Schizophrenia is a severe psychiatric disorder with a chronic course, affecting ~1% of the global population (1). This debilitating disease manifests in early adulthood (2), presenting with positive (hallucinations and delusions), negative (social withdrawal, apathy, and anhedonia), and cognitive symptoms (working memory deficits, attention disorders, and altered information processing) (3). However, the underlying etiological mechanisms remain elusive (4). Similarly, the treatment outcome for schizophrenia remains suboptimal (5), especially with regard to negative and cognitive symptoms (6, 7).

The neurodevelopmental hypothesis has provided a valuable framework for establishing the relationship between pathologic processes during early brain development and the development of schizophrenia later in life (8, 9). This hypothesis suggests an interaction between genetic predisposition and early life environmental vulnerability factors such as malnutrition, substance abuse, obstetric complications, season of birth and infection, and exacerbation by later stresses such as substance abuse, social defeat, and trauma (10–13).

Viral or bacterial maternal infection during pregnancy has been linked to increased risk for developing schizophrenia in the offspring (14, 15), with immune activation rather than the infectious agent, itself, being deemed causal (16, 17). Indeed, trauma is associated with immune activation and increased risk of psychosis (18). Immune adjuvants such as lipopolysaccharide (LPS) (19–21), polyinosinic:polycytidylic acid (poly I:C) (22–24), human influenza virus (25), and cytokines (26, 27) induce diverse biological and behavioral abnormalities in rodents following prenatal maternal exposure. LPS, an endotoxin derived from the cell wall of Gram-negative bacteria, mimics an infection by activating the synthesis and release of pro-inflammatory cytokines, including interleukin-1β (IL-1b), IL-6, and tumor necrosis factor-α (TNF-α) (28–30) and engenders various schizophrenia-like behavioral, neurochemical, and inflammatory changes (31–33).

Oxidative stress underscores various psychiatric conditions (34, 35), in particular, schizophrenia (36). Increased reactive oxygen species (ROS) and reduced antioxidants observed in schizophrenia may contribute to the neuroprogression of the disorder (37) and to the development of cognitive dysfunction (38, 39). Indeed, the antioxidant, N-acetyl cysteine (NAC), has therapeutic benefits in various clinical domains of schizophrenia, but especially negative symptoms (40, 41) and cognition (42), while having also demonstrated efficacy in preclinical animal models (43–45).

There is an increased drive to integrate nutraceuticals and psychotropic herbal medicines into conventional medical practice (46). Co-prescription of certain herbal medicines with traditional pharmaceuticals may display complementary pharmacodynamic actions and so provide a beneficial synergistic effect (46, 47). However, little study has occurred that has directly explored such augmentation effects (46). With raw herbal extracts containing a vast array of potentially bioactive ingredients, the question remains whether the observed pharmacological effect is ingredient specific or a sum effect of the total extract.

The anti-inflammatory and antioxidant activities of herbal bioactive compounds have been widely observed, particularly in a group of polyphenols referred to as xanthones (48). Garcinia mangostana Linn (GML) is a fruit native to Southeast Asia known to contain constituents including xanthones, flavonoids, triterpenoids, and benzophenones (49). Extracts of the fruit have exhibited antioxidant (50, 51), anti-inflammatory (52, 53), antibacterial (54), and antidepressant effects (55). In particular, α-mangostin (AM), a primary component of GML, presents with substantial pharmacological properties (56, 57), including antioxidant activity (58), as well as having moderate inhibitory effects on 5HT_2A receptors and cyclic adenosine monophosphate (cAMP) phosphodiesterase (PDE) (49), actions that hint at possible clinical utility as a pharmacological intervention in psychiatric disorders.

