Neuropharmacological potentials (antipsychotic-, anxiolytic-, and antidepressant-like activities) of methanol leaf extract of Andrographis paniculata Nees in vivo: Possible mechanisms, antioxidant activity, and in silico supportive evidence.

Recent evidence suggests that non-selective cannabinoid receptor agonists may regulate serotonin 2A (5-HT(2A)) receptor neurotransmission in brain. The molecular mechanisms of this regulation are unknown, but could involve cannabinoid-induced enhanced interaction between 5-HT(2A) and dopamine D2 (D₂) receptors. Here, we present experimental evidence that Sprague-Dawley rats treated with a non-selective cannabinoid receptor agonist (CP55,940, 50 µg/kg, 7 days, i.p.) showed enhanced co-immunoprecipitation of 5-HT(2A) and D₂ receptors and enhanced membrane-associated expression of D₂ and 5-HT(2A) receptors in prefrontal cortex (PFCx). Furthermore, 5-HT(2A) receptor mRNA levels were increased in PFCx, suggesting a cannabinoid-induced upregulation of 5-HT(2A) receptors. To date, two cannabinoids receptors have been found in brain, CB1 and CB2 receptors. We used selective cannabinoid agonists in a neuronal cell line to study mechanisms that could mediate this 5-HT(2A) receptor upregulation. We found that selective CB2 receptor agonists upregulate 5-HT(2A) receptors by a mechanism that seems to involve activation of Gα(i) G-proteins, ERK1/2, and AP-1 transcription factor. We hypothesize that the enhanced cannabinoid-induced interaction between 5-HT(2A) and D₂ receptors and in 5-HT(2A) and D₂ receptors protein levels in the PFCx might provide a molecular mechanism by which activation of cannabinoid receptors might be contribute to the pathophysiology of some cognitive and mood disorders.

Atypical Antipsychotics: Mechanism of Action

The physiologic process of aging is known to have an effect on the brain concentration of some monoaminergic receptors. In addition, studies have suggested a causal link between sex hormones and these receptor changes. Accordingly, we were interested to see in our colony of geriatric rhesus monkeys if we could demonstrate an age- and sex-related difference in 5HT2A receptor concentration in the brain using the PET tracer [18F] Setoperone. This tracer has a 10-50x greater affinity for the 5-HT2A receptor than the D2 receptor, but provides a specific signal for both receptors in vivo. Since D2 receptor density is low in cortical regions and 5HT2A is high, while the opposite is true in striatum, [18F] Setoperone can be used to evaluate both receptors. Chase studies with risperidone, a non-selective antagonist of both 5-HT2A and D2 demonstrated significant displacement of the tracer from both 5-HT2A and D2 receptors. Studies were conducted in propofol anesthetized, older (n=5, 26–29 yrs) female Rhesus monkeys and young female (n=2, 8 yrs) and male Rhesus monkeys (n=2, 8 and 10 yrs) as controls in an ECAT HR+ PET camera. At the time of the scans, estradiol levels were 95 11 pg/ml in the young female monkeys compared to 35.3 6.5 pg/ml in the older monkeys. [18F] Setoperone was administered as a bolus followed by a matched constant-rate infusion for 200 minutes, adjusted to compensate for tracer elimination in the bolus, to achieve steady-state brain uptake. Tracer uptake reached a steady state in the brain after 120 min (reflected by constant target-to-cerebellum ratio). Indices of 5HT2A and D2 receptor binding were calculated from the occipital cortex/cerebellum and striatum/cerebellum ratios between 140 and 200 minutes, respectively. The cerebellum was used as a reference region since it is essentially devoid of both 5-HT2A and D2 receptors. The uptake ratios of occipital cortex/cerebellum binding (an index of the 5-HT2A receptor binding potential) and striatum/cerebellum (an index of the D2 receptor binding potential) were calculated. The test-retest reproducibility of this protocol was found to be < 10% in male Rhesus monkeys. Results indicated an 15% and 24% reduction in binding potential for D2 and 5HT2A receptors respectively in the older females compared to the young females. As expected, the binding potential for 5HT2A receptor in young males was comparable to that in the young females. These studies suggest [18F] Setoperone may be used in a convenient protocol to show age related reductions in 5HT2A and D2 receptor binding in post-menopausal woman. An advantage of using such a protocol with [18F] setoperone is that both 5-HT2A and D2 receptors can be evaluated in the same study.

