Ayahuasca, Pain, and Inflammation: A Systematic Review of Preclinical Studies

Abstract

Pain is a protective mechanism that can be classified into acute and chronic types. Ayahuasca is a psychoactive brew rich in dimethyltryptamine or DMT (a 5-HT_2A receptor agonist), and harmine (a monoamine-oxidase (MAO) inhibitor) used for religious and therapeutic purposes. Previous preclinical and anecdotal evidence suggests that ayahuasca and its compounds have antinociceptive and anti-inflammatory effects due to 5-HT_2A agonism and MAO inhibition. Thus, the current study aims to provide a systematic review of the antinociceptive and anti-inflammatory effects of ayahuasca and its alkaloids in preclinical models. All studies published up to December 2024 were screened and evaluated for eligibility. A total of 1535 publications were identified, of which 29 adhered to the predefined criteria. Reviewed articles reported antinociceptive effects of ayahuasca, harmine, and harmaline. Regarding anti-inflammatory effects, the compounds of ayahuasca, especially harmine, have demonstrated a reduction and an increase in pro-inflammatory and anti-inflammatory cytokines, respectively. Although there are promising results regarding the antinociceptive and anti-inflammatory effects of ayahuasca and its alkaloids, further investigation is needed.

1. Introduction

Pain is a sensory and emotional experience that serves as a protective mechanism for the body and can be categorized based on its duration [1,2]. Acute pain lasts for days or weeks and typically occurs as an immediate response to inflammation. In contrast, chronic pain persists for more than three months, indicating persistent inflammation [2]. Pain perception involves interactions between the peripheral and central nervous systems. Specifically, nociceptors in the skin are activated by noxious stimuli, which are then recognized and transmitted to the somatosensory cortex for processing [3,4,5,6]. Together with pain, inflammation also plays an important role in the body’s defense. When an injury occurs, mediators such as prostaglandins, bradykinin, and cytokines contribute to the activation and sensitization of nociceptors, which aid in host defense [7,8].

Classic hallucinogens—such as psilocybin, lysergic acid diethylamide (LSD), N, N-dimethyltryptamine (DMT), and mescaline—are classified as serotonergic due to their agonistic activity at the 5-HT2A receptor [9,10]. Numerous studies have highlighted the potential therapeutic effects of these substances in various psychiatric conditions, including major depression, anxiety, substance use disorders, and obsessive-compulsive disorder [11,12]. Concerning hallucinogens and pain, the literature is more extensive for preclinical research. In humans, several studies have reported analgesic effects of hallucinogens, particularly in the context of chronic pain, including in patients with cancer, fibromyalgia, cluster headaches, and phantom limb pain [13]. Among the analyzed studies, only three were conducted with healthy volunteers. In 2020, participants showed a positive response to the sublingual administration of 20 µg of LSD, with reduced pain perception and increased tolerance to cold-induced pain [14]. However, in 2023, a clinical trial found that a single oral dose of psilocybin (0.22 mg/kg) did not produce an anti-inflammatory effect [15]. Interestingly, another study published the same year reported a reduction in inflammatory cytokines (TNF-α and IL-6) in healthy volunteers following the administration of 0.17 mg/kg of psilocybin [16].

Ayahuasca is a traditional psychoactive brew originating from the Amazon, primarily used in religious and therapeutic contexts [17]. It is prepared by decocting the Banisteriopsis caapi vine and the leaves of the Psychotria viridis shrub, which together produce its psychoactive effects. B. caapi contains β-carboline alkaloids, including harmine, harmaline, and tetrahydroharmine (THH), while P. viridis is a natural source of N, N-dimethyltryptamine (DMT) [18]. The β-carboline alkaloids act as reversible inhibitors of monoamine oxidase A (MAO-A), preventing the breakdown of DMT in the digestive system and thereby allowing its psychoactive properties to take effect [19]. Additionally, THH functions as a selective serotonin reuptake inhibitor, which may contribute to ayahuasca’s antidepressant effects [19]. Lastly, DMT is a potent hallucinogenic tryptamine that exerts its psychoactive properties by acting as an agonist at serotonergic receptors, particularly 5-HT2A, 5-HT2C, and 5-HT1A [20,21].

Research on ayahuasca and its substances has shown promising results, indicating beneficial effects in the treatment of anxiety, depression, and substance use disorders [22,23]. Although ayahuasca pain-related investigations are scarce, existing findings are encouraging and suggest a potential therapeutic effect. A case report published in 2019 described a patient with musculoskeletal pain who experienced significant and long-lasting analgesic effects after using changa, an herbal blend containing DMT [24]. Other studies have shown that β-carbolines and DMT possess anti-inflammatory properties [25,26] and that MAO-A inhibitors have antinociceptive effects [27]. In a clinical setting, a randomized, double-blind, placebo-controlled study also revealed that individuals with depression exhibited a reduction in C-reactive protein (the pro-inflammatory marker) levels following treatment with ayahuasca [28].

Despite this evidence, the mechanism by which hallucinogens act on the nociceptive process is still not fully understood. Some hypotheses suggest that the serotonergic, opioid, and GABAergic systems may be involved in the inhibition of pain transmission at both central and peripheral levels [13,29]. In light of this, the present study systematically reviewed publications regarding the antinociceptive and anti-inflammatory effects of ayahuasca and its alkaloids in preclinical models.

2. Materials and Methods

The data for the present systematic review were obtained following the PRISMA guidelines for systematic reviews and meta-analyses [30]. The protocol for the systematic review of our study was not registered at PROSPERO.

2.1. Search Strategy

The search was conducted using the following databases: PubMed, MEDLINE, SciELO, and LILACS. Studies published up to 5 December 2024 were included with no language restrictions. The following keywords were used as search strategy: (ayahuasca OR carbolines OR N, N-Dimethyltryptamine OR banisteriopsis OR harmaline OR harmine OR DMT OR peganum harmala) AND (pain OR anti-inflammatory OR analgesia OR antinociceptive OR analgesic effect OR antinociceptive activity OR anti-inflammatory effect). Two independent reviewers screened all studies. The following flow diagram illustrates the phases of the systematic review (Figure 1).

2.2. Selection Criteria and Data Extraction

The inclusion criteria were preclinical studies that evaluated the antinociceptive and anti-inflammatory effects of ayahuasca or any of its alkaloids (harmine, harmaline, THH, and DMT). The samples considered for this review were models in vitro (cells, organs) and in vivo (animals).

The following information was extracted: reference with the year of publication, sample (strain, gender or cells), substance (group, dose, and route), model of induced pain or inflammation, and main results (antinociception, acute or chronic effects, anti-inflammatory, biomarkers).

3. Results

3.1. Selected Studies

The research via databases returned 1535 articles, which were selected by title and abstract. After removing duplicates, 1027 records were screened, and among these, 993 were excluded. Three hundred thirty-four full-text articles were assessed; however, only 25 were deemed eligible. Four articles were identified through hand-searching, assessed for eligibility, and subsequently included in this review. In total, 29 articles were included.

Studies were classified based on pre-established inclusion criteria and categorized into two tables according to antinociceptive and anti-inflammatory results. The search yielded six publications assessing antinociceptive effects and 23 that evaluated anti-inflammatory effects. Regarding the compounds analyzed, three studies administered ayahuasca, while the others used its alkaloids (harmaline, harmine, THH, and DMT) or a combination of those. Harmine was the most common substance, appearing in 16 of the 29 references included, followed by harmaline (five studies) and DMT (four studies). Most studies used rodent models, while four used cell cultures, and only one used a plant model.

