperidone failed to icnrease the evoked release of [³H]acetylcholine, whereas oxotremorine and quinpirole caused a concentration-dependent inhibition of transmitter release. Neither the dopamine D₁ receptor antagonist SCH 23390 nor the Dt agonist SKF 38393 in a concentration range of 0.01–1 mol/l changed the evoked [³H]acetylcholine release. The inhibitory effect of the dopamine D₂ receptor agonist quinpirole was virtually abolished in the presence of 0.1 mol/l domperidone and diminished in the presence of 1 mol/l SCH 23390. It remained unchanged in the presence of 1 mol/l SKF 38393. It is concluded that the inhibition of acetylcholine release by dopamine is mediated exclusively via presynaptic dopamine D₂ receptors and that the antagonistic effect of SCH 23390 on the inhibition of acetylcholine release by quinpirole is due to its interaction with dopamine D₂ rather than D₁ receptors located on cholinergic nerve terminals. Send offprint requests to C. Allgaier at the above address     

Modulation of dopamine release from rat striatum by protein kinase C: interaction with presynaptic D2-dopamine-autoreceptors

1. Interactions between dopamine receptors and protein kinase C (PKC) have been proposed from biochemical studies. The aim of the present study was to investigate the hypothesis that there is an interaction between protein kinase C and inhibitory D₂-dopamine receptors in the modulation of stimulation-induced (S-I) dopamine release from rat striatal slices incubated with [³H]-dopamine. Dopamine release can be modulated by protein kinase C and inhibitory presynaptic D₂ receptors since phorbol dibutyrate (PDB) and (−)-sulpiride, respectively, elevated S-I dopamine release.
2. The protein kinase C inhibitors polymyxin B (21 μM) and chelerythrine (3 μM) had no effect on stimulation-induced (S-I) dopamine release. However, when presynaptic dopamine D₂ receptors were blocked by sulpiride (1 μM), an inhibitory effect of both PKC inhibitors on S-I dopamine release was revealed. Thus, sulpiride unmasks an endogenous PKC effect on dopamine release which suggests that presynaptic D₂ receptors normally suppress endogenous PKC activity. This is supported by results in striatal slices which were pretreated with PDB to down-regulate PKC. In this case the facilitatory effect of sulpiride was completely abolished.
3. The inhibitory effect of the dopamine D₂/D₃ agonist quinpirole on S-I dopamine release was partially attenuated by PKC down-regulation. Since the effect of sulpiride was completely abolished under the same conditions, this suggests that exogenous agonists may target a PKC-dependent as well as a PKC-independent pathway. The inhibitory effect of apomorphine was not affected by either polymyxin B or PKC down-regulation, suggesting that it operated exclusively through a PKC-independent mechanism.
4. These results suggest that there are at least two pathways involved in the inhibition of dopamine release through dopamine receptors. One pathway involves dopamine receptor suppression of protein kinase C activity, perhaps through inhibition of phospholipase C activity and this is preferentially utilized by neuronally-released dopamine. The other pathway which seems to be utilized by exogenous agonists does not involve PKC.

     

Type I and II metabotropic glutamate receptors modulate periaqueductal grey glycine release: interaction between mGlu2/3 and A1 adenosine receptors

