L-dopa facilitates the release of endogenous norepinephrine and dopamine via presynaptic beta 1- and beta 2-adrenoceptors under essentially complete inhibition of L-aromatic amino acid decarboxylase in rat hypothalamic slices

In rat hypothalamic slices, L-aromatic amino acid decarboxylase (AADC) was assayed, and the actions of L-DOPA on impulse (2 Hz)-evoked norepinephrine (NE) and dopamine (DA) release were studied under inhibition of AADC. Slices were incubated with L-DOPA, and DA and NE produced by conversion of the precursor were analyzed by high performance liquid chromatography with electrochemical detection (HPLC-ECD). In the slices, the Km and Vmax of AADC were 131 microM and 122 pmol/min/mg protein, respectively. NSD-1015, an AADC inhibitor, caused a noncompetitive type of inhibition, and the K1 value was 0.086 microM. In the presence of 20 microM NSD-1015, which was expected to cause 99.6% inhibition of AADC, L-DOPA (0.01-100 nM) concentration-dependently facilitated the release of NE from the superfused slices, and the L-DOPA (10 nM)-induced facilitation was antagonized by 100 nM ICI 89,406 and 100 nM ICI 118,551, a selective beta 1- and beta 2-adrenoceptor antagonist, respectively. This action of L-DOPA was not modified by 30 microM tropolone, an inhibitor of catechol-O-methyl-transferase. L-DOPA at 0.01-1 nM similarly facilitated the release of DA. A quantitative analysis revealed that the L-DOPA-induced increase in NE and DA release was much higher by a factor of 3 to 4 orders than was the amount of DA and NE converted from L-DOPA. These results add further support to the hypothesis that L-DOPA itself acts as a neuroactive substance in the rat central nervous system.     

L-dopa-like regulatory actions of L-threo-3,4-dihydroxyphenylserine on the release of endogenous noradrenaline via presynaptic receptors in rat hypothalamic slices.

Effects of L-threo-3,4-dihydroxyphenylserine (L-threo-DOPS) on the spontaneous release and the stimulus(2 Hz)-evoked release of endogenous noradrenaline were studied in rat hypothalamic slices with functioning L-aromatic amino acid decarboxylase (AADC) and with AADC inhibition. In non-inhibited slices, spontaneous release was not modified by L-threo-DOPS at 1 pM-100 nM, tended to increase at 1-10 microM and increased at 100 microM. Noradrenaline tissue content slightly increased at 100 microM. Stimulated release was concentration-dependently facilitated at 1-1000 pM and tended to decrease gradually from a maximum at 10 nM-10 microM. Under AADC inhibition, spontaneous release concentration-dependently increased at 10-100 microM by 60% of the increase seen in slices without AADC inhibition. Increase in noradrenaline tissue content was abolished. L-threo-DOPS produced a triphasic pattern on stimulated release; concentration-dependent facilitation at 1-1000 pM similar to that seen in slices with functional AADC, no facilitation at 10-1000 nM, and a concentration-dependent increment at 10-100 microM. The facilitation at 1 nM was stereoselective and was antagonized by (-)-propranolol 10 nM, and no facilitation at 100 nM was restored to the maximum by yohimbine 10 nM, DG-5128 10 nM or S-sulpiride 1 nM. Furthermore, L-threo-DOPS (1-1000 pM)-induced facilitation was competitively antagonized by L-dopa methyl ester, a competitive antagonist for L-dopa, with a pA2 value of 13.6, whereas it was noncompetitively antagonized by (-)-propranolol.     

Nanomolar L-dopa facilitates release of dopamine via presynaptic beta-adrenoceptors: comparative studies on the actions in striatal slices from control and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated C57 black mice, an animal model for Parkinson's disease.

