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International audience; Idiopathic Parkinson's disease (PD) is characterized by mesencephalic dopaminergic neuron cell death and striatal dopamine (DA) depletion. The factors involved in the pathogenesis of the disease are still unknown. Transforming growth factor beta1 (TGFbeta1) is increased in the striatum of patients with PD. However, the effect of this increase is not known. Here, we show that overexpression of TGFbeta1, by recombinant adenovirus TGFbeta1 gene transfer, in the mesostriatal system of an MPTP mouse model of PD decreased the number of mesencephalic dopaminergic neurons. This effect also involved more extensive DA depletion in the striatum. Striatal enkephalin mRNA levels are augmented, suggesting a decrease in dopaminergic transmission to the postsynaptic target. In the absence of MPTP, TGFbeta1 greatly decreased the number of dopaminergic neurons in the ventral mesencephalon of fully mature mice. These results show that an increase in TGFbeta1 levels aggravate the parkinsonian status of MPTP mice and may therefore be a risk factor for the development of PD.

To investigate the contribution of the dopamine (DA) synthesis to both the calcium-dependent and the carrier-mediated, mechanisms of DA release in the striatum, anaesthetized rats were locally superfused in the striatum with a push‐pull cannula supplied with an artificial CSF containing tritiated tyrosine. DA, dihydroxyphenylacetic acid (DOPAC) and their respective specific activity were measured in eCuent and used to evaluate changes in the DA synthesizing rate. Excluding calcium ions from the CSF only partially reduced spontaneous DA release (70%) still leaving a possible carrier-mediated DA release. This eAect was not additive with a local superfusion with 0.1 mM a-methyl-p-tyrosine, a blocker of DA synthesis, suggesting that synthesis could already be reduced by calcium-free superfusion. Local superfusion with 100 mM cadmium in the presence or not of calcium ions, increased the DA release (220 and 350%, respectively), simultaneously reducing DA synthesis. Local application of 1 mM calcium ionophore (A23187) was without eAect on the basal release of DA but enhanced DA synthesis and increased the amphetamine-evoked and carrier-mediated amine release. We conclude that DA synthesis can be a modulatory process of the firing-independent and carrier-mediated amine release while it weakly aAects the classical calcium-dependent release. # 1999 Elsevier Science Ltd. All rights reserved.

A push-pull cannula supplied with artificial CSF was implanted in the striatum of anaesthetized rats, and the basal extracellular DA and DOPAC was assayed in the superfusates using HPLC and electrochemical detection. Simultaneously, a carbon fibre electrode was implanted in close proximity of the cannula and the evoked DA release was detected by differential pulse amperometry during stimulation of the DA axons. Local treatments with cadmium (100 μM) blocked the evoked DA release (−90%), but substantially increased the basal extracellular DA (+ 125%). The effects of glutamate agonists NMDA (1 mM) and kainate (0.1 mM), known to increase basal extracellular DA were confirmed (+150% and +60% respectively). It was, however, simultaneously observed that the evoked DA release was inhibited (−80% and −50%, respectively). Amphetamine (1 μM) released DA (+150%) and produced also an increase (+100%) of the evoked DA release. These results, apparently conflicting, show that the two mechanisms releasing dopamine (firing-dependent and not) can be directly and simultaneously observed. These two releasing processes appear to be not strictly antagonist. They are also differently and independently modulated by calcium and by local influences such those conveyed by glutamate.

The effects of a chronic imipramine treatment on the mesoamygdaloid pathway of rats were examined. Using semiquantitative immunocytochemical techniques, it was observed that the level of TH mRNA was decreased in the ventral tegmental area (VTA). In contrast, the TH protein was increased in both the VTA and amygdala. The TH activity was decreased in the amygdala when assessed under normal conditions but increased after a preincubation to phosphorylate the enzyme, suggesting a lowering of the protein-specific activity in the terminals. These results show that TH protein turnover in the mesoamygdaloid neurons can be reduced by chronic imipramine treatments, thereby producing an accumulation of inactive TH protein in the neurons while also decreasing TH gene activity in the cell bodies.

