Ariane Hornick

The coumarin scopoletin potentiates acetylcholine release from synaptosomes, amplifies hippocampal long-term potentiation and ameliorates anticholinergic- and age-impaired memory.

In a previous study the simple, naturally derived coumarin scopoletin (SCT) was identified as an inhibitor of acetylcholinesterase (AChE), using a pharmacophore-based virtual screening approach. In this study the potential of SCT as procholinergic and cognition-enhancing therapeutic was investigated in a more detailed way, using different experimental approaches like measuring newly synthesized acetylcholine (ACh) in synaptosomes, long-term potentiation (LTP) experiments in hippocampal slices, and behavior studies. SCT enhanced the K+-stimulated release of ACh from rat frontal cortex synaptosomes, showing a bell-shaped dose effect curve (Emax: 4 μM). This effect was blocked by the nicotinic ACh receptor (nAChR) antagonists mecamylamine (MEC) and dihydro-β-erythroidine (DHE). The nAChR agonist (and AChE inhibitor) galantamine induced a similar increase in ACh release (Emax: 1 μM). SCT potentiated LTP in hippocampal slices of rat brain. The high-frequency stimulation (HFS)-induced, N-methyl-D-aspartate (NMDA) receptor dependent LTP of field excitatory postsynaptic potentials at CA3-CA1 synapses was greatly enhanced by pre-HFS application of SCT (4 μM for 4 min). This effect was mimicked by nicotine (2 μM) and abolished by MEC, suggesting an effect on nAChRs. SCT did not restore the total inhibition of LTP by NMDA receptor antagonist d, l-2-amino-5-phosphonopentanoic acid (AP-5). SCT (2 μg, i.c.v.) increased T-maze alternation and ameliorated novel object recognition of mice with scopolamine-induced cholinergic deficit. It also reduced age-associated deficits in object memory of 15–18-month-old mice (2 mg/kg sc). Our findings suggest that SCT possesses memory-improving properties, which are based on its direct nAChR agonistic activity. Therefore, SCT might be able to rescue impaired cholinergic functions by enhancing nAChR-mediated release of neurotransmitters and promoting neural plasticity in hippocampus.

Effects of the coumarin scopoletin on learning and memory, on release of acetylcholine from brain synaptosomes and on long-term potentiation in hippocampus

Background Among the chemical class of coumarins several substances have been described to be effective as cognition enhancers. We have recently characterized the coumarin scopoletin as a compound which fits a pharmacophore model of AChE inhibitors and enhances the release of ACh in rat brain [1]. Now, a comprehensive study was carried out in order to investigate the effects of scopoletin on learning and memory, on release of ACh from synaptosomes and on hippocampal long-term potentiation (LTP) in order to uncover the mechanism of action.

Antagonist of M1 muscarinic acetylcholine receptor prevents neurotoxicity induced by amphetamine via nitric oxide pathway.

The psychostimulant amphetamine (AMPH) has been found to induce striatal acetylcholine release and neurotoxic processes via nitric oxide (NO) and lipid peroxidation (LPO). Our purpose was to determine whether blocking striatal muscarinic (M1) receptors by the selective M1 antagonist toxin 7 (MT 7; bilaterally, 2 μg per side) might attenuate the effects of AMPH (4 × 5 mg/kg, i.p.). Systemic AMPH administration increased NO and LPO in the striatal tissue. Stimulation of M1 receptors by i.c.v. injection of M1 agonist McN‐A‐343 (200 μg) caused a similar enhancement of NO and LPO. Pretreatment with the MT7 prevented the AMPH‐induced NO generation and greatly reduced the LPO caused by the psychostimulant. These results show that M1 acetylcholine receptors are critically involved in neurotoxic processes induced by AMPH via NO and LPO.

