Modifications in DARPP-32 phosphorylation pattern after repeated palatable food consumption undergo rapid habituation in the nucleus accumbens shell of non-food-deprived rats.

In non-food-deprived rats a palatable meal induces a transient increase in dopamine output in the prefrontal cortex and nucleus accumbens shell and core; habituation to this response develops with a second palatable meal, selectively in the shell, unless animals are food-deprived. A palatable meal also induces time-dependent modifications in the dopamine and cAMP-regulated phosphoprotein of Mr 32 000 (DARPP-32) phosphorylation pattern that are prevented when SCH 23390, a selective dopamine D(1) receptor antagonist, is administered shortly after the meal. This study investigated whether dopaminergic habituation in the shell had a counterpart in DARPP-32 phosphorylation changes. In non-food-deprived rats, two consecutive palatable meals were followed by similar sequences of modifications in DARPP-32 phosphorylation levels in the prefrontal cortex and nucleus accumbens core, while changes after the second meal were blunted in the shell. In food-deprived rats two consecutive meals also induced similar phosphorylation changes in the shell. Finally, SCH 23390 administered shortly after the first palatable meal in non-food-deprived rats inhibited DARPP-32 phosphorylation changes in response to the first meal, and prevented the habituation to a second meal in terms of dopaminergic response and DARPP-32 phosphorylation changes. Thus, dopamine D(1) receptor stimulation plays a role in the development of habituation.

Long-term outcomes of treatment-resistant heroin addicts with and without DSM-IV axis I psychiatric comorbidity (dual diagnosis).

OBJECTIVE: The aim of this study was to compare the long-term outcomes of treatment-resistant heroin addicts with and without DSM-IV axis I psychiatric comorbidity (dual diagnosis). METHOD: 129 heroin addicts who also met criteria for treatment resistance, 66 with one or more DSM-IV axis I psychiatric diagnosis (DD patients), and 63 without DSM-IV axis I psychiatric comorbidity (NDD patients) were monitored prospectively (6 years on average, min. 1, max. 9) along a methadone maintenance treatment program (MMTP). RESULTS: The rates for survival-in-treatment were about 50% for NDD patients and about 70% for DD patients. After 4 years of treatment onwards, such rates tended to become stable. DD patients showed better outcome measures than NDD patients. A significantly higher methadone dose was needed to have DD patients stabilized. CONCLUSIONS: Contrary to expectations, treatment-resistant patients with psychiatric comorbidity showed a better long-term outcome than treatment-resistant patients without psychiatric comorbidity.

Behavioral expression of cocaine sensitization in rats is accompanied by a distinct pattern of modifications in the PKA/DARPP-32 signaling pathway.

Repeated cocaine administration induces behavioral sensitization and modifications in the phosphorylation pattern of dopamine and cAMP-regulated phosphoprotein of Mr 32,000 (DARPP-32), characterized by a tonic increase in the Thr75 phosphorylated form, and a decrease in the Thr34 phosphorylated form. This study further investigated the correlations between cocaine sensitization and modifications in the DARPP-32 phosphorylation pattern, cAMP-dependent protein kinase (PKA) activity, and mGluR5 tone in the medial prefrontal cortex and nucleus accumbens. Behavioral sensitization and modifications in these neurochemical markers followed a similar temporal pattern. Moreover, in sensitized rats acute cocaine administration modified phosphorylation levels of Thr75- and Thr34-DARPP-32, GluR1, and NR1 subunits in the nucleus accumbens only at a dose double the efficacious dose in control rats. These results suggest that the high levels of phospho-Thr75 DARPP-32 maintain PKA in a prevalent inhibited state. Furthermore, in sensitized rats the acute administration of 6-methyl-2-(phenylethynyl)-pyridine, a mGluR5 antagonist, reinstated the phosphorylation levels of Thr75- and Thr34-DARPP-32, GluR1, and NR1 to control values, and a subsequent cocaine challenge did not elicit a sensitized response. These data suggest that a tonic increase in mGluR5 transmission in cocaine-sensitized rats sustains both the increase in phospho-Thr75 DARPP-32 levels and the expression of behavioral sensitization.