The aim of this study was to establish whether maternal immune activation (MIA) induced schizophrenia-like behavior and redox-inflammatory alterations in offspring can be reversed with the typical antipsychotic, haloperidol (HAL), GML, and AM separately. Second, since the most common use for nutraceuticals in clinical psychiatry is as an adjunctive treatment (46), we investigated whether adjunctive treatment with GML or AM is able to augment the response to HAL. The inclusion of AM is 2-fold; to investigate whether any observed pharmacological effects of GML may be specific for one of the known bio-active constituents of the extract, i.e., AM, or whether these actions underscore a sum effect of the total extract and, second, to link any effects to a known psychotropic property of AM and/or GML. In order to cover the mood-psychosis continuum, behavioral analyses focused on positive (sensorimotor gating; locomotor hyperactivity) and negative (depression) related symptoms. Moreover, by measuring associated changes in plasma and brain redox-inflammatory markers, it explores possible activity within a key neuropathological feature of the illness, viz. immune-inflammatory dysfunction (35). This study has importance to the field in that a plant extract and one of its known bioactive constituents are compared to a reference control pharmaceutical agent across a range of behavioral and biological parameters of relevance to schizophrenia.

MIA model validation, i.e., LPS vs. saline control, was analyzed separately using T-tests and presented in Figures 2–6. Thereafter, untreated LPS (MIA model) were compared to LPS plus the various drug treatments and analyzed separately using the appropriate ANOVA followed by post hoc Bonferroni analysis. The latter are also presented in Figures 2–6.

MIA induced sensorimotor gating deficits and depressive-like behavior concurrent with elevated cortico-striatal lipid peroxidation and elevated plasma pro-inflammatory cytokines in offspring. HAL reversed the changes in PPI, cortical (not striatal) lipid peroxidation, and elevated plasma IL-6, but failed to reverse depressive manifestations. GML + HAL effectively reversed PPI, while GML, AM, and HAL + AM did not. Conversely, GML and AM alone reversed depressive-like behaviors, more so than HAL, while the GML + HAL combination also reversed depressive-like symptoms. AM reversed both cortical (not striatal) lipid peroxidation and elevated plasma IL-6. GML and HAL + GML reversed elevations in IL-6 and TNF-α. These data present evidence for GML and its active constituent, AM, as being effective as adjunctive treatments but differently active with respect to psychotic and mood-related behaviors.

In line with previous findings (22, 31, 45, 63, 85), prenatal LPS exposure significantly compromised PPI in late adolescent offspring (Figure 2). HAL significantly reversed PPI deficits at 72 and 76 dB, with a similar trend and very large effect sizes at 80 and 84 dB (Figure 2), consistent with data from previous chronic (86) and acute (87–89) treatment studies in animals. Overactive dopaminergic processes are suggested to underlie the reduction in PPI (90), which explains the ability of HAL (D₂ antagonist) to reverse said deficits (91). Although GML + HAL was effective in reversing MIA-induced PPI deficits across all four startle responses, this was not the case for GML or AM separately or for AM + HAL (Figure 2). The latter suggests that AM may abrogate the antipsychotic-like response to HAL, a particular interesting finding. In fact, AM presents with known antioxidant (58), 5HT2A, and a cAMP-PDE inhibitory (49) activity that to varying degrees may underlie these effects. Certainly, 5HT2A receptors play a prominent role in psychotic-like behavior (35) as well as in atypical antipsychotic drug design (35), and hence may play a role in the results described here. On the other hand, HAL is a potent pro-oxidant (92) and pro-inflammatory agent (93). Depending on the stage of disease progression, these actions, together with potent D2 inhibition, may mediate HAL's useful antipsychotic effects. However, these same pro-oxidative actions are purported to cause striatal toxicity and to cause late-onset treatment-related complications (92, 93). Although speculative, could these actions of HAL be countered by the antioxidant and cAMP-PDE inhibitory actions of AM, and possibly even reverse its antipsychotic effects? This opens up new ideas on how antipsychotics work and warrants further study.

