Personality, Psychopathology, Life Attitudes and Neuropsychological Performance among Ritual Users of Ayahuasca: A Longitudinal Study

Ayahuasca is an Amazonian psychoactive plant beverage containing the serotonergic 5-HT(2A) agonist N,N-dimethyltryptamine (DMT) and monoamine oxidase-inhibiting alkaloids (harmine, harmaline and tetrahydroharmine) that render it orally active. Ayahuasca ingestion is a central feature in several Brazilian syncretic churches that have expanded their activities to urban Brazil, Europe and North America. Members of these groups typically ingest ayahuasca at least twice per month. Prior research has shown that acute ayahuasca increases blood flow in prefrontal and temporal brain regions and that it elicits intense modifications in thought processes, perception and emotion. However, regular ayahuasca use does not seem to induce the pattern of addiction-related problems that characterize drugs of abuse. To study the impact of repeated ayahuasca use on general psychological well-being, mental health and cognition, here we assessed personality, psychopathology, life attitudes and neuropsychological performance in regular ayahuasca users (n = 127) and controls (n = 115) at baseline and 1 year later. Controls were actively participating in non-ayahuasca religions. Users showed higher Reward Dependence and Self-Transcendence and lower Harm Avoidance and Self-Directedness. They scored significantly lower on all psychopathology measures, showed better performance on the Stroop test, the Wisconsin Card Sorting Test and the Letter-Number Sequencing task from the WAIS-III, and better scores on the Frontal Systems Behavior Scale. Analysis of life attitudes showed higher scores on the Spiritual Orientation Inventory, the Purpose in Life Test and the Psychosocial Well-Being test. Despite the lower number of participants available at follow-up, overall differences with controls were maintained one year later. In conclusion, we found no evidence of psychological maladjustment, mental health deterioration or cognitive impairment in the ayahuasca-using group.

section_content: Ayahuasca is a psychotropic tea obtained from Banisteripsis caapi and Psychotria viridis, two plant species native to the Amazon Basin [1]. Ayahuasca has traditionally played a central role in Amazonian shamanism and in more recent times it has become the sacrament of various syncretic religious groups that have exported its use and increased its popularity worldwide [2]. Attesting this expansion, the anthropological and ethnographical bibliography on its modern religious use includes more than 400 scientific papers, book chapters, PhD theses and articles in popular magazines written in ten different languages. Some estimate there are around 20,000 regular religious ayahuasca users in the twentythree countries where the so called ''ayahuasca religions'' are present [3]. In the US, Canada, Holland, and Brazil, federal laws protect the religious use of ayahuasca and in Peru it has recently been declared part of the National Cultural Heritage [4]. 

Despite the legal protection enjoyed in some countries, ayahuasca use is not without controversy. P. viridis contains the hallucinogen N,N-dimethyltryptamine (DMT; [5] ), a compound listed in the 1971 Convention on Psychotropic Substances. However, no plants (natural materials) containing DMT are at present controlled under the said convention [6]. Analogously to other serotonergic hallucinogens, DMT is thought to elicit its psychotropic effect through stimulation of the 5-HT 2A receptors [7, 8]. However, unlike other hallucinogens DMT is not active when administered orally, as it is readily metabolized to 3indoleacetic acid by monoamine oxidase [9]. However, B. caapi contains harmala alkaloids that reversibly block the metabolic breakdown of DMT, leading to psychoactivity [10, 11]. 

Clinical studies have shown that ayahuasca induces a modified state of awareness that includes dream-like imagery with eyes closed, increased insight and intense emotions [10, 12]. CNS effects can also evidenced as time-dependent increases in the relative energy of the beta band of the electroencephalogram [11, 13, 14]. Effects start between 30 and 45 minutes, peak between 90 and 120 minutes and are resolved by 240 minutes [10, 12]. Physiological modifications include moderate increases in blood pressure, elevations of blood cortisol and prolactine and lymphocyte redistribution [10, 11, 12, 14]. Pharmacodynamic changes closely follow the plasma concentrations of DMT, which peaks at 90-120 min and shows an elimination half-life of one hour [10]. 