3.2. Antinociceptive Effect

As illustrated in Table 1, only six studies have investigated the antinociceptive effects of ayahuasca and its alkaloids (harmine and harmaline) in rodent models. Among them, one study evaluated ayahuasca, another focused on harmaline, and a third examined the combined effects of ayahuasca and harmine. Notably, Pires et al. (2018) assessed the pharmacological interaction between ayahuasca and either morphine, an opioid analgesic, or propofol, a short-acting anesthetic. To this end, they employed behavioral pain assays, including the hot plate, formalin, and acetic acid tests [31]. Their results revealed that ayahuasca not only exhibited intrinsic antinociceptive properties but also potentiated the analgesic effects of morphine, suggesting a potential synergistic interaction between these compounds [31]. Supporting this finding, other studies demonstrated that the combination of harmaline and morphine not only amplified the analgesic effect of morphine but also increased the pain threshold and induced a significant antinociceptive response in the tail flick and hot plate tests [32,33,34].

Among the six selected studies, only one investigated the antinociceptive effects of ayahuasca in a chronic pain model. Lauria et al. (2024) demonstrated that ayahuasca not only exhibited antinociceptive properties during the acute inflammatory phase but also produced sustained effects following experimental neuropathy [35]. Additionally, harmine was shown to induce a prolonged antinociceptive response, further supporting its potential role in pain modulation [35]. Concerning harmaline, another report revealed that harmaline decreased the paw withdrawal latency in the hot plate test [36].

Table 1. Studies of the antinociceptive effects of ayahuasca and its alkaloids.

Reference	Sample	Group/Dose/Route	Model/Test	Result
Pires et al. (2018) [31]	Male Swiss mice	Control (saline i.p. or water.p.o.) Ayahuasca (120 and 1200 mg/kg i.p. or p.o.; Dimethyltriptamine 0.38 mg/mL, harmine 0.32 mg/mL, tetrahydroharmine 1.67 mg/mL, harmaline 0.12 mg/mL) Morphine (1 and 10 mg/kg i.p.)	Hot plate Acetic acid writhing (0.8%. i.p. of acetic acid) Formalin (20 μL)	Ayahuasca (1200 mg/kg) showed an antinociceptive effect in the acetic acid writhing test and the formalin test (but not in the hot plate test) and intensified the analgesic effect of morphine in the hot plate test and the acetic acid writhing test.
Alijanpour & Zarrindast (2020) [32]	NMRI male mice	Control (saline) Morphine sulfate (2 and 6 mg/kg i.p.) Naloxone hydrochloride (0.5 and 1 mg/kg i.p.) N-methyl-d-aspartate (NMDA) (0.06 and 0.1 μg/mouse; intra-vental tegmental area-VTA) Harmaline (1.25 and 5 mg/kg i.p.) dl-2-amino-5-phosphonoheptanoate (D-AP5) (NMDA receptor antagonist) (0.5 and 1 μg/mouse; intra-VTA)	Tail-flick: Pain threshold mu opioid receptor and analgesia Microinjection intra-VTA of NMDA and D-AP5	No significant effects were observed at low doses of morphine (2 mg/kg) or harmaline (1.25 and 5 mg/kg) on pain threshold; Harmaline (5 mg/kg) intensified the analgesic effect of morphine; after a combination of morphine and harmaline at non-effective doses (2 and 5 mg/kg, respectively), pain threshold was increased and antinociception was induced. Microinjection of NMDA (0.06 and 0.1 μg) reduced the antinociceptive effect induced by that combination. In addition, analgesia induced by morphine and harmaline was inhibited by naloxone (0.5 and 1 mg). No effects were observed on pain threshold after microinjections intra-VTA of D-AP5 (0.5 and 1 μg) and NMDA (0.06 and 0.1 μg); However, D-AP5 increased pain threshold and intensified antinociception induced by morphine and harmaline.
Alijanpour et al. (2021) [33]	Male NMRI mice	Control (saline) Harmaline (0, 1.25, 2.5, 5, and 10 mg/kg i.p.) Morphine sulfate (0, 2, 4, and 6 mg/kg. i.p.) Muscimol (0, 200, and 300 ng/mice.i.ventral hippocampal route-vHIP) Bicuculline (1 μL/mouse.i.vHip route)	Tail-flick	Harmaline group showed an antinociception effect at 10 mg/kg. Harmaline (5 mg/kg) combined with morphine (2 mg/kg) produced an antinociceptive effect and enhanced morphine effects. Muscimol enhanced antinociceptive effects of harmaline combined with morphine.
Alijanpour et al. (2022) [34]	Male NMRI mice	Control (saline 10 mL/kg i.p.) Vehicle (0.6 μL/mouse. Microinjection into the basolateral amygdala (BLA)) Harmaline (4, 6, and 8 mg/kg i.p.) ACPA (0.5, 1, or 1.5 ng/mouse.micorinjection into the BLA) AM251 (0.1, 0.5, or 1 ng/mouse.microinjection into the BLA)	Tail-flick Hot plate	Harmaline (6 and 8 mg/kg) prolonged reaction time to thermal stimuli in tail-flick and hot plate tests. AM251 (0.5 and 1 ng/mouse) attenuated the antinociceptive activity of harmaline (8 ng/mouse) in both tests.
Kadyan & Singh (2024) [36]	Swiss albino mice	Control (vehicle 0.1% carboxymethyl cellulose in distilled water 10 mL/kg i.p.) Pathogenic control-vincristine and vehicle (0.1 mg/kg i.p.; once daily for 10 days) Positive control-vincristine (0.1 mg/kg i.p.) + pregabalin (10 mg/kg p.o.) Harmaline (5 mg/kg i.p.; for 14 days) Harmaline (10 mg/kg i.p.; for 14 days) Substance-P (10 μg/kg i.p.) + harmaline (10 mg/kg i.p) + vincristine (0.1 mg/kg i.p.); for 10 days	Vincristine-induced peripheral neuropathy Hot plate Cold plate Acetone drop Beam balance	Harmaline reduced licking and biting behavior and improved paw withdrawal latency in a dose-dependent manner on days 7 and 14 compared to the vincristine-treated group. It also decreased crossing time, with the 10 mg/kg dose showing a stronger effect than pregabalin. Harmaline (5 mg/kg) increased Nrf-2 levels, while 10 mg/kg reversed vincristine-induced Nrf-2 suppression. Additionally, harmaline countered vincristine-induced elevation of IL-1β, reducing its levels at both doses.
Lauria et al. (2024) [35]	Male Swiss and C57BL/6 mice	Control (water p.o.) Ayahuasca (24, 120, 600, and 3000 μL/kg p.o.) Harmine (0, 0.35, 3.5, and 35 mg/kg p.o.) Indomethacin (5 mg/kg p.o.) Morphine (5 mg/kg i.p.) Dexamethasone (2 mg/kg i.p.) Formalin (20 μL) Complete Freund’s Adjuvant (CFA) (20 μL) Gabapentin (70 mg/kg p.o.) Naloxone (5 mg/kg i.p.) Bicuculline (1 mg/kg i.p.) Methysergide (5 mg/kg i.p.) Rimonabant (10 mg/kg i.p.) Phaclofen (2 mg/kg i.p.)	Acute effect: Formalin test CFA Tail flick test Paw edema Acute and chronic effect: Partial sciatic nerve ligation model of neuropathic pain	Acute effect: Ayahuasca (120, 600, and 3000 μL/kg) and indomethacin (5 mg/kg) demonstrated antinociceptive effects at in the second phase of the formalin test. Morphine (5 mg/kg) was effective in inducing antinociceptive effects in both phases of the formalin test. Ayahuasca (600–3000 μL/kg) decreased CFA-induced allodynia but did not reduce paw edema. Allodynia and edema were both reduced by dexamethasone (2 mg/kg). In the tail flick test, ayahuasca did not show an antinociceptive effect, while morphine (5 mg/kg) caused antinoception. Ayahuasca (24–3000 μL/kg) and gabapentin (70 mg/kg) decreased allodynia in experimental neuropathy. The effects of ayahuasca (3000 μL/kg) lasted 10 h, while the effects of gabapentin (70 mg/kg) lasted only 3 h. At 3.5 mg/kg harmine produced an antinociceptive effect that lasted 10 h. Chronic effect: Ayahuasca (120–600 μL/kg twice a day for 14 days) and gabapentin (70 mg/kg daily for 14 days) induced antinoception in experimental neuropathy. Bicuculline (1 mg/kg) and methysergide (5 mg/kg) reverted the antinociceptive effect of ayahuasca (600 μL/kg), however, naloxone (5 mg/kg), phaclofen (2 mg/kg), and rimonabant (10 mg/kg) did not revert. At 3.5 mg/kg twice a day for 14 days, harmine produced an antinociceptive effect that lasted 10 h.

i.p. Intraperitoneally; p.o. oral administration.

