sr-142948a and levocabastine

Recent evidence suggests that some types of neurotensin receptors may be expressed by astrocytes. In order to explore the function of neurotensin receptors in astrocytes, the effect of a neurotensin receptor agonist, neurotensin(8-13), on intracellular Ca(2+) dynamics in mixed neuronal/glial cultures prepared from rat ventral tegmental area was examined. It was found that neurotensin(8-13) induces a long-lasting rise in intracellular Ca(2+) concentration in a subset of glial fibrilary acidic protein-positive glial cells. This response displays extensive desensitization and appears to implicate both intracellular and extracellular Ca(2+) sources. In the absence of extracellular Ca(2+), neurotensin(8-13) evokes only a short-lasting rise in intracellular Ca(2+). The neurotensin-evoked intracellular Ca(2+) accumulation is blocked by the phospholipase C inhibitor U73122 and by thapsigargin, suggesting that it is initiated by release of Ca(2+) from an inositol triphosphate-dependent store. The Ca(2+)-mobilizing action of neurotensin(8-13) in astrocytes is dependent on at least two receptors, because the response is blocked in part only by SR48692, a type 1 neurotensin receptor antagonist, and is blocked completely by SR142948A, a novel neurotensin receptor antagonist. The finding that the type 2 neurotensin receptor agonist levocabastine fails to mimic or alter the effects of neurotensin(8-13) on intracellular Ca(2+) makes it unlikely that the type 2 neurotensin receptor is involved. In summary, these results show that functional neurotensin receptors are present in cultured ventral tegmental area astrocytes and that their activation induces a highly desensitizing rise in intracellular Ca(2+). The pharmacological profile of this response suggests that a type 1 neurotensin receptor is involved but that another, possibly novel, non-type 2 neurotensin receptor is also implicated. If present in vivo, such signalling could be involved in some of the physiological actions of neurotensin.

Topics: Adamantane; Animals; Animals, Newborn; Astrocytes; Calcium; Cells, Cultured; Estrenes; Gadolinium; Imidazoles; Neurotensin; Peptide Fragments; Piperidines; Pyrrolidinones; Rats; Receptors, Neurotensin; Thapsigargin; Type C Phospholipases; Ventral Tegmental Area

The effect of denervation with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) of the dopamine (DA) nigrostriatal pathway on neurotensin (NT) receptor and DA transporter (DAT) in basal ganglia of monkeys (Macaca fascicularis) was investigated. The MPTP lesion induced a marked depletion of DA (90% or more vs. control) in the caudate nucleus and putamen. The densities of NT agonist binding sites labeled with [125I]NT and the NT antagonist binding sites labeled with [3H]SR142948A decreased by half in the caudate-putamen of MPTP-monkeys. In addition, the densities of [125I]NT and [3H]SR142948A binding sites markedly decreased (-77 and -63%, respectively) in the substantia nigra of MPTP-monkeys. Levocabastine did not compete with high affinity for [125I]NT binding in the monkey cingulate cortex, suggesting that only one class of NT receptors was labelled in the monkey brain. An extensive decrease of [3H]GBR12935 DAT binding sites (-92% vs. Control) was observed in the striatum of MPTP-monkeys and an important loss of DAT mRNA(-86% vs. Control) was observed in substantia nigra. Treatments for 1 month with either the D1 agonist SKF-82958 (3 mg/kg/day) or the D2 agonist cabergoline (0.25 mg/kg/day) had no effect on the lesion-induced decrease in NT and DAT binding sites or DAT mRNA levels. The decrease of striatal NT binding sites was less than expected from the decrease of DA content in this nucleus, suggesting only partial localization of NT receptors on nigrostriatal DAergic projections. These data also suggest that under severe DA denervation, treatment with D1 or D2 DA agonists does not modulate NT receptors and DAT density.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Adamantane; Animals; Autoradiography; Binding Sites; Carrier Proteins; Caudate Nucleus; Denervation; Dopamine; Dopamine Agents; Dopamine Plasma Membrane Transport Proteins; Dose-Response Relationship, Drug; Down-Regulation; Imidazoles; In Situ Hybridization; Macaca fascicularis; Membrane Glycoproteins; Membrane Transport Proteins; Neostriatum; Nerve Tissue Proteins; Neurotensin; Piperidines; Putamen; Receptors, Dopamine; Receptors, Neurotensin; RNA, Messenger; Substantia Nigra

