The aim of the present research was to determine the involvement of the endocannabinoid system, through CB2 receptors, in the hyperlocomotion of mice, provoked by an acute injection of NMDA receptor antagonist, MK-801, as an animal pharmacological model of schizophrenia.

We revealed that an acute injection of AM 630, CB2 receptor antagonist, used in our experiments at the doses 0.1–1.0 mg/kg, but not at the dose of 2.0 mg/kg, decreased locomotion of mice. An acute injection of AM 630 (2.0 mg/kg) of its own had no statistical significant influence on the locomotor activity of mice. However, in the next step, we revealed that an acute injection of AM 630, at the non-effective dose of 2.0 mg/kg, potentiated MK-801-induced hyperactivity. What is of interest, we revealed that an acute injection of JWH 133, CB2 receptor agonist, used in our experiments at the doses 0.05–1.0 mg/kg, but not at the dose of 2.0 mg/kg, also decreased locomotion of mice. An acute injection of JWH 133 (2.0 mg/kg) of its own had no statistical significant influence on the locomotor activity of mice. However, an acute injection of JWH 133 at the non-effective dose of 2.0 mg/kg intensified the MK-801-provoked hyperactivity.

The first CB2 receptor ligand-related effect that should be discussed is associated with the fact that both the CB2 agonist (2.0 mg/kg; not a lower dose) and CB2 antagonist (2.0 mg/kg; not a lower dose) altered MK-801 locomotor effects (similar effects). The second interest CB2 receptor ligand effect should focused on the fact that these cannabinoid compounds had no effect of their own (i.e., without MK-801 injection) on locomotor activity.

The increase of MK-801 induced hyperactivity caused by both CB2 antagonist and CB2 agonist obtained in our studies seems to be very important. The same direction of action we have is revealed in our previous studies. We have shown the enhancement of memory and learning processes by an acute administration of CB2 receptor agonist (JWH 133 at the doses of 0.5; 1.0 and 2.0 mg/kg) and antagonist (AM 630 at the dose of 1.0; 2.0 and 3.0 mg/kg) assessed in the passive avoidance (PA) test (Kruk-Slomka et al. 2016a). Similarly, we have revealed that an acute administration of both JWH 133 (0.5 and 1.0 mg/kg) and AM 630 (0.5 mg/kg) exhibited antidepressant action in the forced swimming test (FST) (Kruk-Slomka et al. 2015). These results confirmed in our present and previous cited experiments may be connected with pharmacokinetic properties of used CB2 receptor ligands, especially the CB2-selective agent, AM 630. The pharmacological properties of AM 630 are complex. It has been shown that AM 630 behaves as inverse agonist rather than as “silent” antagonist. The inverse efficacy at CB2 receptors but also the CB2/CB1 affinity ratio has been indicated for AM 630 (CB2/CB1 affinity = 165). Thus, AM 630 has been found to behave as an inverse agonist at CB2 receptors as well as an inverse agonist at CB1 receptors (Bolognini et al. 2012; Ross et al. 1999).

Naturally, the administration of different doses of selective CB2 receptor agonists/antagonists provides some information for a neurophysiological role of CB2 receptors in the brain, although non-selective effects at these doses cannot be ruled out. What is of interest, it has been revealed that CB2 receptor antagonist, SR144528, causes biphasic effects on the locomotion of mice, increasing spontaneous locomotor activity in the DBA/2 mouse at low doses (1.0–10.0 mg/kg) and decreasing this activity at high doses (20.0 mg/kg) (Onaivi et al. 2008). Although, Sain et al. (2009) as well as Whiteside et al. (2005) revealed that genetic deletion of the CB2 receptor has not been connected with any change in motor effects. On the other hand, increasing doses of the CB2 receptor agonist, JWH-015, decreased locomotor activity and stereotyped behavior what was depended on the gender of animals (Onaivi et al. 2006). Similarly, typical hypolocomotion was observed after administration of an alternative CB2 receptor agonist GW 405833 (Valenzano et al. 2005) but the injection of the selective CB2 receptor agonists HU 308 and AM 1214, at doses provoking significant antinociceptive effects, did not affect locomotor activity (Hanus et al. 1999; Malan et al. 2001).

Additionally, Xi et al. (2011) have revealed that intranasal or intra-accumbens local administration of CB2 receptor agonist, JWH 133, dose-dependently inhibits cocaine-enhanced locomotion in wild-type (WT) and CB1 receptor-knockout (CB1−/−), but not CB2−/− mice. This inhibition is mimicked by GW 405833, another CB2 receptor agonist with a different chemical structure, and is blocked by AM 630, a selective CB2 receptor antagonist. Intra-accumbens administration of JWH 133 alone dose-dependently decreases, while intra-accumbens AM 630 elevates locomotion in WT and CB1−/− mice, but not in CB2−/− mice.