Table 1. Studies of the antinociceptive effects of ayahuasca and its alkaloids.

Reference	Sample	Group/Dose/Route	Model/Test	Result
Pires et al. (2018) [31]	Male Swiss mice	Control (saline i.p. or water.p.o.) Ayahuasca (120 and 1200 mg/kg i.p. or p.o.; Dimethyltriptamine 0.38 mg/mL, harmine 0.32 mg/mL, tetrahydroharmine 1.67 mg/mL, harmaline 0.12 mg/mL) Morphine (1 and 10 mg/kg i.p.)	Hot plate Acetic acid writhing (0.8%. i.p. of acetic acid) Formalin (20 μL)	Ayahuasca (1200 mg/kg) showed an antinociceptive effect in the acetic acid writhing test and the formalin test (but not in the hot plate test) and intensified the analgesic effect of morphine in the hot plate test and the acetic acid writhing test.
Alijanpour & Zarrindast (2020) [32]	NMRI male mice	Control (saline) Morphine sulfate (2 and 6 mg/kg i.p.) Naloxone hydrochloride (0.5 and 1 mg/kg i.p.) N-methyl-d-aspartate (NMDA) (0.06 and 0.1 μg/mouse; intra-vental tegmental area-VTA) Harmaline (1.25 and 5 mg/kg i.p.) dl-2-amino-5-phosphonoheptanoate (D-AP5) (NMDA receptor antagonist) (0.5 and 1 μg/mouse; intra-VTA)	Tail-flick: Pain threshold mu opioid receptor and analgesia Microinjection intra-VTA of NMDA and D-AP5	No significant effects were observed at low doses of morphine (2 mg/kg) or harmaline (1.25 and 5 mg/kg) on pain threshold; Harmaline (5 mg/kg) intensified the analgesic effect of morphine; after a combination of morphine and harmaline at non-effective doses (2 and 5 mg/kg, respectively), pain threshold was increased and antinociception was induced. Microinjection of NMDA (0.06 and 0.1 μg) reduced the antinociceptive effect induced by that combination. In addition, analgesia induced by morphine and harmaline was inhibited by naloxone (0.5 and 1 mg). No effects were observed on pain threshold after microinjections intra-VTA of D-AP5 (0.5 and 1 μg) and NMDA (0.06 and 0.1 μg); However, D-AP5 increased pain threshold and intensified antinociception induced by morphine and harmaline.
Alijanpour et al. (2021) [33]	Male NMRI mice	Control (saline) Harmaline (0, 1.25, 2.5, 5, and 10 mg/kg i.p.) Morphine sulfate (0, 2, 4, and 6 mg/kg. i.p.) Muscimol (0, 200, and 300 ng/mice.i.ventral hippocampal route-vHIP) Bicuculline (1 μL/mouse.i.vHip route)	Tail-flick	Harmaline group showed an antinociception effect at 10 mg/kg. Harmaline (5 mg/kg) combined with morphine (2 mg/kg) produced an antinociceptive effect and enhanced morphine effects. Muscimol enhanced antinociceptive effects of harmaline combined with morphine.
Alijanpour et al. (2022) [34]	Male NMRI mice	Control (saline 10 mL/kg i.p.) Vehicle (0.6 μL/mouse. Microinjection into the basolateral amygdala (BLA)) Harmaline (4, 6, and 8 mg/kg i.p.) ACPA (0.5, 1, or 1.5 ng/mouse.micorinjection into the BLA) AM251 (0.1, 0.5, or 1 ng/mouse.microinjection into the BLA)	Tail-flick Hot plate	Harmaline (6 and 8 mg/kg) prolonged reaction time to thermal stimuli in tail-flick and hot plate tests. AM251 (0.5 and 1 ng/mouse) attenuated the antinociceptive activity of harmaline (8 ng/mouse) in both tests.
Kadyan & Singh (2024) [36]	Swiss albino mice	Control (vehicle 0.1% carboxymethyl cellulose in distilled water 10 mL/kg i.p.) Pathogenic control-vincristine and vehicle (0.1 mg/kg i.p.; once daily for 10 days) Positive control-vincristine (0.1 mg/kg i.p.) + pregabalin (10 mg/kg p.o.) Harmaline (5 mg/kg i.p.; for 14 days) Harmaline (10 mg/kg i.p.; for 14 days) Substance-P (10 μg/kg i.p.) + harmaline (10 mg/kg i.p) + vincristine (0.1 mg/kg i.p.); for 10 days	Vincristine-induced peripheral neuropathy Hot plate Cold plate Acetone drop Beam balance	Harmaline reduced licking and biting behavior and improved paw withdrawal latency in a dose-dependent manner on days 7 and 14 compared to the vincristine-treated group. It also decreased crossing time, with the 10 mg/kg dose showing a stronger effect than pregabalin. Harmaline (5 mg/kg) increased Nrf-2 levels, while 10 mg/kg reversed vincristine-induced Nrf-2 suppression. Additionally, harmaline countered vincristine-induced elevation of IL-1β, reducing its levels at both doses.
Lauria et al. (2024) [35]	Male Swiss and C57BL/6 mice	Control (water p.o.) Ayahuasca (24, 120, 600, and 3000 μL/kg p.o.) Harmine (0, 0.35, 3.5, and 35 mg/kg p.o.) Indomethacin (5 mg/kg p.o.) Morphine (5 mg/kg i.p.) Dexamethasone (2 mg/kg i.p.) Formalin (20 μL) Complete Freund’s Adjuvant (CFA) (20 μL) Gabapentin (70 mg/kg p.o.) Naloxone (5 mg/kg i.p.) Bicuculline (1 mg/kg i.p.) Methysergide (5 mg/kg i.p.) Rimonabant (10 mg/kg i.p.) Phaclofen (2 mg/kg i.p.)	Acute effect: Formalin test CFA Tail flick test Paw edema Acute and chronic effect: Partial sciatic nerve ligation model of neuropathic pain	Acute effect: Ayahuasca (120, 600, and 3000 μL/kg) and indomethacin (5 mg/kg) demonstrated antinociceptive effects at in the second phase of the formalin test. Morphine (5 mg/kg) was effective in inducing antinociceptive effects in both phases of the formalin test. Ayahuasca (600–3000 μL/kg) decreased CFA-induced allodynia but did not reduce paw edema. Allodynia and edema were both reduced by dexamethasone (2 mg/kg). In the tail flick test, ayahuasca did not show an antinociceptive effect, while morphine (5 mg/kg) caused antinoception. Ayahuasca (24–3000 μL/kg) and gabapentin (70 mg/kg) decreased allodynia in experimental neuropathy. The effects of ayahuasca (3000 μL/kg) lasted 10 h, while the effects of gabapentin (70 mg/kg) lasted only 3 h. At 3.5 mg/kg harmine produced an antinociceptive effect that lasted 10 h. Chronic effect: Ayahuasca (120–600 μL/kg twice a day for 14 days) and gabapentin (70 mg/kg daily for 14 days) induced antinoception in experimental neuropathy. Bicuculline (1 mg/kg) and methysergide (5 mg/kg) reverted the antinociceptive effect of ayahuasca (600 μL/kg), however, naloxone (5 mg/kg), phaclofen (2 mg/kg), and rimonabant (10 mg/kg) did not revert. At 3.5 mg/kg twice a day for 14 days, harmine produced an antinociceptive effect that lasted 10 h.

i.p. Intraperitoneally; p.o. oral administration.

