Cyclic-adenosine monophosphate (cAMP) plays an important role in cell signalling and is widely us... more Cyclic-adenosine monophosphate (cAMP) plays an important role in cell signalling and is widely used as a marker for receptor activation and as a target for treating various diseases. In this paper we present the development and validation of a new method for the determination of cAMP and ATP (adenosine triphosphate) and other nucleotides in a biological system by combining zwitterionic hydrophilic interaction liquid chromatography (HILIC) and tandem mass spectrometry (MS/MS). The HILIC-MS/MS method was developed for the simultaneous quantitative analysis of cAMP and ATP, and was validated by assessment of linearity (over a range from 0.5 to 100nM for cAMP and 50 nM to 50 microM for ATP (r(2)>0.999)), resolution, limit of detection (0.5 and 50 nM for cAMP and ATP, respectively) and reproducibility. Furthermore, the method was validated and applied in vitro to determine cAMP accumulation in biological samples. The effect of several dopamine D(2) (partial) agonists and antagonists on cAMP accumulation was assessed by determination of the cAMP/ATP ratio in cells transfected with the human dopamine D(2L) receptor. Quinpirole, dopamine and ropinirole produced agonist effects on cAMP accumulation, with a potency of quinpirole>ropinirole>dopamine. Lisuride, terguride and bifeprunox were found to be partial agonists with efficacies of lisuride>terguride>bifeprunox. As expected, haloperidol, (-)-sulpiride and LY-741626 were antagonists. These results demonstrate that the present analytical method was robust, fast, sensitive, and selective. Moreover, it showed utility in determining cAMP/ATP in biological systems and the ability to study the effect of (partial) agonists and antagonists which makes it a useful tool for drug discovery.
We have previously demonstrated that nicotine withdrawal produces depression-like behavior and th... more We have previously demonstrated that nicotine withdrawal produces depression-like behavior and that serotonin (5-HT)2A/2C receptor ligands modulate that mood-like state. In the present study we aimed to identify the mechanisms (changes in radioligand binding, transcription or RNA-editing) related to such a behavioral outcome. Rats received vehicle or nicotine (0.4 mg/kg, sc) for 5 days in home cages. Brain 5-HT2A/2C receptors were analyzed on day 3 of nicotine withdrawal. Nicotine withdrawal increased [(3) H]ketanserin binding to 5-HT2A receptors in the ventral tegmental area and ventral dentate gyrus, yet decreased binding in the nucleus accumbens shell. Reduction of [(3) H]mesulergine binding to 5-HT2C receptors was seen in the ventral dentate gyrus. Profound decrease in the 5-HT2A receptor transcript level was noted in the hippocampus and ventral tegmental area. Out of five 5-HT2C receptor mRNA editing sites, deep sequencing data showed a reduction in editing at the E site and a ...
Journal of neural transmission (Vienna, Austria : 1996), Jan 22, 2015
Sarizotan 1-[(2R)-3,4-dihydro-2H-chromen-2-yl]-N-[[5-(4-fluorophenyl) pyridin-3-yl]methyl] methen... more Sarizotan 1-[(2R)-3,4-dihydro-2H-chromen-2-yl]-N-[[5-(4-fluorophenyl) pyridin-3-yl]methyl] methenamine, showed an in vivo pharmaco-EEG profile resembling that of methylphenidate which is used in attention deficit/hyperactivity disorder (ADHD). In turn, we tested sarizotan against impulsivity in juvenile rats measuring the choice for large delayed vs. a small immediate reward in a T-maze and obtained encouraging results starting at 0.03 mg/kg (plasma levels of ~11 nM). Results from rats treated neonatally with 6-hydroxydopamine (6-OHDA), also supported anti-ADHD activity although starting at 0.3 mg/kg. However, microdialysis studies revealed that free brain concentration of sarizotan at active doses were below its affinity for 5-HT1A receptors, the assumed primary target. In contrast, electrophysiological experiments in mid-brain Raphé serotonergic cells paralleled by plasma sampling showed that there was ~60 % inhibition of firing rate-indicating significant activation of 5-HT1A rec...
Agmatine [2-(4-aminobutyl)guanidine], is a novel endogenous ligand at alpha2-adrenoceptors, imida... more Agmatine [2-(4-aminobutyl)guanidine], is a novel endogenous ligand at alpha2-adrenoceptors, imidazoline and N-methyl-D-asparate receptors, as well as a nitric oxide synthase inhibitor. The present study tested whether agmatine (5-40 mg/kg, sc) modulated the locomotor, sensitizing, and discriminative stimulus effects of nicotine in male Wistar rats. Agmatine (10-40 mg/kg) affected neither the basal locomotor activity, nor the nicotine (0.4 mg/kg, sc)-evoked hyperactivation. A challenge with saline or nicotine (0.4 mg/kg, sc) on day 10 to rats treated repeatedly (for 5 days) with nicotine (0.4 mg/kg, sc; and exposed to experimental chambers), resulted in the expression of nicotine-evoked conditioned hyperlocomotor response or behavioral sensitization. Given on day 10, agmatine at a dose of 40 mg/kg (but not 20 mg/kg) attenuated nicotine-induced conditioned hyperactivity. However, when this dose was administered to the nicotine-sensitized rats, agmatine failed to alter the effect of th...
