In understanding the neurobiology of addiction and neuropharmacological action of psychoactive plants, CPP model is established to evaluate the reinforcing abilities of a psychoactive substance-paired paradigm against a psychoactive substance-free paradigm (Möller et al., 2018; Wojcieszak et al., 2021). The CCP paradigm is principled on the theory that a textual stimulus can attain secondary appetitive ability when paired with a primary reinforcer, thus indicating abuse liability (Aguilar et al., 2009; Strekalova, 2022). This translates that AECS, AEDS, AENT and AECM have addictive potentials via reinforcing and rewarding effects (Hur et al., 2020). Meanwhile, the significant depletion observed in the ability of alkaloid extracts-administered animals to locate hidden platform, make correct alterations and maintain movement orderliness at high doses during Y-maze and Morris water maze may indicate cognitive dysfunctions, spatial memory and learning impairments (Šlamberová et al., 2015). Inferentially, the present study showed the ability of the psychoactive plants to trigger addiction, however, at high doses, significant cognitive dysfunctions, spatial memory and learning impairments were observed, which are suggestive of neurotoxicity.
The depletion observed with the highest dose (2000 mg/kg) of the plant alkaloid extracts during CPP maybe dependent on the adverse effect observed with Y-maze and MWM tests, a validation that in order to achieve CPP, there is a need to apply memory and learning (Itzhak and Ali, 2006). This corresponds with previous studies that reported that chronic exposure to psychoactive substances at high doses will induce rapid development of tolerance and aversion, despite their potency to induce addictiveness at very low doses (Hur et al., 2020; Fasakin et al., 2022b).
The striatal dopaminergic brain pathway is a key pathway in the psychoactive, addictive and reinforcing effects of all psychoactive substances (Cunha-Oliveira et al., 2008). It is also tasked with learning and motivation; however, its overstimulation has been implicated in hyperactivity, attention deficits and attention deficit hyperactivity disorder (ADHD) (Jones and Miller, 2008; Rudin et al., 2021). Most drugs of abuse have been established to inhibit MAO activity, dopamine deaminating enzyme (Cunha-Oliveira et al., 2008; Fasakin et al. 2021). Therefore, the heightened inhibition of the enzyme activity in the present result may have resulted in elevated striatal dopaminergic concentrations. The elevated LDH activities may have also revealed the effects of the extracts on brain cells via the dopaminergic pathway as elevated LDH release have been established in heightened dopamine-concentration neurons and implicated to be suggestive of the loss of cell membrane integrity (Asanuma et al., 2020).
Glutamate (the major brain excitatory neurotransmitter) and its direct involvement in the addictive, drug seeking and dependence behaviours of drugs of abuse via the reward circuits in brain cells have been established (Danbolt et al., 2016; Yohn et al., 2019). Elevated glutamate concentrations during addiction are attributed to dopamine modulatory effects on the excitatory glutamate-releasing input and γ-aminobutyric acid-releasing output neurons (Wise and Robble, 2020). However, glutamatergic neurotransmission involving N-methyl-D-aspartate receptors (NRs) is directly involved in dependence and reinforcement in humans (Cunha-Oliveira et al., 2008). A functional NMDA receptor (NR) entails a minimum of NR1 subunit that consist a glycine binding site and the other consisting a glutamate binding site each (Ehsanifar et al., 2019). The inhibition of glutamate dehydrogenase activity and the activation of NR2A may be indicative that the stimulation of glutamatergic NMDA receptors is involved in facilitating the addictive-properties of the psychoactive plants (Lobo and Summavielle, 2016; Josiah et al., 2021; Fasakin et al., 2022a). Overstimulation of the glutamatergic NMDA receptors has been implicated to result in mitochondria impairments and excitotoxicity (Myslivecek, 2021; Josiah et al., 2022), which is suggestive of the neurotoxicological potential of AEDS at high dose.
The cholinergic neurotransmission system has been implicated to induce addiction via its involvement in mesolimbic dopamine activities (Yohn et al., 2019). Likewise, the elevation of striatal dopamine concentrations during addiction has been observed to be dependent on muscarinic and nicotinic acetylcholine receptors (mAChRs) activation (Lester et al., 2010; Faro et al., 2019), an indication that dopaminergic and cholinergic systems are intertwined during addiction. The involvement of cholinergic systems in glutamatergic terminals have also been established during the dependence and reinforcing-producing actions of nicotine (Lester et al., 2010; Myslivecek, 2021). The inhibitory effects on acetylcholinesterase may be suggestive of the possible addictive mechanism of the plants, as acetylcholinesterase is the principal factor underlining upregulated cholinergic neurotransmission and homeostatic acetylcholine concentrations (Fasakin et al., 2022a; Ademosun et al. 2023). Extensive acetylcholinesterase inhibition results in excess synaptic acetylcholine concentrations, altered postsynaptic cell function, overstimulated cholinergic receptors and cholinergic toxicity (Pope et al., 2005). AEDS administration elicited the highest inhibitory effects on acetylcholinesterase despite posing a non-significant effect on dopamine release, a probable validator that D. stramonium-induced acetylcholine release may be dependent on interactions between acetylcholine and other several catecholamines rather than solely dopamine (Ferrucci et al., 2019).