Taking these results into consideration, the evidence suggests that both ayahuasca and its alkaloids effectively induce antinociceptive responses in animal models and enhance the analgesic effects of morphine, regardless of administered doses.

3.3. Anti-Inflammatory Effect

Several studies assessed the anti-inflammatory actions of ayahuasca compounds. Harmine stands out as the most extensively studied substance, followed by harmaline, DMT, and THH.

3.3.1. Harmine

Harmine consistently demonstrated immunomodulatory and anti-inflammatory effects across various experimental models, primarily through the reduction of pro-inflammatory cytokines.

In the melanoma cell-induced inflammatory process in a mouse model, the administration of 10 mg/kg of harmine decreased the expression of angiogenic factors such as VEGF, iNOS, and COX-2, suppressed the production of cytokines IL-1β, IL-6, TNF-α, and GM-CSF and increased the levels of the anti-inflammatory cytokine IL-2 [37]. The same findings were observed in 2021, with a report of inhibition of IL-12, IL-6, TNF-α, iNOS, and COX-2 after 30 mg/kg of harmine in an endotoxemia model [25]. Complementary to this result, harmine also suppressed the production of TNF-α, IL-6, and NO in LPS-activated immune cells and acute liver injury [38,39]. While most studies report harmine diminishing pro-inflammatory biomarkers, it has also been shown to increase the concentration of the anti-inflammatory cytokines IL-4 and IL-10 in a peri-prosthetic osteolysis model [40]. Additionally, other key regulators of inflammation have been analyzed. In a cisplatin-induced renal tissue toxicity model, the combination of cisplatin and harmine resulted in increased blood urea nitrogen (BUN) levels and higher concentrations of the oxidative stress biomarker malondialdehyde (MDA), while decreasing oxidative activity and reducing apoptotic cell counts [41].

In 2017, Liu et al. compared the effects of 3 mg/kg of dexamethasone, a steroidal anti-inflammatory drug, with 30 mg/kg of harmine in a model of LPS-induced lung injury. Both treatments resulted in a reduction of TNF-α, IL-6, and IL-1β levels [42]. Another study conducted in 2019 demonstrated that harmine at a dose of 50 mg/kg exhibited anti-inflammatory effects similar to dexamethasone (5 mg/kg) by decreasing the secretion of TNF-α, IL-6, and IL-1β in a model of liver injury [43]. In addition, harmine-loaded ethosomes gel at 1g reduced IL-1β, NO, TNF-α, and PGE-2, as well as promoted a prolonged beneficial effect in animals with paw edema [44].

These anti-inflammatory effects have also been investigated at the brain level, demonstrating a potential neuroprotective action of harmine. In LPS-stimulated BV-2 cells, harmine (20 mg/kg) reduced nitric oxide (NO) production and decreased the expression of inflammatory proteins COX-2 and iNOS in the brains of rodents [45]. Analogous results were observed in a subsequent study, in which harmine suppressed inflammation both in the amygdala and in BV-2 cells [46]. Harmine has also shown benefits in memory impairment models, where it decreased TNF-α, IL-6, and IL-1β levels in the hippocampus in an ethanol-induced memory impairment model [47].

3.3.2. Harmaline

Harmaline has also been shown to be effective in inhibiting inflammatory processes. When compared to dexamethasone, harmaline inhibited the expression of IL-1β, IL-6, COX-2, and iNOS in rheumatoid arthritis induction [48]. In another model, harmaline at 5 mg/kg and 10 mg/kg reduced IL-1β levels in vincristine-induced [36].

3.3.3. DMT

Some studies have demonstrated the anti-inflammatory potential of DMT in animal models. Szabo (2014) showed that DMT increases the production of the anti-inflammatory cytokine IL-10, while Szilagyi (2022) observed that its administration combined with harmaline elevates the levels of the inflammatory protein NFkB p65 [49,50]. In neuroinflammation models, DMT was effective in reducing the volume of cerebral ischemia in rats, decreasing the levels of IL-1β, IL-6, and TNF-α, and increasing IL-10 [51]. Similarly, Kelley et al. (2022) reported that both DMT and pharmahuasca (a combination of DMT with harmaline) reduced inflammatory cytokines and the production of reactive oxygen species (ROS) in the hippocampus and prefrontal cortex of the post-traumatic stress model [52]. In the same year, Borbely (2022) found that DMT decreased the levels of IL-1β and TNF-α in a mouse model of Alzheimer’s disease, reinforcing its potential neuroprotective role [53].

3.3.4. THH

In only one study, THH was isolated, and its effect analyzed regarding cytokine production in a sample of BV-2 cells. Results showed a reduction in the levels of IL-6, TNF-α, and IFN at a dose of 18.5 μM, while there was an increase in the cytokine IL-4 at a dose of 2.3 μM [26].

3.3.5. Ayahuasca

Two studies have evaluated the anti-inflammatory effects of ayahuasca. In one study that utilized a vegetal sample, researchers demonstrated a high level of anti-inflammatory activity associated with ayahuasca [54]. In a more recent study, treatment with ayahuasca was found to be effective in reducing the effects of sepsis, as indicated by an increase in IL-4 levels in the prefrontal cortex [55].

The results for cytokines are summarized in

Table 2. Summary of studies evaluating the effects of ayahuasca and its alkaloids on cytokines.

			Cytokines
		Pro-Inflammatory		Anti-Inflammatory
Reference	Substance	IL-1β IL-6 IL-7 IL-17 IL-12 IFN-γ TNF-α		IL-2 IL- 4 IL-10
[36]	Harmaline	↓
[37]	Harmine	↓		↑
[25]	Harmine	↓
[38]	Harmine	↓
[39]	Harmine	↓
[40]	Harmine	↓		↑
[42]	Harmine	↓
[43]	Harmine	↓
[44]	Harmine	↓
[45]	Harmine	↓
[46]	Harmaline	↓
[47]	Harmine	↓
[49]	DMT	↓		↑
[51]	DMT	↓		↑
[52]	Harmaline + DMT	↓		↓
[53]	DMT	↓
[26]	Harmine, harmaline and THH	↓		↓
[55]	Ayahuasca			↑
[56]	Harmine	↓
[57]	Harmine, harmaline and THH			↓

Decreased ↓ and/or ↑ increased in cytokines. IL interleukin, IFN-ϒ interferon-gamma, TNF- α tumor necrosis factor-alpha, DMT dimethyltryptamine, THH tetrahydroharmine.

, and the main other findings are presented in

Table 3. Studies included in review.

