This animal study (n=8) investigated the efficacy of Banisteriopsis caapi (0.1 -; 0.3 mg/kg harmine) alone and in combination with L-DOPA (4 -; 7 mg/kg) to treat parkinsonian dyskinesia in a marmoset disease model. B. caapi alone has a mild antiparkinsonian effect but does not enhance the L-DOPA response or reduce dyskinesia.
Introduction
Banisteriopsis caapi (B. caapi) contains harmine, harmaline, and tetrahydroharmine, has monoamine oxidase inhibitory activity, and has reported antiparkinsonian activity in humans when imbibed as a tea; however, its effects are poorly documented.
Methods
For this reason, motor function was assessed in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated common marmosets following administration of B. caapi extract (28.4-113.6 mg/kg; po), harmine (0.1 and 0.3 mg/kg; sc), and selegiline (10 mg/kg; sc), alone or with a submaximal dose of L-3,4-dihydroxyphenylalanine (L-DOPA; 4-7 mg/kg).
Results
L-DOPA reversed motor disability, increased locomotor activity, and induced moderate dyskinesia. B. caapi did not increase locomotor activity or induce dyskinesia but at 56.8 and 113.6 mg/kg improved motor disability. The L-DOPA response was unaltered by co-administration of B. caapi. Harmine (0.1 and 0.3 mg/kg) produced a mild improvement in motor disability without affecting locomotor activity or dyskinesia but had no effect on the L-DOPA-induced antiparkinsonian response. Selegiline (10 mg/kg) alone improved motor function to the same extent as L-DOPA, but with only mild dyskinesia, and did not alter the response to L-DOPA, although dyskinesia was reduced.
Discussion
The findings suggest that B. caapi alone has a mild antiparkinsonian effect but does not enhance the L-DOPA response or reduce dyskinesia.
Papers cited by this study that are also in Blossom
Callaway, J. C., Brito, G. S., Neves, E. S. · Journal of Psychoactive Drugs (2011)
Bernschneider‐Reif, S., Poewe, W., Lees, A. · Movement Disorders Clinical Practice (2015)
Schwarz, M. J., Houghton, P. J., Rose, S. et al. · Pharmacology Biochemistry and Behavior (2003)
Parkinson's disease (PD) is a progressive neurodegenerative disorder characterised principally by loss of dopaminergic neurons in the substantia nigra and resulting motor impairments. Symptomatic treatment centres on dopamine replacement, most commonly with L-3,4-dihydroxyphenylalanine (L-DOPA), but chronic L-DOPA use is limited by waning efficacy and the emergence of involuntary movements (dyskinesia). Monoamine oxidase B (MAO-B) inhibitors such as selegiline are used to prolong L-DOPA effects by reducing dopamine metabolism. Banisteriopsis caapi (B. caapi), a plant used traditionally in Amazonian preparations, contains β-carbolines (notably harmine, harmaline, and tetrahydroharmine) with monoamine oxidase inhibitory properties and some clinical reports suggest antiparkinsonian effects when taken as a tea, but controlled preclinical characterisation is limited. Fisher and colleagues set out to determine whether an aqueous extract of B. caapi and one of its principal β-carbolines, harmine, alter motor deficits in a primate model of PD. Using MPTP-treated common marmosets that had been primed with prior L-DOPA to exhibit dyskinesia, the investigators evaluated B. caapi extract, harmine, and the selective MAO-B inhibitor selegiline as monotherapies and in combination with submaximal doses of L-DOPA. The study aimed to establish whether B. caapi or harmine produce antiparkinsonian effects, whether these effects are consistent with MAO inhibition, and whether they modify the motor response or dyskinesia induced by L-DOPA.