Identified cholinergic and a subtype of non-cholinergic, fast-firing neurons were recorded intracellularly in vitro from slices of guinea-pig brain. Recorded neurons were within the boundaries of the medial septum and vertical limb of the diagonal band of the forebrain. The effects of superfused neurotensin and neurotensin receptor antagonists were measured under single-electrode current clamp. Neurotensin consistently caused a dose-dependent, slow depolarization of cholinergic neurons that was accompanied by an increase in membrane resistance and a block of the long-duration (1-10 s) post-spike afterhyperpolarization when present. Neurotensin also blocked a shorter duration, slow afterhyperpolarization, but only in a minority of cholinergic neurons. When present, inhibition of the slow afterhyperpolarization changed the spike pattern from single spikes to short bursts. Inhibition of post-spike afterhyperpolarizations by neurotensin reversed more slowly than did other effects of neurotensin. Tetrodotoxin did not prevent the depolarizing effect of neurotensin. The non-selective neurotensin receptor antagonist, SR142948A, blocked the depolarizing effect of neurotensin but the low-affinity receptor antagonist, levocabastine, did not. A subgroup of noncholinergic, fast-firing neurons (23%) was also depolarized by neurotensin, an effect antagonized by SR142948A but not levocabastine. Neurotensin did not effect post-spike voltage transients or change the firing pattern of non-cholinergic neurons. These data suggest that neurotensin causes a slow depolarization and increased excitability of cholinergic and some noncholinergic neurons in an area of the brain that projects to the hippocampus. Neurotensin type 1 receptors appear to mediate these effects. Neurotensin may modulate hippocampal-dependent learning and memory processes through its effects on septohippocampal neurons.

Topics: Action Potentials; Adamantane; Animals; Dose-Response Relationship, Drug; Female; Guinea Pigs; Histamine H1 Antagonists; Imidazoles; In Vitro Techniques; Male; Membrane Potentials; Neurons; Neurotensin; Patch-Clamp Techniques; Piperidines; Prosencephalon; Receptors, Neurotensin; Tetrodotoxin

The present study describes the characterization of the binding properties and autoradiographic distribution of a new nonpeptide antagonist of neurotensin receptors, [3H]SR 142948A (2-[[5-(2,6-dimethoxyphenyl)-1-(4-(N-(3-dimethylaminopropyl)-N-methyl carbamoyl)-2-isopropylphenyl)-1H-pyrazole-3-carbonyl]-amino]-ad amantane-2-carboxylic acid, hydrochloride), in the rat brain. The binding of [3H]SR 142948A in brain membrane homogenates was specific, time-dependent, reversible and saturable. [3H]SR 142948A bound to an apparently homogeneous population of sites, with a Kd of 3.5 nM and a Bmax value of 508 fmol/mg of protein, which was 80% higher than that observed in saturation experiments with [3H]neurotensin. [3H]SR 142948A binding was inhibited by SR 142948A, the related nonpeptide receptor antagonist, SR 48692 (2-[[1-(7-chloroquinolin-4-yl)-5-(2,6-dimethoxyphenyl)-1H-pyrazole -3-carbonyl]amino]-adamantane-2-carboxylic acid) and neurotensin. Saturation and competition studies in the presence or absence of the histamine H1 receptor antagonist, levocabastine, revealed that [3H]SR 142948A bound with similar affinities to both the levocabastine-insensitive neurotensin NT1 receptors (20% of the total binding population) and the recently cloned levocabastine-sensitive neurotensin NT2 receptors (80% of the receptors) (Kd = 6.8 and 4.8 nM, respectively). The regional distribution of [3H]SR 142948A binding in the rat brain closely matched the distribution of [125I]neurotensin binding. In conclusion, these findings indicate that [3H]SR 142948A is a new potent antagonist radioligand which recognizes with high affinity both neurotensin NT1 and NT2 receptors and represents thus an excellent tool to study neurotensin receptors in the rat brain.

Topics: Adamantane; Animals; Autoradiography; Binding Sites; Brain; Imidazoles; Male; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, Neurotensin; Tritium