The increasing number of individuals using ayahuasca on a regular basis has raised public health concerns [2]. Drugs of abuse such as heroin, cocaine, alcohol or amphetamines share a common neurobiological mechanism which involves the so-called ''neural reward system'', inducing functional changes in brain structures related to pleasure such as the striatum and the dopaminergic ventral-tegmental area [15]. The activation of this neural circuit is considered to play a crucial role in modulating the consequences of drug abuse, which may include psychological, medical, legal, employment and family problems [16]. DMT, on the other hand is a serotonergic drug, binding to postsynaptic 5-HT 2A receptors [17, 18]. Although some studies have found that hallucinogenic drugs like psilocybin or LSD (lysergic acid diethylamide) may also modulate dopaminergic neurotransmission [19] [20] [21], a neuroimaging study using SPECT (Single Photon Emission Computerized Tomography) found that ayahuasca increases regional brain blood flow in frontal and paralimbic areas [22], but did not find any changes in reward-related regions such as the striatum or the midbrain. Nevertheless, in a prior study by our group [23], we assessed addiction severity in two samples of long-term members of the ayahuasca religions and we did not find participants to be ''addicted'' to the tea. Nor did we observe the deleterious psychosocial effects commonly associated with drugs of abuse. 

Despite the above findings, the question remains as to whether the continued use of ayahuasca may have an impact on personality and general mental health (negative effects other than those directly related to addiction) and cognition. On the phenomenological level, the acute cognitive, emotional and perceptual modifications brought about by ayahuasca are quite intense [11] and have been described to potentially cause psychiatric complications in some individuals [24, 25]. From a biological perspective, ayahuasca induces activation of prefrontal and temporal regions of the brain [22, 26], an activatory effect probably mediated by glutamate release [7, 8, 27]. Mental health and cognitive performance of long-term ayahuasca users has not been well-studied, and the few papers published to date on the impact of chronic ayahuasca use have failed to detect negative neuropsychological [28, 29] or psychopathological [30, 31] effects. For a review see Bouso and Riba [32]. 

In this paper we report the results of a study specifically designed to evaluate personality, life attitudes, mental health and neuropsychological performance in a relatively large number of ritual ayahuasca users and their matched controls. The investigation involved one initial assessment and a follow-up one year later.
section_name: Introduction

section_name: Methods

section_content: Participants belonging to ayahuasca-using groups were recruited after a meeting between the research team and members of several Brazilian ayahuasca churches. The inclusion criterion was to have been taking ayahuasca for a minimum of 15 years with a frequency of at least twice a month. Control subjects were recruited to match the age, sex and educational level of ayahuasca users. Controls were only accepted if they had taken ayahuasca a maximum of 5 times. Care was taken to make sure that the majority of individuals in the comparison groups were also actively practicing some form of religion but without the ayahuasca-using component. Participants were distributed as follows:

1. 1. Jungle sample. Ayahuasca group: 56 ayahuasca users from a community within the Amazon rain forest. 

This group was recruited from Ce ´u do Mapia ´, a community of religious ayahuasca users in the Brazilian State of Amazonas. Ce ´u do Mapia ´is the headquarters of the Centro Ecle ´tico da Fluente Luz Universal Raimundo Irineu Serra (CEFLURIS), an important ayahuasca church within the Santo Daime movement, with branches throughout South-America, the US, Canada, Europe and Japan. Ce ´u do Mapia `was established in 1983 by the founder of CEFLURIS, Sebastia ˜o Mota de Melo (known as Padrinho Sebastia ˜o), and it has since received migrants from other parts of Brazil, South-America, and overseas. Its current population is estimated at 600 and it includes men and women of all ages and children. CEFLURIS is a church of the Santo Daime, a syncretic religion that combines Christian, indigenous, Afro-American and esoteric traditions. The experiences attained by followers during rituals are interpreted as contact with 'God' and the 'spirits' and other archetypical 'entities' from their religious beliefs and doctrine (for a comprehensive study of the Santo Daime religion see MacRae [33] ). The mean frequency of ritual attendance in this group was about six times per month. The estimated average lifetime exposure to ayahuasca in this group ranged between 360 and 1080 times. 

Control group: Ce ´u do mapia ´comparison group. A group of 56 controls was recruited from Boca do Acre, the nearest town to the Ce ´u do Mapia ´community. Boca do Acre is located deep in the heart of the Amazon rain forest, has a strong agricultural economy, and is a typical small Amazonian town of about 7,000 inhabitants. Of the 56 controls, only 7 had ever ingested ayahuasca. Five participants had taken it once and the other two had taken it two times. 

1. 2. Urban sample. Ayahuasca group: Urban-based ayahuasca users. 

This group consisted of 71 members of another ayahuasca religious group called Barquinha, located in the city of Rio Branco. The city of Rio Branco, the capital of the State of Acre, has about 150,000 inhabitants and it hosts other branches of the Santo Daime and other ayahuasca churches such as the Unia ˜o do Vegetal and the Alto Santo. The frequency with which Barquinha members attended rituals in our sample was about eight times per month. The estimated average lifetime exposure to ayahuasca in this group ranged between 360 and 1440 times. 

Control group: Urban-based comparison group. Fiftynine controls were recruited in the city of Rio Branco as a comparison group. Of the 59, only 4 had ever ingested ayahuasca and they had all taken it once. 