The study used a within-subjects design in a single cohort of previously MPTP-treated common marmosets (Callithrix jacchus; n = 8, aged 3–10 years) that had been primed with L-DOPA to express dyskinesia. Animals had received MPTP 1–4 years before the experiments and had undergone washout of at least 4 weeks since any previous drug testing. Treatments were administered in a modified Latin square so that each animal received each treatment on a single occasion; experiments occurred twice weekly with at least 72 hr between treatments. For the selegiline crossover experiment, a 1-week washout was used due to selegiline's long half-life. Observers were blinded to treatment during behavioural scoring. An aqueous extract of the dried B. caapi stem was prepared by two sequential 1-hr water refluxes, filtration, freeze-drying, reconstitution in methanol and HPLC analysis. High-performance liquid chromatography with diode-array detection characterised the extract; reported concentrations in the freeze-dried extract were 35.2 mg/g harmine, 17.6 mg/g tetrahydroharmine and no detectable harmaline. Drug treatments comprised oral B. caapi extract (28.4–113.6 mg/kg, po), subcutaneous harmine (0.1 and 0.3 mg/kg, sc), and subcutaneous selegiline (10 mg/kg, sc), each given alone or combined with submaximal oral L-DOPA methyl ester (4 or 7 mg/kg, po) administered together with the peripheral DOPA-decarboxylase inhibitor benserazide (10 mg/kg po). Domperidone (2 mg/kg po) was given 1 hr prior to treatments to prevent vomiting where indicated. Initial dosing included a low B. caapi dose equivalent to harmine 0.3 mg/kg given in the home cage; no effect was noted, informing the choice of higher oral doses on test days. Behavioural testing took place in individual automated test units after 60 min acclimatisation. Baseline and post-dose assessments (every 30 min for up to 5–6 hr) included automated locomotor activity and blinded observer ratings of motor disability and dyskinesia. Motor disability was scored on an established scale combining alertness, checking, posture, balance, reaction to stimuli, vocalisation and motility to produce a summed score (0 = no disability, 18 = severe disability). Dyskinesia was rated on a 0–4 scale (0 = absent to 4 = severe, continuous). Data were summarised as median time courses and area under the curve (AUC) over 5 hr (trapezoid method). Motor disability and dyskinesia scores were square-root transformed prior to analysis. Statistical testing used repeated measures one-way analysis of variance with Newman–Keuls post hoc tests; significance was set at p < .05.
HPLC analysis of the prepared B. caapi extract identified harmine (35.2 mg/g extract) and tetrahydroharmine (17.6 mg/g extract) but not harmaline. All animals were responsive to L-DOPA, and a submaximal dose of L-DOPA (7 mg/kg, po) produced significant reversal of motor disability, increased locomotor activity and induced moderate to severe dyskinesia, confirming the primed MPTP model. B. caapi extract given alone produced a dose-dependent, U-shaped effect on motor disability with a maximal, moderate reversal at 56.8 mg/kg; AUC analyses showed significant improvement versus vehicle at both 56.8 and 113.6 mg/kg. Across doses (28.4–113.6 mg/kg), B. caapi did not increase locomotor activity and did not induce dyskinesia. When co-administered with a submaximal L-DOPA dose (7 mg/kg, po), B. caapi did not significantly alter the L-DOPA-induced reversal of motor disability, and although locomotor activity tended to decline with higher B. caapi dose, this change was not statistically significant. The lowest B. caapi dose (28.4 mg/kg) produced a small, non-significant reduction in total L-DOPA-induced dyskinesia score. Harmine alone at the higher dose (0.3 mg/kg sc) produced a modest but statistically significant reversal of motor disability lasting about 2 hr, whereas harmine did not alter locomotor activity at either dose tested (0.1 and 0.3 mg/kg). Dyskinesia scores after harmine alone were not significantly different from vehicle, although three animals displayed transient abnormal movements and two animals experienced worsened existing action tremor. In combination with submaximal L-DOPA (4 mg/kg, po), harmine did not significantly change the L-DOPA-induced reversal of motor disability or increase in locomotor activity; peak-dose dyskinesia tended to be higher with harmine but this was not statistically significant. Selegiline (10 mg/kg sc) given alone produced a reversal of motor disability and an increase in locomotor activity similar in magnitude to the submaximal L-DOPA (4 mg/kg, po), though with a reduced peak effect; motor disability was significantly improved versus vehicle. When administered prior to L-DOPA (4 mg/kg), selegiline did not alter the overall L-DOPA-induced motor disability score or locomotor activity but extended the duration of improved motor disability (scores < 8) to about 210 min versus 150 min with L-DOPA alone. Selegiline alone produced fleeting low-grade dyskinesia in some animals but significantly reduced the peak dyskinesia induced by L-DOPA.
