Maria Stefania Mascia

Microdialysis measurement of cortical and hippocampal acetylcholine release during sleep-wake cycle in freely moving cats

The variations of Acetylcholine (ACh) release in the cerebral cortex and dorsal hippocampus were monitored by microdialysis during the electroencephalographically recorded sleep-waking cycle in freely moving cats. The results show a state-dependent variation in ACh output in both the cortex and the hippocampus. ACh release increased by approximately 100% during quiet waking (QW) and by 175% during active waking (AW) as referred to slow wave sleep (SWS) baseline. In contrast, a clear difference between the two areas was observed during REM sleep. During this stage ACh release in the cortex reached approximately the same values observed during QW, while in the hippocampus ACh release rose to about 4-fold the level obtained during SWS or twice that of QW. The results support the idea that the increase in ACh release in the cortex reflects the desynchronized EEG of wakefulness and REM sleep, while the marked increase of ACh during REM in the hippocampus may be related to the sustained theta activity in this area.

Cannabinoids decrease acetylcholine release in the medial-prefrontal cortex and hippocampus, reversal by SR 141716A

The effect of delta9-tetrahydrocannabinol, the psychoactive principle of marijuana, and [R-(+)-(2,3-dihydro-5-methyl-3-[[4-morpholinylmethyl]pyrol[1,2,3-d e-]-1,4-benzoxazin-6y)(1-naphthalenyl)methanone monomethanesulfonate] (WIN 55,212-2), a synthetic cannabinoid receptor agonist, on the acetylcholine output in the medial-prefrontal cortex and hippocampus was studied by microdialysis in freely moving rats. The administration of delta9-tetrahydrocannabinol (1 and 5 mg/kg i.p.) and WIN 55,212-2 (5 and 10 mg/kg i.p.) produced a long lasting inhibition of acetylcholine release in both areas. The inhibitory effect of delta9-tetrahydrocannabinol and WIN 55,212-2 was suppressed in both areas by the specific cannabinoid CB1 receptor antagonist, [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-me thyl-1H-pyrazole-3carboxamide]HCl (SR 141716A), at the dose of 0.1 mg/kg i.p., per se ineffective to modify basal acetylcholine release. Most interestingly, SR 141716A alone at higher doses increased acetylcholine release both in the medial-prefrontal cortex (3 mg/kg i.p.) and hippocampus (1 and 3 mg/kg i.p.), suggesting that acetylcholine output is tonically inhibited by endogenous cannabinoids. Since the inhibitory effect of delta9-tetrahydrocannabinol is produced by doses within those relevant to human use of marijuana, our results suggest that the negative effects of the latter on cognitive processes may be explained by its ability to reduce acetylcholine release in the medial-prefrontal cortex and hippocampus. Conversely, cannabinoid receptor antagonists may offer potential treatments for cognitive deficits.

Lack of morphine-induced dopamine release in the nucleus accumbens of cannabinoid CB(1) receptor knockout mice

Morphine (10 and 20 mg/kg, s.c.) does not modify dopamine release in the nucleus accumbens of cannabinoid CB(1) knock-out mice under conditions where it dose-dependently stimulates the release of dopamine in the corresponding wild-type mice. These results demonstrate that cannabinoid CB(1) receptors, regulate mesolimbic dopaminergic transmission in brain areas known to be involved in the reinforcing effects of morphine.

Inhibition of hippocampal acetylcholine release by cannabinoids: reversal by SR 141716A

Two synthetic cannabinoids, WIN 55,212-2 {R-(+)-(2,3-dihydro-5-methyl-3-[{4-morpholinylmethyl]pyrol [1,2,3-de]-1,4-benzoxazin-6-yl)(1-naphthalenyl)methanone monomethanesulfonate} (5.0 and 10 mg/kg i.p.) and CP 55,940 {[1a,2-(R)-5-(1.1-dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxypropyl) cyclohexyl]-phenol} {[1a,2-(R)-5-(1,1-dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxypropyl) cyclohexyl]-phenol} (0.5 and 1.0 mg/kg i.p.), inhibited acetylcholine release in the rat hippocampus. The inhibition was prevented by the cannabinoid receptor antagonist, SR 141716A {N-(piperidin-1-yl)-5-(4- chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide} HCl, at the dose of 0.1 mg/kg i.p. Higher doses of SR 141716A (1.0 and 3.0 mg/kg i.p.) themselves increased hippocampal acetylcholine release, suggesting that acetylcholine output is tonically inhibited by endogenous cannabinoids. The results also suggest that the negative effects of marijuana on learning and memory may depend on cannabinoid receptor-mediated inhibition of acetylcholine release.