Our results indicating that CB2 receptor ligands did not affect locomotor activity in the dose used (2.0 mg/kg) are consistent also with other literature data (Ishiguro et al. 2010; Kim and Li 2015; Khella et al. 2014; Onaivi 2009; Ortega-Alvaro et al. 2011). The data indicating the role of CB2 receptor agonist in the MK-801-induced disruptions of PPI in mice seem to be interesting (Khella et al. 2014). In this cited experiment, JWH 015, a CB2 receptor agonist had no significant effect on prepulse inhibition (PPI) alone but reversed disruptions in PPI induced by MK-801. This effect was attenuated by the injection of AM 630, but not by AM 251, CB1 receptor antagonist, suggesting an involvement of CB2 receptors in these effects. Additionally, JWH 203, the mixed CB1/CB2 receptor agonist only partially reduced MK-801-induced PPI disruptions but neither AM 630 nor AM 251 had any significant effect alone or on MK-801-induced disruptions in PPI (Khella et al. 2014). In turn, Ishiguro et al. (2010) have confirmed that AM 630 exacerbated MK-801- or methamphetamine-induced disturbance of PPI and hyperactivity in C57BL/6JJmsSlc mice. Thus, we can suspect that the inhibition of the activity of CB2 receptor by AM 630 at the dose of 2.0 mg/kg is not able to contribute to the emergence of symptoms of schizophrenia. However, the inhibition of the function of receptor CB2 in combination with other risk factors (e.g., the impairment of glutamatergic transmission by the use of NMDA receptor antagonist—MK-801) may provoke or potentiate the symptoms of schizophrenia, perhaps in the people predisposed to schizophrenia. Perhaps, the possible explanation of our results is connected with the fact that pharmacological ligands targeting CB2 receptor (JWH 133 and AM 630) in the normal conditions (without injection of MK-801) and in the highest dose used (2.0 mg/kg) had no influence on the locomotor activity in mice. But in the presence of some risk factor of schizophrenia in animals (e.g., administration of MK-801), CB2 expression in the brain is changed and CB2 receptor ligands exhibit potent antipsychotic (anti-hyperactivity) effects. Naturally, we can only support these conclusions and more detailed knowledge of these effects needs further investigations.

The mechanism of action of CB2 receptor ligands in psychosis-like responses in mice is still unclear. Although, a number of studies have investigated the association between cannabis and psychosis, many questions remain unanswered. It has been known that endocannabinoid system, mainly via CB1 receptors, is involved in pathomechanisms of schizophrenia (Barzegar et al. 2015; Kruk-Slomka et al. 2016b; Kucerova et al. 2014; Levin et al. 2012; Roser and Haussleiter 2012), while the CB2 receptors locating outside the CNS are involved in the processes related to the function of the immune system (Fukuda et al. 2014; Pertwee 2010; Wright et al. 2008). However, recent studies have provided evidence that CB2 receptors may also be arranged outside the cells of the immune system. Literature data suggested that these receptors can also be found in various brain regions of humans and many animal species (Benito et al. 2008; Zhang et al. 2014). The studies on the biochemical and histological level have revealed that the CB2 receptors are located in the neuronal progenitor cells, neurons, glial chambers, and endothelium. There is also evidence for the expression of CB2 receptors in areas of the brain that are particularly pertinent to the context of schizophrenia, such as regions involved in sensorimotor gating (Onaivi 2009; Racz et al. 2008; Zhang et al. 2014), but the functional significance of this discovery has not been established yet.

Following that, the functional role of central CB2 receptors is not fully elucidated yet. Recent evidence obtained from pharmacological (Ishiguro et al. 2010; Kim and Li 2015; Onaivi 2009; Ortega-Alvaro et al. 2011) and genetic studies (Ortega-Alvaro et al. 2011) suggest that centrally expressed CB2 receptors are involved in many processes of neuropsychiatric disorders, such as behavior characteristic of schizophrenia, impulsive behavior, locomotor activity, stereotyped behaviors, anxiety-, pain-, and memory-related processes (Khella et al. 2014; Kim and Li 2015; Kruk-Slomka et al. 2016a). Moreover, literature data (Kruk-Slomka et al. 2015; Onaivi 2009) revealed the possible involvement of CB2 receptors in depression, which may be of importance in many psychiatric disorders connecting with emotional imbalance, including schizophrenia. What is more, the literature data indicated that the deletion of the gene encoding CB2 receptors may lead to neurochemical changes which can be a consequence of a behavioral disorder mentioned above (Ishiguro et al. 2010; Ortega-Alvaro et al. 2011).

We confirmed in results presented in this paper that both stimulation and blockade of CB2 receptors is able to modulate locomotor activity of mice that correlates with the neuropsychiatric effects. It has been known that the neuropsychiatric effects of endocannbinoid system, via CB1 and CB2 receptors, are associated with the modulation of different neurotransmitter systems, such as dopaminergic system, glutamatergic system, or GABA-related system (Broome et al. 2005; Carlsson et al. 2004). A variety of pre-clinical and clinical studies have indicated that mainly CB1 receptors participate in many central pathways connected with psychosis-like state through influence on the glutamatergic transmission (Barzegar et al. 2015; Kruk-Slomka et al. 2016b). However, in the case of CB2 receptors, many literature data suggested an important role of CB2 receptors, locating in the specific brain areas, mainly on the dopaminergic neurons in the ventral tegmental area (VTA). Electrophysiological studies have demonstrated that CB2 receptor activation by JWH 133 or other CB2 receptor agonists leads to inhibition of dopaminergic neuron activation in the VTA. Therefore, these CB receptors play an important role in the modulation of dopaminergic system and are involved in the behaviors connecting with dopaminergic system-related disorders, such as schizophrenia, anxiety, depression, and Parkinson disease (Zhang et al. 2014). Additionally, the endocannabinoid system via CB2 receptors can be associated with schizophrenia-like response, due to the fact that this type of receptors is responsible for the increase of amount of annandamide, the main endocannabinoid occurring in the brain. The clinical studies confirmed that the clinical remission of schizophrenia may be connected with the significant decrease in the level of anandamide and messenger ribonucleic acid (mRNA) encoding CB2 receptors (Ishiguro et al. 2010; Vigano et al. 2008). However, there are significant species differences in CB2 receptor mRNA splicing and expression, protein sequences, and CB2 receptor ligand-related in mice and rats (Zhang et al. 2015) indicating that the cellular mechanisms underlying these actions are still unclear; therefore, the function of cannabinoid CB2 receptors in the brain have been subject to debate. Following that, the influence of CB2 agonists and antagonists on hyperlocomotion of mice is controversial and inconsistent, for this reason need further investigation.