Twenty weeks after ibotenic acid lesions of the striatum, the amount of tyrosine hydroxylase (TH) in this structure was markedly increased. This was accompanied by a 3-fold increase in TH mRNA levels in the ipsilateral subtantia nigra (SN). Striatal levels of dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) were markedly reduced. In the nucleus accumbens, spared by the lesion, DA neurotransmission was also altered, as evidenced by a reduction of DA and DOPAC, but no increase in TH could be detected. TH mRNA levels were moderately enhanced in the ventral tegmental area (VTA). Thus, lesioning in the striatum induces TH gene activation in both SN and VTA neurones, not strictly related to DA function at the terminal level.

The short-term inhibition by estradiol of tyrosine hydroxylase (TH) in tuberoinfundibular dopaminergic neurons was examined in vitro on hypothalamic slices from ovariectomized rats. TH activity (determined by L-3,4-dihydroxyphenylalanine accumulation in the median eminence after blockade of decarboxylase with NSD 1055) showed a 30-40% decrease within 1 h of incubation with estradiol. To determine whether a dephosphorylation process was involved in this decline in TH activity, we studied the sensitivity of the enzyme to dopamine (DA) feedback inhibition: In controls, we observed that two kinetically different forms of TH coexisted, with one exhibiting a Ki(DA) of 26.4 +/- 2 microM and the other being approximately 10-fold more sensitive to DA inhibition, with a Ki(DA) of 2.56 +/- 0.17 microM, likely corresponding to a phosphorylated and active form and to a nonphosphorylated and poorly active form, respectively. Conversely, after estradiol treatment all TH molecules exhibited the same Ki(DA) of 2.5 +/- 0.3 microM. This effect was stereospecific, because 17 alpha-estradiol could not promote it, whereas with 17 beta-estradiol, it could be observed at only 10(-11) M and after a short delay (30 min). Finally, this decrease in the Ki(DA) of the purported active form of TH could be prevented by okadaic acid (an inhibitor of protein phosphatases). These results suggest that estradiol can act directly on the mediobasal hypothalamus to trigger a rapid decline in TH activity and that this action may involve a decrease in TH phosphorylation.

We have analyzed some functional aspects of the promoter of the human dopamine beta-hydroxylase (DBH) gene. A fragment of 1,247 bp directly 5' to the transcriptional start was progressively shortened, placed in front of a reporter gene, and tested in a human neuroblastoma cell line expressing DBH (SK-N-SH-TFM) and in a monkey kidney cell line (CV-1). A remarkably short region (267 bp), directly upstream from the transcription start, was sufficient to confer activity and tissue-specific expression. Furthermore, the expression of the DBH gene was shown to be inducible by cyclic AMP in SK-N-SH-TFM cells. This effect was demonstrated to occur at the transcriptional level, as shown by run-on assays, and was due to the presence of a near-consensus cyclic AMP-responsive element located in the untranscribed 5' regulatory region of the gene.

A possible interference of the GnRH-associated peptide (GAP) with the metabolism of dopamine in the rat striatum was investigated. The presence of the precursor of the peptide in this central region of dopaminergic terminals was first evidenced using specific RIA. The action of GAP on dopamine release was investigated in the caudate nucleus using the local superfusion with a push-pull cannula supplied with an artificial CSF containing the tritiated precursor of dopamine [( 3H]tyrosine). Addition of GAP (1 microM) to the superfusing fluid resulted in an increase of the release of the newly synthesized dopamine without a significant modification of the total amine release. In situ neutralization of GAP by addition in the CSF of a rabbit serum containing antibodies directed against the GAP produced opposite effects evidencing a tonic function for this peptide. In addition to the increased specific activity of the dopamine released during GAP treatment, the alterations observed in the efflux (and the specific activity) of dihydroxyphenyl acetic acid and the activation of dopamine synthesis obtained in vitro in striatal slices in the presence of GAP led us to conclude that the GAP system could be considered as a positive control of dopamine synthesis.