Extra-cellular dopamine increases in the paraventricular nucleus of male rats during sexual activity

Dopamine and 3,4‐dihydroxyphenylacetic acid (DOPAC) concentrations were measured in the dialysate obtained with vertical microdialysis probes implanted into the paraventricular nucleus of the hypothalamus of sexually potent male rats. Animals showed noncontact erections when put in the presence of, and copulated with a receptive (ovarietomized oestrogen and progesterone primed) female rat. Dopamine and DOPAC concentrations in the paraventricular dialysate increased 140% and 19%, respectively, above baseline values during exposure to the receptive female and 280% and 31%, respectively, during copulation. No changes in dopamine and DOPAC concentrations were detected in the paraventricular dialysate when sexually potent male rats were exposed to nonreceptive (ovariectomized not oestrogen plus progesterone primed) female rats. These results confirm the involvement of the paraventricular nucleus in control of erectile function and copulatory behaviour and show for the first time that dopamine neurotransmission is increased in this hypothalamic nucleus when erection occurs in physiological contexts.

PD-168077, a selective dopamine D4 receptor agonist, induces penile erection when injected into the paraventricular nucleus of male rats

The effect of PD-168077 (N-methyl-4-(2-cyanophenyl)piperazynil-3-methylbenzamide maleate), a selective D4 dopamine receptor agonist, injected into the paraventricular nucleus of the hypothalamus on penile erection was studied in male rats. PD-168077 (1-200 ng) induced penile erection in a dose-dependent manner. The minimal effective dose was 50 ng, while the maximal response was found with 200 ng of the compound, which increased penile erection episodes from 0.3+/-0.03 to 1.7+/-0.21. The proerectile effect of PD-168077 was reduced almost completely by L-745,870 (3-(4-[chlorophenyl]piperazin-1-yl)-methyl-1H-pyrrolo[2,3-B]pyridine trihydrochloride), a selective D4 dopamine receptor antagonist, (1 microg) given into the paraventricular nucleus before the D4 dopamine agonist, and by other nonselective dopamine receptor antagonists, such as haloperidol (1 microg) and clozapine (1 microg), which block all dopamine receptor subtypes. The pro-erectile effect of PD-168077 was also reduced by the NO synthase inhibitor NG-nitro-L-arginine methylester (25 microg), but not by the oxytocin receptor antagonist d(CH2)5Tyr(Me)2-Orn8-vasotocin (1 microg), when given into the paraventricular nucleus. In spite of its inability to prevent the pro-erectile effect of PD-168077 when given in the paraventricular nucleus, d(CH2)5Tyr(Me)2-Orn8-vasotocin (1 microg) reduced almost completely PD-168077-induced penile erection when given into the lateral ventricles. The present results show that D4 dopamine receptors present in the paraventricular nucleus may influence penile erection by modulating the activity of paraventricular oxytocinergic neurons mediating erectile function.

Antagonism of cannabinoid CB1 receptors in the paraventricular nucleus of male rats induces penile erection

The effect of cannabinoid CB1 receptor agonists and antagonists on penile erection was studied in male rats when injected into the paraventricular nucleus of the hypothalamus. The CB1 receptor antagonist SR 141716A [N-(piperidin-1-yl)-5-(4-chlorophenyl)-4-methyl-1H-pyrazole-3-carboxyamide] (0.5-5 microg) induced penile erection in a dose-dependent manner. The minimal effective dose was 1 microg, while the maximal response was found with 5 microg of the compound. In contrast, the CB1 receptor agonists WIN 55,212-2 [4,5-dihydro-2-methyl-4(4-morpholinylmethyl)-1-(1-naphthalenyl-carbonyl)-6H-pyrrolo[3,2,1-I,j]quinolin-6-one] (0.5-5 microg) and CP 55,940 [1alpha,2beta-(R)-5alpha]-5-(1,1-dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxy-propyl)cyclohexyl]phenol (0.5-5 microg) were ineffective at all the doses tested. Nevertheless, both compounds reduced the enhancing effect of SR 141716A on penile erection when given into the paraventricular nucleus at the above doses before SR 141716A. The pro-erectile effect of SR 141716A was also reduced by the non-competitive NMDA receptor antagonist dizolcipine (MK-801) (0.2 microg) and by the NO synthase inhibitor NG-nitro-l-arginine methylester (L-NAME) (20 microg) but not by the dopamine receptor antagonist cis-flupenthixol (10 microg) or the oxytocin receptor antagonist d(CH2)5Tyr(Me)2-Orn8-vasotocin (0.1 microg), when given into the paraventricular nucleus. In spite of its inability to prevent the pro-erectile effect of SR 141716A when given in the paraventricular nucleus, d(CH2)5Tyr(Me)2-Orn8-vasotocin) (1 microg) reduced almost completely SR 141716A-induced penile erection when given into the lateral ventricles. The present results show that cannabinoid CB1 receptors present in the paraventricular nucleus may influence erectile function and sexual activity by modulating paraventricular oxytocinergic neurons mediating erectile function.