Schizophrenia has been associated with a dysfunction of brain dopamine (DA). This, so called, DA ... more Schizophrenia has been associated with a dysfunction of brain dopamine (DA). This, so called, DA hypothesis has been refined as new insights into the pathophysiology of schizophrenia have emerged. Currently, dysfunction of prefrontocortical glutamatergic and GABAergic projections and dysfunction of serotonin (5-HT) systems are also thought to play a role in the pathophysiology of schizophrenia. Refinements of the DA hypothesis have lead to the emergence of new pharmacological targets for antipsychotic drug development. It was shown that effective antipsychotic drugs with a low liability for inducing extra-pyramidal side-effects have affinities for a range of neurotransmitter receptors in addition to DA receptors, suggesting that a combination of neurotransmitter receptor affinities may be favorable for treatment outcome.This review focuses on the interaction between DA and 5-HT, as most antipsychotics display affinity for 5-HT receptors. We will discuss DA/5-HT interactions at the l...
5-HT (20 microM) enhanced dopamine (DA) D2-like receptor mediated reduction of the firing rate of... more 5-HT (20 microM) enhanced dopamine (DA) D2-like receptor mediated reduction of the firing rate of DA neurons in the substantia nigra pars compacta (A9) and ventral tegmental area (A10) in a rat midbrain slice preparation. Quinpirole (30 nM) induced a mean reduction of the firing rate in A9 and A10 DA neurons to 64 +/- 4%, respectively, 71 +/- 5% of the baseline value. Bath application of 5-HT in the presence of quinpirole further reduced the firing rate to 37 +/- 7% in A9 and 33 +/- 13% in A10. The 5-HT2 receptor agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI, 500 nM) enhanced quinpirole-induced reduction of firing rate of A10 DA neurons, but not of A9 DA neurons, suggesting that different 5-HT receptor subtypes are involved in modulation of dopamine D2-like receptor mediated inhibition in the two regions. The selective 5-HT2A receptor antagonist MDL100907 and the selective 5-HT2C receptor antagonist SB242084 (50 and 500 nM) both abolished the enhance...
We have previously demonstrated that serotonin (5-HT)2A and 5-HT2C receptor ligands modulate the ... more We have previously demonstrated that serotonin (5-HT)2A and 5-HT2C receptor ligands modulate the sensitizing effects of nicotine. In the present study we used male rats to verify the hypothesis that the binding pattern of 5-HT2A and 5-HT2C receptors in the brain is altered by chronic nicotine treatment in different environments. Rats were given repeatedly vehicle or nicotine in different sensitizing regimens (home or experimental cages). On day 10, animals were challenged with nicotine (expression of nicotine sensitization) or vehicle in either home or experimental cages, and were sacrificed immediately after the experiment. Repeated treatment with nicotine in home cages evoked significant increases in [(3)H]ketanserin binding to 5-HT2A receptors in the prefrontal cortex, striatal subregions and ventral tegmental area as well as reductions in [(3)H]mesulergine binding to 5-HT2C receptors in subregions of the prefrontal cortex. In contrast, nicotine paired with environmental context produced robust increases in 5-HT2A receptor labeling in the infralimbic cortex and decreased [(3)H]ketanserin binding in striatal subregions and ventral tegmental area; 5-HT2C receptor labeling in the prefrontal cortex fell. The present data indicate that chronic nicotine administration in home cages induces bi-directional neuroplastic changes within 5-HT2A and 5-HT2C receptors in the prefrontal cortex. Pairing the nicotine with environmental context potentiates the neuroplastic response in the latter region and evokes opposite changes in 5-HT2A receptor binding in striatal and tegmental regions compared with nicotine administered in the absence of the context, indicating a modulatory role of environmental context in the expression of nicotine-induced sensitization.