Effects of L-DOPA (0.1-10,000 nM) on spontaneous release (Sp), evoked release (S) and tissue content (C) of dopamine (DA) were studied comparatively in superfused striatal slices from control and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated C57 black mice to obtain evidence for L-DOPA-induced facilitation of S via presynaptic beta-adrenoceptors. In control slices, isoproterenol-induced concentration-dependent increases in S were propranolol-sensitive. L-DOPA at 0.1-3 nM tended to increase the S of DA with a concomitant tendency of increases in Sp. L-DOPA at 10-1 x 10(4) nM concentration-dependently increased Sp. L-DOPA at 1-10 microM tended to increase S and 10 microM increased C. In slices from MPTP-treated mice, the absolute amounts of Sp, S and C decreased by half compared to those in control slices. L-DOPA at 3 nM facilitated S without increasing Sp. This facilitation was antagonized by propranolol at 3 nM. L-DOPA at 30 nM decreased S from the peak facilitation, which contrasted with no effect in the control slices. However, 10-100 nM L-DOPA increased Sp more markedly than that in the control slices. L-DOPA at 100 nM increased S and C, which contrasted with no effect in the control slices. In conclusion, nanomolar L-DOPA facilitates the S of DA via presynaptic beta-adrenoceptors at concentrations lower than those required to induce conversion to DA even in striatal slices from the MPTP-treated mice model for Parkinson's disease.(ABSTRACT TRUNCATED AT 250 WORDS)     

L-dopa methyl ester antagonizes competitively L-dopa-induced facilitation of noradrenaline release from rat hypothalamic slices

In rat hypothalamic slices, antagonism by L-DOPA methyl ester and (-)-propranolol against L-DOPA-induced facilitation of endogenous noradrenaline (NA) release was characterized under the inhibition of dopadecarboxylase. L-DOPA at 10 nM to 1 microM facilitated the evoked NA release in a concentration-dependent manner. L-DOPA methyl ester (3, 10 and 30 nM) progressively shifted the concentration-release curve for L-DOPA to the right: Schild plots gave a straight line with a slope of 1.00 and pA2 was 8.9. This antagonistic action was not mimicked by L-phenylalanine, a substrate for L-DOPA transport system. In contrast, 10 and 100 nM propranolol concentration-dependently reduced the maximal effect of L-DOPA without rightward shift of the concentration-release curve. L-DOPA methyl ester is a potent competitive antagonist for the action of L-DOPA, and the recognition site of L-DOPA differs from presynaptic beta-adrenoceptors.     

Carteolol is a useful tool to prove the tonically functioning nature of presynaptic beta-adrenoceptors on peripheral noradrenergic neurons but not on central catecholaminergic neurons

Effects of carteolol on norepinephrine (NE) release were studied at 2 Hz mainly in rat hypothalamic slices. Isoproterenol at 1 and 10 nM concentration-dependently facilitated NE release. Isoproterenol (10 nM)-induced facilitation was antagonized by 1 and 10 nM dl-carteolol, but not antagonized by 1 nM d-carteolol. dl-Carteolol alone at 1 nM to 10 microM did not inhibit NE release. In brainstem slices, 10 nM isoproterenol also facilitated NE release, and this facilitation tended to be antagonized by 1 nM dl-carteolol. Nanomolar concentrations of carteolol stereoselectively antagonized isoproterenol-induced facilitation of NE release via presynaptic beta-adrenoceptors in rat hypothalamic slices.     

Stereoselectivity of presynaptic autoreceptors modulating dopamine release

The effects of the (R)- and (S)-enantiomers of sulpiride and butaclamol were studied on the spontaneous and field stimulation-evoked release of total radioactivity from slices of rabbit caudate nucleus prelabelled with [3H]dopamine. (S)-Sulpiride in concentrations ranging from 0.01--1 microM enhanced the electrically evoked release of [3H]dopamine while (R)-sulpiride was 10 times less potent than (S)-sulpiride. Exposure to (S)-butaclamol (0.01--1 microM) but not to (R)-butaclamol (0.1--10 microM) enhanced the field-stimulated release of [3H]dopamine. The facilitatory effects of (S)- and (R)-sulpiride and (S)-butaclamol on the stimulated release of the labelled neurotransmitter were observed under conditions in which these drugs did not modify the spontaneous outflow of radioactivity. Only the active enantiomers of sulpiride and butaclamol antagonized the inhibition by apomorphine (1 microM) of the stimulated release of [3H]dopamine. Our results indicate that the presynaptic inhibitory dopamine autoreceptors modulating the stimulation-evoked release of [3H]dopamine in the caudate nucleus are, like the classical postsynaptic dopamine receptors, chemically stereoselective.     