Fisher and colleagues interpret their findings as evidence that B. caapi extract and its constituent harmine exert a modest antiparkinsonian effect in the MPTP-treated marmoset when given as monotherapy. The extract improved motor disability without increasing locomotor activity or eliciting dyskinesia, a profile that differs from L-DOPA and may be of interest for early-stage symptomatic treatment. However, B. caapi did not potentiate the antiparkinsonian effect of submaximal L-DOPA nor substantially reduce L-DOPA-induced dyskinesia when co-administered. The authors note that harmine reproduced some of the extract's effects, consistent with a contribution from β-carbolines. HPLC data showed harmine and tetrahydroharmine present in the tea-derived extract but no detectable harmaline; the absence of harmaline may reflect extraction-method differences. Because harmine is a relatively selective MAO-A inhibitor and has been reported to increase dopamine efflux, the investigators propose that direct facilitation of dopamine release rather than MAO-B inhibition is a plausible mechanism for the observed motor benefit. Comparison with selegiline—a selective MAO-B inhibitor—supports this view: selegiline produced a more marked reversal of disability and an increase in locomotion than B. caapi or harmine, suggesting that the latter do not exert strong MAO-B inhibition at the doses used. Selegiline extended the duration of L-DOPA's effect and reduced peak dyskinesia, findings the authors note were somewhat unexpected in the primate model and not entirely concordant with prior animal literature, though they align with clinical reports of MAO-B inhibitors extending L-DOPA benefit. The investigators acknowledge limitations and uncertainties: the extent of central MAO inhibition achieved by the doses of harmine and extract is not directly measured, the selectivity between MAO isoforms may be critical, and other pharmacological actions cannot be excluded. They also flag variability in extraction and analytic methods as a factor influencing constituent profiles. Finally, the authors point out clinical relevance: the lowest effective extract dose approximated a human-consumable tea volume and aligns with previous clinical observations of single-dose benefit, supporting the idea that B. caapi or harmine might offer mild symptomatic benefit in early PD but are unlikely to provide additive advantages when used alongside L-DOPA in later disease stages.
Parkinson's disease (PD) is a progressive, neurodegenerative disease that affects motor function resulting primarily from the degeneration of dopaminergic neurons in the substantia nigra pars compacta. Treatment is based on dopamine replacement therapy, and the amino acid precursor of dopamine, L-3,4-dihydroxyphenylalanine (L-DOPA), is the most widely used and efficacious symptomatic treatment; however, chronic use commonly leads to a loss of efficacy and the development of unwanted involuntary movements (dyskinesia;. To extend the duration of effect of L-DOPA and reduce the dose required, monoamine oxidase B (MAO-B) inhibitors, such as selegiline and rasagiline, are employed to inhibit the metabolism of dopamine derived from L-DOPA. Banisteriopsis caapi (B. caapi) is a liana, endemic to the Amazon, basin known to contain a number of β-carbolines such as harmine, harmaline, and tetrahydroharmine that possess MAO inhibitory properties. Extracts of B. caapi have been shown to be effective in treating PD when used in combination with L-DOPA by extending its duration of action possibly as a result of reduced dopamine metabolism and enhanced release of dopamine from dopaminergic neurons. It is thought that these effects of B. caapi are mediated through its constituent β-carbolines. Indeed,90 years ago, Lewin and Beringer both reported the successful use of harmine as monotherapy in the pre-L-DOPA era for the treatment