Forskolin (FSK) was locally injected into the substantia nigra (SN) of anesthetised rats. The day after injection (24 and 36 hr), tyrosine hydroxylase (TH) activity increased locally in this structure but remained unmodified in the ipsilateral caudate nucleus (CN). The amount of messenger RNA for TH (TH-mRNA) was also increased in the SN 24 hr after the injection. However, TH protein content was modified neither locally in the SN nor in the ipsilateral CN. In addition, the decrease of the ratio between dopamine and its first metabolite in the CN and the SN suggested a decreased activity of the dopaminergic nigral cells. The absence of increase of the protein synthesis in spite of the fact that TH-gene transcription was initiated could be the consequence of the inhibition of dopaminergic cells by the drug. These results confirm that, in vivo, TH induction is cAMP-dependent and demonstrate that the TH-gene activity is not strictly coupled to the activity of dopaminergic cells in the SN.

A push-pull cannula supplied with an artificial cerebrospinal fluid containing the tritiated precursor of dopamine, [ 3 H]tyrosine, was implanted in the caudate nucleus of rats anesthetized with halothane. The extracellular dopamine and dihydroxyphenylacetic acid were measured in successive 20 min fractions (both in their tritiated and unlabeled form) and the ratio between the two forms calculated. Glutamate was added to the superfusing cerebrospinal fluid to investigate its role in the process of dopamine release. The release of dopamine and the efflux of dihydroxyphenylacetic acid were activated by a low concentration (10 −8 M) of glutamate. In contrast, a higher concentration (10 −4 M) of the amino acid reduced the release of dopaminc. These results first confirmed the presence of a dual mechanism of control, by glutamate, of the dopamine release in the striatum depending on the extracellular concentration. Secondly, these treatments affected the dihydroxyphenylacetic acid amount and predominantly the tritiated form of dopamine, suggesting that the glutamate induces an important increase of the amine synthesis, in spite of a moderate effect on the release. The reversal of the inhibition by applications of tetrodotoxin (5 × 10 −7 M) and bicuculline (10 −4 M) confirmed that it was mediated by an indirect mechanism involving a GABAergic neurotransmission. In addition, the increase of the spontaneous dopamine release during bicuculline application suggested the existence of a tonic mechanism of inhibition of dopamine release in the striatum. This was confirmed by the fact that local xylocaine-induced anesthesia of the sensory motor cortex increased the spontaneous release of dopamine in the striatum. Antagonists of glutamate receptors, either 2-amino-7-phosphono-heptanoic acid or 6-cyano-7-nitroquinoxaline-2,3-dione, were used to characterize the involvement of a particular receptor type. Local application of 2-amino-7-phosphono-heptanoic arid (10 −4 M) did not block the inhibition produced by a high glutamate concentration but did induce an increased spontaneous release of [ 3 H]dopamine. The quisqualate/kainate antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (5 × 10 −5 M) was responsible for the disappearance of low concentration-dependent effects (10 −8 M). Facilitation is thus likely to be mediated by a 6-cyano-7-nitroquinoxaline-2, 3-dione-dependent mechanism when 2-amino-7-phosphono-heptanoic acid-sensitive receptors control the tonic inhibitory influence. It is concluded that under cortical control a glutamatergic neurotransmission mediates a double influence on the release of dopamine in the striatum: a tonic inhibitory influence and a phasic activation.