The study was conducted in accordance with the Declarations of Helsinki, as amended in Edinburgh 2000, and subsequent updates. All subjects signed an informed consent form prior to participation. The study was approved by the human research committee of UNINORTE University (Rio Branco, Acre State, Brazil).
section_name: Participants

section_content: 2. 1. Sociodemographic variables. Age (years), sex (male/ female) and years of education were used to match study and control groups. Additional sociodemographic indicators such as employment status (according to Hollingshead's categories [23] ), race, marital status and religion were recorded for comparison purposes.
section_name: Study Variables

section_content: inventory -TCI. The TCI is based on the psychobiological model of personality developed by Cloninger and coworkers [34]. The temperament dimensions are assumed to be independently inheritable and to manifest in early development, while the character dimensions are assumed to be more influenced by sociocultural learning and maturation. The TCI has 240 items with a true/false option response. The four primary dimensions of temperament and their facets are: Harm Avoidance (HA): HA1-Anticipatory Worry vs. Uninhibited Optimism; HA2-Fear of Uncertainty vs. Confidence; HA3-Shyness with Strangers vs. Gregariousness; HA4-Fatigability and Asthenia vs. Vigor; Novelty Seeking (NS): NS1-Exploratory Excitability vs. Stoic Rigidity; NS2-Impulsiveness vs. Reflection; NS3-Extravagance vs. Reserve; NS4-Disorderliness vs. Regimentation; Reward Dependence (RD): RD1-Sentimentality vs. Insensitivity; RD3-Attachment vs. Detachment; RD4-Dependence vs. Independence; and Persistence (P). The three dimensions of character are: Self-Directedness (SD): SD1-Responsibility vs. Blaming; SD2-Purposefulness vs. Lack of Goal-Direction; SD3-Resourcefulness; SD4-Self-Acceptance vs. Self-Striving; SD5-Congrugent Second Nature; Cooperativeness (C): C1-Social Acceptance vs. Social Intolerance; C2-Empathy vs. Social Disinterest; C3-Helpfulness vs. Unhelpfulness; C4-Compassion vs. Revengefulness; C5-Integrated Conscience; and Self-Transcendence (ST): ST1-Self-Forgetfulness vs. Self-Conscious Experience; ST2-Transpersonal Identification vs. Self-Isolation; ST3-Spiritual Acceptance vs. Rational Materialism. In this study we used the Brazilian Portuguese version of the TCI adapted by Fuentes and coworkers [35].
section_name: Personality traits: Temperament and character

section_content: The Symptom Check-List-90-Revised -SCL-90-R. The SCL-90-R [36] is a self-report questionnaire that assesses 9 psychopathological symptomatic dimensions including 90 likert-type items that are scored from 0 to 4: Somatization (SOM), Obsessive-Compulsive (O-C), Interpersonal Sensitivity (I-S), Depression (DEP), Anxiety (ANX), Hostility (HOS), Phobic Anxiety (PHOB), Paranoid Ideation (PAR), and Psychoticism (PSY). The scale also provides 3 additional psychopathological indices: General Severity Index (GSI), Positive Symptoms Distress Index (PSDI), and Positive Symptoms Total (PST). For all the scales higher scores imply worse symptomatology. In this study we used the Brazilian Portuguese version adapted by Tosello [37].
section_name: Psychopathological status:

section_content: Stroop Color and Word Test. The Stroop test [38] assesses conflict monitoring and resolution (resistance to interference), cognitive tasks involving the anterior cingulate-dorsolateral prefrontal system and the rostroventral prefrontal cortex [39]. In this test subjects must first read a list of color names (''red'', ''green'', ''blue'') written in black ink. When this is completed, a list of ''X'' printed in different colors (red, green, blue) is presented and the subject must indicate the color in which each element is printed. 

Finally, a third list is presented to the participant containing a list of color names (''red'', ''green'', ''blue'') but printed this time in an incongruent ink color. As with the second list, the participant is asked to indicate the color in which each element is printed. The numbers of correctly read (first list) and correctly reported items (lists two and three) in 45 seconds are recorded. Dependent variables are the total number of words read (W), the total number of correctly identified colors in the second list (C) and the total number of color incongruent words read (IW). Finally, a ''Resistance to Interference'' measure (RI) is calculated according to the following formula: RI = IW-(CxW/C+W). Better performance is reflected as higher scores on IW and RI.
section_name: Neuropsychological performance and behavior. The