Reference	Sample	Group/Dose/Route	Model/Test	Result
Hamsa & Kuttan (2010) [37]	Male C57BL/6 mice	Harmine (10 mg/kg i.p.) TNP-470 (reference compound) (30 mg/kg i.p.)	Melanoma cell-induced inflammatory processes (intradermally)	Harmine reduced serum levels of the pro-inflammatory cytokines IL-1β, IL-6, TNF-α, and GM-CSF, of the pro-inflammatory transcription factor NF-κB, and of iNOS. Harmine also increased levels of the anti-inflammatory/anti-tumor cytokine IL-2.
Yamazaki & Kawano (2010 [38]	Mouse RAW264 macrophage cells/human THP-1 cells	Harmine (0.1–10 μM) LPS (100 μg/mL)	Lipopolysaccharide (LPS)-stimulated	The production of IL-6 and TNF-α in RAW264 cells were suppressed by harmine. Harmine also suppressed the production of TNF-α and NO in LPS-stimulated immune cells.
Szabo et al. (2014) [49]	Human primary monocyte-derived dendritic cell (moDCs) coats	N, N-dimethyltryptamine (DMT) (100 μM) 5-methoxy-N, N-dimethyltryptamine (5-MeO-DMT) (100 μM) PRE-084 (high affinity sigma-1 agonist) (100 μM)	Induction of inflammatory responses induced by LPS, polyI:C or pathogen-derived stimuli (influenza virus and Escherichia coli)	Pre-treatment of LPS- and polyI:C-stimulated moDCs with DMT, 5-MeO-DMT, and PRE- reduced the mRNA expression and secreted levels of the pro-inflammatory cytokines IL-1b, IL-6, TNF-α, and also of the chemokine IL-8/CXCL8 (only with LPS). DMT increased the production of the anti-inflammatory cytokine IL-10. No significant differences between DMT and 5-MeO-DMT were observed. DMT and 5-MeO also reduced the CD4+ helper T cell-priming capacity of moDCs [increased expression of IFNϒ (Th1) and IL-17(Th17)] induced by influenza virus and Escherichia coli. Sigmar-1 gene expression silencing significantly reduced these effects.
Khor et al. (2015) [56]	Murines: Balb/c, C57Bl/6, Cd45.1+/+, Rag1−/−, Foxp3IRES-GFP, Il17IRES-GFP, NOD-scid and NOD-BDC2.5 Cells: CD4 + CD62L + naive T cells (murine) CD4 + CD45RA + naive T cells (human)	Harmine HCl (i.n.) 1 mg for 5 days	Immune homeostasis via Th17/T-regulatory cell balance (T-regulatory suppression assay, TD1, CD45RB colitis, and airway inflammation models)	It was shown through diverse experiments that harmine prevents inflammation through modulation and restoring of Th17/T-regulatory cell proportion. This is achieved specifically via inhibition of Dyrk1a activity, which in turn enhances T-regulatory cell differentiation.
Liu et al. (2017) [42]	ICR male mice, mouse RAW 264.7 macrophage cells, HEK-293T cells	Control LPS (10 mg/kg i.p.) + vehicle LPS (10 mg/kg i.p.) + dexamethason (3 mg/kg) LPS (10 mg/kg i.p.) + harmine (30 mg/kg)	LPS-induced inflammatory processes	Harmine has an anti-inflammatory effect by reducing NF-kB transcription activity in HEK-293T cells, decreasing TNF-α, IL-6, and IL-1β levels in RAW 264.7 cells, and reducing the production of NO in RAW 264.7 cells. Harmine (and dexamethason) also reduced the protein levels of TNF-α, IL-1β, and IL-6 and suppressed pulmonary congestion, alveolar wall and inflammatory cell infiltration in an acute lung injury mouse model (LPS-induced inflammation).
Pires et al. (2018) [31]	Male mice	Control (saline i.p. or water p.o.) Ayahuasca (120 and 1200 mg/kg i.p. or p.o; DMT 0.38 mg/mL, harmine 0.32 mg/mL, tetrahydroharmine 1.67 mg/mL, harmaline 0.12 mg/mL) Morphine (1 and 10 mg/kg i.p.)	Hot plate Acetic acid writhing (0.8% i.p. of acetic acid) Formalin (20 μL)	Ayahuasca (1200 mg/kg) showed an antinociceptive effect in the acetic acid writhing test and the formalin test (but not in the hot plate test) and intensified the analgesic effect of morphine in the hot plate test and the acetic acid writhing test.
Alijanpour & Zarrindast (2020) [32]	NMRI male mice	Control (saline) Morphine sulfate (2 and 6 mg/kg i.p.) Naloxone hydrochloride (0.5 and 1 mg/kg i.p.) N-methyl-d-aspartate (NMDA) (0.06 and 0.1 μg/mouse; intra-vental tegmental area-VTA) Harmaline (1.25 and 5 mg/kg i.p.) dl-2-amino-5-phosphonoheptanoate (D-AP5) (NMDA receptor antagonist) (0.5 and 1 μg/mouse; intra-VTA)	Tail-flick: Pain threshold mu opioid receptor and analgesia Microinjection intra-VTA of NMDA and D-AP5	No significant effects were observed at low doses of morphine (2 mg/kg) or harmaline (1.25 and 5 mg/kg) on pain threshold; harmaline (5 mg/kg) intensified the analgesic effect of morphine; after a combination of morphine and harmaline at non-effective doses (2 and 5 mg/kg, respectively), pain threshold was increased and antinociception was induced. Microinjection of NMDA (0.06 and 0.1 μg) reduced the antinociceptive effect induced by that combination. In addition, analgesia induced by morphine and harmaline was inhibited by naloxone (0.5 and 1 mg). No effects were observed on pain threshold after microinjections intra-VTA of D-AP5 (0.5 and 1 μg) and NMDA (0.06 and 0.1 μg); however, D-AP5 increased pain threshold and intensified antinociception induced by morphine and harmaline.
Alijanpour et al. (2021) [33]	Male NMRI mice	Control (saline) Harmaline (0, 1.25, 2.5, 5, and 10 mg/kg i.p.) Morphine sulfate (0, 2, 4, and 6 mg/kg i.p.) Muscimol (0, 200, and 300 ng/mice.i.ventral hippocampal route-vHIP) Bicuculline (1 μL/mouse.i.vHip route)	Tail-flick	Harmaline group showed antinociception effect at 10 mg/kg. Harmaline (5 mg/kg) combined with morphine (2 mg/kg) produced antinociceptive effect and enhance morphine effects. Muscimol enhanced antinociceptive effects of harmaline combined with morphine.
Alijanpour et al. (2022) [34]	Male NMRI mice	Control (saline 10 mL/kg i.p.) Vehicle (0.6 μL/mouse. Microinjection into the basolateral amygdala (BLA)) Harmaline (4, 6, and 8 mg/kg i.p.) ACPA (0.5, 1, or 1.5 ng/mouse.micorinjection into the BLA) AM251 (0.1, 0.5, or 1 ng/mouse.microinjection into the BLA)	Tail-flick Hot plate	Harmaline (6 and 8 mg/kg) prolonged reaction time to thermal stimuli in tail-flick and hot plate tests. AM251 (0.5 and 1 ng/mouse) attenuated the antinociceptive activity of harmaline (8 ng/mouse) in both tests.
Jiang et al. (2019) [44]	Sprague-Dawley rats/pathogen free (SPF) Balb/c nude mice	Control (saline) Edema treatment: Carragenan without drug (1 g.s.i) Harmine solution gel-HSG (1 g.t.a) Harmine loaded ethosomes gel-HLEG (1 g.t.a) Positive control (dexamethasone acetate cream((CDXMC): 0.4 g.t.a)	Carragenan induced-paw edema	The levels of inflammation cytokines (IL-1β, NO, PGE-2, and TNF-α) were low after administration of HLEG. The degree of paw swelling was reduced after administration of HLEG, HSG, and CDXMC. HLE promotes prolonged therapeutic effect.
Niu et al. (2019) [43]	Male Kunming mice	Control (sodium carboxymethycellulose 0.5%) LPS (20 mg/kg i.p.) Dexamethasone + LPS (5 mg/kg of DEM) Harmine + LPS (25, 50 mg/kg of harmine i.g.)	LPS-induced kidney damage	At 50 mg/kg, harmine reduced the levels of TNF-α, IL-6, IL-1β and myeloperoxidase activity. Harmine also suppressed the expression of toll-like receptor 4 (TLR4), phosphorylation of nuclear factor-kappa B (NF-κB) p65, inhibitor of κBα (IκBα), NLRP3, caspase-1, and interleukin-1β (IL-1β).