In this study we investigated the effects of type I and II mGlu receptors ligands in glycine extracellular concentrations at the periaqueductal gray (PAG) level by using in vivo microdialysis, in conscious rats. An agonist of type I mGlu receptors, (S)-3,5-DHPG (1 and 5 mM), but not a selective agonist for mGlu5 receptors, CHPG (3 and 5 mM), was noticed to increase the dialysate glycine levels in a concentration-dependent manner (60+/-15% and 136+/-13%, respectively). CPCCOEt (1mM), a selective mGlu1 receptor antagonist, perfused in combination with (S)-3,5-DHPG, counteracted the effect induced by (S)-3,5-DHPG, but did not change per se the extracellular PAG glycine values, even at the highest dosage used (2 mM). MPEP (1 and 2 mM), a selective antagonist of mGlu5 receptor, did not modify extracellular glycine level. An agonist of type II mGlu receptors, 2R,4R-APDC (25 and 50 microM), decreased the dialysate glycine in a concentration-dependent manner (-26+/-4% and -54+/-6%, respectively). The 2R,4R-APDC-induced decrease in extracellular glycine was prevented by EGlu (0.5 mM), a selective type II mGlu receptors antagonist. EGlu (0.5 and 1 mM), per se, led to a significant decrease (-56+/-7% and -57+/-2%, respectively) in extracellular PAG glycine too. This effect was prevented by DPCPX (100 microM), a selective antagonist for A1 adenosine receptors, but was not affected by CPA (1 mM), a selective A1 adenosine receptors agonist. Intra-PAG perfusion of CPA (0.1-1 mM) decreased the extracellular PAG glycine values (-47+/-13%) with 1 mM concentration. The CPA-induced effect was prevented by DPCPX (100 microM), and resulted to be additive with the 2R,4R-APDC-induced decrease in glycine values. DPCPX (1 mM) increased per se extracellular glycine (48+/-7%) at the highest dose used. Dipyridamole (100 microM), an inhibitor of both adenosine reuptake and phosphodiesterases, decreased extracellular glycine (-28+/-7%). Extracellular concentrations of glutamine never changed throughout this study. These data show opposing effects of type I and II mGlu receptors in the regulation of PAG glycine values. Moreover, functional interaction between type II mGlu and adenosine A1 receptors, which possibly operate through a common transductional pathway, may be relevant in the physiological control of glycine release in awake, freely moving rats at the periaqueductal gray matter.     

Neuropeptide Y-induced enhancement of the evoked release of newly synthesized dopamine in rat striatum: mediation by Y2 receptors

The purpose of the present study was to determine whether or not activation of neuropeptide Y (NPY) receptors resulted in an enhancement or attenuation of the KCl (50 mM) evoked release of [3H]dopamine newly synthesized from [3H]tyrosine in superfused striatal slices and, if so to identify the NPY receptor subtype mediating the effect. Rat striatal slices were prepared and placed in microsuperfusion chambers and continuously superfused with physiological buffer containing 50 microCi/ml of l-3-5-[3H]tyrosine. Superfusate effluents were collected and analyzed for [3H]dopamine by liquid scintillation spectrometry following amberlite CG50 and alumina chromatography. NPY agonists (NPY and PYY3-36) were added 6 min prior to the addition of KCl, while the Y1, Y2, and Y5 antagonist BIBO3304, BIIE0246 and CGP71683A, respectively were added 6 min prior to the agonists. Continuous superfusion with [3H]tyrosine resulted in the production of [3H]dopamine which reached a steady state at approximately 48 min. Depolarization with KCl resulted in a 2- to 3-fold increase in [3H]dopamine overflow. NPY and PYY3-36 produced a concentration dependent enhancement in the KCl induced increase in newly synthesized [3H]dopamine overflow. The Y2 antagonist BIIE0246 produced an attenuation of both the NPY and PYY3-36 induced enhancement while the Y1 antagonist BIBO3304 and theY5 antagonist CGP71683A failed to alter the NPY or PYY3-36 induced enhancement. These results are consistent with the NPY-Y2 receptor subtype mediating the facilitatory effect.     

Noradrenaline release from rat sympathetic neurons evoked by P2-purinoceptor activation