section_content: The WCST [40] is considered a measure of executive function in that it requires strategic planning, organized searching, the ability to use environmental feedback to shift cognitive set, goaloriented behavior, and the ability to modulate impulsive responding [41]. Anatomically, the WCST involves the dorsoand ventrolateral prefrontal cortices [42]. The test consists of 4 stimulus cards placed in front of the subject. The first has a printed red triangle, the second two green stars, the third three yellow crosses, and the fourth four blue circles. Subjects are then given two decks each containing 64 response cards, which have designs similar to those on the stimulus card, varying in color, geometric form, and number. Subjects are told to match each of the cards and are given feedback as to whether they are doing right or wrong. The sorting rule is changed at fixed intervals but no warning is provided that the sorting rule has changed. There is no time limit to perform the test.
section_name: The Wisconsin Card Sorting Test (WCST)

section_content: The LNS [43] is a measure of working memory, a task involving dorso-, ventrolateral and orbitofrontal prefrontal cortices [44]. Subjects are verbally presented with a random series of numbers and letters which they have to report back in a specified order, i. e, numbers in ascending order and letters in alphabetical order. Series of increasing length are presented to the subject until an error is committed. The score is the maximum number of elements in the series correctly reported by the participant. Higher scores indicate better performance.
section_name: The Letter-NumberSequencing (LNS) from the WAIS-III

section_content: The FrSBe [45] is a rating scale designed to measure behaviors associated with damage to the frontal lobes and systems of the brain. This questionnaire was used to assess hypothetical frontal lobe alterations that could potentially go undetected with classical neuropsychological tests but that could have an impact on everyday life. The questionnaire comprises 46 likert-type items with 5 response options. The items are distributed into 3 subscales: Apathy/Akinesia (14 items), Disinhibition/Emotional Dysregulation (15 items), and Executive Dysfunction (17 items). Higher scores reflect worse frontal function. A global score is computed adding up the scores of the individual scales. We used the self-report version of the scale adapted to Brazilian Portuguese by our team.
section_name: The Frontal Systems Behaviour Scales (FrSBe)

section_content: The spiritual orientation inventory (SOI). The SOI [46] is a measure of spirituality based on the humanistic model and is designed to assess the spirituality of those affiliated with traditional religion. It is a 85-Likert-type item self-report questionnaire. Items are distributed into nine major components: Transcendent Dimension, Meaning and Purpose in Life, Mission in Life, Sacredness of Life, Material Values, Altruism, Idealism, Awareness of the Tragic, and Fruits of Spirituality. Each item has 7 response options The questionnaire was adapted into Brazilian Portuguese by our team. 

The purpose in life test (PLT). The PLT [47] is a measure of a subject's perceived ''meaning of life'' versus ''existential vacuum'' and is based on Victor Frankl's Logotherapy. It consists of 20 items, each rated on a 7-point scale ranging from 1 (low purpose) to 7 (high purpose). The total score can range from 20 (low purpose) to 140 (high purpose). The questionnaire was adapted into Brazilian Portuguese by our team.
section_name: Life Attitudes and Psychosocial Well-Being

section_content: Psicosocial). The BIEPS [48] is a measure of psychosocial well-being composed of a global and four specific dimensions: Self-Acceptance, Autonomy, Psychosocial Bonds, and Projects. It consists of 13 items with three response options (agree, nor agree nor disagree, disagree). The questionnaire was adapted into Brazilian Portuguese by our team.
section_name: The psychosocial well-Being (BIEPS-Bienestar

section_content: 3. 1. Sociodemographic variables. In order to match the samples, age and years of education were compared using independent samples Student's t test. Although not a matching variable, employment status was also compared between users and controls by means of Student's t test. The distribution of gender, race, marital status and religion between ayahuasca users and controls in each sample were analyzed by means of x2. 

3. 2. Personality, psychopathology, neuropsychology and life attitude variables. Due to the longitudinal nature of the design, we were unable to contact all the participants at the second assessment. Also, due to the field nature of the study, data from some tests was lost for some subjects. In order to maximize sample size and statistical power, we used the data from all subjects available for a given test. 

Individual and group scores were obtained for the different variables. For each variable a two-way analysis of variance (ANOVA) was performed with two between-subjects factors, i. e., Group (ayahuasca users vs. controls) and Sample (jungle vs. urban). Each ANOVA was performed in the first assessment and in the second assessment 8-12 months later. Results were considered significant for p values,0. 05.
section_name: Statistical Analysis

section_name: Results

section_content: Results concerning the sociodemographic characteristics of the samples are presented in Table 1. 