of PD and although marketed in the 1920s for this purpose, it fell out of use because of low efficacy (see. More recently,reported that short-term intravenous administration of harmine used in combination with L-DOPA increased its plasma concentration and raised dopamine levels in the striatum of rats and rabbits. This supported the idea that MAO inhibition by B. caapi extracts underlies the antiparkinsonian activity observed, but this has never been formally tested. The purpose of this study was to determine the effects of B. caapi extracts and one of its constituent β-carbolines, harmine, on motor dysfunction in a primate model of PD. To that end, we have used the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated common marmoset primed by previous chronic treatment with L-DOPA to exhibit dyskinesia, an established model of the motor symptoms of PD. The treatments were evaluated both as monotherapy and in combination with L-DOPA and compared with the established MAO-B inhibitor, selegiline. An extract from the dried whole stem of B. caapi was prepared as previously described, approximating to the method of preparation of the extract used in Ecuador for the earlier clinical trials in PD. The stem was ground into a powder and then refluxed (50 g/500 ml distilled water) for 1 hr. After filtration, the remaining plant material was refluxed (50 g/500 ml distilled water) for 1 hr as before and filtered, and the two filtrates were combined and freezedried. The freeze-dried aqueous extracts were then reconstituted in methanol and analysed by high-performance liquid chromatography (HPLC) with diode array for levels of harmine, harmaline, and tetrahydroharmine.
The samples were analysed on a HP1050 HPLC system equipped with an autosampler, a quaternary pump, and a diode-array detector. A Zorbax SB C-18 2.1 mm × 10 cm (particle size 3.5 micron) column was employed. A linear gradient of mobile Phase B (acetonitrile containing 0.1% trifluoroacetic acid) over mobile Phase A (0.1% trifluoroacetic acid in water) from 0% to 50% B in 50 min was employed at a flow rate of 0.2 ml/min, and the eluents were monitored at wavelengths between 210 and 280 nm. Data were collected and compared with standard curves harmine, harmaline, tetrahydroharmine, and N,N-dimethyltryptamine using ChemStation software.
All studies were performed in the same group (n = 8) of previously MPTP-treated common marmosets (Callithix jaccus; aged 3-10 years;. Animals were housed in mixed sex or femalefemale pairs under standard housing conditions (23 °C, humidity 50%). They have access to water and marmoset pellets ad libitum and received meals of fresh fruit, cereals, and protein (e.g., egg and chicken) twice daily. Vitamin D supplement is given once per week. Animals were treated with MPTP between 1 and 4 years prior to the start of the study. Following MPTP treatment, the animals became bradykinetic and rigid and some showed action tremor. They exhibited reduced vocalization; however, they were able to eat, drink, and move around the home cage unaided. All animals were responsive to L-DOPA and had been primed with L-DOPA (8-12.5 mg/kg po + benserazide (10 mg/kg po) for up to 28 days) to express dyskinesia between 1 and 4 years prior to these studies. All experiments were performed according to the Animals (Scientific Procedures Act) 1986 under Project Licence No 70/7146, with local ethical approval. All animals had previously been involved in studies involving the assessment of novel drugs for PD and received a drug washout period of at least 4 weeks prior to the start of this study. Drug treatments were performed according to a modified Latin square schedule such that all animals were given all treatments on a single occasion in a randomized manner. Experiments were performed twice weekly with at least 72 hr between treatments to ensure appropriate animal welfare consideration.