Locomotor hyperactivity represents positive symptom schizophrenia, especially psychotic agitation (94). Prenatal LPS-exposed offspring demonstrated increased locomotor activity in the OFT (Figure 3), a behavioral response that has been ascribed to hyperdopaminergia (95). HAL lowered LPS-induced locomotor hyperactivity, albeit not significantly, in line with its antidopaminergic/antipsychotic actions. Only AM effectively reduced hyperlocomotion (Figure 3), yet it failed to alter PPI deficits (noted above). AM is a selective, competitive histamine antagonist (96) with sedative properties (97) that may adversely affect startle response in the PPI test. That said, the study design did not allow us to assess the effects of drug treatments on startle response in healthy controls, although rats exposed to LPS + AM did not differ significantly to rats exposed to LPS + vehicle. This and the above-noted HAL + AM findings prompt further research into the putative “antipsychotic-like” effects of AM. No other treatment had any noteworthy effects on locomotor activity (Figure 3), although the locomotor effects described for AM may complicate interpretation of swimming behavior in the FST (see below).

The negative symptoms of schizophrenia comprise affective flattening, alogia, anhedonia, asociality, and avolition (lack of motivation) (98), congruent with the basic symptoms of depression (99). Although the FST assesses behavioral despair, it has been suggested to signify an absence of motivational behavior which is commonly seen in schizophrenia (100). Indeed, LPS-exposed offspring presented with significant depressive-like behaviors (increased immobility) (Figure 4A) and reduced active coping (swimming and climbing) (Figures 4B,C), in agreement with earlier studies (101, 102). Especially, atypical antipsychotics may present with antidepressant activity (103, 104), while clinical (105) and preclinical (106) studies have described the pro-depressant effects of HAL (105, 106). In the present study, however, HAL showed a small, albeit negligible, effect to reverse MIA-associated immobility in the FST and reduced swimming (Figures 4A,C), although significantly increased struggling/climbing behavior (Figure 4B). Importantly, GML significantly decreased immobility and increased swimming behaviors (Figures 4A,C), antidepressant effects congruent with an earlier study in Flinders Sensitive Line (FSL) rats (55). Interestingly, the latter study described GML's prominent serotonergic actions and therapeutic equivalence with imipramine, which is shown here, too, but less emphatically (elevated swimming) (Figure 4C). Moreover, GML is a superior antidepressant to HAL (Figures 4A,C). Similarly, AM also exhibited significant antidepressant-like properties regarding its effects on immobility and swimming behavior, although not as marked as GML alone (immobility) (Figure 4A). Of note, GML and AM augmented the actions of HAL on immobility (Figure 4A) with GML bolstering the HAL effect on struggling (Figure 4B), suggesting a bolstering of HAL's action via mechanisms other than D₂ receptor blockade. Despite AM suppressing locomotor activity, as noted above, this action did not affect its ability to reduce immobility and to increase swimming in the FST, thus highlighting a psychogenic action to bolster escape-driven behavior that is not related to, or mediated by, an increase in locomotor activity. Given the noted antioxidant actions of GML and its constituents, other antioxidants like NAC (107) are also antidepressant in the FST. Interestingly, NAC seems to have specific benefit in especially negative-symptom schizophrenia (40, 41), thus highlighting that antioxidants may have preferential psychopharmacological actions as antidepressants, which is borne out in this study as well.