A series of novel bicyclic 1-heteroaryl-4-[omega-(1H-indol-3-yl)alkyl]piperazines was synthesized... more A series of novel bicyclic 1-heteroaryl-4-[omega-(1H-indol-3-yl)alkyl]piperazines was synthesized and evaluated on binding to dopamine D(2) receptors and serotonin reuptake sites. This class of compounds proved to be potent in vitro dopamine D(2) receptor antagonists and in addition were highly active as serotonin reuptake inhibitors. Some key representatives showed potent pharmacological in vivo activities after oral dosing in both the antagonism of apomorphine-induced climbing and the potentiation of 5-HTP-induced behavior in mice. On the basis of the preclinical data, 8-{4-[3-(5-fluoro-1H-indol-3-yl)propyl]piperazin-1-yl}-4H-benzo[1,4]oxazin-(R)-2-methyl-3-one (45c, SLV314) was selected for clinical development. In vitro and in vivo studies revealed that 45c has favorable pharmacokinetic properties and a high CNS-plasma ratio. Molecular modeling studies showed that the bifunctional activity of 45c can be explained by its ability to adopt two different conformations fitting either...
The treatment of dyskinesia in Parkinson׳s disease remains poor but H3 receptor agonists have bee... more The treatment of dyskinesia in Parkinson׳s disease remains poor but H3 receptor agonists have been suggested as a novel pharmacological approach. We examined the effects of the H3 agonist, immepip, in 6-OHDA-lesioned rats exhibiting AIMs (abnormal involuntary movements), a rat analogue of dyskinesia, in response to l-dopa compared to the known anti-dyskinetic agents amantadine, MK-801 and 8-OHDPAT. We then attempted to extend these studies in to dyskinetic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treated common marmosets. Amantadine, MK-801 and 8-OHDPAT all dose-dependently reduced l-dopa-induced axial, lingual and oral (ALO) AIMs in 6-OHDA-lesioned animals accompanied by a reduction in contralateral rotation with higher doses of amantadine and MK-801. By contrast, immepip had no effect on AIMs expression or contralateral rotation. In the MPTP-treated common marmoset exhibiting dyskinesia to l-dopa, immepip alone induced retching and in combination with l-dopa administered subcutaneously or orally induced the rapid onset of retching and vomiting which was not controlled by pretreatment with domperidone. Administration of the unrelated H3 agonist, imetit had the same effect. Despite causing negative side-effects, it appears that both agonists reduced the antiparkinsonian response to l-dopa resulting in reduced dyskinesia. H3 agonists appear unlikely candidates for the treatment of dyskinesia in PD based on lack of evidence of efficacy and potential adverse effects.
The objective of this study was to evaluate the efficacy of varenicline, a novel partial agonist ... more The objective of this study was to evaluate the efficacy of varenicline, a novel partial agonist at alpha 4 beta 2 and full agonist at alpha 7 nicotinic acetylcholine receptor (nAChR) subtypes, in blocking the locomotor effects of acute or repeated treatments with nicotine (0.4 mg/kg, s.c.) in rats. Varenicline (0.3-3 mg/kg, s.c.) by itself enhanced the basal locomotor activity in naive rats while it had an inhibitory effect on acute nicotine-induced hyperlocomotion. Varenicline (0.3-3 mg/kg) did not change the nicotine-evoked conditioned locomotion, but when administered to nicotine-sensitized rats (0.1 and 1 mg/kg), reduced the expression of nicotine sensitization. In another set of experiments, varenicline (1 mg/kg) administered during the second withdrawal period (days 11-14) to nicotine-treated rats, attenuated the reestablishment of the expression of nicotine sensitization. Our pharmacological analyses further support the hypothesis that varenicline might be a useful treatment for smoking cessation considering its actions on the locomotor and reinforcing effects of nicotine without inhibition of conditioned locomotion.
Present Parkinson&amp... more Present Parkinson's disease treatment strategies are far from ideal for a variety of reasons; it has therefore been suggested that partial dopamine receptor agonism might be a potential therapeutic approach with potentially fewer side effects. In the present study, we describe the in vitro characterization of the nonergot ligand SLV308 (7-[4-methyl-1-piperazinyl]-2(3H)-benzoxazolonemonohydrochloride). SLV308 binds to dopamine D(2), D(3), and D(4) receptors and 5-HT(1) (A) receptors and is a partial agonist at dopamine D(2) and D(3) receptors and a full agonist at serotonin 5-HT(1) (A) receptors. At cloned human dopamine D(2,L) receptors, SLV308 acted as a potent but partial D(2) receptor agonist (pEC(50) = 8.0 and pA(2) = 8.4) with an efficacy of 50% on forskolin stimulated cAMP accumulation. At human recombinant dopamine D(3) receptors, SLV308 acted as a partial agonist in the induction of [(35)S]GTPgammaS binding (intrinsic activity of 67%; pEC(50) = 9.2) and antagonized the dopamine induction of [(35)S]GTPgammaS binding (pA(2) = 9.0). SLV308 acted as a full 5-HT(1) (A) receptor agonist on forskolin induced cAMP accumulation at cloned human 5-HT(1) (A) receptors but with low potency (pEC(50) = 6.3). In rat striatal slices SLV308 concentration-dependently attenuated forskolin stimulated accumulation of cAMP, as expected for a dopamine D(2) and D(3) receptor agonist. SLV308 antagonized the inhibitory effect of quinpirole on K(+)-stimulated [(3)H]-dopamine release from rat striatal slices (pA(2) = 8.5). In the same paradigm, SLV308 had antagonist properties in the presence of quinpirole (pA(2) = 8.5), but the partial D(2) agonist terguride had much stronger antagonistic properties. In conclusion, SLV308 combines high potency partial agonism at dopamine D(2) and D(3) receptors with full efficacy low potency serotonin 5-HT(1) (A) receptor agonism and is worthy of profiling in in vivo models of Parkinson's disease.
