Universidade de São Paulo Biblioteca Digital da Produção Intelectual - BDPI Departamento de Patologia - FMVZ/VPT Artigos e Materiais de Revistas Científicas - FMVZ/VPT 2011 Alstonine as an Antipsychotic: Effects on Brain Amines and Metabolic Changes EVIDENCE-BASED COMPLEMENTARY AND ALTERNATIVE MEDICINE, p.1-7, 2011 http://producao.usp.br/handle/BDPI/15416 Downloaded from: Biblioteca Digital da Produção Intelectual - BDPI, Universidade de São Paulo Hindawi Publishing Corporation Evidence-Based Complementary and Alternative Medicine Volume 2011, Article ID 418597, 7 pages doi:10.1093/ecam/nep002 Original Article Alstonine as an Antipsychotic: Effects on Brain Amines and Metabolic Changes Viviane M. Linck,1, 2 Ana P.Herrmann,1 Angeloˆ L. Piato,1, 2 Bernardo C. Detanico,1 Micheli Figueiro,´ 1 Jorge Florio,´ 3 Maurice M. Iwu,4, 5 Christopher O. Okunji,4, 5 Mirna B. Leal,1 and Elaine Elisabetsky1, 2 1 Laborat´orio de Etnofamacologia, ICBS, Universidade Federal do Rio Grande do Sul, Rua Sarmento Leite 500/202, 90050-170 Porto Alegre, RS, Brazil 2 Programa de P´os-Graduac¸˜ao em Ciˆencias Farmacˆeuticas, UFRGS, Porto Alegre, Brazil. Av. Ipiranga, 2752, 1◦ andar, 90610-000 Porto Alegre, RS, Brazil 3 Departamento de Patologia, Faculdade de Medicina Veterin´aria e Zootecnia, Universidade de S˜ao Paulo, USP, S˜ao Paulo-SP 05508-900, Brazil 4 International Centre for Ethnomedicine and Drug Development, University of Nigeria, Nsukka, Nigeria 5 Bioresources Development and Conservation Programme, University of Nigeria, Nsukka, Nigeria Correspondence should be addressed to Viviane M. Linck, [email protected] Received 4 August 2008; Accepted 12 January 2009 Copyright © 2011 Viviane M. Linck et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Managing schizophrenia has never been a trivial matter. Furthermore, while classical antipsychotics induce extrapyramidal side effects and hyperprolactinaemia, atypical antipsychotics lead to diabetes, hyperlipidaemia, and weight gain. Moreover, even with newer drugs, a sizable proportion of patients do not show significant improvement. Alstonine is an indole alkaloid identified as the major component of a plant-based remedy used in Nigeria to treat the mentally ill. Alstonine presents a clear antipsychotic profile in rodents, apparently with differential effects in distinct dopaminergic pathways. The aim of this study was to complement the antipsychotic profile of alstonine, verifying its effects on brain amines in mouse frontal cortex and striatum. Additionally, we examined if alstonine induces some hormonal and metabolic changes common to antipsychotics. HPLC data reveal that alstonine increases serotonergic transmission and increases intraneuronal dopamine catabolism. In relation to possible side effects, preliminary data suggest that alstonine does not affect prolactin levels, does not induce gains in body weight, but prevents the expected fasting-induced decrease in glucose levels. Overall, this study reinforces the proposal that alstonine is a potential innovative antipsychotic, and that a comprehensive understanding of its neurochemical basis may open new avenues to developing newer antipsychotic medications. 1. Introduction potential to increase treatment adherence. Unfortunately, however, it is now recognized that atypical antipsychotics The management of schizophrenic symptoms has never been often induce diabetes, hyperlipidaemia and weight gain a trivial matter, further complicated by the low adherence [5–7]. For instance, patients on clozapine have a 4 times to treatments and the serious side effects of available drugs. higher chance of developing diabetes, hyperglycaemia, and Classical antipsychotics, blocking D2 dopamine receptors, hyperlipidaemia than patients on classical antipsychotics [8]. lead to extrapyramidal effects related to antagonism in the The mechanisms by which these metabolic alterations are nigrostriatal pathway [1], and hyperprolactinaemia due to produced are not entirely understood [9]. antagonism in the tuberoinfundibular pathway [2]. In the Post marketing surveillance has shown that the so- early 1990s a new class of antipsychotics was introduced in called atypical antipsychotic drugs are indeed quite heteroge- the clinic, with the alleged advantage of causing no or mini- neous pharmacologically, varying in effectiveness, as well as mal extrapyramidal (EPS) side effects [3, 4], and the resulting metabolic and EPS side effects [10]. It is thought that atypical 2 Evidence-Based Complementary and Alternative Medicine of Nigeria, Nsukka, and authenticated by Mr A. Ozioko of the Department of Botany of the same university. The fruit N+ rind was separated, dried, and pulverized. The powdered