The effects of moderate changes in extracellular dopamine concentrations on the in vivo binding of specific dopaminergic D2 radioligands with different affinities and kinetics were investigated in rats. Either ( 125 I)NCQ298 (Kd 5 19 pM), or ( 125 I)iodolisuride (Kd 5 0.27 nM) or ( 3 H)raclopride (Kd 5 1.5 nM) were administered intravenously (IV) to animals 1 h after the intraperitoneal (IP) injection of either a-methyl-p-tyrosine (AMPT) (250 mg/kg) or nomifensine (15 mg/kg), or saline. The kinetics of radioactivity concentration in the striatum, cerebellum, and plasma were measured for up to 4 h after ( 125 I)NCQ298 or ( 125 I)iodolisuride injection and up to 1.5 h after ( 3 H)raclopride injection. For each tracer, the striatum-to- cerebellum radioactivity concentration ratios (S/C) and the binding potential (BP), calculated as the association to dissociation binding rate constant ratios (k3/k4), were assessed and related to the changes in extracellular dopamine concentration induced by drug treatments. Results show that S/C and BP of ( 3 H)raclopride were significantly diminished by pretreatment with nomifensine, a drug that increases extracellular dopamine concentration. Nomifensine pretreatment induced no changes in the in vivo binding indexes of the high affinity ( 125 I)NCQ298 and a slight but not significant decrease of the binding indexes of ( 125 I)iodolisuride. Treatment with AMPT, which induced a 40% reduction in dopamine concentration, did not change ( 125 I)NCQ298 binding indexes but slightly increased those of ( 3 H)raclopride and ( 125 I)iodoli- suride. In conclusion, the change of dopamine concentration induces modification of radiotracer kinetics. Thus, the combined use of tracers with high and low affinities could allow us to obtain information both on receptor density and neurotransmitter release in vivo. However, as indicated by the ( 3 H)raclopride study with AMPT, small changes in the concentration of intrasynaptic dopamine cannot be easily detected. NUCL MED BIOL 26;1:91-98, 1999. © 1998 Elsevier Science Inc.

The acute effect of physiological doses of estradiol (E2) on the dopaminergic activity in the striatum was studied. In a first series of experiments, ovariectomized rats were injected with 17 alpha or 17 beta E2 (125, 250, or 500 ng/kg of body weight, s.c.), and in situ tyrosine hydroxylase (TH) activity (determined by DOPA accumulation in the striatum after intraperitoneal administration of NSD 1015) was quantified. A dose-dependent increase in striatal TH activity was observed within minutes after 17 beta (but not 17 alpha) E2 treatment. To examine whether E2 acts directly on the striatum, in a second series of experiments, anesthetized rats were implanted in the striatum with a push-pull cannula supplied with an artificial CSF containing [3H]tyrosine. The extracellular concentrations of total and tritiated dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) were measured at 20-min intervals. Addition of 10(-9) M 17 beta (but not 17 alpha) E2 to the superfusing fluid immediately evoked an approximately 50% increase in [3H]DA and [3H]DOPAC extracellular concentrations, but total DA and DOPAC concentrations remained constant. This selective increase in the newly synthesized DA and DOPAC release suggested that E2 affects DA synthesis rather than DA release. Finally, to determine whether this rapid E2-induced stimulation of DA synthesis was a consequence of an increase in TH level of phosphorylation, the enzyme constant of inhibition by DA (Ki(DA)) was calculated. Incubation of striatal slices in the presence of 10(-9) M 17 beta (but not 17 alpha) E2 indeed evoked an approximate twofold increase in the Ki(DA) of one form of the enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

The release of dopamine (DA) from striatal terminals appears to be a rather complex and misunderstood phenomenon. It has been observed from many years that this release is only partially dependent on calcium ions in the extracellular space (Arbilla and Langer, 1978; Raiteri et al., 1979; Schwartz et al, 1980; Okuma et al., 1983; Woodward et al., 1988). However, it is now well known that the synthesis of DA is modulated by means of alterations of tyrosine hydroxylase (TH) activity under the control of calcium-dependent phosphorylating mechanisms (Fung and Uretsky, 1982; El Mestikawy et al., 1983; Albert et al., 1984; Chowdhury and Fillenz, 1988; Zigmond et al. 1989). In addition, the synthesis of DA is the basis of its release (Costa et al., 1972; Roth et al., 1973; Schwarz et al., 1980) probably through an increase of both the vesicular and the cytosolic stores. Thus, in the striatum calcium is clearly related to the extracellular DA concentration both by its action on synthesis and release.