The Jungle sample consisted in the first assessment of 56 regular ayahuasca users and 56 controls. No significant differences were found between ayahuasca users and controls in sex, age, years of education or income either in the first or second assessment. In the Jungle sample, a statistical difference was noted in employment, with the comparison group being more qualified according to the Hollingshead categories [23]. A total of 88 volunteers from the Jungle sample were assessed in a follow-up one year later: 39 from the ayahuasca group and 49 from the comparison group. No statistical differences were found. Other demographic data such as race, marital status, and religion are also shown in the table. Both ayahuasca users and controls were mainly of whites and mestizos. The predominant marital status was ''never married'' in the ayahuasca users, and ''married'' in the controls. All ayahuasca users in the Jungle sample were members of CEFLURIS, and all but 3 individuals from the control group were followers of other Christian religions (Catholicism followed by Protestantism). 

The Urban sample consisted in the first assessment of 71 ayahuasca users and 59 controls. In the second assessment 58 volunteers (39 ayahuasca users and 19 comparisons) were evaluated. We found no significant differences between the groups for sex, age, years of education, income, employment status or income variables, either in the first or second assessment. Participants in both groups were mainly whites and mestizos. Regarding marital status, most volunteers in both groups were either ''never married'' or ''married''. All ayahuasca users in the Urban sample were members of Barquinha. Most comparison subjects defined themselves as followers of traditional Christian religions (Catholicism followed by Protestantism).
section_name: Sociodemographic Variables

section_content: Mean (SD) scores on the different TCI subscales for each sample, group, and time point are shown in Table 2. 

In the first assessment, the two-way ANOVA on temperament dimensions showed a main effect of Group (ayahuasca users vs. controls) for Harm Avoidance [F(1,223) = 17. 73; p,0. 001], with lower values for ayahuasca users than controls and for Reward Dependence [F(1,223) = 6. 98; p = 0. 009], with higher values for ayahuasca users. Despite lower mean values for Novelty Seeking, the overall comparison was not significant. No significant main effect was found either for Persistence and a trend to significance was found for the interaction between Group and Sample The analysis of first assessment scores on TCI character dimensions showed significantly lower scores for ayahuasca users in Self-Directedness [F(1,223) = 9. 56; p = 0. 002], no differences in Cooperativeness and significantly higher scores in Self-Transcendence. 

The significant effect on Self-Directedness was driven by lower scores in Responsibility Character dimensions again showed lower scores for ayahuasca users on Self-Directedness [F(1,116) = 11. 87; p = 0. 001], no differ-ences on Cooperativeness and significantly higher scores on Self-Transcendence [F(1,116) = 7. 3; p = 0. 008]. 

Lower Self-Directedness was due to lower scores on Responsibility [F(1,116) = 24. 75; p,0. 001] and Self-Acceptance [F(1,116) = 12. 23; p = 0. 001]. Again, despite no overall effect on Cooperativeness, Helpfulness remained higher in the ayahuasca-using group [F(1,116) = 4. 20; p = 0. 043] but no differences were seen in Compassion. 

The maintained higher scores on Self-Transcendence could be attributed to Transpersonal Identification [F(1,116) = 7. 56; p = 0. 007] and Spiritual Acceptance [F(1,116) = 17. 94; p,0. 001], but not to Self-Forgetfulness.
section_name: Personality Traits: Temperament and Character Inventory -TCI

section_content: The Symptom Check-List-90-Revised -SCL-90-R Mean (SD) scores on the 9 SCL-90-R dimensions for each sample, group, and time point are shown in Table 3. In the first assessment, ayahuasca users showed significantly lower scores on all 9 psychopathological dimensions, as reflected by a significant Group effect on Somatization In the second assessment, lower scores in the ayahuasca-using group were again observed relative to the control subjects for 7 of the 9 dimensions, i.
section_name: Psychopathological Status:

section_content: Mean (SD) scores on neuropsychological tests for each sample, group, and time point are shown in Table 4.
section_name: Neuropsychological Performance and Behavior

section_content: In the first assessment, ayahuasca-using subjects obtained higher scores on total words [F(1,235) = 21. 00; p,0. 001], total colors [F(1,235) = 29. 38; p,0. 001], number of correctly read incongru-
section_name: The Stroop Color and Word Test

section_content: The statistical analysis of the WCST in the first assessment showed a significantly lower number of Total Errors
section_name: The Wisconsin Card Sorting Test (WCST)

section_content: The first assessment showed that ayahuasca users scored significantly higher on this task than their controls [F(1,237) = 21. 27; p,0. 001]. This difference was larger in the Urban sample, as reflected in the Group by Sample interaction F(1,241) = 5. 86; p = 0. 016]. 