Male Wistar rats were used to verify the hypothesis that serotonin (5-HT)(2A) or 5-HT(2C) recepto... more Male Wistar rats were used to verify the hypothesis that serotonin (5-HT)(2A) or 5-HT(2C) receptors may control the locomotor effects evoked by nicotine (0.4 mg/kg). The 5-HT(2A) receptor antagonist (M100,907), the 5-HT(2A) receptor agonist (DOI), the 5-HT(2C) receptor antagonist (SB 242,084), and the 5-HT(2C) receptor agonists (Ro 60-0175 and WAY 163,909) were used. M100,907 (0.5-2mg/kg) did not alter, while DOI (1 mg/kg) enhanced the nicotine-induced hyperlocomotion. The effect of DOI was antagonized by M100,907 (1 mg/kg). SB 242,084 (0.25-1 mg/kg) augmented, while Ro 60-0175 (1 and 3 mg/kg) and WAY 163,909 (1.5 mg/kg) decreased the overall effect of acute nicotine; effects of Ro 60-0175 and WAY 163,909 were attenuated by SB 242,084 (0.125 mg/kg). In another set of experiments, M100,907 (2 mg/kg) on Day 10 attenuated, while DOI (0.1-1 mg/kg) enhanced the nicotine-evoked conditioned hyperlocomotion in rats repeatedly (Days 1-5) treated with nicotine in experimental chambers. SB 242,084 (0.125 or 1 mg/kg) did not change, while Ro 60-0175 (1 mg/kg) or WAY 163,909 (1.5 mg/kg) decreased the expression of nicotine-induced conditioned hyperactivity. Only DOI (0.3 and 1 mg/kg) and SB 242,084 (1 mg/kg) enhanced the basal locomotion. The present data indicate that 5-HT(2A) receptors are significant for the expression of nicotine-evoked conditioned hyperactivity. Conversely, 5-HT(2C) receptors play a pivotal role in the acute effects of nicotine. Pharmacological stimulation of 5-HT(2A) receptors enhances the conditioned hyperlocomotion, while activation of 5-HT(2C) receptors decreases both the response to acute nicotine and conditioned hyperactivity.
We verified the hypothesis that serotonin (5-HT)(2) receptors control the locomotor effects of ni... more We verified the hypothesis that serotonin (5-HT)(2) receptors control the locomotor effects of nicotine (0.4 mg kg(-1)) in rats by using the 5-HT(2A) receptor antagonist M100907, the preferential 5-HT(2A) receptor agonist DOI, the 5-HT(2C) receptor antagonist SB 242084, and the 5-HT(2C) receptor agonists Ro 60-0175 and WAY 163909. Repeated pairings of a test environment with nicotine for 5 days, on Day 10 significantly augmented the locomotor activity following nicotine administration. Of the investigated 5-HT(2) receptor ligands, M100907 (2 mg kg(-1)) or DOI (1 mg kg(-1)) administered during the first 5 days in combination with nicotine attenuated or enhanced, respectively, the development of nicotine sensitization. Given acutely on Day 10, M100907 (2 mg kg(-1)), Ro 60-0175 (1 mg kg(-1)), and WAY 163909 (1.5 mg kg(-1)) decreased the expression of nicotine sensitization. In another set of experiments, where the nicotine challenge test was performed on Day 15 in animals treated repeatedly (Days: 1-5, 10) with nicotine, none of 5-HT(2) receptor ligands administered during the second withdrawal period (Days: 11-14) to nicotine-treated rats altered the sensitizing effect of nicotine given on Day 15. Our data indicate that 5-HT(2A) receptors (but not 5-HT(2C) receptors) play a permissive role in the sensitizing effects of nicotine, while stimulation of 5-HT(2A) receptors enhances the development of nicotine sensitization and activation of 5-HT(2C) receptors is essential for the expression of nicotine sensitization. Repeated treatment with the 5-HT(2) receptor ligands within the second nicotine withdrawal does not inhibit previously established sensitization.