Gonçalves et al. (2020) [54]	Vegetal samples: Commercial mixture (without composition information) Psychotria viridis Banisteriopsis caapi Mimosa hostilis Peganum harmala Decoctions: Psychotria viridis + Banisteriopsis caapi (ayahuasca) Psychotria viridis + Peganum harmala Mimosa hostilis + Banisteriopsis caapi Mimosa hostilis + Peganum harmala	0.210 g of each vegetal sample	Protein denaturation	Higher anti-inflammatory activity: P. harmala, M. hostilis, P. harmala + M. hostilis, comercial mixture Lower anti-inflammatory activity: P. viridis, B. caapi, P. viridis + B. caapi (ayahuasca)
Nardai et al. (2020) [51]	Male Wistar rats	Control (vehicle bolus) N, N-dimethyltryptamine (DMT) (1 mg/kg/2 mg/kg bw/h. i.p.) 1-piperazine dihydrochloride (BD-1063) (2 mg/kg bw/24 h. i.p./1 mg/kg-body weight + DMT)	Ischemia-reperfusion injury/inflammation in the rat brain	DMT decreased ischemic lesion volume and downregulated mRNA expression of the pro-inflammatory citokines IL-1β and IL-6 bilaterally (contralateral and in peri-infarct cortex), while levels of TNF-α were reduced only in the contralateral cortex; mRNA expression of the anti-inflammatory citokine IL10 was upregulated in the peri-infarct cortex. Serum concentrations of IL-1β, IL-6 and TNF-α were reduced, and IL-10 increased.
Jin et al. (2021) [25]	Male Balb/c mice or C57BL/6 mice Mouse peritoneal macrophages	Control (saline i.p.) Harmine (30 mg/kg, i.p.) LPS (1 mg/kg i.p.) Poly(I:C) at 50 and 100 μg/mL for 24 h Lipoteichoic acid (TLR2 agonist) 10 μg/mL CpG DNA (TLR9 agonist) 2 μM for 6 h	LPS Polyinosinic:polycytidylic acid (Poly(I:C)) TLR agonists Endotoxemia model (in vivo)	Harmine inhibited IL-12 (IL-12 p70), TNF-, IL-6, COX-2, iNOS, JNK, and STAT1 levels and marker genes in a dose-dependent manner in macrophages (LPS, poly(I:C), TLR agonists). In the endotoxemia model, harmine also reduced liver inflammation by inhibiting the expression of COX-2 and iNOS genes (but not TNF-α).
Wang et al. (2021) [40]	Male C57BL/J6 mice/Murine macrophage RAW 264.7 cells	Sham operation Phosphate-buffered saline + titanium particles (30 mg) (vehicle) Titanium particles (30 mg) + harmine (5 mg/kg/day or 10 mg/kg/day)	Peri-prosthetic osteolysis (PPO)	Harmine presence significantly reduced the inflamation area, TNF-α, IL-1β (all on both concentrations) and Il-6 (high concentration). On the contrary, harmine increased levels of IL-4 and IL-10. These results also corroborate that harmine treatment diminishes inflammation in PPO by changing the polarization of macrophages from M1(phenotype pro-inflammatory) to M2 (phenotype anti-inflammatory). Lastly, Ti-particle-induced activation of JNK was suppressed by harmine.
Borbély et al. (2022) [53]	Male C57BL/6 wild-type mice	Control DMT (1 mg/kg i.p.) PRE-084 (1 mg/kg i.p.)	Amyloid-β-(Aβ-) induced neurotoxicity C28	DMT reduced the effect of Aβ1-42 induced of astrocytes but not effects on microglial activation. DMT also reduced cytokines pro-IL-1β and TNF-α, however, only the change for TNF-α was significant and DMT negatively influenced neurogenesis.
Ju et al. (2022) [45]	Immortalized murine BV2 microglia lines/male ICR mice	Control (2 μL of saline) LPS challenge (1 μg/μL, 2 μL; injected into the left lateral ventricles) LPS + harmine (20 mg/kg i.p.)	LPS-stimulated BV2 cells	Harmine attenuated nitric oxide (NO), reactive oxygen species, tumor necrosis factor-α, cyclooxygenase 2 and inducible nitric synthase. The expression of inflammatory proteins (COX2 and iNOS) in the mice’s brains were reduced by harmine (20 mg/kg). Harmine at 3, 4, and 5 μM doses decreased the NO release in LPS- challenged BV2 cells.
Ghandari et al. (2022) [41]	Balb/c male mice	Control (saline) Cisplatin (5.5 mg/kg i.p.) Harmine (5.10 and 15 mg/kg i.p.) Cisplatin (5.5 mg/kg) + harmine (5, 10 and 15 mg/kg)	Cisplatin-induced renal tissue toxicity	The cisplatin group showed an increase in the serum levels of blood urea nitrogen (BUN) and creatinine. No difference was observed between harmine and control groups when analyzing BUN and creatinine levels. In the cisplatin + harmine (5 mg/kg) group, the serum levels of BUN were high. The kidney damage was reduced at all doses of cisplatin with harmine. The groups who received harmine showed a decrease in malondialdehyde (MDA) levels compared to the cisplatin group. Cisplatin + harmine (5 mg/kg) and cisplatin + harmine (10 mg/kg) increased MDA levels. This parameter was also increased in the cisplatin groups. Total antioxidant capacity (TAC) level was reduced in the cisplatin group. Harmine (5, 10 and 15 mg/kg) and cisplatin + harmine (5, 10, and 15 mg/kg) groups demonstrated an increase in the TAC level compared to the cisplatin group. Cisplatin + harmine (5 mg/kg) and cisplatin + harmine (10 mg/kg) groups demonstrated a decrease in TAC levels compared to the control group. A reduction in the apoptosis cells was observed in the harmine and cisplatin + harmine (5, 10, and 15 mg/kg) groups compared to the cisplatin group. Harmine group (5, 10, and 15 m/kg) did not show effect on apoptotic cells.
Kelley et al. (2022) [52]	Male Sprague Dawley rats/Human embryonic kidney	Control Predator exposure/psychosocial stress (PE/PSS) PE/PSS + DMT (2 mg/kg i.p.) PE/PSS + DMT (4 mg/kg i.p.) PE/PSS + harmaline (1.5 mg/kg i.p.) PE/PSS + harmaline + DMT (Pharmahuasca) The administration occurred 30 days post stress every other day for five days	Traumatic stress (PE/PSS)	DMT and pharmahuasca decreased ROS production in the prefrontal cortex and in the hipoccampus, as well as reduced the transcription of inflammatory cytokines (TLR3, TLR4, TLR6, TLR7; IL-1R, IL-1,IL-10, IL-12, IL-13, IL-15, IL-17, IL-18, IL-21, IL-23, IL-33; NFkB2, and IFNGR1). Pharmahuasca downregulated NFkB2.
Santos et al. (2022) [26]	BV-2 microglial cells	Banisteriopsis caapi (6.917 mg/g harmine, 1.583 mg/g harmaline, and 3.084 mg/g Tetrahidroharmine-THH) extract (4 μL) Harmine (2.4; 9.4; 18.9; 37.7, and 75.5 μM) Harmaline (2.4; 9.3; 18.7; 37.4, and 74.8 μM) THH (2.3; 9.3; 18.5; 37.0, and 74.1 μM)	Cytokine production	Harmine at high doses (18.9 to 75.5 μM) reduced TNF and IL-2 (18.9 and 75.5 μM). Harmaline reduced IL-7A, IFNϒ and TNF levels at high doses (18.7; 37.4 and 74.8 μM). Harmaline (18.9 μM) reduced IL-10. THH (18.5 μM μg/mL) reduced IL-6, IFNϒ, and TNF, and increased IL-4 (2.3 μM).
Szilágyi et al. (2022 [50]	Male Sprague–Dawley rats	Control (vehicle-treated) N, N-dimethyltryptamine (DMT) (10 mg/kg/day; osmotic mini pum.i.s) + harmaline-treated (10 mg/kg/day.gavage) Harmaline-treated (10 mg/kg/day.gavage)	Retinal ischemia–reperfusion injuries	DMT + harmaline-treated group showed higher levels of NFkBp65protein compared to harmaline-treated and healthy untreated groups. No difference was observed between harmaline-treated and healthy untreated groups.