In vivo benzodiazepine receptor occupancy by increasing doses of CL 218,872 has been evaluated in the baboon Papio papio, using (11C) RO 15-1788 as specific radioligand and positron emission tomography as external detection system. Although BZR heterogeneity has been previously demonstrated in the brain of the living baboon using PET, we did not observe in our studies that CL 218,872 interacts preferentially with one of the BZR subtypes. The monophasic pattern of the dose dependent CL 218,872 displacement curve and the corresponding “in vivo Hill coefficient” near unity suggest that CL 218,872 binds in cerebral baboon cortex with a similar affinity with BZ1 as well as BZ2 subtypes. The anticon-vulsant properties of CL 218,872 against bicuculline and allylglycine-induced seizures were correlated with benzodiazepine receptor occupancy by assessment of electroencephalographic activity during positron emission tomography studies. Our data confirmed in vivo the hypothesis of a partial agonist anticonvulsant activity of CL 218,872. At the same time, the use of a GABAantagonist (bicuculline) or an inhibitor of the GABA synthesis (allylglycine) suggested the existence of an allosteric interaction between benzodiazepine receptors and GABA receptors.

It is now well admitted that intraterminal dopamine (DA) is distributed between various compartments. Numerous studies have been devoted to the problem of the DA compartmentation (Javoy and Glowinski, 1971; Groppetti et al., 1977, Mc Millen et al., 1980) and we have recently proposed a new functional model of the intraterminal organization of the metabolic pathways leading to the synthesis and the release of DA (Leviel et al., 1989). In brief, the greatest part of the amine appears to be stored; besides, the newly synthesized molecules constitute a second pool under close dependence of the synthesis and being the first to be released under basal conditions (Besson et al.,1969; Leviel and Guibert, 1987). In consequence, an evoked overflow of DA could result from the involvement of either the stored or the newly synthesized amine.

Rats were anesthetized with fluothane and implanted in the caudate nucleus with a push-pull cannula supplied with artificial CSF containing the tritiated precursor of dopamine (DA), [3H]tyrosine. Total DA and dihydroxyphenylacetic acid (DOPAC) were measured in successive 20 min fractions using high performance liquid chromatography and electrochemical detection. Radioisotopic counting of the peaks permitted the calculation of the specific activity of both DA and DOPAC released into the extracellular space. Local applications of potassium (8, 16 and 32 mM) induced a dose-dependent increase of the release of DA with a decrease of its specific activity as evidence of the involvement of a stored DA pool. Base release of DOPAC was increased by repeating potassium applications with a temporary decrease during the applications. Superfusion with alpha-methyl-p-tyrosine produced a decrease of both the [3H]DA and total DA with a simultaneous decrease of its specific activity. This decrease was considered to be an indicator of the involvement by synthesis inhibition of the stored amine, but the simultaneous decrease of the specific activity of DOPAC suggests that this release was intraterminal. These results constitute the first direct observation that DA terminals act with two separate pools (stored and releasable) and suggest that the stored amine is preferentially released intraterminally. Systemic injection of reserpine induced a decrease of the release of DA and DOPAC without alteration of DA-specific activity when the specific activity of DOPAC was lowered. From these results it is concluded that the releasable compartment of the amine is located, in part, in vesicles different in nature from the vesicles containing the stored amine.(ABSTRACT TRUNCATED AT 250 WORDS)

Little is known about the molecular events mediating neurotransmitter release, a crucial step in synaptic transmission. In this paper, the biosynthesis and release of L-beta-3,4-dihydroxyphenylalanine (L-DOPA) and dopamine were analyzed in three heterologous cell lines after retroviral-mediated gene transfer of tyrosine hydroxylase (EC 1.14.16.2), the rate-limiting enzyme in catecholamine synthesis. A recombinant retrovirus encoding human tyrosine hydroxylase type I as well as neomycin-resistance gene was used to infect a fibroblast (NIH 3T3), a neuroblastoma (NS20 Y), and a neuroendocrine (AtT-20) cell line. After selection in the presence of neomycin and in tyrosine-free medium, high levels of exogenous tyrosine hydroxylase activity were detected in extracts of the three cell lines. High-performance liquid chromatography of cell extracts and culture supernatants confirmed that the three cell lines hydroxylated tyrosine to form L-DOPA and released this metabolite into the culture medium. Interestingly, the neuroendocrine cell line AtT-20 synthesized not only L-DOPA but also dopamine. Evoked secretion studies established that AtT-20 cells released the transmitter upon depolarization in a regulated, calcium-dependent way. We discuss the implication of this approach for the analyses of neurotransmitter release as well as in the context of degenerative disorders such as Parkinson disease.