One year later, in the second assessment, this overall effect was observed [F(1,132) = 5. 52; p = 0. 020], but the interaction was not.
section_name: The Letter-NumberSsequencing (LNS) from the WAIS-III

section_content: In the first assessment, ayahuasca users showed lower values on the total FrsBe score [F(1,216) = 31. 81; p,0. 001], on the Apathy/ Transcendent F = 153. 54; p,0. 001 The ANOVA column shows results for the main effect of Group (ayahuasca users vs. controls). df = degrees of freedom. Aya. = ayahuasca-using group.
section_name: The Frontal Systems Behaviour Scales (FrSBe)

section_content: Mean (SD) scores on the different subscales of the 3 life attitudes tests for each sample, group and time point are shown in Table 5.
section_name: Subjective Life Attitudes

section_content: In the first assessment, ayahuasca users showed significantly higher scores on all 9 components of the SOI, as revealed by a main Group effect on Transcendent Dimension In the second assessment, the pattern of results remained unchanged, with higher scores on all components in the ayahuasca-using subjects. Thus, significant Group effects were found on Transcendent Dimension
section_name: The Spiritual Orientation Inventory (SOI)

section_content: Higher scores on this test were found for the ayahuasca-using subjects in the first assessment [F(1,216) = 14. 10; p,0. 001] but not in the second assessment.
section_name: The Purpose in Life Test (PLT)

section_content: The statistical analysis in the first assessment showed significantly higher values for ayahuasca users on the global BIEPS score [F(1,213) = 16. 17 In the second assessment, the Group effect on the global score remained [F(1,115) = 6. 33; p = 0. 013], as did the effect on Psychosocial Bonds [F(1,115) = 4. 10; p = 0. 045], but no other main effect was found. However, a significant Group by Sample interaction was found for Projects [F(1,19) = 4. 75; p,0. 05] revealing lower values for the ayahuasca users as compared to controls in the Urban sample.
section_name: The Psychosocial Well-Being (BIEPS)

section_content: In this paper we present data from a field research study in which personality, mental health, life attitudes and neuropsychological performance were assessed in a large number of ritual ayahuasca users and their matched controls.
section_name: Discussion

section_content: The TCI [49] was used to assess personality. Differences between ayahuasca users and controls were found in several of the temperament dimensions, which are believed to be genetically determined. Higher scores on Reward Dependence (RD) may reflect a feature allowing the group to adapt to a demanding environment such as the tropical rainforest. This interpretation is supported by the significant scores on the RD subdimensions Attachment (RD3) and Dependence (RD4), but not Sentimentality (RD1). This profile is probably useful for life in a small community and in a hostile ecological environment. Participants in the Jungle sample showed a trend to higher scores on Persistence than their urban counterparts. Higher scores on this temperament dimension could explain the adaptation capacity shown by these people to their environment, and the ability to persist as a group despite isolation. Additionally, Harm Avoidance (HA) was lower in the ayahuasca-using subjects, probably reflecting the strength in personality required to undergo regular ayahuasca sessions for long periods of time. It is interesting to note that there were no differences between groups in Novelty Seeking (NS) scores nor in its subscales, including Impulsiveness (NS2). Since high scores in NS and Impulsiveness have been associated with drug use [50, 51], the mere search for new experiences may not be the underlying reason of their involvement with ayahuasca. On the contrary, members of the ayahuasca religions report that the experiences transcend the merely perceptual or recreational aspects of psychoactive drug effects. 

The analysis of Character dimensions showed that ayahuasca users scored significantly higher in Self-Transcendence (ST). Since all participants (users and controls) actively practiced some religion, and Character traits can be influenced by personal experience and culture, this finding could be interpreted as a direct effect of ayahuasca use. Self-Directedness (SD), another Character dimension, is consistently lower in the ayahuasca groups, and may also be related to ayahuasca intake. Used in a religious context, the potent psychotropic effects of ayahuasca may strengthen adherence to the doctrine. The lower Self-Directedness (SD) scores found may reflect the greater relevance of the community over the individual. At the same time, there were no differences between users and controls in Cooperativeness (C). So despite greater Self-Transcendence and spirituality in the ayahuasca-using group, willingness to cooperate with others was not different from that seen in more conventional religions. It would be very interesting to assess if subjects who have decided to leave the group and discontinue ayahuasca use share personality traits with the longterm users. 

In a group of 15 long term urban ayahuasca users, Grob et al. [28] found lower scores on NS and HA and no differences in RD compared to 15 matched non-users, in line with our own results. The higher RD scores in our study, driven mainly by the Jungle sample, may reflect the difference in environment mentioned above. Another research group has found changes in the Temperament dimensions of the TCI after 6 months of regular ayahuasca use in a religious setting in subjects who were initially naı ¨ve to ayahuasca. However, these same subjects did not show changes in the Character dimensions [52]. Based on these findings, a less conservative explanation for the differences observed in Temperament traits in the present study would be that they are a consequence and not the cause of ritual ayahuasca use. This would mean that ayahuasca may induce changes in personality traits traditionally considered inherited. A recent study in which high doses of psilocybin were administered in a supportive setting showed positive long term changes in Openness to Experiences [53]. This temperament trait is considered to be the most substantially heritable trait in the Big Five personality model, and relatively stable through adulthood [54].
section_name: Personality