Cyclic-adenosine monophosphate (cAMP) plays an important role in cell signalling and is widely us... more Cyclic-adenosine monophosphate (cAMP) plays an important role in cell signalling and is widely used as a marker for receptor activation and as a target for treating various diseases. In this paper we present the development and validation of a new method for the determination of cAMP and ATP (adenosine triphosphate) and other nucleotides in a biological system by combining zwitterionic hydrophilic interaction liquid chromatography (HILIC) and tandem mass spectrometry (MS/MS). The HILIC-MS/MS method was developed for the simultaneous quantitative analysis of cAMP and ATP, and was validated by assessment of linearity (over a range from 0.5 to 100nM for cAMP and 50 nM to 50 microM for ATP (r(2)>0.999)), resolution, limit of detection (0.5 and 50 nM for cAMP and ATP, respectively) and reproducibility. Furthermore, the method was validated and applied in vitro to determine cAMP accumulation in biological samples. The effect of several dopamine D(2) (partial) agonists and antagonists on cAMP accumulation was assessed by determination of the cAMP/ATP ratio in cells transfected with the human dopamine D(2L) receptor. Quinpirole, dopamine and ropinirole produced agonist effects on cAMP accumulation, with a potency of quinpirole>ropinirole>dopamine. Lisuride, terguride and bifeprunox were found to be partial agonists with efficacies of lisuride>terguride>bifeprunox. As expected, haloperidol, (-)-sulpiride and LY-741626 were antagonists. These results demonstrate that the present analytical method was robust, fast, sensitive, and selective. Moreover, it showed utility in determining cAMP/ATP in biological systems and the ability to study the effect of (partial) agonists and antagonists which makes it a useful tool for drug discovery.
We have previously demonstrated that nicotine withdrawal produces depression-like behavior and th... more We have previously demonstrated that nicotine withdrawal produces depression-like behavior and that serotonin (5-HT)2A/2C receptor ligands modulate that mood-like state. In the present study we aimed to identify the mechanisms (changes in radioligand binding, transcription or RNA-editing) related to such a behavioral outcome. Rats received vehicle or nicotine (0.4 mg/kg, sc) for 5 days in home cages. Brain 5-HT2A/2C receptors were analyzed on day 3 of nicotine withdrawal. Nicotine withdrawal increased [(3) H]ketanserin binding to 5-HT2A receptors in the ventral tegmental area and ventral dentate gyrus, yet decreased binding in the nucleus accumbens shell. Reduction of [(3) H]mesulergine binding to 5-HT2C receptors was seen in the ventral dentate gyrus. Profound decrease in the 5-HT2A receptor transcript level was noted in the hippocampus and ventral tegmental area. Out of five 5-HT2C receptor mRNA editing sites, deep sequencing data showed a reduction in editing at the E site and a ...
Journal of neural transmission (Vienna, Austria : 1996), Jan 22, 2015
Sarizotan 1-[(2R)-3,4-dihydro-2H-chromen-2-yl]-N-[[5-(4-fluorophenyl) pyridin-3-yl]methyl] methen... more Sarizotan 1-[(2R)-3,4-dihydro-2H-chromen-2-yl]-N-[[5-(4-fluorophenyl) pyridin-3-yl]methyl] methenamine, showed an in vivo pharmaco-EEG profile resembling that of methylphenidate which is used in attention deficit/hyperactivity disorder (ADHD). In turn, we tested sarizotan against impulsivity in juvenile rats measuring the choice for large delayed vs. a small immediate reward in a T-maze and obtained encouraging results starting at 0.03 mg/kg (plasma levels of ~11 nM). Results from rats treated neonatally with 6-hydroxydopamine (6-OHDA), also supported anti-ADHD activity although starting at 0.3 mg/kg. However, microdialysis studies revealed that free brain concentration of sarizotan at active doses were below its affinity for 5-HT1A receptors, the assumed primary target. In contrast, electrophysiological experiments in mid-brain Raphé serotonergic cells paralleled by plasma sampling showed that there was ~60 % inhibition of firing rate-indicating significant activation of 5-HT1A rec...
Agmatine [2-(4-aminobutyl)guanidine], is a novel endogenous ligand at alpha2-adrenoceptors, imida... more Agmatine [2-(4-aminobutyl)guanidine], is a novel endogenous ligand at alpha2-adrenoceptors, imidazoline and N-methyl-D-asparate receptors, as well as a nitric oxide synthase inhibitor. The present study tested whether agmatine (5-40 mg/kg, sc) modulated the locomotor, sensitizing, and discriminative stimulus effects of nicotine in male Wistar rats. Agmatine (10-40 mg/kg) affected neither the basal locomotor activity, nor the nicotine (0.4 mg/kg, sc)-evoked hyperactivation. A challenge with saline or nicotine (0.4 mg/kg, sc) on day 10 to rats treated repeatedly (for 5 days) with nicotine (0.4 mg/kg, sc; and exposed to experimental chambers), resulted in the expression of nicotine-evoked conditioned hyperlocomotor response or behavioral sensitization. Given on day 10, agmatine at a dose of 40 mg/kg (but not 20 mg/kg) attenuated nicotine-induced conditioned hyperactivity. However, when this dose was administered to the nicotine-sensitized rats, agmatine failed to alter the effect of th...