The high oxidizability of dopamine alongside high concentrations of dopamine observed in this study may have resulted in the aggravated ROS concentrations observed in the study. The inappropriate sequestering of dopamine into vesicles results in high cytosolic dopamine concentrations that has been implicated to result in elevated quinones, ROS and other toxic intermediates production via autoxidation, enzymatic reactions and metabolisms (Carvalho et al. 2012; Goldstein et al. 2012; Masoud et al. 2015). Quinones undergo redox cycle to form semiquinone radicals that result in superoxide radicals’ generation, which may then react with the elevated nitric oxide levels observed in the present study to potentiate neurotoxin peroxynitrite production which have been implicated in cellular proteins and DNA damages via interactions with thiol groups (Rudin et al. 2021). Additionally, dopamine autoxidation results in hydrogen peroxide (H2O2) and superoxide (O2.–) production, with the later having ability to react with elevated nitric oxide concentrations to form highly neurotoxic peroxinitrite (Cunha-Oliveira et al., 2008).
The induced-oxidative stress can trigger inflammatory responses characterized by heightened upregulation of proinflammatory mediators (such as TNF-α, NF-κB, IL-1β) and inhibition of anti-inflammatory mediators (such as IL-10) to stimulate apoptotic pathway (Fasakin et al., 2022a; Josaih et al., 2022). Neuroimmune response to addiction is characterized by the stimulation of microglia that leads to elevated proinflammatory cytokines production that interacts with neurotransmitters in related brain circuits (Flores-López et al. 2021; Gipson et al., 2021). In fact, brain pro-inflammatory neuroimmune signalling have been established to regulate GABA and AMPA receptor trafficking during addiction (Gipson et al., 2021). The oxidative stress stimulated-inflammatory responses can then initiate programmed-apoptosis via the activation of NFκB-dependent p53 signalling pathway and thereby, upregulated p53 gene expression which have been established to trigger loss and death of striatal dopamine-producing cells (Yan et al. 2014; Josiah et al., 2022). Elevated IL-1β and TNF-α concentrations can also directly induce neuronal apoptosis by stimulating adaptive and innate immune cells, and excessive glial hyper-activation (Tjalkens et al., 2017). This cascade coupled with the stimulation of NFκB and p53 gene expression of experimental rats’ striatum at the high doses of the alkaloid extracts may be indicative of addiction-induced neurodegeneration and cell death.
Purinergic neurotransmitters have also been implicated in the aetiology of addiction and neurotoxicity (Rozisky et al., 2010; Wu and Li, 2020). ATP and ADP are hydrolysed by E-NTPDases while AMP are hydrolysed by ecto-5’-nucleotidase to adenosine. Dopamine has also been postulated to act on appropriate dopamine receptors to elevate adenosine concentrations (Kombian et al., 2003). Resulting adenosine can then act via specific G-protein-coupled P1 purinoceptors, A2A-D2 receptor-receptor and A2AR interactions in brain’s dorsal striatum to mediate addiction and habitual substance seeking behaviours (Pintsuk et al., 2016; Burnstock, 2017). However, simulation of the striatal A2AR have been implicated to mediate fear, and elevate glutamate induced-excitotoxicity and interleukin-1β concentrations; indicators of neuroinflammation, depression and anxiety (Mayhew et al., 2018; Fasakin et al., 2022a). Observed E-NTPDase activity in this study is suggestive of elevated ATP hydrolysis, an indication of depleted bioenergetics (Leong et al. 2020). Also, the elevation of E-NTPDase activity resulting in reduced cAMP formation, information loss and cognitive dysfunction have also been established (Ademosun et al., 2022). Contra-wise, the inhibition of ecto-5’-nucleotidase and E-NTPDase activities by AEDS have been postulated to induce neurotoxicity via excessive inhibition of purinergic neurotransmission systems, a cascade that has proven more neurotoxic than that experienced with overstimulation of the adrenergic neurotransmission system (Fasakin et al., 2021).