Benserazide is a DOPA-decarboxylase inhibitor and prevents the peripheral conversion of L-DOPA to dopamine. Benserazide HCl (10 mg/kg, Sigma, UK) was dissolved in 10% sucrose and then used as the vehicle for dissolving L-DOPA methyl ester (ME) such that benserazide and L-DOPA-ME were then dosed concomitantly in the same syringe. L-DOPA-ME (Sigma, UK) was used throughout this study as it is easily soluble and rapidly hydrolysed to the parent compound and produces the same effect as using the freebase. L-DOPA-ME was dissolved in 10% sucrose (containing benserazide as above) using a salt:base ratio of 1:1.25 to provide the freebase dose (L-DOPA 4 or 7 mg/kg). All doses were corrected for freebase and referred to as L-DOPA. L-DOPA was administered at a submaximal dose determined from dose ranging studies (data not shown) and performed in the animals immediately prior to the present studies. The dose volume was 2 ml/ kg administered by oral gavage (po). Domperidone HCl (2 mg/kg, Sigma, UK) was prepared as a suspension in 10% sucrose and administered orally at a dose volume of 2 ml/ kg, 1 hr prior to treatment to prevent vomiting. B. caapi was analysed for content of harmine, harmaline, and tetrahydroharmine, which are known to be the three main chemicals in this plant. The stem extracts contained 35.2 mg/g extract of harmine, 17.6 mg/g extract of tetrahydroharmine, and no harmaline.6 mg/kg) was dissolved in 20% Kleptose HPB (hydroxypropyl-beta-cyclodextrin-HPBCD, Roquette pharma, France) and administered by oral gavage (po) at a dose volume of 2 ml/kg. Harmine HCL (Santa Cruz Biotechnology, Germany) was dissolved in 0.9% sterile saline (Baxter Healthcare Ltd., UK) using a salt: base ratio of 1:1.17 to provide the freebase dose (Harmine 0.1 and 0.3 mg/kg). The dose volume was 1 ml/kg administered by subcutaneous injection (sc). Selegiline HCl (Sigma, UK) was dissolved in 0.9% sterile saline (Baxter Healthcare Ltd., UK) using a salt base ratio of 1:1.195 to provide the freebase dose of selegiline (10 mg/kg). The dose volume was 1 ml/kg administered by subcutaneous injection (sc). Initially a dose of B. caapi extract (8.5 mg/2 ml po), equivalent to harmine (0.3 mg/kg), was given to all animals in their home cage. No changes in activity or disability were noted (data not shown). Importantly, the animals drank the concentrated tea with no adverse effect. Therefore, the lowest dose of B. caapi extract chosen (28.4 mg/kg) was equivalent to drinking 2.36 ml/kg of the original tea and contained the equivalent of 1 mg/2 ml harmine. On the day of the study, B. caapi extract or vehicle was administered immediately prior to L-DOPA (7 mg/kg, po) or vehicle at t = 0 hr. All drug combinations were administered in a randomized manner according to a modified Latin square such that all animals received all treatments with at least 2 days between each treatment.
The effect of selegiline (10 mg/kg, sc) on basal and L-DOPA (4 mg/kg, po)-induced motor activity was investigated as positive control in a separate experiment. Animals were treated with either selegiline (10 mg/kg sc) plus vehicle (10% sucrose po) or selegiline (10 mg/kg sc) plus a submaximal dose of L-DOPA (4 mg/kg, po). Selegiline was administered 60 min prior to L-DOPA (4 mg/kg, po) or vehicle at t = -60 min. Treatments were administered in a crossover design with 1 week between each treatment to allow washout of drug due to the long half-life of selegiline.
On test days, the animals were removed from their home cages before presentation of breakfast and were placed individually into automated test units. They were allowed a 60-min acclimatization period prior to treatment during which basal locomotor activity, motor disability, and dyskinesia were assessed before (baseline) and for up to a maximum of 6 hr as described below.
Motor disability was assessed simultaneously with locomotor activity through a one-way mirror by experienced observers blinded to treatment. Basal disability was assessed before and once every 30 min after drug treatment for 5 hr using an established motor disability rating scale; alertness (normal = 0, reduced = 1, sleepy = 2); checking (present = 0, reduced = 1, absent = 2); posture (normal = 0, abnormal trunk +1, abnormal tail +1, abnormal limbs +1, flexed = 4); balance (normal = 0, impaired = 1, unstable = 2, spontaneous falls = 3); reaction to stimuli (normal = 0, reduced = 1, slow = 2, absent = 3); vocalization (normal = 0, reduced = 1, absent = 2); motility (normal = 0, bradykinesia = 1, akinesia = 2). These values were summed, a maximum score of 18 indicating severe motor disability, a minimum score of 0 indicating maximum reversal of motor disability.