The authors' objective was to assess the potential efficacy of fananserin (RP62203), a potent antagonist at the D4 and serotonin2A (5-HT2A) receptors, on symptoms of schizophrenia. A double-blind, placebo-controlled study was conducted in 97 patients. Doses of fananserin reached 250 mg b.i.d. over 28 days, starting with an 8-day escalation. Most of the patients were men with paranoid schizophrenia; they were approximately 38 years old. The primary outcome measure was the total Positive and Negative Syndrome Scale score. The patients' mean score on the Positive and Negative Syndrome Scale at entry was 91.8 (SD=16.5). A low dropout rate was observed in both groups of patients (19 [30%] of those given fananserin and nine [27%] of those given placebo). The total Positive and Negative Syndrome Scale score of the patients given fananserin decreased at endpoint by a mean of 4.2 points (SD=15.4); the score of the patients given placebo decreased by 6.7 points (SD=19.6). No differences between treatments were found on secondary measures such as the Clinical Global Impression, Positive and Negative Syndrome Scale subscores or individual items, and Brief Psychiatric Rating Scale total score. The patients' extrapyramidal symptoms did not worsen during the trial, but the patients given fananserin had an increase in akathisia. The safety profile was good in both groups of patients. The results of this study do not support the prediction that a selective D4 antagonist associated with strong 5-HT2A antagonism will exhibit an antipsychotic effect.

Our previous studies showed that xanthone derivatives with N-(2-methoxyphenyl)piperazine fragment have an affinity to the 5-HT1A receptor and show antidepressant-like properties in rodents. In this study, we tested three xanthone derivatives, HBK-1 (R, S) and its enantiomers, in which we increased the distance between the piperazine and xanthone fragments by using a hydroxypropoxy linker. We hypothesized that this would increase the binding to the 5-HT1A receptor and consequently, pharmacological activity. We aimed to assess the in vitro and in vivo pharmacological activity of the xanthone derivatives. We evaluated the in vitro affinity for serotonin 5-HT1A and 5-HT2A receptors and serotonin transporter. We also determined the intrinsic activity at the 5-HT1A receptor. We investigated the antidepressant-like properties and safety after acute administration (dose range: 1.25-20 mg/kg) using the forced swim, tail suspension, locomotor activity, rotarod and chimney tests in mice. We also evaluated the basic pharmacokinetic parameters. Our results indicated that the compounds showed a high affinity for the 5-HT1A receptor but very weak antagonistic properties in the Ca2+ mobilization assay; however, they showed significant agonistic properties in the β-arrestin recruitment assay. In both behavioural tests the studied xanthone derivatives showed antidepressant-like activity. Pre-treatment with p-chlorophenylalanine or WAY-100635 abolished their antidepressant-like activity. None of the compounds caused motor impairments at antidepressant-like doses. The racemate penetrated the blood-brain barrier and had a relatively high bioavailability after intraperitoneal administration. Xanthone derivatives with N-(2-methoxyphenyl)piperazine fragment and hydroxypropoxy linker show increased binding to the 5-HT1A receptor and may represent an attractive putative treatment candidate for depression.