The effects of ATP and analogues on the release of previously incorporated ³H-noradrenaline were studied in cultured sympathetic neurons derived from superior cervical ganglia of neonatal rats. Electrical field stimulation (40 mA at 3 Hz) of the neurons for 10 s markedly enhanced the outflow of tritium. ATP applied for 5 s to 2 min at concentrations of 0.01 to 1 mmol/l caused a time- and concentration-dependent overflow with half maximal effects at about 10 s and 100 mol/l, respectively. 2-Methylthio-ATP was equipotent to ATP in inducing ³H-overflow. ADP (100 mol/l), when applied for 2 min, also caused a small ³H-overflow, but , -methylene-ATP (100 mol/l), AMP (100 mol/l), R(–)N⁶-(2-phenylsiopropyl)-adenosine (R(–)-PIA; 10 mol/l) and 5-N-ethylcarboxamidoadenosine (NECA; 1 mol/l) did not. The ³H-overflow induced by 10 s applications of 100 mol/l ATP was abolished by suramin (100 mol/l) and reduced by about 70% by reactive blue 2 (3 mol/l). Electrically evoked overflow, in contrast, was slightly enhanced by suramin, but not modified by reactive blue 2. Xanthine amine congener (10 mol/l) and hexamethonium (10 mol/l) did not alter ATP-evoked release. Removal of extracellular Ca²⁺ from the medium reduced ATP- and electrically induced overflow by about 95%. Tetrodotoxin (1 mol/l) abolished electrically evoked ³H-overflow but inhibited ATP-induced overflow by only 70%. The ₂-adrenoceptor agonist UK 14,304 at a concentration of 1 mol/l diminished both electrically and ATP-evoked tritium overflow by approximately 70%. These results indicate that activation of P₂-purinoceptors stimulates noradrenaline release from rat sympathetic neurons. The release resembles electrically induced transmitter release, but additional mechanisms may contribute. Correspondence to: S. Boehm at the above address     

3- and 4-O-sulfoconjugated and methylated dopamine: highly reduced binding affinity to dopamine D2 receptors in rat striatal membranes

Summary The binding properties of 3- and 4-O-sulfoconjugated dopamine (DA-3-0-S, DA-4-0-S) as well as 3-O-methylated dopamine (MT) to rat striatal dopamine D₂ receptors were investigated. ³H-spiperone was used as a radioligand in the binding studies. In saturation binding experiments (+)butaclamol, which has been reported to bind to dopaminergic D₂ and serotoninergic 5HT₂ receptors, was used in conjunction with ketanserin and sulpiride, which preferentially label 5HT₂ and D₂ receptors, respectively, in order to discriminate between ³H-spiperone binding to D₂ and to 5HT₂ receptors. Under our particular membrane preparation and assay conditions, ³H-spiperone binds to D₂ and 5HT₂ receptors with a maximal binding capacity (B _max) of 340 fmol/mg protein in proportions of about 75%:25% with similar dissociation constants K _D (35 pmol/l; 43 pmol/l). This result was verified by the biphasic competition curve of ketanserin, which revealed about 20% high (K _D = 24 nmol/l) and 80% low (K _D = 420 nmol/l) affinity binding sites corresponding to 5HT₂ and D₂ receptors, respectively. Therefore, all further competition experiments at a tracer concentration of 50 pmol/l were performed in the presence of 0.1 mol/l ketanserin to mask the 5HT₂ receptors. DA competition curves were best fitted assuming two binding sites, with high (K _H = 0.12 mol/l) and low (K_L = 18 mol/l) affinity, present in a ratio of 3:1. The high affinity binding sites were interconvertible by 100 mol/l guanyl-5-yl imidodiphosphate [Gpp(NH)p], resulting in a homogenous affinity state of DA receptors (K _D = 2.8 mol/l). Competition experiments with various compounds confirmed the binding of ³H-spiperone to D₂ receptors. DA-3-O-S, DA-4-O-S, and MT were more than 5,000-, more than 10,000-, and 530-fold less potent in competing for ³H-spiperone binding when compared with DA at the high affinity binding site which mediates biological effects. Therefore, it is concluded that these DA metabolites are biologically ineffective at central D₂ receptors. Send offprint requests to E. Werle at the above address     

Ammonium acetate challenge in experimental chronic hepatic encephalopathy induces a transient increase of brain 5-HT release in vivo

Ammonia has been shown to cause release of neurotransmitters such as serotonin (5-hydroxytryptamine; 5-HT) from synaptosomal preparations in vitro. In the present study, frontal neocortical extracellular levels of 5-HT and its major metabolite, 5-hydroxyindole-3-acetic acid (5-HIAA), were determined in vivo by the use of microdialysis in portacaval shunted (PCS) rats, an experimental model of chronic hepatic encephalopathy (HE), prior to and after an acute coma-inducing administration of ammonium acetate (NH₄Ac; 5.2 mmol/kg, i.p.). PCS rats displayed elevated (P<0.01) 5-HIAA but unaltered 5-HT extracellular levels compared with controls, supporting the contention of an increased neocortical 5-HT metabolism but unaltered neuronal 5-HT output in chronic HE. However, a transient elevation of extracellular 5-HT levels was observed when PCS-NH₄ Ac rats were in coma. Increased brain ammonia may thus augment neuronal 5-HT release in chronic HE, which in turn could be a causative factor for precipitation of more severe stages of HE.     