Reduction of dopamine release and synthesis by repeated amphetamine treatment : Role in behavioral sensitization

Changes in extracellular dopamine concentration in the ventral striatum during repeated amphetamine administration and over the first 7 days of withdrawal were studied by transversal microdialysis in freely moving rats. 2 days after fiber implantation rats were treated with either amphetamine (1.5 mg/kg i.p.) or saline every 12 h for 14 days. In amphetamine-treated rats, the baseline extracellular dopamine concentration, preceding the morning treatment, increased from 0.43 +/- 0.01 on day 1 up to 0.59 +/- 0.02 pmol/40 microliters sample on day 3 of treatment. Thereafter, dopamine fell rapidly on day 5(0.16 +/- 0.01 pmol/40 microliters) and remained at approximately the level reached on day 7(0.11 +/- 0.01 pmol/40 microliters) throughout the treatment and also over the 7 days of withdrawal. In contrast, in control rats, the extracellular dopamine concentration (0.40 +/- 0.01 pmol/40 microliters, on day 1) decreased progressively during the first days of treatment to reach a fairly stable value on day 4 (0.25 +/- 0.01 pmol/40 microliters sample). Thereafter, dopamine remained stable at this level throughout the remaining period of experimentation. Challenge with amphetamine (1.5 mg/kg i.p.) of animals treated with amphetamine for 10 days or withdrawn for 7 days produced a potentiated motor response compared to that in control rats but much less marked dopamine releasing effects. Dopamine synthesis in the ventral striatum, measured as L-dihydroxyphenylalanine formation after blockade of dihydroxyphenylalanine decarboxylase, was found to be reduced by approximately 60% after 2 weeks of amphetamine treatment and in animals withdrawn for 1 day or 7 days. These results indicate that repeated amphetamine treatment causes persistent inhibition of dopamine synthesis and release in the ventral striatum. Such inhibition may be a compensatory response to the repeated stimulation of postsynaptic dopamine receptors by the endogenously released dopamine and also the cause of postsynaptic sensitization to dopamine action.

Ghrelin injected into the paraventricular nucleus of the hypothalamus of male rats induces feeding but not penile erection

The effect of ghrelin, a recently characterized endogenous receptor agonist for growth hormone (GH) secretagogue receptors, on feeding and penile erection was compared with that of EP 80661, a peptide analogue of the GH secretagogue hexarelin, previously identified for its pro-erectile activity when injected into the paraventricular nucleus of the hypothalamus of male rats. Ghrelin (0.01-1 microg), but not EP 80661 (0.02-1 microg), was found to be particularly effective in enhancing feeding. The minimal effective dose of ghrelin was 0.1 microg, which increased food intake by 88%, while the maximal response (355% above control values) was found with 1 microg of the peptide. The enhancing effect of ghrelin on feeding was prevented by the prior administration of the neuropeptide Y Y5 receptor antagonist (DTyr(2), DThr(32)) neuropeptide Y (NPY, 10 microg), but not by the GH-RH receptor antagonist MZ-4-71 (10 microg), or by EP 91073, a hexarelin analogue that antagonizes the pro-erectile effect of EP 80661 (10 microg), given into the lateral ventricles. In contrast, ghrelin failed to induce penile erection at all doses tested, while EP 80661 induced penile erection in a dose-dependent manner. The pro-erectile effect of EP 80661 was prevented by EP 91073 (10 microg), but not by (DTyr(2), DThr(32)) NPY (10 microg) or by the GH-RH receptor antagonist MZ 4-71 (10 microg), given into the lateral ventricles. The present results provide further support to the hypothesis that the GH secretagogue receptors mediating feeding are different from those mediating penile erection and activated by pro-erectile EP peptides.

Opposite effects of stress on dopamine release in the limbic system of drug-naive and chronically amphetamine-treated rats

The effects of repeated amphetamine administration on stress-induced dopamine release in the ventral striatum were examined in male Wistar rats treated with D-amphetamine (1.5 mg/kg; i.p./injection) or saline at 12 h intervals for 14 days. After 12 h as well as 7 days of amphetamine withdrawal, dopamine release was monitored by transverse microdialysis under basal conditions and during exposure to 60 min of restraint stress. Basal dopamine release was significantly suppressed relative to saline-pretreated controls after both 12 h and 7 days of amphetamine withdrawal. In control rats, restraint stress resulted in significantly increased dopamine efflux. In contrast, exposure to this stressor was associated with a significant suppression of dopamine release in rats chronically exposed to amphetamine. This effect was observed at both post-amphetamine test points. The results suggest that chronic amphetamine impairs the dopaminergic response to stress and that this dopaminergic deficit may play a role in stress-induced drug-seeking behavior and relapse.