Serotonin 2A (5-HT2A) receptors and dopamine D2 receptors are intimately related to the physiology and pathophysiology of neuropsychiatric disorders. A large number of studies have reported the effectiveness of psychotropic agents targeting 5-HT2A and D2 receptors in these disorders. In addition to the individual functions of these receptors, the interaction between the two neurotransmitter systems has been studied in the living brain. However, little is known about their regional relationship in individual human brains. We investigated regional relationships between 5-HT2A and D2 receptors using positron emission tomography (PET) and a bicluster analysis of the correlation matrix of individual variation in the two receptor densities to identify groups of distinctive regional correlations between the two receptors.MethodsSeven healthy volunteers underwent PET scans with [18F]altanserin and [11C]FLB 457 for 5-HT2A and D2 receptors, respectively. As a measure of receptor density, a binding potential (BP) was calculated from PET data for 76 cerebral cortical regions. A correlation matrix was calculated between the binding potentials of [18F]altanserin and [11C]FLB 457 for those regions. The regional relationships were investigated using a bicluster analysis of the correlation matrix with an iterative signature algorithm.ResultsWe identified two clusters of regions. The first cluster identified a distinct profile of correlation coefficients between 5-HT2A and D2 receptors, with the former in regions related to sensorimotor integration (supplementary motor area, superior parietal gyrus, and paracentral lobule) and the latter in most cortical regions. The second cluster identified another distinct profile of correlation coefficients between 5-HT2A receptors in the bilateral hippocampi and D2 receptors in most cortical regions.ConclusionsThe observation of two distinct clusters in the correlation matrix suggests regional interactions between 5-HT2A and D2 receptors in sensorimotor integration and hippocampal function. A bicluster analysis of the correlation matrix of these neuroreceptors may be beneficial in understanding molecular networks in the human brain.

(-)-OSU6162 has promise for treating Parkinson's disease, Huntington's disease and schizophrenia. Behavioral tests evaluating the locomotor effects of (-) and (+)-OSU6162 on 'low activity' animals (reserpinized mice and habituated rats) and 'high activity' animals (drug naive mice and non-habituated rats) revealed that both enantiomers of OSU6162 had dual effects on behavior, stimulating locomotor activity in 'low activity' animals and inhibiting locomotor activity in 'high activity' animals. To elucidate a plausible mechanism of action for their behavioral effects, we evaluated the intrinsic actions of (-)- and (+)-OSU6162, and a collection of other antipsychotic and antiparkinsonian agents at 5-HT2A and D2 receptors in functional assays with various degrees of receptor reserve, including cellular proliferation, phosphatidyl inositol hydrolysis, GTPγS and beta-arrestin recruitment assays. We also tested for possible allosteric actions of (-)-OSU6162 at D2 receptors. Both enantiomers of OSU6162 were medium intrinsic activity partial agonists at 5-HT2A receptors and low intrinsic activity partial agonists at D2 receptors. (+)-OSU6162 had higher efficacy at 5-HT2A receptors, which correlated with its greater stimulatory activity in vivo, but (-)-OSU6162 had higher potency at D2 receptors, which correlated with its greater inhibitory activity in vivo. (-)-OSU6162 did not display any convincing allosteric properties. Both (+)- and (-)-OSU6162 were significantly less active at 27 other monoaminergic receptors and reuptake transporters tested suggesting that D2 and 5-HT2A receptors play crucial roles in mediating their behavioral effects. Compounds with balanced effects on these two receptor systems may offer promise for treating neuropsychiatric diseases.