Study of mirtazapine antidepressant effects in rats.

Mirtazapine is a widely used antidepressant and the aim of this study was to further investigate its antidepressant activity in rats. Thus, the efficacy of long-term mirtazapine treatment was assessed in three models of depressive symptoms induced by stress exposure: the acute escape deficit, the chronic escape deficit, and the stress-induced disruption of the acquisition of an appetitive behaviour sustained by a palatable food (vanilla sugar). Administration of mirtazapine for 2 wk prevented the escape deficit development induced by acute exposure to unavoidable stress. This protective effect was antagonized by the administration of a beta-adrenergic or a 5-HT1A receptor antagonist just before stress exposure; that is, mirtazapine effect was dependent on functional beta-adrenergic and 5-HT1A receptor systems. Repeated stress exposure indefinitely prolongs the condition of escape deficit and a 40-d mirtazapine treatment reversed this model of chronic escape deficit. In a Y-maze satiated rats learn to choose the arm baited with vanilla sugar, and exposure to stress during Y-maze training prevents this learning. Repeated mirtazapine administration completely antagonized the disrupting effect of chronic stress on the acquisition of this instrumental behaviour. We consider these effects to be crucial in the definition of antidepressant activity.

The mesolimbic dopaminergic response to novel palatable food consumption increases dopamine-D1 receptor-mediated signalling with complex modifications of the DARPP-32 phosphorylation pattern.

Acute cocaine administration increases extraneuronal dopamine and Thr34 phosphorylation of dopamine- and cAMP-regulated phosphoprotein (M(r) 32 kDa; DARPP-32) in striatal and cortical areas. Novel palatable food consumption increases extraneuronal dopamine in the same areas. We examined the DARPP-32 phosphorylation pattern in food non-deprived rats at different times after vanilla sugar consumption. The phosphorylation state of DARPP-32 and two cAMP-dependent protein kinase (PKA) substrates, GluR1 and NR1, were detected by immunoblotting. Thirty to 45 min after vanilla sugar consumption, phospho-Thr34 DARPP-32, GluR1 and NR1 levels increased in the nucleus accumbens, and phospho-Thr75 DARPP-32 levels decreased. At 60 min, all parameters returned to baseline values. However, 2 and 3 h after vanilla sugar consumption, phospho-Thr34 DARPP-32 levels decreased, while phospho-Thr75 DARPP-32 levels increased. In contrast to the pattern observed in the NAcS, no delayed changes in DARPP-32 phosphorylation were observed in the mPFC. Both early and delayed DARPP-32, GluR1 and NR1 phosphorylation changes were prevented by a dopamine D1 receptor antagonist administration. The delayed modifications in nucleus accumbens DARPP-32 phosphorylation were prevented by an mGluR5 antagonist administration. The mesolimbic dopaminergic response to an unfamiliar taste is correlated to a gustatory memory trace development, and the observed changes in DARPP-32 phosphorylation may be part of this process.

QTc interval prolongation in patients on long-term methadone maintenance therapy.

OBJECTIVE: The aim of the present study was to assess the incidence of abnormal QTc interval values in a population of subjects on a long-term methadone maintenance treatment, as a single therapy, and with methadone dosages ranging between 10 and 600 mg/daily (mean+/-SD=87+/-76). METHOD: Basal ECG recordings were carried out in 83 former heroin addicts on long-term successful methadone maintenance therapy for at least 6 months, while no other known QT-prolonging agent was being administered. RESULTS: Eighty-three percent of the subjects had a more prolonged QT interval than the reference values for persons of the same sex and age. Only 2 patients displayed a QTc interval of >500 ms. No correlation emerged between QTc values and methadone dosages. CONCLUSION: Patients on long-term methadone maintenance treatment show longer than expected QTc interval values. This data, associated with the finding that methadone is a rather potent inhibitor of HERG potassium channels and that it may induce torsade de pointes in predisposed subjects, supports the recommendation that patients entering methadone treatment (MT) are screened for cardiac risk factors. ECG might be considered in ongoing MT patients especially before starting QT-prolonging medications.