Evidence for the existence of stereoselective presynaptic beta 1-adrenoceptors on noradrenergic and dopaminergic neurons in the rat hypothalamus

Using high performance liquid chromatography with an electrochemical detector, we examined antagonistic effects of l- and d-acebutolol and atenolol against isoproterenol-induced facilitation of impulse-evoked release of endogenous norepinephrine and dopamine in rat hypothalamic slices. Isoproterenol (10 nM) increased the release of the both catecholamines, and this facilitation was similarly antagonized by 100 nM l-acebutolol and 100 nM atenolol, but was not antagonized by 100 nM d-acebutolol. In conclusion, there exist stereoselective presynaptic beta 1-adrenoceptors on noradrenergic and dopaminergic neurons in the rat hypothalamus.     

The adenosine receptor-mediated inhibition of noradrenaline release possibly involves an N-protein and is increased by alpha 2-autoreceptor blockade.

The stimulation-evoked overflow of [3H]-noradrenaline from slices of the rabbit hippocampus is inhibited by alpha 2-autoreceptors as well as by adenosine (A1)-receptors. Slices of rabbit hippocampus were labelled with [3H]-noradrenaline, superfused continuously and stimulated twice electrically (rectangular pulses; 2 ms, 3 Hz, 24 mA, 5 V cm-1). Treatment of hippocampal slices with N-ethylmaleimide (NEM, 30 microM; 30 min), which functionally disturbs certain N-proteins, decreased the inhibitory action of adenosine receptor agonists like (-)-N6-(R-phenylisopropyl)-adenosine ((-)-PIA) and adenosine on noradrenaline release. Release inhibition caused by (-)-PIA (0.03-1 microM) was antagonized by NEM in a non-competitive manner in the absence and in the presence of the alpha 2-adrenoceptor antagonist yohimbine. The adenosine receptor antagonist 8-phenyltheophylline significantly increased the evoked noradrenaline release by about 15% in control slices by diminishing the inhibitory action of endogenous adenosine. In NEM-treated slices this effect of 8-phenyltheophylline was not seen. In the presence of (-)-PIA (0.1 microM), i.e. under conditions of an increased inhibitory tone, release facilitation by 8-phenyltheophylline was decreased by NEM compared to that in the respective controls. Occupation of the A1-receptor with (-)-PIA prior to and during the NEM treatment did not protect the A1-receptor-coupled signal transduction system from being affected by NEM. In the presence of the alpha 2-adrenoceptor antagonist yohimbine, the inhibitory action of (-)-PIA was strongly increased. The above results suggest the involvement of a regulatory N-protein in the A1-receptor-mediated inhibition of noradrenaline release and an interaction between the alpha 2-autoreceptor and the A1-receptor-coupled signal transduction system, possibly at the level of a N-protein.     

Newly synthesized noradrenaline mediates the alpha 2-adrenoceptor inhibition of [3H]5-hydroxytryptamine release induced by beta-phenylethylamine in rat hippocampal slices

In slices of the rat hippocampus, alpha 2-adrenoceptors located presynaptically on serotonergic nerve terminals modulate the electrically evoked calcium-dependent release of [3H]serotonin [( 3H]5HT). We have investigated the effects of a naturally occurring trace amine, beta-phenylethylamine (beta-PEA), on noradrenergic transmission in the rat hippocampus. Under experimental conditions in which MAO of type B is inhibited by deprenyl-exposure to beta-PEA (0.1-10 microM) facilitates the spontaneous outflow of [3H]noradrenaline and inhibits the electrically evoked release of [3H]5HT. The inhibitory effect of beta-PEA (3 microM) on the evoked release of [3H]5HT was antagonized by the alpha 2-adrenoceptor antagonist idazoxan at 1 microM, and by pretreatment with alpha-methyl-p-tyrosine (alpha-MpT, 300 mg/kg i.p., 2 h). The inhibition of tyrosine hydroxylase activity by alpha-MpT does not modify the inhibition of the evoked release of [3H]5HT by the alpha 2-adrenoceptor agonist, 6-fluoronoradrenaline, or the serotonin receptor agonist, 5-methoxytryptamine. Pretreatment with the neurotoxin DSP4 (50 mg/kg i.p., 10 days) markedly antagonized the inhibitory action of beta-PEA on [3H]5HT release. These results indicate that the noradrenaline-releasing action of beta-PEA inhibits the electrically evoked release of [3H]5HT through the activation of alpha 2-adrenoceptors. This inhibitory effect appears to be mediated exclusively through the release of newly synthesized noradrenaline, and does not involve the direct activation by beta-PEA of the inhibitory 5HT autoreceptors which modulate [3H]5HT release in the rat hippocampus.     