Histamine H3 receptor-mediated inhibition of serotonin release in the rat brain cortex

Summary Rat brain cortex slices preincubated with ³H-serotonin were superfused with physiological salt solution (containing citalopram, an inhibitor of serotonin uptake) and the effect of histamine on the electrically (3 Hz) evoked ³H overflow was studied. Histamine decreased the evoked overflow in a concentration-dependent manner. The inhibitory effect of histamine was antagonized by impromidine and burimamide, but was not affected by pheniramine, ranitidine, metitepine and phentolamine. Given alone, impromidine facilitated the evoked overflow, whereas burimamide, pheniramine and ranitidine had no effect. The results suggest that histamine inhibits serotonin release in the rat brain cortex via histamine H3 receptors, which may be located presynaptically. Send offprint requests to E. Schlicker at the above address     

A partial noradrenergic lesion induced by DSP-4 increases extracellular noradrenaline concentration in rat frontal cortex: a microdialysis study in vivo

The effect of systemic administration of the selective neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) on noradrenaline efflux in the frontal cortex was studied in freely-moving rats using microdialysis in vivo. Five days after treatment with DSP-4 (40 mg/kg IP), the noradrenaline content of the frontal cortex was reduced by 75%. Yet, noradrenaline efflux in the frontal cortex was nearly two-fold greater in DSP-4 treated rats than in saline-injected controls. Local infusion of the noradrenaline-selective uptake blocker, desipramine (5 μM), via the microdialysis probe, increased noradrenaline efflux in rats from both groups. Perfusion of Ringer’s solution, containing 80 mM K⁺, also increased noradrenaline efflux in both groups, but the increase after DSP-4 pretreatment was greater than in the controls. In contrast, removal of Ca²⁺ from the infusion medium reduced noradrenaline efflux in both treatment groups. These results indicate that, at this dose, DSP-4 increases the extracellular concentration of noradrenaline in rat frontal cortex despite causing a partial lesion of noradrenergic neurones. This is due to an increase in the release of noradrenaline, although reduced clearance is also likely. These data challenge the assumption that depletion of noradrenaline content after treatment with DSP-4 invariably translates into diminished noradrenergic transmission. Received: 4 September 1997/Final version: 7 September 1997     

Inhibition of noradrenaline release in the rat brain cortex via presynaptic H3 receptors