Dyskinesia was assessed simultaneously with motor disability by experienced observers blinded to treatment. Basal dyskinesia was assessed before and once every 30 min after drug treatment for 5 hr using an established dyskinesia rating scale; 0 = absent; 1 = mild, fleeting, and rare dyskinetic postures and movements; 2 = moderate: more prominent abnormal movements, but not significantly affecting normal behaviour; 3 = marked, frequent and at times continuous dyskinesia affecting the normal pattern of activity; 4 = severe, virtually continuous dyskinetic activity, disabling to the animal and replacing normal behaviour.
Data are presented as median in time course and area under curve (AUC) (calculated over 5 hr using the trapezoid method from the time course curves, Graphpad Prism®). Motor disability and dyskinesia scores were transformed using the formula Y = √(Y) prior to statistical analysis. Changes in the AUC for locomotor activity, reversal of motor disability, and dyskinesia expression were then analysed using a repeated measures one-way analysis of variance (ANOVA) and Newman-Keuls test. Significance was set at p < .05 for all analyses and performed using GraphPad Prism Version 5.00 for Windows (GraphPad Software, San Diego California USA, www.graphpad.com).
The effect of B. caapi alone and combined with L-DOPA (7 mg/kg, po) on locomotor activity, motor disability, and dyskinesia Neither vehicle treatments (Kleptose or sucrose) had any effect on motor disability, locomotor activity and did not induce dyskinesia (Figure); however, the submaximal dose of L-DOPA (7 mg/kg, po) produced a significant reversal of motor disability, an increase in locomotor activity and induced moderate to severe dyskinesia. B. caapi improved overall motor disability scores with a U-shaped dose response curve, with a maximum, but moderate reversal of motor disability following 56.8 mg/kg. This reversal was significantly different to vehicle following both 56.8 and 113.6 mg/kg B. caapi alone (Figureb). No one aspect of motor disability was specifically affected by B. caapi treatment. By contrast, at all doses investigated (28.4 to 113.6 mg/kg), B. caapi alone had no effect on locomotor activity and did not induce dyskinesia (Figures). When given in combination with a submaximal dose of L-DOPA (7 mg/kg, po), B. caapi produced no significant alteration to the L-DOPA (7 mg/kg, po) response, although L-DOPA (7 mg/kg, po)-induced locomotor activity was somewhat reduced with increasing dose of B. caapi, but this was not statistically significant (Figure). The lowest dose of B. caapi (28.4 mg/kg) caused a slight reduction in L-DOPA (7 mg/kg, po)-induced total dyskinesia score; however, this reduction was not statistically different when compared with L-DOPA (7 mg/kg, po) alone (Figure).
The effect of treatment with harmine alone and in combination with L-DOPA (4 mg/kg, po) on locomotor activity, motor disability, and dyskinesia As expected, the submaximal dose of L-DOPA (4 mg/kg, po) produced a significant reversal of motor disability, an increase in locomotor activity, and expression of mild to severe dyskinesia (Figure). The higher dose of harmine (0.3 mg/kg sc) alone produced a small reversal of motor disability that lasted for about 2 hr with total scores significantly improved compared with vehicle (Figure). By contrast, harmine had no effect on locomotor activity at any dose given alone (Figure). Although three animals showed a fleeting display of abnormal movements, the dyskinesia scores were also not significantly different to control (Figures). L-DOPA (4 mg/kg, po)-induced reversal of motor disability or increase in locomotor activity was not altered by any dose of harmine (0.1 and 0.3 mg/kg sc) (Figures). The severity of L-DOPA (4 mg/kg, po)-induced peak dose dyskinesia seen 1 hr after dosing tended to be increased by harmine (0.1 and 0.3 mg/kg; Figure), although the scores were not significantly different to L-DOPA (4 mg/kg, po) alone. No one individual component of the motor disability scores was specifically altered by harmine except in two animals where existing action tremor was worsened.