Effect of a sulfonium analog of dopamine on the depolarization-induced release of [3H]acetylcholine from mouse striatal slices

P3 have synthesized a chemical analog or dopamine in which the amino group has been replaced by a charged dimethylsulfonium group. The dopaminergic activity of this drug was evaluated by determining its ability to inhibit the depolarization-evoked release of [3H]acetylcholine from mouse striatal slices. The slices were preincubated with [3H]choline (0.1 microM) and then superfused in physiological medium. [3H]Acetylcholine release was induced by exposure of the slices to a high potassium medium (12.5 mM) for 5 min. The sulfonium analog of dopamine, dopamine, and apomorphine inhibited the evoked [3H]acetylcholine release with IC50 values of approximately 10, 2.0, and 0.3 microM respectively. The inhibition by the sulfonium analog was reversed by fluphenazine (1 microM), suggesting that the inhibition of [3H]acetylcholine release was due to the activation of dopaminergic receptors. The sulfonium analog also inhibited the uptake of [3H]dopamine into striatal slices and caused the release of exogenously taken up [3H]dopamine from these slices. The release of [3H]dopamine by the sulfonium analog was inhibited by cocaine (3 microM), suggesting that the drug-induced release of [3H]dopamine was dependent on the carrier-mediated uptake of the sulfonium analog into dopaminergic neurons. The inhibition of the evoked [3H]acetylcholine release by high concentrations (30 and 60 microM) of the sulfonium analog did not appear to be mediated by endogenous dopamine release, since the analog still inhibited [3H]acetylcholine release from slices after reserpine-alpha-methyl-p-tyrosine treatment. However, the inhibitory effect of the sulfonium analog at 10 microM was reduced by reserpine-alpha-methyl-p-tyrosine treatment, suggesting that the inhibition at lower concentrations was mediated through endogenous DA release. These results suggest that a charged compound can act as a substrate for the dopamine carrier and can activate the dopamine receptor regulating acetylcholine release. They also indicate that the nitrogen on the dopamine molecule is not essential for dopamine agonist activity.     

Mu-, delta- and kappa-opioid receptor-mediated inhibition of neurotransmitter release and adenylate cyclase activity in rat brain slices: studies with fentanyl isothiocyanate

We investigated the effects of [D-Ala2,D-Leu5]enkephalin (DADLE). [D-Ala2,MePhe4,Gly-ol5]enkephalin (DAGO), [D-Pen2,D-Pen5]enkephalin (DPDPE) (0.01-1 microM) and bremazocine (0.001-0.3 microM) on the electrically evoked release of radiolabelled neurotransmitters and on the dopamine (DA)-stimulated cyclic AMP efflux from superfused rat brain slices. The differential inhibitory effects of these agonists on the evoked neurotransmitter release indicate that the opioid receptors mediating presynaptic inhibition of [3H]noradrenaline (NA, cortex), [14C]acetylcholine (ACh, striatum) and [3H]DA (striatum) release represent mu, delta and kappa receptors, respectively. In agreement with this classification, preincubation (60 min) of the slices with the delta-opioid receptor-selective irreversible ligand, fentanyl isothiocyanate (FIT, 0.01-1 microM), antagonized the inhibitory effects of DADLE and DPDPE on striatal [14C]ACh release only. On the other hand, the D-1 DA receptor-stimulated cyclic AMP efflux from striatal slices appeared to be inhibited by activation of mu as well as of delta receptors. In this case, the reversible mu antagonist, naloxone (0.1 microM), fully antagonized the inhibitory effect of the mu agonist, DAGO, without changing the effect of the delta agonist DPDPE but was ineffective as an antagonist in slices pretreated with FIT (1 microM). The inhibitory effect of DAGO on the electrically evoked [3H]NA release was antagonized by naloxone whether the receptors were irreversibly blocked by FIT or not. These data not only further support the existence of independent presynaptic mu-, delta- and kappa-opioid receptors in rat brain but also evidence strongly that mu and delta receptors mediating the inhibition of DA-sensitive adenylate cyclase could share a common binding site (for naloxone and FIT) and, therefore, may represent constituents of a functional opioid receptor complex.     