Schizophrenia has been associated with a dysfunction of brain dopamine (DA). This, so called, DA ... more Schizophrenia has been associated with a dysfunction of brain dopamine (DA). This, so called, DA hypothesis has been refined as new insights into the pathophysiology of schizophrenia have emerged. Currently, dysfunction of prefrontocortical glutamatergic and GABAergic projections and dysfunction of serotonin (5-HT) systems are also thought to play a role in the pathophysiology of schizophrenia. Refinements of the DA hypothesis have lead to the emergence of new pharmacological targets for antipsychotic drug development. It was shown that effective antipsychotic drugs with a low liability for inducing extra-pyramidal side-effects have affinities for a range of neurotransmitter receptors in addition to DA receptors, suggesting that a combination of neurotransmitter receptor affinities may be favorable for treatment outcome.This review focuses on the interaction between DA and 5-HT, as most antipsychotics display affinity for 5-HT receptors. We will discuss DA/5-HT interactions at the l...
5-HT (20 microM) enhanced dopamine (DA) D2-like receptor mediated reduction of the firing rate of... more 5-HT (20 microM) enhanced dopamine (DA) D2-like receptor mediated reduction of the firing rate of DA neurons in the substantia nigra pars compacta (A9) and ventral tegmental area (A10) in a rat midbrain slice preparation. Quinpirole (30 nM) induced a mean reduction of the firing rate in A9 and A10 DA neurons to 64 +/- 4%, respectively, 71 +/- 5% of the baseline value. Bath application of 5-HT in the presence of quinpirole further reduced the firing rate to 37 +/- 7% in A9 and 33 +/- 13% in A10. The 5-HT2 receptor agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI, 500 nM) enhanced quinpirole-induced reduction of firing rate of A10 DA neurons, but not of A9 DA neurons, suggesting that different 5-HT receptor subtypes are involved in modulation of dopamine D2-like receptor mediated inhibition in the two regions. The selective 5-HT2A receptor antagonist MDL100907 and the selective 5-HT2C receptor antagonist SB242084 (50 and 500 nM) both abolished the enhance...
We have previously demonstrated that serotonin (5-HT)2A and 5-HT2C receptor ligands modulate the ... more We have previously demonstrated that serotonin (5-HT)2A and 5-HT2C receptor ligands modulate the sensitizing effects of nicotine. In the present study we used male rats to verify the hypothesis that the binding pattern of 5-HT2A and 5-HT2C receptors in the brain is altered by chronic nicotine treatment in different environments. Rats were given repeatedly vehicle or nicotine in different sensitizing regimens (home or experimental cages). On day 10, animals were challenged with nicotine (expression of nicotine sensitization) or vehicle in either home or experimental cages, and were sacrificed immediately after the experiment. Repeated treatment with nicotine in home cages evoked significant increases in [(3)H]ketanserin binding to 5-HT2A receptors in the prefrontal cortex, striatal subregions and ventral tegmental area as well as reductions in [(3)H]mesulergine binding to 5-HT2C receptors in subregions of the prefrontal cortex. In contrast, nicotine paired with environmental context produced robust increases in 5-HT2A receptor labeling in the infralimbic cortex and decreased [(3)H]ketanserin binding in striatal subregions and ventral tegmental area; 5-HT2C receptor labeling in the prefrontal cortex fell. The present data indicate that chronic nicotine administration in home cages induces bi-directional neuroplastic changes within 5-HT2A and 5-HT2C receptors in the prefrontal cortex. Pairing the nicotine with environmental context potentiates the neuroplastic response in the latter region and evokes opposite changes in 5-HT2A receptor binding in striatal and tegmental regions compared with nicotine administered in the absence of the context, indicating a modulatory role of environmental context in the expression of nicotine-induced sensitization.
