M.L. Kemel

Glutamatergic Control of Dopamine Release in the Rat Striatum: Evidence for Presynaptic N-Methyl-D-Aspartate Receptors on Dopaminergic Nerve Terminals

Abstract: The N‐methyl‐D‐aspartate (NMDA) receptor‐mediated regulation of the release of newly synthesized [3H]dopamine ([3H]DA) was studied in vitro, both on rat striatal slices using a new microsuperfusion device and on rat striatal synaptosomes. Under Mg2+‐free medium conditions, the NMDA (5 × 10−5M)‐evoked release of [3H]DA from slices was found to be partly insensitive to tetrodotoxin (TTX). This TTX‐resistant stimulatory effect of NMDA was blocked by either Mg2+ (10−3M) or the noncompetitive antagonist MK‐801 (10−6M). In addition, the TTX‐resistant NMDA‐evoked response could be potentiated by glycine (10−6M) in the presence of strychnine (10−6M). The coapplication of NMDA (5 × 10−5M) and glycine (10−6M) stimulated the release of [3H]DA from striatal synaptosomes. This effect was blocked by Mg2+ (10−3M) or MK‐801 (10−5M). These results indicate that some of the NMDA receptors involved in the facilitation of DA release are located on DA nerve terminals. These presynaptic receptors exhibit pharmacological properties similar to those described in electro‐physiological studies for postsynaptic NMDA receptors.

Regulation of dopamine release by presynaptic nicotinic receptors in rat striatal slices: Effect of nicotine in a low concentration

Abstract Rat striatal slices were continuously superfused with 3H-tyrosine to study the effect of a low concentration of nicotine on the spontaneous release of newly synthesized 3H-dopamine (3H-DA). Nicotine (10−6M) stimulated the calcium-dependent spontaneous release of 3H-DA. This effect was prevented by nicotinic blockers such as pempidine (10−5M) and d-tubocurarine (5×10−6M). The stimulatory effect of nicotine and its blockade by pempidine were still observed in the presence of tetrodotoxin (5×10−7M). This demonstrates for the first time that a low concentration of nicotine is effective in releasing DA by acting on presynaptic nicotinic receptors located on terminals of the nigro-striatal dopaminergic neurons.

The vesicular glutamate transporter VGLUT3 synergizes striatal acetylcholine tone

Three subtypes of vesicular transporters accumulate glutamate into synaptic vesicles to promote its vesicular release. One of the subtypes, VGLUT3, is expressed in neurons, including cholinergic striatal interneurons, that are known to release other classical transmitters. Here we showed that disruption of the Slc17a8 gene (also known as Vglut3) caused an unexpected hypocholinergic striatal phenotype. Vglut3−/− mice were more responsive to cocaine and less prone to haloperidol-induced catalepsy than wild-type littermates, and acetylcholine release was decreased in striatum slices lacking VGLUT3. These phenotypes were associated with a colocalization of VGLUT3 and the vesicular acetylcholine transporter (VAChT) in striatal synaptic vesicles and the loss of a synergistic effect of glutamate on vesicular acetylcholine uptake. We propose that this vesicular synergy between two transmitters is the result of the unbalanced bioenergetics of VAChT, which requires anion co-entry for continuing vesicular filling. Our study reveals a previously unknown effect of glutamate on cholinergic synapses with potential functional and pharmacological implications.

Distinct presynaptic regulation of dopamine release through NMDA receptors in striosome- and matrix-enriched areas of the rat striatum

Striosome- and matrix-enriched striatal zones were defined in coronal and sagittal brain sections of the rat, on the basis of 3H-naloxone binding to mu-opiate receptors (a striosome-specific marker). Then, using a new in vitro microsuperfusion device, the NMDA (50 microM)- evoked release of newly synthesized 3H-dopamine (3H-DA) was examined in these four striatal areas under Mg(2+)-free conditions. The amplitudes of the responses were different in striosomal (171 +/- 6% and 161 +/- 5% of the spontaneous release) than in matrix areas (223 +/- 6% and 248 +/- 12%), even when glycine (1 or 100 microM) was coapplied (in the presence of 1 microM strychnine). In the four areas, the NMDA-evoked release of 3H-DA was blocked completely by Mg2+ (1 mM) or (+)-5-methyl- 10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine maleate (MK-801; 1 microM) and almost totally abolished by kynurenate (100 microM). Because the tetrodotoxin (TTX)-resistant NMDA-evoked release of 3H-DA was similar in striosome- (148 +/- 5% and 152 +/- 6%) or matrix- enriched (161 +/- 5% and 156 +/- 7%) areas, the indirect (TTX- sensitive) component of NMDA-evoked responses, which involves striatal neurons and/or afferent fibers, seems more important in the matrix- than in the striosome-enriched areas. The modulation of DA release by cortical glutamate and/or aspartate-containing inputs through NMDA receptors in the matrix appears thus to be partly distinct from that observed in the striosomes, providing some functional basis for the histochemical striatal heterogeneity.