Oxidative damage is implicated in the pathophysiology and neuroprogression of schizophrenia (35–37). Schizophrenia patients present with increased plasma lipid peroxidation (108–110), possibly correlated with certain clinical features (111). Adjunctive treatment with antioxidants improve symptoms in animal models (43, 112) as well as patients with schizophrenia (40). Prenatal LPS exposure significantly increased cortical and striatal lipid peroxidation in offspring (Figure 5), in agreement with previous findings (45, 113). Mouse models of oxidative stress are associated with cognitive and motivational deficits, as well as dysfunction of the prefrontal cortex (114). Schizophrenia is a hyperdopaminergic state (35) where dopamine metabolism contributes to oxidative stress by lowering glutathione (GSH) levels, which in turn is abrogated by D₁/D₂ receptor antagonists (36, 115). Importantly, HAL treatment significantly reduced lipid peroxidation in the frontal cortex (Figure 5A), while not having any marked effect in the striatum (Figure 5B). However, total striatum was analyzed here, whereas it is predominantly the ventral striatum encompassing the ventral tegmentum that is more relevant in rodents for an association with schizophrenia (116). The fact that HAL did not significantly reduce striatal lipid peroxidation may also be explained by HAL's known pro-oxidant actions in the striatum following chronic treatment (117) and which is associated with its long-term locomotor side effects. This action may otherwise obscure any possible antioxidant abilities in reducing MDA levels as was evident in the frontal cortex. This differential pro-oxidant action for HAL in these two brain regions is not new. In fact, Martins et al. (118) found that chronic HAL treatment increases oxidative stress in the striatum but decreases such levels in the cerebral cortex. Importantly, HAL still reversed LPS-induced PPI deficits (Figure 2), reiterating the importance of the frontal cortex in antipsychotic action (119). Moreover, here, we also show a frontal cortical role for its redox modulatory actions and how this may affect behavior. The frontal cortex is involved in cognitive processes such as working memory, behavioral flexibility, and attention (120). With regard to the antidepressant-like effects of GML and AM, the frontal cortex is also implicated in the development of depression and, hence, in antidepressant response (121, 122).

Although GML possesses antioxidant activity in vitro (123–126), GML had no effect on striatal lipid peroxidation, although it prompted a large effect size reduction in the frontal cortex (Figure 5A), thus qualitatively similar to that observed with HAL. Earlier, we found that chronic GML reversed elevated hippocampal lipid peroxidation in FSL rats (55), although the discrepancy between these two studies may be due to the different translational models used and the brain region assayed. AM also presents with antioxidant activity (58), here, significantly and again selectively reducing LPS-induced lipid peroxidation in the frontal cortex (Figure 5A). The absence of obvious antioxidant actions for GML and AM in the striatum is noteworthy, but may be related to assaying the whole striatum, as noted for HAL earlier. The diverse antioxidant actions of AM, viz. modulating GSH levels (127), free radical scavenging (128), inhibiting low-density lipoprotein oxidation (129), may afford it a more prominent antioxidant action than raw GML in vivo. As AM is the dominant bioactive xanthone in GML pericarp (55), it probably provides the dominant antioxidant activity observed with the raw extract. However, the more pronounced antioxidant action of AM does not translate into an improved behavioral outcome for AM over GML (Figures 2, 3), while GML and AM only offered small (GML) to negligible (AM) benefits in combination with HAL with regard to redox markers (Figure 5A). This suggests that GML may be offering beneficial effects through mechanisms other than antioxidant activity alone.

Immune-inflammatory dysfunction has been extensively reported in schizophrenia (35), specifically elevated levels of pro-inflammatory cytokines (130), while being a protagonist for oxidative stress (131). Pro-inflammatory cytokines have a developmental role in the brain (132, 133) and are implicated in the pathogenesis of neurodevelopmental disorders such as schizophrenia (134, 135). IL-6 and TNF-α levels are elevated in schizophrenia (136) and animal models (45, 137). Likewise, IL-6 and TNF-α were elevated in MIA offspring (Figures 6A,B) together with increased cortico-striatal lipid peroxidation (Figure 5). HAL-associated reversal of elevated IL-6 levels (Figure 6A) is consistent with clinical findings (138, 139), although it did not alter elevated plasma TNF-α levels (Figure 6B). Here, both GML and AM treatment reduced elevated plasma IL-6 levels (Figure 6A), with GML, but not AM, also reducing TNF-α levels (Figure 6B). This suggests a broader immunosuppressant action for GML vs. HAL or AM. AM has been shown to decrease inflammatory cytokines following LPS induction (140), to inhibit IL-2 release (141) and to suppress IL-6 expression (142). HAL + GML, but not HAL + AM, successfully reversed elevated IL-6 and TNF-α levels, although not more so than HAL alone (Figures 6A,B). In fact, HAL has immunosuppressive effects (143) as does GML have anti-inflammatory properties (125, 144). However, that neither GML nor AM bolstered the antioxidant effects of HAL again asserts that any beneficial effects offered by adjunctive GML treatment may involve mechanisms over and above inflammatory-redox processes. This warrants further study.