A series of novel bicyclic 1-heteroaryl-4-[omega-(1H-indol-3-yl)alkyl]piperazines was synthesized... more A series of novel bicyclic 1-heteroaryl-4-[omega-(1H-indol-3-yl)alkyl]piperazines was synthesized and evaluated on binding to dopamine D(2) receptors and serotonin reuptake sites. This class of compounds proved to be potent in vitro dopamine D(2) receptor antagonists and in addition were highly active as serotonin reuptake inhibitors. Some key representatives showed potent pharmacological in vivo activities after oral dosing in both the antagonism of apomorphine-induced climbing and the potentiation of 5-HTP-induced behavior in mice. On the basis of the preclinical data, 8-{4-[3-(5-fluoro-1H-indol-3-yl)propyl]piperazin-1-yl}-4H-benzo[1,4]oxazin-(R)-2-methyl-3-one (45c, SLV314) was selected for clinical development. In vitro and in vivo studies revealed that 45c has favorable pharmacokinetic properties and a high CNS-plasma ratio. Molecular modeling studies showed that the bifunctional activity of 45c can be explained by its ability to adopt two different conformations fitting either...
The treatment of dyskinesia in Parkinson׳s disease remains poor but H3 receptor agonists have bee... more The treatment of dyskinesia in Parkinson׳s disease remains poor but H3 receptor agonists have been suggested as a novel pharmacological approach. We examined the effects of the H3 agonist, immepip, in 6-OHDA-lesioned rats exhibiting AIMs (abnormal involuntary movements), a rat analogue of dyskinesia, in response to l-dopa compared to the known anti-dyskinetic agents amantadine, MK-801 and 8-OHDPAT. We then attempted to extend these studies in to dyskinetic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treated common marmosets. Amantadine, MK-801 and 8-OHDPAT all dose-dependently reduced l-dopa-induced axial, lingual and oral (ALO) AIMs in 6-OHDA-lesioned animals accompanied by a reduction in contralateral rotation with higher doses of amantadine and MK-801. By contrast, immepip had no effect on AIMs expression or contralateral rotation. In the MPTP-treated common marmoset exhibiting dyskinesia to l-dopa, immepip alone induced retching and in combination with l-dopa administered subcutaneously or orally induced the rapid onset of retching and vomiting which was not controlled by pretreatment with domperidone. Administration of the unrelated H3 agonist, imetit had the same effect. Despite causing negative side-effects, it appears that both agonists reduced the antiparkinsonian response to l-dopa resulting in reduced dyskinesia. H3 agonists appear unlikely candidates for the treatment of dyskinesia in PD based on lack of evidence of efficacy and potential adverse effects.
The objective of this study was to evaluate the efficacy of varenicline, a novel partial agonist ... more The objective of this study was to evaluate the efficacy of varenicline, a novel partial agonist at alpha 4 beta 2 and full agonist at alpha 7 nicotinic acetylcholine receptor (nAChR) subtypes, in blocking the locomotor effects of acute or repeated treatments with nicotine (0.4 mg/kg, s.c.) in rats. Varenicline (0.3-3 mg/kg, s.c.) by itself enhanced the basal locomotor activity in naive rats while it had an inhibitory effect on acute nicotine-induced hyperlocomotion. Varenicline (0.3-3 mg/kg) did not change the nicotine-evoked conditioned locomotion, but when administered to nicotine-sensitized rats (0.1 and 1 mg/kg), reduced the expression of nicotine sensitization. In another set of experiments, varenicline (1 mg/kg) administered during the second withdrawal period (days 11-14) to nicotine-treated rats, attenuated the reestablishment of the expression of nicotine sensitization. Our pharmacological analyses further support the hypothesis that varenicline might be a useful treatment for smoking cessation considering its actions on the locomotor and reinforcing effects of nicotine without inhibition of conditioned locomotion.
Present Parkinson&amp... more Present Parkinson's disease treatment strategies are far from ideal for a variety of reasons; it has therefore been suggested that partial dopamine receptor agonism might be a potential therapeutic approach with potentially fewer side effects. In the present study, we describe the in vitro characterization of the nonergot ligand SLV308 (7-[4-methyl-1-piperazinyl]-2(3H)-benzoxazolonemonohydrochloride). SLV308 binds to dopamine D(2), D(3), and D(4) receptors and 5-HT(1) (A) receptors and is a partial agonist at dopamine D(2) and D(3) receptors and a full agonist at serotonin 5-HT(1) (A) receptors. At cloned human dopamine D(2,L) receptors, SLV308 acted as a potent but partial D(2) receptor agonist (pEC(50) = 8.0 and pA(2) = 8.4) with an efficacy of 50% on forskolin stimulated cAMP accumulation. At human recombinant dopamine D(3) receptors, SLV308 acted as a partial agonist in the induction of [(35)S]GTPgammaS binding (intrinsic activity of 67%; pEC(50) = 9.2) and antagonized the dopamine induction of [(35)S]GTPgammaS binding (pA(2) = 9.0). SLV308 acted as a full 5-HT(1) (A) receptor agonist on forskolin induced cAMP accumulation at cloned human 5-HT(1) (A) receptors but with low potency (pEC(50) = 6.3). In rat striatal slices SLV308 concentration-dependently attenuated forskolin stimulated accumulation of cAMP, as expected for a dopamine D(2) and D(3) receptor agonist. SLV308 antagonized the inhibitory effect of quinpirole on K(+)-stimulated [(3)H]-dopamine release from rat striatal slices (pA(2) = 8.5). In the same paradigm, SLV308 had antagonist properties in the presence of quinpirole (pA(2) = 8.5), but the partial D(2) agonist terguride had much stronger antagonistic properties. In conclusion, SLV308 combines high potency partial agonism at dopamine D(2) and D(3) receptors with full efficacy low potency serotonin 5-HT(1) (A) receptor agonism and is worthy of profiling in in vivo models of Parkinson's disease.