Effects of the central analgesic tramadol on the uptake and release of noradrenaline and dopamine in vitro.

1. The centrally acting analgesic, tramadol, has low affinity for opioid receptors and therefore presumably other mechanisms of analgesic action. Neurotransmitter release and uptake experiments were used to characterize the effects of tramadol on the central noradrenergic and dopaminergic systems. 2. Tramadol inhibited the uptake of [3H]-noradrenaline into purified rat hypothalamic synaptosomes with an IC50 of 2.8 microM; the (-)-enantiomer was about ten times more potent than the (+)-enantiomer. Results with the principal metabolite O-desmethyltramadol were very similar. Inhibition of dopamine uptake into purified rabbit caudate nucleus synaptosomes was very weak with 62% inhibition of 100 microM. 3. Rat occipital cortex slices were preincubated with [3H]-noradrenaline and rabbit caudate nucleus slices with [3H]-dopamine, then superfused and stimulated electrically. Tramadol, 1 and 10 microM, enhanced the stimulation-evoked [3H]-noradrenaline overflow by 25 and 69%, respectively; the (-)-enantiomer was more potent than the racemate or the (+)-enantiomer. Tramadol, 10 microM, had no effect on dopamine release. 4. The effects of tramadol on the stimulation-evoked [3H]-noradrenaline release were abolished when uptake sites were already blocked by a high concentration of cocaine. 5. The metabolite O-desmethyltramadol showed a slight facilitation of the stimulation-evoked noradrenaline release; the effect was more pronounced in the presence of a high concentration of naloxone. In the presence of cocaine, inhibition of the release was observed similar to the effect of morphine but less potent. 6. The results show that tramadol blocks noradrenaline uptake with selectivity as compared to dopamine uptake. The interaction with the noradrenaline transporter is stereoselective.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of amphetamine-induced dopamine release on radiotracer binding to D1 and D2 receptors in rat brain striatal slices

The in vivo binding of positron emission tomography (PET) and single photon emission computer tomography (SPECT) radiotracers to dopamine D2 receptors in the striatum can be influenced by competition with endogenous dopamine. The present study was undertaken to determine if a similar inhibition of radiotracer binding to dopamine receptors could be observed following pharmacologically-evoked dopamine release in rat brain striatal slices. Striatal slices were incubated in a large volume of oxygenated Krebs saline and exposed to amphetamine or methamphetamine to evoke dopamine release within the slice. Amphetamine and methamphetamine, at concentrations up to 30 microM, reduced [3H]raclopride binding in the slices by 77% and 86%, respectively, with 50% inhibition at 1.6 microM amphetamine or 3.0 microM methamphetamine. Neither drug produced a significant effect on binding of [3H]SCH 23390 in the slices. This suggests that dopamine was able to interfere with radiotracer binding to D2 but not D1 receptors. The dopamine uptake blockers, cocaine and methylphenidate, had relatively little effect by themselves on [3H]raclopride binding but, by inhibiting amphetamine-induced dopamine release, significantly reduced inhibition of [3H]raclopride binding by a low (3 microM) amphetamine concentration. At a higher (30 microM) amphetamine concentration the inhibition of [3H]raclopride binding was not antagonized by uptake blockers and data obtained from homogenate binding experiments indicated a direct displacement of [3H]raclopride binding by amphetamine at this concentration. In conclusion the data obtained in the present study demonstrate that the effects of amphetamine on striatal radiotracer accumulation observed in PET and SPECT can also be observed in brain slices in vitro and, at least at low amphetamine concentrations, are mediated by competition with released dopamine.     