Summary The effects of histamine and related drugs on the evoked tritium overflow from superfused rat brain cortex slices preincubated with³H-noradrenaline were determined. Tritium overflow was stimulated electrically (3 Hz; slices superfused with normal physiological salt solution) or by introduction of CaCl₂ 1.3 mmol/l (slices superfused with Ca²⁺-free medium containing K⁺ 20 mmol/l).Histamine slightly decreased the electrically evoked^H overflow in slices superfused in the presence of desipramine. The degree of inhibition obtained with histamine was doubled when both desipramine and phentolamine were present in the superfusion medium (pIC₁₅ 6.46). Under the latter condition, the evoked overflow was inhibited by the H₃ receptor agonist R-(–)--methylhistamine and its S-(+) enantiomer (pIC₁₅ 7.36 and 5.09, respectively), but was not affected by the H₂ receptor agonist dimaprit and the H₁ receptoragonist 2-thiazolylethylamine (both at up to 32 µmol/l). The concentration-response curve of histamine was shifted to the right by the H3 receptor antagonists thioperamide, impromidine and burimamide (apparent pA₂ 8.37, 6.86 and 7.05, respectively), by the H₂ receptor antagonist ranitidine (apparent pA₂ 4.27) and was not affected by the H₁ receptor antagonist dimetindene (32 µmol/l). The inhibitory effect of R-(–)--methylhistamine on the evoked overflow was also counteracted by thioperamide. Given alone, none of the five histamine receptor antagonists affected the evoked overflow. In the absence of desipramine plus phentolamine, impromidine and burimamide facilitated the electrically evoked³H overflow whereas thioperamide had no effect. The facilitatory effects of impromidine and burimamide were abolished by phentolamine, but not affected by desipramine. The concentration-response curve of noradrenaline for its inhibitory effect on the evoked overflow was shifted to the right by impromidine and burimamide, but not influenced by thioperamide (apparent pA₂ 5.24, 5.04 and <6.5, respectively; experiments carried out in the presence of desipramine). In slices superfused with Ca²⁺-free K⁺-rich medium containing tetrodotoxin, desipramine plus phentolamine, the tritium overflow evoked by introduction of Ca²⁺ was inhibited by histamine; the concentration-response curve of histamine was shifted to the right by thioperamide.The present study shows that the inhibitory effect of histamine on noradrenaline release in the rat brain cortex involves presynaptic H₃ receptors and that the degree of inhibition is increased in the presence of phentolamine. The H₃ receptor antagonists impromidine and burimamide are weak ₂-adrenoceptor antagonists. Send offprint requests to E. Schlicker at the above address     

Inhibitory muscarinic receptors modulate the potassium-evoked release of [3H]serotonin from rat hypothalamic slices

The effects of oxotremorine, acetylcholine and nicotine have been investigated on the potassium-evoked release of [³H]serotonin from slices of rat hypothalamus. Oxotremorine and acetylcholine in the presence of physostigmine inhibited potassium-evoked tritium release without affecting the spontaneous release. Nicotine had no effect. The response to oxotremorine was unaffected by tetrodotoxin but was blocked by atropine and hyoscine suggesting that the muscarinic receptor mediating the response to oxotremorine was located on the serotonergic nerve terminal.     

Withdrawal from repeated morphine sensitises mice to the striatal dopamine release enhancing effect of acute morphine

The effects of morphine withdrawal and challenge on the a-methyl-p-tyrosine (MT)-induced depletion of dopamine (DA) as well as on DA metabolism and ³H-SCH 23390 and ³H-spiperone binding were studied in the striata of male mice. Morphine was given s.c. 3 times daily for 5 days followed by 1 to 3 days' withdrawal.The MT induced DA depletion was retarded in mice withdrawn for 1 day from repeated morphine. At this time point the striatal concentrations of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) fell, too. In mice withdrawn for 3 days from morphine neither DA depletion nor DOPAC or HVA concentrations differed from those of control mice. In control mice acute morphine challenge accelerated the DA depletion at the dose 10 mg/kg but not at the dose 30 mg/kg. Both doses elevated striatal DOPAC and HVA. In mice withdrawn from repeated morphine for 1 day acute morphine partially counteracted the withdrawal-induced retardation of DA depletion and elevated striatal DOPAC and HVA clearly less than in control mice. However, in mice withdrawn for 3 days 10 mg/kg of morphine clearly enhanced DA depletion and its effect on striatal HVA was significantly augmented. In these mice as in controls the 30 mg/kg dose did not alter striatal DA depletion and elevated HVA less than in controls. Acute morphine did not alter striatal 3-methoxytyramine (3-MT) concentration in control mice but at the dose 10 mg/kg increased it in mice withdrawn for 3 days. Morphine withdrawal did not significantly affect striatal 3H-SCH 23390 binding, but slightly decreased ³H-spiperone binding in mice withdrawn for 3 days indicating a down-regulation of D₂ receptors.Our results by using three different indices of DA release (DA depletion after aMT, HVA and 3-MT) show that long enough withdrawal from repeated morphine treatment augments the morphine-induced release of striatal DA in mice. We propose that the striatal DA release in mice is regulated by two opposite opioid sensitive mechanisms with different dose-dependencies and different tolerance development. Correspondence to: L. Ahtee at the above address