Zheng et al. (2023) [46]	Male C57BL/6J mice/BV-2 microglial cells	Control (Saline i.p.) LPS (0.83 mg/kg i.p.) Harmine (20 mg/kg i.p.) Positive control: Fluoxetine (10 mg/kg i.p.)	LPS-induced neuroinflammation	Harmine inhibited LPS-induced microglia neuroinflammation in basolateral amygdala and BV-2 microglial cells.
De Camargo et al. (2024) [55]	Male Wistar rats	Sham + saline Cecal ligation and puncture (CLP) + saline Sham + ayahuasca (1 mL/kg.gavage) Sham + ayahuasca (2 mL/kg.gavage) Sham + ayahuasca (4 mL/kg.gavage) CLP + ayahuasca (1 mL/kg.gavage) CLP + ayahuasca (2 mL/kg.gavage) CLP + ayahuasca (4 mL/kg.gavage) (3 days of treatment for all groups) (ayahuasca alkaloid concentration: 1.40 mg/mL DMT, 0.10 mg/mL harmaline, 2.43 mg/mL harmine, and 1.43 mg/mL THH)	Sepsis induction (CLP) or Sham procedure Elevated plus maze Open field test Object recognition test	Ayahuasca pretreatment (1 mL/kg) increased the levels of IL-4 in the prefrontal cortex and cortex and increased the levels of BDNF in the cortex. In general, the results show antioxidative and anti-inflammatory effects of ayahuasca in sepsis-induced damage.
Graça et al. (2024) [57]	Male Wistar rats	Banisteripsis caapi extract (380 μg/mL THH, 126 μg/mL harmaline, 650 μg/mL harmine; 1.5 mL/kg.gavage 3 times a week for 4 weeks) 6-hydroxydopamine (6-OHDA.injection into the Locus coeruleus) LC lesion group LC lesion group under the effect of the extract Vehicle group Vehicle group under the effect of the extract	Open field test Elevated plus maze Rotarod test Histology	B. caapi extract reduced the concentration of IL-10 and expression of BDNF in the hippocampus compared to the control group and the group only under the effect of the extract. The B. caapi extract also increased the number and activation of inflammatory cells (microglia) in the LC.
Kadyan & Singh (2024) [36]	Swiss albino mice	Control (vehicle 0.1% carboxymethyl cellulose in distilled water 10 mL/kg i.p.) Pathogenic control-vincristine and vehicle (0.1 mg/kg i.p.; once daily for 10 days) Positive control-vincristine (0.1 mg/kg i.p.) + pregabalin (10 mg/kg p.o.) Harmaline (5 mg/kg i.p.; for 14 days) Harmaline (10 mg/kg i.p.; for 14 days) Substance-P (10 μg/kg i.p.) + harmaline (10 mg/kg i.p.) + vincristine (0.1 mg/kg i.p.); for 10 days	Vincristine- induced peripheral neuropathy Hot plate Cold plate Acetone drop Beam balance	Harmaline reduced licking and biting behavior and improved paw withdrawal latency in a dose-dependent manner on days 7 and 14 compared to the vincristine-treated group. It also decreased crossing time, with the 10 mg/kg dose showing a stronger effect than pregabalin. Harmaline (5 mg/kg) increased Nrf-2 levels, while 10 mg/kg reversed vincristine-induced Nrf-2 suppression. Additionally, harmaline countered vincristine-induced elevation of IL-1β, reducing its levels at both doses.
Lauria et al. (2024) [35]	Male Swiss and C57BL/6 mice	Control (water p.o.) Ayahuasca (24, 120, 600, and 3000 μL/kg p.o.) Harmine (0, 0.35, 3, 5, and 35 mg/kg p.o.) Indomethacin (5 mg/kg p.o.) Morphine (5 mg/kg i.p.) Dexamethasone (2 mg/kg i.p.) Formalin (20 μL) Complete Freund’s Adjuvant (CFA) (20uL) Gabapentin (70 mg/kg p.o.) Naloxone (5 mg/kg i.p.) Bicuculline (1 mg/kg i.p.) Methysergide (5 mg/kg i.p.) Rimonabant (10 mg/kg i.p.) Phaclofen (2 mg/kg i.p.)	Acute effect: Formalin test CFA Tail flick test Paw edema Acute and chronic effect: Partial sciatic nerve ligation model of neuropathic pain	Acute effect: Ayahuasca (120, 600, and 3000 μL/kg) and indomethacin (5 mg/kg) demonstrated antinociceptive effects at in the second phase of the formalin test. Morphine (5 mg/kg) was effective in inducing antinociceptive effects in both phases of the formalin test. Ayahuasca (600–3000 μL/kg) decreased CFA-induced allodynia, but did not reduce paw edema. Allodynia and edema were both reduced by dexamethasone (2 mg/kg). In the tail flick test, ayahuasca did not show an antinociceptive effect, while morphine (5 mg/kg) caused antinoception. Ayahuasca (24–3000 μL/kg) and gabapentin (70 mg/kg) decreased allodynia in experimental neuropathy. The effects of ayahuasca (3000 μL/kg) lasted 10 h, while the effects of gabapentin (70 mg/kg) lasted only 3 h. At 3.5 mg/kg, harmine produced an antinociceptive effect that lasted 10 h. Chronic effect: Ayahuasca (120–600 μL/kg twice a day for 14 days) and gabapentin (70 mg/kg daily for 14 days) induced antinoception in experimental neuropathy. Bicuculline (1 mg/kg) and methysergide (5 mg/kg) reverted the antinociceptive effect of ayahuasca (600 μL/kg), however, naloxone (5 mg/kg), phaclofen (2 mg/kg), and rimonabant (10 mg/kg) did not revert. At 3.5 mg/kg twice a day for 14 days, harmine produced an antinociceptive effect that lasted 10 h.
Ma et al. (2024) [39]	Male Kunming mice	Control Harmine (50 mg/kg p.o.) Model (0.2% CCl4, v/v, dissolved in rapeseed oil, 10 mL/kg body weight i.p.) Medium dose (harmine, 25 mg/kg; 0.2% CCl4 i.p.) High dose (harmine, 50 mg/kg; 0.2% CCl4 i.p.) Positive (bifendate, 200 mg/kg; 0.2% CCl4 p.o.)	Carbon tetrachloride (CCL4)-induced acute liver injury	Harmine reduced the serum levels of aspartate and alanine aminotransferase and the levels of TNF-α and IL-6. The levels of glutathione and superoxide dismutase were restored and the production of nitric oxide and malondialdehyde was suppressed by harmine. Harmine down-regulated LC3B II/I, p38 MAPK, TLR4, and NF-κB levels, while upregulating p62, Bcl-2, Beclin1, ULK1, and p-mTOR expression.
Xie et al. (2024) [47]	Male C57BL/6 J mice	Control (0.5% CMC-Na.gavage) Model (0.5% CMC-Na.gavage) Donepezil (5 mg/kg.gavage) Harmine (5 mg/kg; 10 mg/kg and 20 mg/kg.gavage) Ethanol (0.1 mL/10 g.gavage)	Morris water maze test Ethanol-induced memory impairment	Harmine reduced the levels of IL-6, IL-β and TNF-α while raising BDNF levels in plasma and hippocampus. From a cellular perspective, harmine promoted synaptic vesicle fusion and protected the myelin sheath, demonstrating beneficial effects against ethanol-induced memory impairment.
Zheng et al. (2024) [48]	Balb/c mice/RAW 264.7 cells	In vitro Control group Phosphate-buffered saline (PBS) (100 μL injection) Negative group 0.1% dexamethasone Positive group 25 and 50 μM dexamethasone Natural harmaline (HML) (25 and 50 μM) LPS 1 μg/mL (24 h after treatment) In vivo Test and positive control group (harmaline 10 mg/kg i.p. and physiological saline every 12 h for 3 days) λ-carrageenan (2.0 mg/mL, 100 µL)	Rheumatoid arthritis (RA) induction	Harmaline inhibited production of nitric oxide and the secretion of IL-1β and IL-6 in cells activated by LPS. Moreover, the thickness and extent of swelling of paw were reduced by HML. In cells activated by LPS, the expression of iNOS and COX-2 was decreased by HML, as well as the expression of p65 and p65 phosphorylation.