The effects of dopamine and dopamine agonists on the release of 3H-GABA and 3H-5HT from rat nigral slices

Only high micromolar concentrations of dopamine and dopamine agonists altered spontaneous and KCl-evoked release of 3H-GABA and 3H-5HT from rat nigral slices in vitro. Apomorphine (100 microM) and dopamine (100 microM) enhanced the spontaneous release of 3H-5HT but the effect of dopamine was not reversed by haloperidol (1 microM). Both apomorphine (100 microM) and dopamine (100 microM) enhanced the KCl-evoked release of 3H-5HT but these effects were not reversed by haloperidol (1 microM). Apomorphine (10-250 microM) and dopamine (10-250 microM) inhibited 3H-5HT uptake into nigral synaptosomal preparations in a concentration-dependent manner. Accordingly, a major portion of the apparent effect of these drugs on 3H-5HT release may be due to inhibition of 3H-5HT uptake. Dopamine (100 and 1000 microM), amphetamine (100 microM), apomorphine (100 microM) and 2-amino-6,7-dihydroxytetralin (ADTN; 100 microM) were without effect on the spontaneous release of 3H-GABA from nigral slices. Apomorphine (100 microM) and ADTN (100 microM) reduced the KCl-evoked release of 3H-GABA from substantia nigra, an effect antagonized by haloperidol (1 microM). However, amphetamine (100 microM) and dopamine (100-1000 microM) were without effect on KCl-evoked 3H-GABA release. These results suggest that only high concentration of some dopamine agonists can modulate 3H-5HT and 3H-GABA release in substantia nigra. However, dopamine either had no effect, or its actions were not reversed by dopamine receptor blockade, so it appears unlikely that dendritic dopamine release will influence GABA and 5HT release in substantia nigra.     

Further characterization of the ORL1 receptor-mediated inhibition of noradrenaline release in the mouse brain in vitro.

Mouse brain slices preincubated with [3H]-noradrenaline or [3H]-serotonin were superfused with medium containing naloxone 10 microM; we studied whether nociceptin (the endogenous ligand at ORL1 receptors) affects monoamine release. Furthermore, the affinities of ORL1 ligands were determined using [3H]-nociceptin binding. The electrically (0.3 Hz) evoked tritium overflow in mouse cortex slices preincubated with [3H]-noradrenaline was inhibited by nociceptin and [Tyr14]-nociceptin (maximally by 80%; pEC50 7.52 and 8.28) but not affected by [des-Phe1]-nociceptin (pEC50<6). The ORL1 antagonist naloxone benzoylhydrazone antagonized the effect of nociceptin and [Tyr14]-nociceptin. The effect of nociceptin did not desensitize, was not affected by blockade of NO synthase, cyclooxygenase and P1-purinoceptors and was decreased by the alpha2-adrenoceptor agonist talipexole. Nociceptin also inhibited the evoked overflow in mouse cerebellar, hippocampal and hypothalamic slices in a manner sensitive to naloxone benzoylhydrazone. The electrically (3 Hz) evoked tritium overflow in mouse cortex slices preincubated with [3H]-serotonin was inhibited by nociceptin; naloxone benzoylhydrazone antagonized this effect. The affinities (pKi) for [3H]-nociceptin binding to mouse cortex membranes were: nociceptin, 8.71; [Tyr14]-nociceptin, 9.82; [des-Phe1]-nociceptin, <5.5; naloxone benzoylhydrazone, 5.85; naloxone, <4.5. In conclusion, nociceptin inhibits noradrenaline release in the mouse cortex via ORL1 receptors, which interact with presynaptic alpha2-autoreceptors on noradrenergic neurones. The effect of nociceptin does not desensitize nor does it involve NO, prostanoids or adenosine. Nociceptin also attenuates noradrenaline release from several subcortical regions and serotonin release from cortical slices by a naloxone benzoylhydrazone-sensitive mechanism.     