Male Wistar rats were used to verify the hypothesis that serotonin (5-HT)(2A) or 5-HT(2C) recepto... more Male Wistar rats were used to verify the hypothesis that serotonin (5-HT)(2A) or 5-HT(2C) receptors may control the locomotor effects evoked by nicotine (0.4 mg/kg). The 5-HT(2A) receptor antagonist (M100,907), the 5-HT(2A) receptor agonist (DOI), the 5-HT(2C) receptor antagonist (SB 242,084), and the 5-HT(2C) receptor agonists (Ro 60-0175 and WAY 163,909) were used. M100,907 (0.5-2mg/kg) did not alter, while DOI (1 mg/kg) enhanced the nicotine-induced hyperlocomotion. The effect of DOI was antagonized by M100,907 (1 mg/kg). SB 242,084 (0.25-1 mg/kg) augmented, while Ro 60-0175 (1 and 3 mg/kg) and WAY 163,909 (1.5 mg/kg) decreased the overall effect of acute nicotine; effects of Ro 60-0175 and WAY 163,909 were attenuated by SB 242,084 (0.125 mg/kg). In another set of experiments, M100,907 (2 mg/kg) on Day 10 attenuated, while DOI (0.1-1 mg/kg) enhanced the nicotine-evoked conditioned hyperlocomotion in rats repeatedly (Days 1-5) treated with nicotine in experimental chambers. SB 242,084 (0.125 or 1 mg/kg) did not change, while Ro 60-0175 (1 mg/kg) or WAY 163,909 (1.5 mg/kg) decreased the expression of nicotine-induced conditioned hyperactivity. Only DOI (0.3 and 1 mg/kg) and SB 242,084 (1 mg/kg) enhanced the basal locomotion. The present data indicate that 5-HT(2A) receptors are significant for the expression of nicotine-evoked conditioned hyperactivity. Conversely, 5-HT(2C) receptors play a pivotal role in the acute effects of nicotine. Pharmacological stimulation of 5-HT(2A) receptors enhances the conditioned hyperlocomotion, while activation of 5-HT(2C) receptors decreases both the response to acute nicotine and conditioned hyperactivity.
We verified the hypothesis that serotonin (5-HT)(2) receptors control the locomotor effects of ni... more We verified the hypothesis that serotonin (5-HT)(2) receptors control the locomotor effects of nicotine (0.4 mg kg(-1)) in rats by using the 5-HT(2A) receptor antagonist M100907, the preferential 5-HT(2A) receptor agonist DOI, the 5-HT(2C) receptor antagonist SB 242084, and the 5-HT(2C) receptor agonists Ro 60-0175 and WAY 163909. Repeated pairings of a test environment with nicotine for 5 days, on Day 10 significantly augmented the locomotor activity following nicotine administration. Of the investigated 5-HT(2) receptor ligands, M100907 (2 mg kg(-1)) or DOI (1 mg kg(-1)) administered during the first 5 days in combination with nicotine attenuated or enhanced, respectively, the development of nicotine sensitization. Given acutely on Day 10, M100907 (2 mg kg(-1)), Ro 60-0175 (1 mg kg(-1)), and WAY 163909 (1.5 mg kg(-1)) decreased the expression of nicotine sensitization. In another set of experiments, where the nicotine challenge test was performed on Day 15 in animals treated repeatedly (Days: 1-5, 10) with nicotine, none of 5-HT(2) receptor ligands administered during the second withdrawal period (Days: 11-14) to nicotine-treated rats altered the sensitizing effect of nicotine given on Day 15. Our data indicate that 5-HT(2A) receptors (but not 5-HT(2C) receptors) play a permissive role in the sensitizing effects of nicotine, while stimulation of 5-HT(2A) receptors enhances the development of nicotine sensitization and activation of 5-HT(2C) receptors is essential for the expression of nicotine sensitization. Repeated treatment with the 5-HT(2) receptor ligands within the second nicotine withdrawal does not inhibit previously established sensitization.
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Papers by Andrew McCreary