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4. Discussion

This systematic review assessed the antinociceptive and anti-inflammatory effects as the main outcome after the administration of ayahuasca or its alkaloids. Several tests for pain induction were described, such as the tail flick, formalin, acetic acid, hot plate, and rotarod. Despite the different approaches concerning substances, dosages, and induction models, the results were similar in promoting antinociceptive effects. Furthermore, the modulatory role of both ayahuasca and its alkaloids in the inflammatory process is particularly evident. The results obtained were positive regarding the reduction and increase in the levels of pro- and anti-inflammatory cytokines, respectively.

The antinociceptive effect induced by ayahuasca or its alkaloids has been attributed to interactions with various neurochemical systems involved in nociception. Although the exact mechanisms underlying this effect remain unclear, several hypotheses have been proposed. There is evidence to suggest that the opioid system may play a role in the antinociceptive effects of certain compounds [58]. Supporting this, the extract of Peganum harmala—a plant rich in harmaline—has shown notable antinociceptive properties, which are likely mediated by these same opioid receptors [59]. Furthermore, experimental models such as the tail-flick and hot plate tests (which are commonly used to evaluate centrally mediated analgesia) are highly sensitive to the effects of opioid drugs [60]. The results are consistent with previous reports, in which harmaline with morphine promoted antinociceptive effect [32], prolonged the time of tail flick and hot plate tests [34], and reduced vincristine-induced hyperalgesia in the hot plate test [36]. In the remaining studies analyzed, where ayahuasca was the primary substance investigated, no antinociceptive effect was observed during the hot plate test [31]. However, ayahuasca demonstrated significant efficacy in both the acetic acid and formalin tests and was capable of potentiating the effects of morphine in the acetic acid and hot plate models [31]. A similar pattern was observed in a recent study by Lauria, in which ayahuasca again proved effective in the formalin test, despite showing no apparent effect in the tail-flick assay [35]. Although these results differ from those previously reported, they suggest that ayahuasca may exert its analgesic effects by acting directly on inflammatory mediators involved in the pain responses triggered by the formalin and acetic acid tests [61].

Another hypothesis is related to the serotonergic system, since serotonin (5-HT) plays a crucial role in pain modulation. This neurotransmitter acts at the peripheral level, influencing descending inhibitory pathways, and at the central level, where it plays a role in pain perception and interpretation [13,62]. To investigate the possible mechanisms of action, antagonist drugs were administered before ayahuasca treatment. The results demonstrated that the antinociceptive effect of ayahuasca was reversed by bicuculline, a GABA_A receptor antagonist, and by methysergide, a serotonergic receptor antagonist [35]. In contrast, naloxone, an opioid receptor antagonist, did not affect the action of ayahuasca. It can be inferred that GABAergic and serotonergic receptors play a key role in mediating the antinociceptive effects of ayahuasca [35]. Furthermore, the activation of the 5-HT_2A receptor can disrupt existing neural networks, leading to the reorganization of pain-processing pathways and the formation of new functional connections. This shift could allow pain to be experienced differently, potentially altering its perception and emotional processing [13,62].

Serotoninergic hallucinogens (receptor agonists 5-HT_2A), such as lysergic acid diethylamide (LSD), psilocybin, and DMT, were also found to suppress the inflammatory responses [63,64]. Bufotenine, an alkaloid structurally analogous to DMT, has demonstrated anti-inflammatory and antinociceptive properties. This alkaloid was shown to reduce elevated levels of COX and PGE2, indicating an anti-inflammatory effect, and increased latency in the hot plate test, suggesting central antinociceptive activity [63]. It has been proposed that bufotenine mediates nociceptive modulation at the central level, as this compound interacts with sigma-1 and 5-HT₃ receptors [63]. Similarly, psilocybin has shown potential in attenuating neuroinflammation. When combined with eugenol, it reduced inflammatory markers IL-6 and IL-8, as well as the expression of COX-2 and TNF-α [64]. Psilocybin also effectively decreased TNF-α and IFN-γ levels in intestinal tissue [65]. These findings align with the results reported by Teksen, who also demonstrated the neuroprotective properties of P. harmala, including increased expression of the antioxidant transcription factor Nrf2 and a reduction in the levels of the pro-inflammatory factor NF-κB in the brain [66]. These findings corroborate those presented previously in studies that used both harmine and DMT. Based on this evidence, the anti-inflammatory effects of harmine may be linked to the inhibition of the TLR4/NF-κB signaling pathway, which plays a central role in mediating inflammatory responses [42,43]. Additionally, the immunomodulatory effects observed with classical psychedelics may be partly explained by their interaction with 5-HT2A and sigma-1 receptors (Sig-1Rs), both of which are known to influence neuroimmune regulation [29].

Clinical studies have also reported the anti-inflammatory effects of hallucinogens. In healthy volunteers administered psilocybin, acute levels of pro-inflammatory cytokines, such as TNF-α, and chronic markers, including IL-6 and C-reactive protein (CRP), were reduced [16]. However, another study involving psilocybin administration in healthy participants did not find significant changes in inflammatory biomarkers [15]. Regarding ayahuasca, healthy volunteers showed an increase in natural killer cells and a decrease in the percentage of CD4 and CD3 lymphocytes [67]. In contrast, a study evaluating the effects of ayahuasca in both healthy and depressed individuals demonstrated a reduction in C-reactive protein levels in both groups [28]. Given these findings, along with preclinical evidence, both antinociceptive and anti-inflammatory effects could be modulated by serotonergic receptors.

Taken together, the evidence reviewed in this study suggests that ayahuasca and its alkaloids possess significant antinociceptive and anti-inflammatory properties, mediated by complex interactions within multiple neurochemical systems. While the opioid pathway may contribute to the analgesic effects of β-carbolines such as harmaline, the most consistent findings point toward a central role of the serotonergic and GABAergic systems, mainly modulating 5-HT2A and possibly sigma-1 receptors.

Although some clinical findings are inconsistent, both preclinical and clinical data support their potential as therapeutic agents for pain and inflammation. Further research is needed to better understand their mechanisms, safety, and clinical utility. Specifically, in future studies, other pain models could be used in association with the more commonly used tail-flick and hot plate tests, as well as the acetic acid and formalin tests. These models could be directly compared and used in association with specific agonists and antagonists to better understand neurochemical mechanisms behind these effects. Moreover, although acute pain models give us important information on possible analgesic and anti-inflammatory effects, chronic pain models would have more translational value and should be used in future studies.

5. Conclusions

This review highlights the antinociceptive and anti-inflammatory potential of ayahuasca and its alkaloids. Although the mechanism by which ayahuasca compounds produce nociceptive and inflammatory processing is unclear, preclinical studies indicate promising results, suggesting a possible therapeutic approach. Further preclinical studies, comparing different models and using chronic pain models alongside antagonists and agonists, are needed to elucidate the effects and mechanisms of ayahuasca and its alkaloids in pain and inflammation, and human studies using pain models should confirm the translational potential.