Spatial organization of patch and matrix compartments in the rat striatum.

The visualization of mu opiate receptors by [3H]naloxone binding was used to determine precisely the spatial organization of the patch compartment in the rat striatum and its reproducibility in different animals. Three-dimensional reconstruction of the patch network was made using maps of autoradiographic data obtained from successive coronal, sagittal or horizontal sections. The extreme rostral pole of the striatum (A 11) was characterized by a large patch territory exhibiting complex and tortuous fields with several extensions. In the intermediate part of the structure (A 9.0-10.0), about 20 serial parallel continuous patch channels running in a mediolateral axis, obliquely oriented and displaying in some cases connecting branches, could be observed. However, no channels could be distinguished in the rostrocaudal direction. More caudally, patches were rare and of small size. In addition, the laterocaudal region of the striatum was almost exclusively represented by a large matrix field. Finally, a fine discontinuous band of [3H]naloxone binding was seen in all sections, bordering and limiting the dorsolateral part of the striatum. The topographical and spatial distribution of the patch compartment was similar in all animals investigated. However, due to the tortuous shape and the labyrinthine organization of the patches, the precise degree of reproducibility from one animal to another could not be established. Nevertheless, the prominent patch compartment observed in the rostral pole of the striatum, the patch channels, oriented in the mediolateral axis as well as the large laterocaudal matrix field were observed in all cases. These results were compared with previous data obtained in the cat in which patch (striosome) channels oriented along a rostrocaudal axis are also observed.

Three-dimensional organization of the striosomal compartment and patchy distribution of striatonigral projections in the matrix of the cat caudate nucleus

Acetylcholinesterase staining on successive frontal or sagittal sections was used to determine the three-dimensional organization of the striosomal and matrix compartments in the adult cat caudate nucleus. Reconstruction drawings of the acetylcholinesterase-poor zones (striosomes) indicated that the striosomal compartment is a labyrinthine network organized in the rostrocaudal and mediolateral axis which is reproducible from one animal to another. Four main anteroposterior channels converging in the mediorostral pole of the caudate nucleus were distinguished. Seven to eight diagonally oriented channels crossing the previous ones were seen also in the mediolateral axis on the central core of the caudate nucleus. The pattern of organization of the numerous and tortuous striosomal channels was more complicated medially, while the lateral part of the caudate nucleus was represented mainly by the matrix compartment. In addition, a sub-compartmentation of the matrix was demonstrated by retrograde tracing studies made by injecting either horseradish peroxidase-wheat germ agglutinin, [14C]amino acids or a mixture of horseradish peroxidase-wheat germ agglutinin and [14C]amino acids in several areas of the substantia nigra pars reticulata. Labelled patches were seen with both tracers, their topographical localization depended on the nigral injection site but reconstruction analysis indicated that the populations of cells which innervate the substantia nigra pars reticulata originate in the two third lateral parts of the caudate nucleus all along its rostrocaudal extension. Examination of horseradish peroxidase-wheat germ agglutinin labelled cells indicated that not all cells were labelled in patches suggesting a further sub-compartmentation of these patches. Finally, a comparison of the topographical distributions of labelled patches and of striosomes revealed that most patches were located in the extrastriosomal matrix.

A 25 year adventure in the field of tachykinins

Several aspects of our 25 year adventure in the field of tachykinins will be successively described. They concern: substance P (SP) synthesis and release in the basal ganglia, the identification and pharmacological characterization of central tachykinin NK(1), NK(2) and NK(3) binding sites and their topographical distribution, the description of some new biological tests for corresponding receptors, the identification of tachykinin NK(1) receptor subtypes or conformers sensitive to all endogenous tachykinins (substance P, neurokinin A (NKA), neurokinin B (NKB), neuropeptide gamma (NP gamma) and neuropeptide K (NPK)) and finally, the functional involvement of these receptors and their subtypes in tachykinin-induced regulations of dopamine and acetylcholine release in the striatum.