Alterations in dopamine release from striatal slices of rats after chronic treatment with haloperidol

The effects of chronic treatment with haloperidol on spontaneous and electrically evoked dopamine (DA) release from striatal slices of rats were investigated in vitro. DA was measured by high-performance liquid chromatography coupled to an electrochemical detector. The superfusion with haloperidol caused a dose-dependent (100 nM-100 microM) reduction in the electrically evoked DA release from striatal slices of rats, which was not antagonized by the superfusion with apomorphine. Chronic administration of haloperidol (1 mg/kg per day for 21 days) caused a significant reduction in electrically evoked DA release as well as in spontaneous DA release from striatal slices 24 h after the last injection. Moreover, pretreatment with haloperidol prevented the reduction of the DA release evoked in response to haloperidol superfusion (1 microM). These results indicated that chronic administration of haloperidol reduced DA release from striatal slices of rats, accompanied by tolerance for the inhibitory effect of drug superfusion on evoked DA release.     

Inhibition by L-3,4-dihydroxyphenylalanine of hippocampal CA1 neurons with facilitation of noradrenaline and gamma-aminobutyric acid release

Electrophysiological studies were performed to elucidate whether L-3,4-dihydroxyphenylalanine (L-DOPA) acted on hippocampal CA1 neurons, since this drug has been reported to act as a neurotransmitter in the hypothalamus and striatum. Hippocampal slices (450 microM thick) obtained from male Wistar rats (4-7 weeks of age) were placed in a bath (maintained at 30+/-1 degrees C) continuously perfused with artificial cerebrospinal fluid. The population spikes elicited by electrical stimuli applied to the Schaffer collateral/commissural fibers were recorded in the hippocampal CA1 region, using a glass micropipette filled with 3 M NaCl. Drugs were applied in the bath through a perfusion system. The population spikes were inhibited by L-DOPA (1 nM-10 microM) with a bell-shaped concentration-response curve (n=7-15). Maximum inhibitory effects were obtained at 100 nM. L-DOPA cyclohexyl ester, a putative L-DOPA recognition site antagonist, antagonized the L-DOPA-induced inhibition of population spike. However, the inhibition remained unaffected in the presence of 3-hydroxybenzylhydrazine, an aromatic amino acid decarboxylase inhibitor. Furthermore, bath application of either phentolamine, an alpha-adrenoceptor antagonist, or bicuculline, a GABA(A) receptor antagonist, antagonized the inhibitory effects of L-DOPA on population spikes. In addition, bicuculline (1 microM) antagonized the inhibition of population spike induced by 6-fluoronorepinephrine (10 microM), an alpha-adrenoceptor agonist, while phentolamine (10 microM) did not affect the muscimol (1 microM)-induced inhibition. These results suggested that L-DOPA itself acted on L-DOPA recognition sites to release noradrenaline, and that the latter facilitates gamma-aminobutyric acid (GABA) release via alpha-adrenoceptors located on the GABA-containing cells and/or their nerve terminals, thereby inhibiting the population spikes in the hippocampal CA1 field.     

Evoked noradrenaline release in the rabbit ear artery: enhancement by purines, attenuation by neuropeptide Y and lack of effect of calcitonin gene-related peptide.

1. Adenosine (30 microM) and its analogues 5'-N-ethylcarboxaminoadenosine (5 and 30 microM) and L-phenylisopropyladenosine (5 and 30 microM), potentiated the evoked but not spontaneous release of tritiated noradrenaline in the rabbit central ear artery. 2. Prejunctional inhibition of the evoked but not spontaneous release of tritiated noradrenaline by 100 nM neuropeptide Y is greater at 2 min than at 10 min after superfusion of the peptide. 3. Calcitonin gene-related peptide (2.63 to 263 nM) did not affect the evoked or spontaneous release of tritiated noradrenaline in this preparation. 4. These results are discussed in terms of prejunctional modulation of sympathetic transmission in the rabbit central ear artery.     