In vivo occupation of dopamine D1, D2 and serotonin (5-HT)2A receptors by a novel antipsychotic drug, SM-9018 (perospirone hydrochloride; cis-N- [4- [4- (1,2-benzisothiazol-3-yl)- 1-piperazinyl]butyl]cyclohexane-1,2-dicarboximide monohydrochloride) and its major metabolite (ID-15036; N-[4-[4-(1,2-benzisothiazol-3-yl)-1-piperazinyl]butyl]-1-hydroxy-1 , 2-cyclohexanedicarboximide) was measured in rat brain using N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), an irreversible antagonist, at these receptor sites. SM-9018 and its metabolite, ID-15036, dose-dependently reversed EEDQ-induced 5-HT2A and D2 receptor inactivation, but not D1 receptor inactivation. At lower doses (0.1 mg/kg i.p.), SM-9018 showed a preferential occupation of the 5-HT2A receptors, with only a small effect on the D2 receptors; while at higher doses (1.0 and 5.0 mg/kg i.p.), it was nearly equipotent in its occupation of both the D2 (77.8%) and the 5-HT2A receptors (78.6%). On the other hand, ID-15036 was more potent in occupying the 5-HT2A than the D2 receptors even at higher doses (1.0 and 5.0 mg/kg i.p.). We previously reported that atypical antipsychotic drugs, such as clozapine, were characterized by a high occupancy of the 5-HT2A receptors, with a low or minimum occupancy of the D2 receptors in vivo. The present study suggests that SM-9018 and its metabolite ID-15036 show a preferential tendency to occupy 5-HT2A receptors, and that the clozapine-like atypical properties of SM-9018 may be due to some pharmacological action of both the SM-9018 itself and its metabolite, ID-15063.

In this study, by homology modelling and molecular dynamics (MD) simulation, models of l-stepholidine (l-SPD) activating the 5-HT1A and D1 receptors were constructed. In 100-ns MD simulations, the D1 and 5-HT1A receptors were activated by the partial agonist l-SPD, conforming with the global toggle switch activation model and the sequential activation model. The residues Y7.53 and Y5.58 swing significantly between different transmembrane (TM) domains after activation. Similarities between D1 and 5-HT1A included (1) the outward motion of TM-5; (2) the ionic lock was independent of the tilt of TM-6 and (3) there was an apparent bending of TM-6, and the ring of l-SPD formed strong π–π interactions with residue W6.48. Differences between the two included the following: (1) in 5-HT1A, l-SPD formed a hydrogen bond with Ala1725.46 of TM-5, and the intracellular end of TM-5 moved outward slowly; that hydrogen bond did not form with the D1 receptor; (2) l-SPD formed stronger interactions with D3.32 and W6.48 in the D1 receptor than in the 5-HT1A receptor and (3) the hydrogen bonding network was somewhat different in SPD-5-HT1A and SPD-D1 receptors. We propose the interaction between l-SPD and D3.32 or/and W6.48 is the original driving force during the whole activation process.

It is known that hetaryl(aryl)piperazines possess important neuropharmacological (anxiolytic, antidepressant, neuroleptic, etc.) properties. In the process of studying the relationship between the structure and properties of N-(arylpiperazinyl) butylimides bicyclo[2.2.1]gept-5-en-endo-endo-2,3-dicarbonic acid (compounds 1-5) the neuropharmacological properties and their affinity for D2 and 5-HT1A receptors have been studied in this work. It has been determined by the radioligands method that arylpiperazines 1-5 possess the high affinity for D2 and 5-HT1A receptors. Compounds 1-5 have been found to have the marked sedative and neuroleptic properties. The compounds (1-5) synthesized revealed a dose-dependent pharmacological effect. In lower doses (5 mg/kg) on the model of the conflict test in rats m-tolyl derivatives had the same anxiolytic effect as buspirone (10 mg/kg); in higher doses (10 mg/kg) all compounds revealed the neuroleptic activity on the model of “Waxy flexibility” catalepsy induction. Compounds of this series in the dose of 10 mg/kg reduced the apomorphine-induced stereotypic behaviour by 33.7-86.5% in rats, as well as a refrence drug haloperidol. All of these compounds are nontoxic, the value of their LD50 ≥ 300 mg/kg.