The effects of various sagittal sections on the asymmetric changes in [3H]dopamine release from nerve terminals and dendrites of the two nigrostriatal dopaminergic pathways induced by the application of α-methylparatyrosine ord-amphetamine in the left substantia nigra were examined in halothane-anaesthetized cats implanted with push-pull cannulae in both caudate nuclei and both substantiae nigrae. [3H]tyrosine was continuously delivered to each push-pull cannula and [3H]dopamine was estimated in serial fractions of superfusates. In cats without a lesion, α-methylparatyrosine (10−4M) reduced the release of [3H]dopamine in the left substantia nigra and induced an opposite effect in the left caudate nucleus. In contrast, [3H]dopamine release was enhanced in the right substantia nigra and reduced in the right caudate nucleus. Opposite asymmetric responses were observed in the four structures during application ofd-amphetamine (10−6M) into the left substantia nigra. The sagittal section of the mesencephalic decussations was without effect on the changes in [3H]dopamine release induced either by α-methylparatyrosine ord-amphetamine. The section of the corpus callosum and of the commissura anterior potentiated, whereas the section of the thalamic massa intermedia prevented thed-amphetamine-induced reduction of [3H]dopamine release in the left caudate nucleus. This could indicate that neuronal circuits under the control of contralateral structures may contribute to the regulation of the activity of ipsilateral dopaminergic neurons. Only the section of the thalamic massa intermedia prevented the contralateral responses evoked by the unilateral nigral application of α-methylparatyrosine ord-amphetamine. These results suggest that nigrothalamic neurons and thalamic nuclei are involved in the reciprocal regulation of both nigrostriatal dopaminergic pathways.

The release of newly synthesized [ 3 H]adenosine has been studied in vivo in cat caudate nucleus and substantia nigra, using a push pull cannula. In the presence of [ 3 H]adenosine as precursor, spontaneously released [ 3 H]adenosine was easily detectable in superfusates of the push pull cannula. In the caudate nucleus, potassium and veratridine caused a marked and reversible increase in [ 3 H]adenosine release. The effect of veratridine was completely blocked by tetrodotoxin (TTX) although TTX had no action by itself. Ouabain as well as glutamate, also markedly increased the release of [ 3 H]adenosine. The specific 5′ nucleotidase inhibitor α,β-methylene ADP, did not alter the increase in the amount of [ 3 H]adenosine obtained by veratridine, although it diminished the spontaneous release of [ 3 H]adenosine by about 20%. Push pull cannulae were also implanted simultaneously into the caudate nucleus and substantia nigra. Potassium applied into the caudate nucleus increased the local release of adenosine but did not change that observed in the substantia nigra. When potassium was applied into the substantia nigra, it also increased the local release of adenosine but did not change that observed in the caudate nucleus. The results are discussed in term of the possible existence of “purinergic neurons” and of the relation between the adenosine release and central nervous activity.