The effects of central aromatic amino acid DOPA decarboxylase inhibition on the motor actions of L-DOPA and dopamine agonists in MPTP-treated primates

1. Endogenous L-DOPA may act as a neuromodulator contributing to the production of motor activity. We now investigate the effects of the centrally acting aromatic amino acid dopa decarboxylase (AADC) inhibitor NSD-1015 (3-hydroxybenzyl hydrazine) on the motor actions of L-DOPA and dopamine agonist drugs in MPTP treated common marmosets. 2. Pretreatment with NSD-1015 (10 - 50 mg kg(-1); i.p.) worsened baseline motor deficits in MPTP-treated common marmosets. Similarly, it abolished L-DOPA (5 - 18 mg kg(-1) s.c.) induced locomotor activity and reversal of disability. NSD-1015 pretreatment inhibited dopamine formation and elevated L-DOPA levels in plasma. 3. The increase in locomotor activity and improvement in disability produced by the administration of the D-1 agonist A-86929 (0.03 - 0. 04 mg kg(-1) s.c.) or the D-2 agonist quinpirole (0.05 - 0.3 mg kg(-1) i.p.) was abolished by NSD-1015 (25 mg kg(-1) i.p.) pretreatment. While the effects of a low dose combination of A-86929 (0.04 mg kg(-1) s.c.) and quinpirole (0.05 mg kg(-1) i.p.) were inhibited by NSD-1015 (25 mg kg(-1) i.p.), there was little effect on the action of a high dose combination of these drugs (0.08 mg kg(-1) A-86929 and 0.1 mg kg(-1) quinpirole). 4. Following central AADC inhibition with NSD-1015 (25 mg kg(-1) i.p.), locomotor behaviour induced by administration of high dose combinations of A-86929 (0.08 mg kg(-1) s.c.) and quinpirole (0.1 mg kg(-1) i.p.) was unaffected by L-DOPA (5 mg kg(-1) s.c.) pretreatment. 5. These results do not support a role for endogenous L-DOPA in spontaneous or drug induced locomotor activity. Rather, they strengthen the argument for the importance of endogenous dopaminergic tone in the motor actions of dopamine agonists.     

Muscarinic inhibition of [3H]-noradrenaline release on rabbit iris in vitro: effects of stimulation conditions on intrinsic activity of methacholine and pilocarpine.

1. Rabbit isolated irides were loaded with [3H]-noradrenaline and superfused with Tyrode solution. The inhibition by the muscarinic agonists (+/-)-methacholine and pilocarpine of the [3H]-noradrenaline overflow into the superfusate evoked by field stimulation (pulses of 1 ms duration, 75 mA) was measured as an index of activation of presynaptic muscarinic receptors. 2. The fractional rate of release per pulse during the first stimulation period (S1) was low with 360 pulses at 3 Hz, intermediate with 360 pulses at 10 Hz and high with 1200 pulses at 10 Hz. Upon repetitive stimulation (7 periods at 20 min intervals), the fractional rates of release per pulse during S7 no longer differed, suggesting a 'long-term' regulation of [3H]-noradrenaline release depending on the stimulation conditions. 3. The evoked [3H]-noradrenaline overflow was depressed by (+/-)-methacholine in a concentration-dependent manner. The EC50 ranged from 0.29 to 0.42 microM. Methacholine nearly abolished the transmitter release evoked at 3 Hz but reduced that induced at 10 Hz by only 50%. Under the latter condition the methacholine concentration-inhibition curve was bell-shaped and no muscarinic inhibition was observed in the presence of methacholine 30 microM. After washout of methacholine the evoked [3H]-noradrenaline release was temporarily enhanced. 4. Atropine 0.1 microM enhanced the [3H]-noradrenaline overflow (evoked by stimulation with 360 or 1200 pulses at 10 Hz), probably antagonizing a presynaptic inhibition by endogenous acetylcholine. The inhibition by methacholine was competitively antagonized by atropine 0.1 microM (apparent -log KB = 8.5-9.0). 5. Depending on the concentration, pilocarpine reduced the [3H]-noradrenaline overflow evoked by 360 pulses at 3 Hz up to 63%. However, at 10 Hz stimulation frequency the compound was inactive as an agonist but competitively antagonized the presynaptic inhibition induced by methacholine. The KB under the latter condition (0.95 microM) was very close to the EC50 value determined at 3 Hz (0.85 microM). 6. The results demonstrate a muscarinic inhibition of noradrenaline release from the rabbit isolated iris. The activation by pilocarpine of the presynaptic receptors provides an alternative explanation for the miosis induced in the rabbit in vivo, which might be the result of a decreased sympathetic tone in the iris dilator muscle.