Functional mu opioid receptors are expressed in cholinergic interneurons of the rat dorsal striatum: territorial specificity and diurnal variation

Striatal cholinergic interneurons play a crucial role in the control of movement as well as in motivational and learning aspects of behaviour. Neuropeptides regulate striatal cholinergic transmission and particularly activation of mu opioid receptor (MOR) inhibits acetylcholine (ACh) release in the dorsal striatum. In the present study we investigated whether this cholinergic transmission could be modulated by an enkephalin/MOR direct process. We show that mRNA and protein of MORs are expressed by cholinergic interneurons in the limbic/prefrontal territory but not by those in the sensorimotor territory of the dorsal striatum. These MORs are functional because potassium‐evoked release of ACh from striatal synaptosomes was dose‐dependently reduced by a selective MOR agonist, this effect being suppressed by a MOR antagonist. The MOR regulation of cholinergic interneurons presented a diurnal variation. (i) The percentage of cholinergic interneurons containing MORs that was 32% at the beginning of the light period (morning) increased to 80% in the afternoon. (ii) The MOR‐mediated inhibition of synaptosomal ACh release was higher in the afternoon than in the morning. (iii) While preproenkephalin mRNA levels remained stable, enkephalin tissue content was the lowest (−32%) in the afternoon when the spontaneous (+35%) and the N‐methyl‐d‐aspartate‐evoked (+140%) releases of enkephalin (from microsuperfused slices) were the highest. Therefore, by acting on MORs present on cholinergic interneurons, endogenously released enkephalin reduces ACh release. This direct enkephalin/MOR regulation of cholinergic transmission that operates only in the limbic/prefrontal territory of the dorsal striatum might contribute to information processing in fronto‐cortico‐basal ganglia circuits.

Spontaneous and potassium-evoked release of 3H-GABA newly synthesized from 3H-glutamine in slices of the rat substantia nigra

Abstract A technique has been developed to measure 3 H-GABA not only in tissues but also in medium of slices of the rat substantia nigra (SN) incubated for 15 min with 3 H-glutamine. The quantity of 3 H-GABA in tissues was about 30 to 35 times that released in the medium. Nevertheless, the amount of the 3 H-transmitter spontaneously released was about 10 to 15 times the blank value. GAD activity in the SN was decreased by 40 and 80% respectively ten days after the kainic acid lesion of the ipsilateral striatum or hemitransection. These effects were associated with parallel reductions in the amounts of 3 H-GABA accumulated in tissues and released in medium. The spontaneous release of newly synthesized 3 H-GABA was increased in absence of calcium and reduced with an excess of calcium (10 −2 M). Tetrodotoxin (10 −5 , 5.10 −6 M) reduced by 40% the spontaneous release of 3 H-GABA. These various effects were not associated with significant change in the total accumulation of 3 H-GABA in tissues + medium. Finally depolarization of the slices with potassium (30 mM) increased the release of 3 H-GABA (300%). This effect was abolished in absence of calcium and was not associated with a significant change in the amount of 3 H-GABA accumulated in tissues.

Role of dynorphin-containing neurons in the presynaptic inhibitory control of the acetylcholine-evoked release of dopamine in the striosomes and the matrix of the cat caudate nucleus

The roles of acetylcholine and dynorphin (1-13) in the presynaptic control of the release of [3H]dopamine continuously synthesized from [3H]tyrosine were examined in a prominent striosomal enriched area and in an adjacent matrix enriched area of the cat caudate nucleus. This was achieved using microsuperfusion devices applied vertically onto coronal slices of cat brain. These devices were placed in a striosomal enriched area located in the core of the structure (acetylcholinesterase-poor zone) and in an adjacent matrix enriched area (acetylcholinesterase-rich zone). [3H]Tyrosine was delivered continuously to each microsuperfusion device and [3H]dopamine released was estimated in the superfusate. As previously shown, in the presence of tetrodotoxin (1 microM), acetylcholine (50 microM) induces a prolonged stimulation of [3H]dopamine release in both compartments through an interaction with muscarinic receptors. Our present study indicates that both dynorphin 1-13 (1 microM) and the selective kappa agonist trans-3,4-dichloro-N-methyl-N[2-(1-pyrrolidinyl)cyclohexyl]benzeneace tamine (U50488) (1 microM) inhibit the tetrodotoxin-resistant acetylcholine-evoked release of [3H]dopamine, these effects being slightly more pronounced in the matrix than in the striosomal enriched area. Naloxone (1 microM) reversed the inhibitory effect of U50488 in both areas. These results suggest that dynorphin exerts an inhibitory presynaptic control of dopamine release through kappa opioid receptors located on dopamine nerve terminals in the striosome as well as in the matrix. However, the presynaptic cholinergic control of dopamine release is much more complex in the matrix than in the striosomal enriched area. Besides its tetrodotoxin-resistant stimulatory effect, acetylcholine exerts two opposing tetrodotoxin-sensitive effects on [3H]dopamine release, one facilitatory and the other inhibitory. We demonstrate here that in the superfused matrix enriched area, the indirect acetylcholine inhibitory response is mediated by dynorphin-containing neurons. Indeed, the short-lasting stimulatory effect of acetylcholine on [3H]dopamine release was converted into a long-lasting response in the presence of naloxone (1 microM), and, in this latter condition, the co-application of dynorphin 1-13 (1 microM) restored the short-lasting stimulatory effect.