Design, synthesis and evaluation of benzo[a]thieno[3,2-g]quinolizines as novel l-SPD derivatives possessing dopamine D1, D2 and serotonin 5-HT1A multiple action profiles

The atypical antipsychotic olanzapine was compared to other atypical as well as typical antipsychotic agents for in vivo occupancy of D1, D2, D3, 5HT2, and muscarinic receptors in rat brain. Blockade of D2 receptors was determined by measuring the levels of the dopamine metabolite 3,4-dihydroxyphenylacetic acid (DOPAC). To assess the interaction with phosphoinositide (PI)-coupled 5HT2A and muscarinic receptors in vivo, we used a novel radiometric technique to measure in vivo PI hydrolysis. The antagonism of olanzapine and other antipsychotic agents on 5HT2A and muscarinic receptors was determined by in vivo blockade of PI hydrolysis, stimulated by the 5HT2 agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) or the muscarinic agonist pilocarpine. Olanzapine inhibited 5HT2, D2, and D3 in vivo binding with high potency (ID50=0.15, 0.6 and 1.2 mg/kg, IP, respectively), while inhibiting D1 and muscarinic in vivo binding with much less potency (ID50 > 10 mg/kg, IP). The binding of olanzapine to D2 receptors in neostriatum was well correlated with the increase of DOPAC (ED200 = 0.8 mg/kg, IP) in vivo, indicating dopamine D2 antagonism. In vivo PI hydrolysis was increased by DOI in frontal cortex and by pilocarpine in hippocampus up to 2- and 7-fold above the basal level, respectively. The agonist-induced increases in PI hydrolysis were fully blocked by the 5HT2A antagonist MDL100907 and the muscarinic antagonist scopolamine, indicating the mediation by 5HT2A receptors in frontal cortex and PI-coupled muscarinic receptors (ml, m3, and m5) in hippocampus, respectively. Olanzapine was about 8-fold more potent in vivo in blocking DOI-induced stimulation of PI hydrolysis (ID50 = 0.1 mg/kg, IP) than pilocarpine-induced stimulation of PI hydrolysis (ID50 = 0.8 mg/kg, IP). In conclusion, olanzapine is more potent in blocking the 5HT2A receptor than D1, D2, D3 and muscarinic receptors in vivo, consistent with its favorable clinical profiles. In addition, the novel in vivo PI hydrolysis assay proved to be a useful and reliable in vivo method to assess the functional efficacy of compounds that interact with the 5HT2 and muscarinic receptors.

Introduction: Cariprazine, a dopamine D3-preferring D3/D2 receptor partial agonist and serotonin 5-HT1A receptor partial agonist, has two major human metabolites, desmethyl-cariprazine (DCAR) and didesmethyl-cariprazine (DDCAR). The metabolite pharmacology was profiled to understand the contribution to cariprazine efficacy.MethodsIn vitro receptor binding and functional assays, electrophysiology, animal models, microdialysis, and kinetic-metabolism approaches were used to characterize the pharmacology of DCAR and DDCAR. ResultsSimilar to cariprazine, both metabolites showed high affinity for human D3, D2L, 5-HT1A, 5-HT2A, and 5-HT2B receptors, albeit with higher selectivity than cariprazine for D3 versus D2 receptors. In [35S]GTPγS binding assays, cariprazine and DDCAR were antagonists in membranes from rat striatum and from cells expressing human D2 and D3 receptors, and were partial agonists in membranes from rat hippocampus. In cAMP signaling assays, cariprazine, DCAR, and DDCAR acted as partial agonists at D2 and D3 receptors; cariprazine and DDCAR were full agonists, whereas DCAR was a partial agonist at 5-HT1A receptors. Cariprazine, DCAR, and DDCAR were pure antagonists at human 5-HT2B receptors. Cariprazine and DDCAR increased rat striatal dopamine and reduced cortical serotonin turnover. Cariprazine and DDCAR showed similar in vivo D3 receptor occupancy in rat brain; however, cariprazine was more potent for D2 receptor occupancy. Both cariprazine and DDCAR dose-dependently but partially suppressed the spontaneous activity of midbrain dopaminergic neurons in rats, with the parent compound being more potent but shorter acting than its metabolite. Consistent with the D2 receptor occupancy profile, DDCAR was 3- to 10-fold less potent than cariprazine in rodent models of antipsychotic-like activity. Following acute cariprazine administration, DDCAR was detected in the rodent brain but at much lower levels than cariprazine.ConclusionOverall, in vitro and in vivo pharmacological profiles of DCAR and DDCAR demonstrated high similarity with cariprazine, suggesting that the major metabolites of cariprazine contribute significantly to its clinical efficacy.