Repeated acetyl-l-carnitine administration increases phospho-Thr34 DARPP-32 levels and antagonizes cocaine-induced increase in Cdk5 and phospho-Thr75 DARPP-32 levels in rat striatum.

Abstract Acute cocaine administration increases phosphorylation of dopamine and cAMP-regulated phosphoprotein (M(r) 32 kDa) (DARPP-32) at threonine (Thr)-34, whereas repeated cocaine administration increases DARPP-32 phosphorylation at Thr-75 in Sprague-Dawley rat striatum. Repeated acetyl-l-carnitine (ALCAR) administration persistently increases dopamine outflow in the nucleus accumbens. The present study examined the effect of repeated ALCAR administration on the DARPP-32 phosphorylation pattern in the nucleus accumbens and caudate-putamen. ALCAR increased phosphoThr-34 DARPP-32 levels and decreased phosphoThr-75 DARPP-32 levels, after 1 and 10 days of washout. We compared the effects of repeated cocaine and repeated ALCAR administrations on the behavioural response to cocaine challenge and on the DARPP-32 phosphorylation pattern and cyclin-dependent kinase 5 (Cdk5) levels in the striatum. We also studied whether ALCAR administered daily during or after cocaine sensitization procedure would interfere with the effects of cocaine. When the response to the cocaine challenge was assessed, cocaine- and ALCAR-treated rats showed a similar sensitized behavioural response, and rats receiving combined cocaine and ALCAR treatments, irrespective of treatment order, also showed a sensitized response. A week after the cocaine challenge, the two drugs had induced opposite modifications in DARPP-32 phosphorylation, as cocaine increased phosphorylation at Thr-75, while ALCAR increased phosphorylation at Thr-34. In cocaine plus ALCAR treated rats, irrespective of treatment order, ALCAR administration antagonized cocaine effects on DARPP-32 phosphorylation. Moreover, cocaine, but not ALCAR, increased DeltaFosB and Cdk5 expression, and the increase in Cdk5 was antagonized by ALCAR administration in rats receiving combined treatments. These effects were relatively persistent, as they were still present 7 days after the last treatment.

Sardinian alcohol-preferring rats show low 5-HT extraneuronal levels in the mPFC and no habituation in monoaminergic response to repeated ethanol consumption in the NAcS.

Sardinian ethanol-preferring (sP), non-preferring (sNP), and Wistar rats show similar dopaminergic response to vanilla sugar consumption in nucleus accumbens shell (NAcS) and medial prefrontal cortex (mPFC), and similarly learn a vanilla sugar-sustained appetitive behavior. In this study we investigated whether in satiated sP, sNP, and Wistar rats vanilla sugar would also elicit a serotonergic response in NAcS and mPFC, and whether in these areas voluntary ethanol consumption would elicit dopaminergic and/or serotoninergic responses. In the NAcS, all rats showed similar serotonin increases in response to the two meals and similar development of rapid habituation. In the mPFC, Wistar and sNP rats showed similar serotonin increases after two vanilla sugar meals, while sP rats, which had low serotonin basal levels, did not show a serotonergic response. When presented with a 10% ethanol solution, Wistar and sP rats rapidly consumed it, while sNP rats did not. In the NAcS, Wistar and sP rats presented dopamine and serotonin increases in response to ethanol. However, while Wistar rats showed habituation in their response, sP rats did not. In the mPFC, ethanol induced similar dopamine increases in Wistar and sP rats; serotonin increases were observed only in Wistar rats. In conclusion, all three lines showed increased serotonin release in response to palatable food, but they profoundly differed in their response to ethanol. In fact, only Wistar and sP rats drank ethanol, Wistar rats showed a monoaminergic response similar to that obtained after palatable food, while sP rats did not develop habituation, suggesting that they perceived ethanol as a more relevant stimulus.