section_content: The analysis of psychopathology indicators showed the important finding that ayahuasca users scored significantly lower on all nine dimensions of the SCL-90-R. The two immediate explanations for this finding are that either ayahuasca has a low potential to induce psychopathology, or that samples of long-term users suffer from a self-selection bias by which only those who do not experience adverse psychological effects continue ayahuasca use. Regarding the second explanation it is worth mentioning that at follow-up lower scores were still seen on most dimensions, despite the loss in sample size. Similar findings have been reported in the literature. In a study where a group of 32 long term US ayahuasca users were assessed with the same instrument, scores were significantly lower than normative data for 7 of the 9 dimensions [31]. Halpern et al. [55] did not find evidence either of psychopathology in a group of peyote (a mescaline-containing cactus) users when compared to controls. Grob et al. [28] did not find evidence of psychopathology in their ayahuasca-taking sample using the CIDI (Composite International Diagnostic Interview), despite the fact that in the retrospective assessment most subjects met criteria for psychiatric disorders prior to their religious use of ayahuasca. Another study with teenage members of an ayahuasca church did not find differences with the control group, but rather showed a tendency to an improvement in some measures of psychopathology [30]. Barbosa et al. [52, 56] also failed to find psychophatological symptoms both in the short-term after a first ritual ayahuasca experience, and at follow up 6 months after continued use. Some participants even showed a decrease in minor psychopathological symptoms. 

In summary, though there are case reports describing psychiatric complications following ayahuasca intake [24, 25], it appears that current long-term users do not show higher psychopathology. One study reported that some experienced users even show reduced scores of panic and hopelessness while under the effects of the tea [57]. Future research should assess not only long-term users but also ex-users to evaluate whether adverse psychological effects play any role in the decision to discontinue use. The apparent contradiction between reports of psychiatric crisis after acute ayahuasca and the absence of psychopathology in many chronic users should be studied in more detail. 

One last consideration is the potential bias introduced by the self-assessment nature of the SCL-90-R. Subjects may have been inclined to give socially acceptable responses. However, scores on the PST subscale were always higher than 3-4. According to the interpretation norms for the SCL-90-R [58], low scores on this subscale would be indicative of a social desirability bias. Further support for the validity of our present findings is derived from results in the neuropsychological assessment (see below). Psychiatric disorders are commonly accompanied by neuropsychological deficits [59, 60], but these were not observed in the ayahuascausing subjects in the present study.
section_name: Psychopathology

section_content: Based on the administered tests and the Frontal Systems Behaviour Scales, we found no evidence of neuropsychological impairment in the ayahuasca-using group. Furthermore, in general terms they scored better than their respective comparison groups and these differences were maintained one year later. 

These results do not fit the hypothesis of potential frontal impairment secondary to 5-HT 2A receptor activation, and are more in line with prior observations in users of psychedelics. Grob et al. [28] found no working memory deficits in their sample of ayahuasca users, but rather an improvement in one memory subset. Da Silveira et al. [29] did not find deficits in the stroop and other neuropsychological tests in their group of adolescent ritual ayahuasca users. These users did not score differently than their control group in most variables. They did fare worse on some memory subtests, but results were within the normalcy range. Halpern et al. [55] did not find neuropsychological impairment in a group of long term peyote users from the Native American Church. Tests included the Stroop test, the Wisconsin Card Sorting Test and working memory tests. Although more research is needed before definite conclusions can be drawn regarding this drug class, based on the available evidence chronic use of psychedelics does not seem to cause cognitive impairment. 

The lack of cognitive impairment in our ayahuasca users can not be attributed to a lack of sensitivity of the neuropsychological tests administered, as they were sensitive enough to differentiate between users and non users. The Stroop task and the Wisconsin Card Sorting Test tap various cognitive functions such as selective attention, behavioral inhibition, working memory and goaldirected behavior, and are sensitive to PFC damage [61]. Also, these same tests have been found to detect neuropsychological impairment in various groups of drug abusers. For example, the Wisconsin Card Sorting Test has proven sensitive to detect flexibility impairments in non-addicted cocaine polydrug users (between 1-4 gr. of cocaine per month). This population showed more Perseverative Errors, fewer Categories Completed and worse Conceptual Level Responses than matched controls [62]. Also, the Stroop test was sensitive to detect executive dysfunctions in individuals using alcohol [63], cocaine [64] and amphetamines [65]. The same applies to the Letter-Number Sequencing task [66]. However, the detection of differences between users and non-users is known to be influenced by the length of the abstinence period, the severity and duration of the addiction, the use of multiple drugs and the presence of associated psychopathology [67]. In any case, better performance in the drug-using group is rarely found in the literature other than for the psychedelics. Animal research has shown that 5-HT 2A receptor activation plays a role in normal neuropsychological and memory functioning [68] [69] [70] [71]. Another explanation for the present results has to do with motivation. There is evidence that motivation may in fact improve performance of drug users in neuropsychological tasks [72]. While the recruited ayahuasca users may have been motivated to demonstrate the safety of ayahuasca to researchers, the controls did not obtain any specific benefit from their participation in the study. 