Combining antagonist/partial agonist activity at dopamine D2 and agonist activity at serotonin 5-HT1A receptors is one of the approaches that has recently been chosen to develop new generation antipsychotics, including bifeprunox, SSR181507 and SLV313. There have been, however, few comparative data on their pharmacological profiles. Here, we have directly compared a wide array of these novel dopamine D2/5-HT1A and conventional antipsychotics in rat models predictive of antipsychotic activity. Potency of antipsychotics to antagonize conditioned avoidance, methylphenidate-induced behaviour and D-amphetamine-induced hyperlocomotion correlated with their affinity at dopamine D2 receptors. Potency against ketamine-induced hyperlocomotion was independent of affinity at dopamine D2 or 5-HT1A receptors. Propensity to induce catalepsy, predictive of occurrence of extrapyramidal side effects, was inversely related to affinity at 5-HT1A receptors. As a result, preferential D2/5-HT1A antipsychotics displayed a large separation between doses producing 'antipsychotic-like' vs. cataleptogenic actions. These data support the contention that 5-HT1A receptor activation greatly reduces or prevents the cataleptogenic potential of novel antipsychotics. They also emphasize that interactions at 5-HT1A and D2 receptors, and the nature of effects (antagonism or partial agonism) at the latter has a profound influence on pharmacological activities, and is likely to affect therapeutic profiles.

Serotonin 5-HT1A receptors are promising targets in the management of schizophrenia but little information exists about affinity and efficacy of novel antipsychotics at these sites. We addressed this issue by comparing binding affinity at 5-HT1A receptors with dopamine rD2 receptors, which are important targets for antipsychotic drug action. Agonist efficacy at 5-HT1A receptors was determined for G-protein activation and adenylyl cyclase activity. Whereas haloperidol, thioridazine, risperidone and olanzapine did not interact with 5-HT1A receptors, other antipsychotic agents exhibited agonist properties at these sites. E(max) values (% effect induced by 10 microM of 5-HT) for G-protein activation at rat brain 5-HT1A receptors: sarizotan (66.5), bifeprunox (35.9), SSR181507 (25.8), nemonapride (25.7), ziprasidone (20.6), SLV313 (19), aripiprazole (15), tiospirone (8.9). These data were highly correlated with results obtained at recombinant human 5-HT1A receptors in determinations of G-protein activation and inhibition of forskolin-stimulated adenylyl cyclase. In binding-affinity determinations, the antipsychotics exhibited diverse properties at r5-HT1A receptors: sarizotan (pK(i)=8.65), SLV313 (8.64), SSR181507 (8.53), nemonapride (8.35), ziprasidone (8.30), tiospirone (8.22), aripiprazole (7.42), bifeprunox (7.19) and clozapine (6.31). The affinity ratios of the ligands at 5-HT1A vs. D2 receptors also varied widely: ziprasidone, SSR181507 and SLV313 had similar affinities whereas aripiprazole, nemonapride and bifeprunox were more potent at D2 than 5-HT1A receptors. Taken together, these data indicate that aripiprazole has low efficacy and modest affinity at 5-HT1A receptors, whereas bifeprunox has low affinity but high efficacy. In contrast, SSR181507 has intermediate efficacy but high affinity, and is likely to have more prominent 5-HT1A receptor agonist properties. Thus, the contribution of 5-HT1A receptor activation to the pharmacological profile of action of the antipsychotics will depend on the relative 5-HT1A/D2 affinities and on 5-HT1A agonist efficacy of the drugs.