7-nitroindazole, nNOS inhibitor, attenuates amphetamine-induced amino acid release and nitric oxide generation but not lipid peroxidation in the rat brain

Summary.The aim of the present study was to elucidate whether amphetamine modulates the output of the neurotransmitters glutamate, aspartate, GABA and acetylcholine (ACh) in nucleus accumbens (NAc) as well as the formation of lipid peroxidation (LPO) and nitric oxide (NO). D,L-amphetamine (AMPH, 5 mg/kg, i.p., 4 times every 2 h) was injected into anaesthetized rats and the release of neurotransmitters in the NAc, tissue content of NO and LPO products were determined.While AMPH increased the release of aspartate, GABA and ACh in the NAc, the glutamate release was not affected. Levels of NO and LPO products were elevated in striatum and cortex. Pretreatment with the neuronal NO synthase inhibitor 7-nitroindazole (50 mg/kg, i.p.) was highly effective in abating the rise of the neurotransmitter release and NO generation but failed to influence the intensity of LPO elicited by the AMPH administration.These findings suggest that activation of NO synthesis is a potent factor in the AMPH-induced neurotransmitter release and that activation of NO synthesis and LPO by AMPH are not parallel processes.

Origin of endogenous nitric oxide released in the nucleus accumbens under real-time in vivo conditions.

AIMS Nitric oxide (NO), is a simple but multifarious molecule. It is implicated in physiological and pathological processes within the striatum, mainly in the nucleus accumbens (NAc). The aim of the present study was to determine the origin of NO in the NAc of anaesthetized rats by applying various compounds known to modulate the release of NO when applied either systemically or locally. MAIN METHODS Real-time monitoring of NO was carried out by introducing an amperometric NO sensor into the outer tubing of a push-pull cannula. For local application of substances, the push-pull superfusion technique was used. KEY FINDINGS An overdose of urethane (i.p.) or superfusion of the NAc with tetrodotoxin (TTX) led to a fall of NO release in the NAc. The NO synthase (NOS) inhibitors 7-nitroindazolmonosodiumsalt (7-NINA, neuronal NOS selective) and N-nitro-L-arginine (L-NNA, NOS selective) decreased release of NO when applied i.p. or locally. Superfusion of the NAc with N-methyl-D-aspartate (NMDA) elicited a dose dependent increase of NO release. SIGNIFICANCE Combination of an amperometric NO sensor for real-time monitoring of NO release with the push-pull superfusion technique showed that NO released in the NAc is, at least to a great extent, of neuronal origin. The enhanced release of NO elicited by locally applied NMDA demonstrates that activation of NMDA receptors facilitates NO synthesis, thus underlining the functionality of NO targets within the NAc.

Comparative effects of NO-synthase inhibitor and NMDA antagonist on generation of nitric oxide and release of amino acids and acetylcholine in the rat brain elicited by amphetamine neurotoxicity

Abstract: The aim of this study was to clarify the role of nitric oxide (NO) and lipid peroxidation (LPO) processes as well as the contribution of various neurotransmitters in pathophysiological mechanisms of neurotoxicity induced by amphetamine (AMPH). NO level was determined directly in brain tissues using electron paramagnetic resonance spectroscopy technique. The content of the products of lipid peroxidation (LPO) was measured spectrophotometrically as thiobarbituric acid reactive species (TBARS). The output of neurotransmitter amino acids (glutamate, aspartate, and GABA) and acetylcholine (ACH) was monitored in nucleus accumbens (NAc) by push‐pull technique with HPLC detection. Repeated, systemic application of AMPH elevated striatal and cortical NO generation and LPO production. Moreover, administration of AMPH led to a marked and long‐lasting increase of ACH release. Surprisingly, while glutamate output was not affected, aspartate release was enhanced 30 to 50 min after each AMPH injection. The release rate of GABA was also elevated. The selective NO‐synthase inhibitor 7‐nitroindazole (7‐NI) was highly effective in abating the rise in the neurotransmitter release induced by the AMPH. The NOS inhibitor also abolished the increase of NO generation produced by AMPH, but did not influence the intensity of LPO elicited by the AMPH administration. Pretreatment with the noncompetitive NMDA receptor antagonist dizocilpine (MK‐801) completely prevented increase of NO generation and TBARS formation induced by multiple doses of AMPH. Dizocilpine also abolished the effect of the psychostimulant drug on the release of neurotransmitters ACH, glutamate, aspartate, and GABA in the NAc. Our findings suggest a key role of NO in AMPH‐induced transmitter release, but not in the formation of LPO products. It appears that AMPH enhances release of ACH and neurotransmitter amino acids through increased NO synthesis and induces neurotoxicity via NO and also by NO‐independent LPO.