Sceletium tortuosum (L.) N.E.Br. (ST) is an alkaloid-rich succulent plant with various mechanisms of action that infer psychotropic effects. These actions correlate with clinical evidence suggesting efficacy in the treatment of depression and anxiety, in line with its use by indigenous populations. Its low side effect profile suggests potential of ST to improve the overall wellbeing and compliance of millions of patients that experience severe side effects and/or do not respond to current prescription medication. However, to elucidate specific physiological effects of ST extracts, it is necessary to first understand which of its constituents are the major contributors to beneficial effects demonstrated for ST in this context. To determine an anxiolytic- and antidepressant-like effective concentration of a ST extract by means of a dose response in zebrafish (ZF) larvae, and to assess relative contributions of equivalent concentrations of isolated alkaloids contained in the effective concentration(s). A dose response study employing a light-dark transition test (LDTT) was done in ZF larvae (<5 days post fertilization) to track locomotor activity in terms of anxiety-like (hyperlocomotion) and depression-like (hypolocomotion) behaviour. Larvae were treated for 1 h directly before the LDTT with escalating concentrations of a ST extract commercially known as Zembrin® (Zem) ranging from 0.25 to 500 μg/mL and compared to an untreated control group (n = 12 per treatment concentration). LDTT was repeated after 24 h to evaluate long-term exposure toxicity. The concentration that best attenuated hyperlocomotion during the dark phase following light-dark transition was identified as the anxiolytic-like concentration. This concentration, plus one higher and one lower concentration, were used for subsequent tests. The percentage content of each alkaloid (mesembrine, mesembrenone, mesembrenol, and mesembranol) in these concentrations were calculated and applied to additional larvae to identify the most effective anxiolytic-like alkaloid in the LDTT. To identify antidepressant-like therapeutic concentration and equivalent alkaloid concentration, the same treatment concentrations were tested in larvae (n = 12 per treatment concentration) pre-exposed to reserpine for 24 h. Depending on normality of data distribution, Brown-Forsythe and Welch, or Kruskal-Wallis ANOVA were used, with Dunnett or Dunn's multiple comparisons tests. Only the extreme concentration of Zem (500 μg/mL) elicited toxicity after treatment for 24 h. Zem 12.5 μg/mL was the most effective anxiolytic-like concentration as it significantly decreased locomotor activity (P = 0.05) in the LDTT. Low (5 μg/mL), optimal (12.5 μg/mL) and high (25 μg/mL) Zem concentrations, as well as treatment solutions of single alkaloids (mesembrine, mesembrenone, mesembranol and mesembrenol), prepared to contain equivalent concentrations of each major alkaloid contained within these three concentrations of Zem, were tested further. Only mesembrine concentrations equal to that contained within the optimal and high dose of Zem (12.5 and 25 μg/mL) showed significant anxiolytic-like effects (P < 0.05). Only the highest Zem concentration (25 μg/mL) reversed the effects of reserpine - indicating antidepressant-like properties (P < 0.05) - while isolated alkaloids failed to induce such effects when administered in isolation. Current data provide evidence of both anxiolytic- and antidepressant-like effect of whole extract of Zem, with relatively higher concentrations required to achieve antidepressant-like effect. Of all alkaloids assessed, only mesembrine contributed significantly to the anxiolytic-like effects of Zem. No alkaloid alone could be pinpointed as a contributor to the antidepressant-like activity observed for higher concentration Zem. This may be due to synergistic effects of the alkaloids or may be due to other components not tested here. Current data warrants further investigation into mechanisms of action, as well as potential synergy, of ST alkaloids in suitable mammalian in vivo models.

Sceletium tortuosum (ST) induces antidepressant and anxiolytic effects, purportedly by monoamine regulation, anti-inflammatory and antioxidant properties, and phosphodiesterase 4 (PDE4) inhibition. These multimodal actions have not been demonstrated in an animal model of major depressive disorder. Wistar rats (both sexes) were subjected to 8-week unpredictable chronic mild stress, subsequently receiving saline, a standardized ST extract, Zembrin^® 25 and 12.5 mg/kg (ZEM25 and ZEM12.5), its primary alkaloid mesembrine (MES), or escitalopram (20 mg/kg) for 36 days. Sucrose preference, open field, Barnes maze, and forced swim tests were performed, with cortico-hippocampal monoamines, inflammatory and oxidative stress markers analyzed post-mortem. Male, but not female rats, presented with increased anhedonia and anxiety but not despair. Males presented with increased hippocampal PDE4B expression, increased dopamine metabolites, and decreased cortical serotonin. In males, ZEM12.5 decreased anhedonia- and anxiety-like behavior, decreased cortical and hippocampal PDE4B, and increased plasma interleukin-10. MES induced a transient decrease in anhedonia-like behavior and increased hippocampal serotonergic and cortical dopaminergic activity, whilst decreasing hippocampal PDE4B. ZEM25 increased plasma interleukin-10 but decreased cortical glutathione, indicating paradoxical anti-inflammatory and prooxidant effects. ZEM12.5 and MES more effectively addressed anxious–depressive-like behavior and stress-induced inflammation and monoaminergic alterations, respectively. Multitargeted actions on monoamines, redox-inflammation, and PDE4 may provide ST with antidepressant effects across multiple symptom domains, although mutually synergistic/antagonistic effects of constituent alkaloids should be considered.