Concerning the capacity of the Frontal Systems Behaviour Scales, a self-report questionnaire, to detect impairment, it is worth noting that it has revealed deficits in non-addicted [73] and addicted polydrug users [74, 75]. The lower scores found for our ayahuasca subjects on this measure of prefrontal deficits is consistent with their better neuropsychological performance. This result was found for both samples in the first assessment and in the second.
section_name: Neuropsychological Functions

section_content: All SOI scores were consistently higher in both samples and along time for the ayahuasca users, in consonance with scores on the Self-Transcendence subscale of the TCI. Although a recent study showed no significant differences in spiritually after an ayahuasca session, the magnitude of the observed change was positively correlated with the intensity of the peak of the experience [76]. The qualitative data recorded revealed common spiritual themes among participants [76]. In our first assessment, ayahuasca users showed higher scores on Purpose in Life, although this finding was not replicated one year later. This difference in Purpose of Life may be understood as a consequence of the religious use of ayahuasca, and is compatible with adherence to a religious belief [77]. In line with the above results, ayahuasca users scored higher on subjective psychological well-being. In a previous report where these same participants were assessed on frequency and degree of illicit drug use, ayahuasca users scored lower on the different dimensions of the Addition Severity Index (ASI; [23] ). Taken together, the data point at better general mental health and bio-psycho-social adaptation in the ayahuasca-using group compared to the control subjects.
section_name: Life Attitude and Psychosocial Well-being

section_content: The present study has several limitations. A relevant limitation is that groups were not matched in premorbid IQ, so it is not possible to know whether the differences found in the neuropsychological tests are due to preexisting differences in cognitive abilities or whether ayahuasca used in a ritual context is responsible for the differences observed. Since neuropsychological tasks, and especially working memory tests, are influenced by IQ [78], future studies should control for this variable. From the statistical point of view, the many variables analyzed may have increased the occurrence of type I error. However, given the difficulty of accessing ayahuasca-using populations we chose to administer a comprehensive battery of tests and questionnaires. In addition to the problems associated with self-report questionnaires and the motivational aspects discussed above, a serious limitation, at least in terms of psychopathology, may have been the selfselection bias previously mentioned. Potentially, the assessed individuals may have been those who did not experience any negative neuropsychiatric consequences derived of their continued ayahuasca use. Subjects experiencing adverse consequences may have given up ayahuasca use altogether and may consequently not be among the long-term users accessible to researchers. Future investigation into the neuropsychiatric effects of ayahuasca use should ideally also include people who used ayahuasca regularly in the past but decided to discontinue its use.
section_name: Limitations

section_content: The assessment of the impact of long-term ayahuasca use on mental health from various perspectives (personality, psychopathology, neuropsychology, life attitudes and psychosocial wellbeing) did not find evidence of pathological alterations in any of the spheres studied. Although ayahuasca-using subjects differed in some personality traits, differences did not fit with a pathological profile. Furthermore, ayahuasca users showed a lower presence of psychopathological symptoms compared to controls. They performed better in neuropsychological tests, scored higher in spirituality and showed better psychosocial adaptation as reflected by some attitudinal traits such as Purpose in Life and Subjective Well-Being. Overall differences with the control group were still observable at follow-up one year later.
section_name: Conclusion

section_name: Acknowledgments

section_content: The authors thank all volunteers for their participation. DG and SF thank their former academic advisor Dr Jose ´Luis Linaza, from the Universidad Auto ´noma de Madrid (UAM). JCB thanks his former academic advisor Dr Gregorio Go ´mez-Jarabo, from the UAM. This paper is dedicated to our friends and colleagues Xavier Ferna ´ndez and Manel J. Barbanoj, in memoriam.
section_name: Acknowledgments

section_content: Funding for this study was provided by IDEAA, Instituto de Estopsicologı ´a Amazo ´nica Aplicada, Barcelona (Spain)/Prato Raso (Brazil). We also would like to thank the International Center for Ethnobotanical Education, Research & Service (ICEERS Foundation ; www. iceers. org)for funding the edition costs of this publication. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

section_name: Author Contributions