These findings have significance as a catalyst for future pre-clinical and clinical studies. Considering the important role of inflammation in the progression of mood and psychotic disorders, there is a growing interest in nutraceuticals with anti-inflammatory/antioxidant activity in psychiatry. That GML and AM have evinced therapeutic efficacy in the MIA model, as well as possess anti-inflammatory and antioxidant properties, suggests potential as a novel adjunctive treatment for these disorders (145). However, there appears to be distinct differential effects with respect to the mood–psychosis continuum, with a bias in favor of a depressed mood component. This prompts further investigation into GML's clinical benefits as an antidepressant vs. an antipsychotic. These aspects need deeper consideration in further animal studies but also in controlled clinical trials (146).

Certain limitations to this study are worth noting. Given the less-than-adequate antipsychotic-like effects for GML and AM, it would have been informative to include another schizophrenia-like behavioral assessment in the protocol to confirm these findings, e.g., memory, social interaction. Moreover, a dose titration analysis for GML and AM may have revealed a dose-dependent association in their behavioral effects, especially since the dose used for GML was based on a prior antidepressant study in another animal model (55). We also did not explore synergistic effects with atypical agents, which could differ to HAL. Biological analysis could have benefitted from regional striatal analysis, i.e., ventral, rostral, as opposed to assay of the whole striatum, as was done here. The study design did not allow for the assessment of drug treatments in healthy controls, which may have allowed for more in-depth explanation of treatment effects in LPS-exposed animals. Finally, having the same number of animals in both the saline-treated and LPS-exposed groups could have been an added benefit.

Schizophrenia is plagued by poor treatment outcomes and the limited efficacy of currently available antipsychotics (147–149). Supplementary treatment with nutraceutical anti-inflammatory agents and antioxidants such as GML may offer distinct therapeutic benefits (46). Plant extracts invariably contain a rich mixture of various bioactive constituents, yet little is known whether the pharmacological properties of a given extract are the sum of one or a group of inherent constituents or the result of the unique mix that the raw extract offers. This study has attempted to highlight this important question. Unlike the reference antipsychotic, HAL, chronic treatment with GML or AM failed to impact on sensorimotor gating deficits in the MIA model. However, both GML and AM not only displayed significant antidepressant-like properties but also bolstered the anti-immobility response to HAL. This is noteworthy as unlike atypical antipsychotics, HAL is not a recognized antidepressant, while here, it only marginally reduced immobility in the FST. GML and AM were both anti-inflammatory in the model, which may underlie their antidepressant effects. AM and, to a lesser degree, GML abrogated frontal cortical oxidative stress. This study confirms the antidepressant-like effects of GML described in another translational model of depression, the FSL rat (55). Having performed this study in a MIA model supports the use of GML and AM to address depressive symptoms in schizophrenia. However, their ability to address broader psychotic manifestations of the illness, e.g., PPI deficits, requires further study. Whether GML or AM is able to confer therapeutic benefits remains to be confirmed in dose–response and other clinical studies (150).

The datasets generated for this study are available on request to the corresponding author.

The animal study was reviewed and approved by AnimCare animal research ethics committee (National Health Research Ethics Council reg. no. AREC-130913-015) of the North-West University (Ethical approval numbers: NWU-00376-16-A5 and NWU-00147-14-A5).

JL: all laboratory work, data collection and formal analysis, validation, and first draft of the manuscript. MM: all figures, supervised behavioral methods and analysis, and statistical analysis. MB and OD: study design, data interpretation, and manuscript review. BH: conceptualization, methodology and statistics, writing—review and editing, supervision of JL, project administration, and funding acquisition.