In order to study the mechanisms regulating the dopaminergic nigroamygdaloid cells, the release of dopamine was observed in the central nucleus of the amygdaloid complex. Halothane anesthetized rats were implanted, according to the experiment, with one or two push-pull cannulae in the central nuclei of the amygdala (ACE), the substantia nigra (SN) and/or the caudate nucleus (CN). Cannulae were supplied with artificial cerebrospinal fluid (CSF) containing tritiated tyrosine, and labeled dopamine [3H]DA was evaluated in successive superfusate fractions. Electrical stimulation of the medial forebrain bundle with an implanted bipolar electrode induced an increase of the [3H]DA release in the ipsi- and contralateral ACE. Electrical stimulation of the SN produced only a very delayed effect in the ipsilateral ACE but an immediate and large increase of [3H]DA release in the contralateral structure. Superfusion of unlabeled DA and α-methyl-p-tyrosine in the SN remained ineffective on the [3H]DA release in the ipsilateral ACE. In this structure the release of [3H]DA was, however, decreased by nigral superfusion with γ-amino-butyric acid (GABA). d -(+)-Amphetamine (1 μM), when superfused in the CN, induced a large enhancement of the [3H]DA release in the ipsilateral ACE simultaneously with the local increase of [3H]DA release. The results presented here are in agreement with the previous studies concerning the anatomical organization of the dopaminergic nigroamygdaloid pathway. The DA cell bodies located in the SN appear insensitive to a local action of DA, perhaps due to a lack of autoreceptors. They are, however, powerfully inhibited by GABA and the relation observed between the [3H]DA release in the CN and ACE support the hypothesis that the SN can act as a relay between the extrapyramidal and limbic systems.

The release of dopamine in the substantia nigra and striatum was investigated in halothane anaesthetized rats by means of the pushpull cannula method. Electrical stimulation of the subthalamic nucleus produced a marked enhancement of dopamine release in the ipsilateral substantia nigra. This effect is likely to be mediated by subthalamic efferent neurons since the application of acetylcholine in the subthalamic nucleus produced a similar effect. A later decrease of dopamine release was always observed in the ipsilateral striatum and was attributed to the autoregulation mechanisms of nigro-striatal dopaminergic neurons.

Positron emission tomography (PET) was used to investigate, in the living baboon, the in vivo modulation of [11C]Ro 15-1788 binding to benzodiazepine receptors in brain and the changes with ligands acting at the supramolecular complex during status epilepticus induced by pentylenetetrazole. The central type benzodiazepine receptors were labelled in vivo by intravenous injection of [11C]Ro 15-1788. Simultaneous positron emission tomography and electroencephalographic activity recording evidenced a modulation of the brain binding of [11C]Ro 15-1788 during pentylenetetrazole-induced status epilepticus. We investigated the changes in the modulation of radioligand kinetics and in seizure activity after intravenous administration of a benzodiazepine agonist (diazepam, 1.5 mg/kg), a benzodiazepine antagonist (Ro 15-1788, 2 mg/kg), a GABA agonist (progabide, 50 mg/kg) and a ligand of the picrotoxin/barbiturate binding sites (LY81067, 3.5 mg/kg). The results showed that there is an in vivo competitive interaction of pentylenetetrazole with the benzodiazepine receptors, as reflected by the low displacement of [11C]Ro 15-1788 in the first 10 min of the status epilepticus. However, in contrast to diazepam, progabide and LY81067, a dose (2 mg/kg) of Ro 15-1788 that saturates the benzodiazepine receptors was unable to block the seizures induced by pentylenetetrazole. This indicates that the benzodiazepine receptors play only a minor role in the status epilepticus induced by pentylenetetrazole. The contribution of other binding sites within the supramolecular complex is assessed.

Intravenous administration of 1-methyl-4-phenyl-1,2,3,6,-tetrahydropyridine (MPTP) leads to the progressive development of a model of Parkinson's disease in the primate. The development of damage occuring in the striatal area during MPTP-treatment was followed “in vivo” in a baboon by positron emission tomography (PET). Spiperone labelled with a positron emitter 76Br (76Br-BSP) was used for the quantitative “in vivo” imaging of D2 dopamine receptors. The decrease in the striatal binding of 76Br-BSP measured “in vivo”, after three series of MPTP injections, paralleled the increase in the severity of behavioral symptoms seen immediately after administration of the neurotoxin. At the end of the MPTP-treatment when neurological symptoms were the most important, a 36 % decrease in the 76Br-BSP specific binding was measured. Between the series of MPTP injections a partial recovery in the quantitative measurement of the 76Br-BSP specific binding occuring in the striatum was well correlated with the disappearance of the neurological syndrome. Post-mortem histological and biochemical studies in nigro-striatal anatomical structures of MPTP-intoxicated primates compared with control animals showed a 80 % loss of neuronal cell bodies in the substantia nigra compacta and a 42 % decrease in the density (Bmax) of D2 receptors (in vitro 3H-spiperone binding). All these results showed that the use of PET and 76Br-BSP allow to follow in a noninvasive way both the degenerative processes and the subsequent partial recovery which occur in dopaminergic striatal receptor function during MPTP-treatment.