Synthetic peptides in the form of dendrimers become resistant to protease activity.

In previous papers, we observed that dendrimers of peptide mimotopes of the nicotinic receptor ligand site are strong antidotes against the lethality of the nicotinic receptor ligand alpha-bungarotoxin. Although their in vitro activity is identical to that of dendrimers, the corresponding monomeric peptide mimotopes are not effective in vivo. Because the higher in vivo efficiency of dendrimers could not in this case be related to polyvalent interaction, the stability to blood protease activity of monomeric versus tetrabranched dendrimeric mimotope peptides was compared here by incubating three different mimotopes with human plasma and serum. Unmodified peptides and cleaved sequences were followed by high pressure liquid chromatography and mass spectrometry. Tetrabranched peptides were shown to be much more stable in plasma and also in serum. To assess the notable stability of multimeric peptides, different bioactive neuropeptides, including enkephalins, neurotensin and nociceptin, were synthesized in monomeric and tetrabranched forms and incubated with human plasma and serum and with rat brain membrane extracts. All the tetrabranched neuropeptides fully retained biological activity and generally showed much greater stability to blood and brain protease activity. Some tetrabranched peptides were also resistant to trypsin and chymotrypsin. Our findings provide new insights into the possible therapeutic use of bioactive peptides.

Effects of long-term acetyl-L-carnitine administration in rats--II: Protection against the disrupting effect of stress on the acquisition of appetitive behavior.

Long-term acetyl-L-carnitine (ALCAR) administration prevents the development of escape deficit produced by acute exposure to unavoidable stress. However, it does not revert the escape deficit sustained by chronic stress exposure. Rats exposed to chronic stress show a low dopamine (DA) output in the nucleus accumbens shell (NAcS) and do not acquire an appetitive behavior sustained by the earning of vanilla sugar (VS) made contingent on the choice of one of the two divergent arms of a Y-maze (VS-sustained appetitive behavior, VAB), while control rats consistently do. The present study shows that ALCAR treatment in rats exposed to a 7-day stress protocol prevented a decrease in DA output in the NAcS and medial prefrontal cortex (mPFC) of rats, and that it strengthened the DA response to VS consummation in the same two areas. Moreover, rats treated with long-term ALCAR or exposed to chronic stress while treated with ALCAR acquired VAB as efficiently as control rats. Moreover, VAB acquisition in stressed rats treated with ALCAR coincided with the reversal of the deficits in escape and in dopaminergic transmission in the NAcS. Thus, repeated ALCAR treatment preserved the DA response to VS in chronically stressed rats and this effect appeared to be predictive of the rat's competence to acquire VAB.

Effects of long-term acetyl-L-carnitine administration in rats: I. increased dopamine output in mesocorticolimbic areas and protection toward acute stress exposure.

Acetyl-L-carnitine (ALCAR) is the acetyl ester of carnitine that has been reported to be beneficial in depressive disorders and Alzheimer's disease. A 7-day administration of ALCAR in rats increased dopamine and serotonin output in the nucleus accumbens shell and it prevented the development of escape deficit produced by acute exposure to unavoidable stress. No tolerance developed to this protective effect, which appeared to be mediated by (1) the activation of 5-HT(1A) receptors, as it was antagonized by the administration of WAY100635 30 min before stress exposure; and (2) a process of neuronal plasticity dependent on NMDA receptor activity, as subcutaneous dizocilpine infusion during ALCAR treatment prevented the development of the protective effect on stress. Chronic stress exposure maintains an escape deficit condition that is reverted by a long-term treatment with antidepressants, but the same condition was not modified by long-term ALCAR administration. Thus, ALCAR cannot be defined as an antidepressant.