N material was successively extracted with n-hexane, methylene H H chloride, and methanol. Extracts were concentrated under CH3 vacuum using a rotary evaporator. H O O 2.2. Isolation and Identification of Alstonine. Pure alsto- C nine hydrochloride used for this investigation was isolated CH3 from the fruit rind of P. nitida Stampf Th. et H.Dur. O (Apocynaceae). The separations were performed using pH- zone-refining counter-current chromatography as previously Figure 1: Alstonine. described [17]. Briefly, the experiment was performed with a two-phase solvent system composed of methyl tert-butyl ether (MtBE)-acetonitrile-water (2 : 2 : 3, v/v), where triethy- lamine (TEA) was added to the upper organic stationary ff antipsychotics fail to induce significant EPS e ects due to phase as a retainer, and hydrochloric acid (HCl) to the their weakened blockade of D2 dopamine (DA) receptors aqueous mobile phase as an eluter. The basic organic phase combined with interactions with various neurotransmitters, was used as the stationary phase and the acidic lower especially serotonin (5HT) and in particular the 5HT1A, phase was used as the mobile phase. The separation was 5HT2A and 5HT2C receptors [1]. Accordingly, specific DA initiated by filling the entire column with the stationary pathways may be modulated by serotonin receptors, depend- phase using the LC pump, and then loading the sample. ing on the presence of serotonergic receptor sub-types [11]. The sample solution was prepared by dissolving 15.0 g of Given that positive and negative symptoms of schizophrenia alkaloid fraction of the methylene chloride extract of P. nitida are thought to reflect an imbalance in DA mesolimbic and in 100 mL of a phase mixture consisting of equal volumes mesocortical pathways [12], and the problems seen with of each phase. The mobile phase was then pumped into merely blocking DA receptors, it is an attractive idea that the column at 2 mL min−1 while the column was rotated ff 5HT receptors can precisely modulate di erent DA pathways. at 834 rpm in the combined head to tail elution mode The indole alkaloid alstonine (Figure 1)wasidentifiedas [18, 19]. The absorbance of the eluate was continuously the major component of a plant-based treatment given to monitored at 280 nm and 4 mL fractions were collected. The mentally ill patients in Nigeria [13]. Alstonine shows a clear pH of each eluted fraction was measured with a pH meter antipsychotic profile in rodents, closer to atypical than to and fractions were dried using a Speed Vac. Identification classical agents [14]. Apparently, alstonine induces dissim- of pH-zone refining counter-current chromatography pure ff ilar e ects in dopaminergic pathways: while apomorphine- fractions was carried out by using thermospray liquid induced stereotypy and amphetamine-induced lethality were chromatography-mass spectrometry (LC-MS) and by TLC significantly reduced by alstonine, suggesting a decrease co-elution experiments with reference alstonine samples in mesolimbic DA, alstonine reversed haloperidol-induced provided by InterCEDD, Nsukka, Nigeria. catalepsy, indicating that nigrostriatal dopamine transmis- sion is not lessened [14]. Relevant for the treatment of nega- tive symptoms, alstonine also presents anxiolytic properties, 2.2.1. Drugs. Clozapine was purchased from Sigma Chem- involving 5HT2A,C receptors, and reverses interaction deficits ical Co. (St Louis, MO, US), and haloperidol was used as induced by MK801 [15, 16]. commercial Haldol (Janssen Farmaceuticaˆ Ltda, SP, Brazil). In order to better understand the basis of alstonine’s Clozapine was solubilized in HCl (0.1N) and its pH adjusted antipsychotic profile, the aim of this study was to verify to 6.0 with NaOH 0.5N; alstonine and haloperidol were alstonine effects on brain amines in mouse frontal cortex and diluted in water. All drugs were administered intraperi- striatum. Additionally, we examined if alstonine causes the toneally (ip) in a volume of 0.1 mL/10 g of body weight. same hormonal and metabolic changes induced by classical and atypical antipsychotics. 2.2.2. Animals. Experiments were performed using male (CF1) adult albino mice, received from Fundac¸ao˜ Estadual 2. Materials and Methods de Produc¸ao˜ e Pesquisa em Saude´ (FEPPS) at 2 months of age (40–45 g). Animals were maintained in our own animal 2.1. Plant Material. Mature fruits of Picralima nitida Stampf facility under controlled environmental conditions (22 ± Th. et H.Dur. were collected by staff from the Interna- 1◦C, 12 h-light/dark cycle, free access to food (Nuvilab CR1) tional Centre of Ethnomedicine and Drug Development and water), for at least 2 weeks before the experiments. (InterCEDD) in February