Tritiated dopamine synthesized from tritiated tyrosine was estimated simultaneously in the two caudate nuclei and the two substantia nigrae of cats anaesthesized with halothane. In control animals, the electrical stimulation of the right forelimb enhanced dopamine release in the right caudate nucleus and decreased dopamine release in the right substantia nigra. Opposite effects were observed in the contralateral structures. Left nigral application of d-amphetamine produced the same effect. However in cats with extensive lesions of the left pericruciate cortex, an increase in the release of dopamine in the left substantia nigra was the only detectable effect of these two treatments. These results suggest that the cortical structures are involved not only in the transfer of information between the two dopaminergic pathways but are also involved with regulation of the release of dopamine in the striatum originating in the substantia nigra. With regard to the role of the thalamic structures in this transfer of information, it is proposed that the thalamostriatal control of the release of dopamine previously suggested is closely dependent on cortical activity.

The selective benzodiazepine antagonist RO 15-1788, labelled with carbon 11 [11C] RO 15-1788, as a specific marker, together with positron emission tomography, allows the in vivo study of benzodiazepine receptors in primates. In addition, when coupled with recordings of electroencephalographic activity, this method offers the feasibility of studying the correlation between occupancy of benzodiazepine receptors and the convulsant action of drugs acting at the benzodiazepine-GABA receptor complex in vivo. The present study showed that convulsant doses of pentylenetetrazol (PTZ) could affect the binding of [11C] RO 15-1788 in vivo in two ways, depending on the doses tested: at concentrations of 20 and 30 mg/kg, pentylenetetrazol increased the binding of [11C] RO 15-1788 whereas larger concentrations displaced the binding of [11C] RO 15-1788. The direct correlation between the occupancy of respective benzodiazepine receptors, afforded by increasing convulsant doses of pentylenetetrazol, revealed that competitive interaction with benzodiazepine receptors was not necessary for pentylenetetrazol to induce the appearance of seizures in vivo.

A push-pull perfusion technique was used to study the release of endogenously synthesized [3H]-adenosine from caudate nucleus in the anaesthetized cat. The spontaneous release of [3H]adenosine newly synthesized from [3H]adenine reached a steady state level 40 min after the beginning of superfusion and continued for 4 h. Potassium and veratridine increased the release of newly synthetized [3H]-adenosine. The action of veratridine was completely blocked by tetrodotoxin. We conclude that spontaneous and evoked release of adenosine occurs in the cat striatum and might potentially affect central nervous function.

The convulsant actions of methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) and of methyl beta-carboline-3-carboxylate (beta-CCM) were evaluated in the baboon (Papio papio). DMCM, 0.6-4 mg/kg, induced epileptic seizures with short latency. DMCM convulsive seizures could be blocked by i.v. administration of the benzodiazepine agonist diazepam (10 mg). Similarly, beta-CCM, 0.3-3 mg/kg i.v., provoked generalized seizures in the baboons. These seizures were also reversed by the administration of propyl beta-carboline-3-carboxylate (3 mg/kg) or of diazepam (5 mg/kg). Combining the results from Positron Emission Tomography and the EEG assessments, benzodiazepine receptor occupancy by beta-CCM and DMCM was directly correlated with their convulsant actions in the living baboon. beta-CCM exerted its convulsant action in the living baboon at 76 and 74% benzodiazepine receptor occupancy in, respectively, occipital and temporal cortices whereas DMCM displayed a similar convulsive activity when only 58 and 65% of these receptors in the above regions were occupied.

The nucleus centralis of the amygdaloid complex was examined for the localization of the release of newly synthesized dopamine. Results are in good agreement with histochemical descriptions. The functional role of this release remains to be established.