The effect of treatment with selegiline alone and in combination with L-DOPA (4 mg/kg, po) on locomotor activity, motor disability, and dyskinesia Selegiline (10 mg/kg, sc) alone produced a similar reversal of motor disability and increase in locomotor activity as the submaximal dose of L-DOPA (4 mg/kg, po), although with a reduced peak effect. Motor disability was significantly improved compared with vehicle (Figures). Overall motor disability reversal and locomotor activity induced by L-DOPA (4 mg/kg, po) was not altered by selegiline (Figures); however, the duration of effect was increased with improved motor disability scores (score < 8) up to 210 min compared with 150 min with L-DOPA alone. Interestingly, when given alone, selegiline produced fleeting dyskinesia in some animals (scores 1-2), which did not persist but significantly reduced the L-DOPA (4 mg/kg, po)-induced dyskinesia (Figure). Previous studies suggest that extracts of B. caapi may possess antiparkinsonian activity based on limited clinical experience in humans. This activity may relate to the monoamine oxidase inhibitory actions of the β-carboline content of these extracts and to the subsequent effects that their actions have on dopamine metabolism and release in the striatum. However, alternative mechanisms have been proposed. For example, The MPTP-treated primate is highly responsive to L-DOPA treatment, and as expected from previous studies, administration of a submaximal dose (4-7 mg/kg) of the drug reversed the bradykinesia shown by these animals and normalized motor function as indicated by the reversal of motor disability scores. The drug also induced involuntary movements in the animals as they had been primed by prior L-DOPA exposure to exhibit dyskinesia. In a similar manner, the higher doses of B. caapi administered alone also produced some reversal of motor disability although not to the same extent as L-DOPA (7 mg/kg, po). In contrast to L-DOPA (7 mg/kg, po), there was no increase in locomotor activity and no induction of dyskinesia. This is of interest as increased locomotor activity parallels more closely the induction of dyskinesia than an improvement in motor function related to antiparkinsonian efficacy. This selective effect on motor disability suggests that it may exert an antiparkinsonian action in humans in the absence of the expression of established dyskinesia and would distinguish B. caapi extract from the effects of currently used dopaminergic medications. However, when combined with a submaximal dose of L-DOPA (7 mg/kg, po), the effects of the B. caapi extract were not additive as no improvement in motor disability was seen over and above that produced by L-DOPA (7 mg/kg, po) alone. Similarly, there was no change in the increased locomotor activity or dyskinesia induction produced by L-DOPA (7 mg/kg, po). These results suggest that the mild antiparkinsonian effect of B. caapi may be relevant to early monotherapy in PD but not to the later stages of the disease where adjunct therapy with L-DOPA would be employed. Importantly, the doses of the extract of B. caapi were clinically relevant as the lowest dose was equivalent to drinking approximately 150 ml of the original tea in humans. This is consistent with a study in patients with PD where a significant improvement in motor function was reported following a single dose (200 ml) of B. caapi tea. The question then arises as to whether the modest effects seen with B. caapi extracts reflect the monoamine oxidase inhibitory actions of the constituent β-carbolines. The tea was analysed for content of harmine, harmaline, and tetrahydroharmine and found that harmine and tetrahydroharmine, but not harmaline, was present in the tea. This was surprising as previously it has been reported that harmaline is present in the vine; however, levels of harmaline have previously been reported as low (10% of that of harmine;.The reason for this difference is not clear but may be due to differences in the extraction processes involved in the analysis. Indeed, in this study, we prepared the B. caapi as a tea as it was prepared in water as for earlier clinical trials, which was freeze-dried and then reconstituted with methanol, whereas in the analysis by, the samples were solely extracted by methanol. Administration of the major β-carboline component of B. caapi, harmine, at a dose equivalent to that found in the tea extract, produced a modest but significant reversal of motor disability without increased locomotor activity or dyskinesia. This would support a MAOI-based effect of B. caapi extracts but cannot exclude the possibility of other pharmacological effect. Harmine has been shown to increase striatal dopamine efflux both in vitroand in vivo in rats at equivalent doses. As harmine is a selective inhibitor of MAO-A, and this isoform does not contribute to the metabolism of dopamine in the striatum, it is more likely that this direct effect on DA release explains the reversal of motor disability. Interestingly, the modest improvement in motor disability was not obvious when harmine was combined with a submaximal dose of L-DOPA (4 mg/kg, po). This is in line with the reported effect of combined treatment with a MAOBI and L-DOPA in PD patients but contradicts findings in 6-OHDA lesioned rats where selegiline potentiated the effects of L-DOPA. However, it would depend on the extent to which the doses of harmine used blocked MAO activity in the brain and the relative selectivity for MAO-A and MAO-B, as it is the latter that is believed to control synaptic dopamine concentrations in the denervated striatum in PD, and prior studies have indicated that harmine has little inhibitory effect on MAO-B. To assess what changes in motor function would be expected if inhibition of MAO-B activity contributed to the effects of B. caapi extracts or harmine, we compared the changes observed to those produced by the selective MAO-B inhibitor, selegiline. In PD, selegiline is used both as monotherapy for its mild symptomatic effects and in later disease as an adjunct to L-DOPA therapy. In the MPTP-treated primate, administration of selegiline reversed motor disability and increased locomotor activity with little or no expression of dyskinesia. These effects were more marked than seen with either B. caapi extracts or harmine and suggest that the actions of B. caapi and harmine are not wholly dependent on MAO-B inhibition or that they have a weak inhibitory effect at the doses used. This is in line with the evidence suggesting that both harmine and extracts of B. caapi have selective inhibitory activity at MAO-A, with little or no effect on MAO-B but that both have modest dopamine releasing properties. As for B. caapi and harmine, selegiline administration did not potentiate the effects of a submaximal dose of L-DOPA (4 mg/kg, po) on motor disability and locomotor activity, and although it reduced the peak effects of L-DOPA (4 mg/kg, po), it tended to increase the duration of response, while reducing the expression of L-DOPA-induced dyskinesia. This was unexpected as it does not reflect the clinically accepted effect of selegiline in enhancing dopamine's effect in humans. A MAO-B inhibitor might be expected to potentiate the effects of dopamine formed from L-DOPA at the striatal level. Indeed, although selegiline did not alter the overall score for motor disability, the duration of activity, as measured by a score less than 8, was increased by about 1 hr, reflecting the extension of L-DOPA's response and reduced end of dose deterioration reported clinically. To our surprise, we could find no prior investigation of the symptomatic effects of selegiline in MPTP-treated primates, with the exception of one review that suggests that administration of selegiline increased the antiparkinsonian effects of levodopa and decreased dopamine metabolites. Although there is some suggestion that some β-carbolines can be neurotoxic (Haghdoost-Yazdi, Hosseini, Faraji, Nahid, & Jahanihashemi, 2010;, evidence suggests that harmine is, if anything, neuroprotective with antiinflammatory and antiapoptotic activitythat could be beneficial in the chronic treatment of PD. In addition, these results in the MPTP-treated primate provide support for the reports of the benefits of B. caapi and harmine monotherapy as a mild symptomatic treatment for early PD, as there was little or no evidence to show that there was any additive or synergistic action in conjunction with L-DOPA that is indicated for mid to late stages of the disease. The effects seen may be due at least in part to the MAO inhibitory actions of some of the constituents of B. caapi extract although a direct action on dopamine release cannot be excluded. drug-naive MPTP-treated common marmosets: Effect of dose, frequency of administration, and brain exposure. Movement disorders : Official Journal of the Movement Disorder. Repeated administration of piribedil induces less dyskinesia than L-dopa in MPTP-treated common marmosets: A behavioural and biochemical investigation.
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