Effects of the cannabinoid receptor 1 positive allosteric modulator GAT211 and acute MK-801 on visual attention and impulsivity in rats assessed using the five-choice serial reaction time task
Introduction
Schizophrenia is a chronic neuropsychiatric disorder affecting 0.4% of the global population (Nicholl et al., 2010). The majority of antipsychotics in use today are dopamine antagonists, though many of these drugs cause serious side effects, including heart disease, diabetes, and extrapyramidal motor symptoms (Li et al., 2016; Goff et al., 2017). In addition, the efficacy of these dopamine antagonists are questionable in certain populations, and they tend to treat only the positive symptoms of schizophrenia while having little or no effect on the negative and cognitive symptoms (Keefe et al., 2007; Stafford et al., 2015). As a result, some researchers suggest that abnormalities in dopamine signaling are likely the “final common pathway” to schizophrenia and result from the dysregulation of other neurotransmitter systems, including at N-methyl-d-aspartate (NMDA) receptors, a subtype of ionotropic glutamate receptors (Coyle, 2012; Cannon, 2015). Indeed, non-competitive NMDA receptor antagonists are routinely used as a drug-induced model for schizophrenia, as they can replicate the aforementioned NMDA receptor hypofunction and induce a state resembling some aspects of acute psychosis.
Cannabinoid receptor 1 (CB1R) is the most abundant G protein-coupled receptor (GPCR) in the central nervous system and its activation regulates neurotransmitter release in glutamatergic neurons in the cortico-striatal-limbic circuitry involved in schizophrenia (Robson et al., 2014; Pertwee, 2015; Laprairie et al., 2017; Alaverdashvili and Laprairie, 2018). The 2 main endogenous cannabinoids acting on CB1R are 2-arachidonoylglycerol (2-AG, a full CB1R agonist) and anandamide (AEA, a partial CB1R agonist). 2-AG and AEA primarily act via retrograde signaling, meaning they are released by postsynaptic neurons and bind to the CB1Rs expressed on presynaptic neurons to modulate presynaptic neurotransmitter release (Van Winkel and Kuepper, 2014). Recent studies show that patients with schizophrenia have lower CB1R availability in many brain regions when compared to controls (Ranganathan et al., 2016; Volk and Lewis, 2016; Borgan et al., 2019). Therefore, compounds which enhance CB1R activity may be promising therapeutic agents. However, the roles of cannabinoids and CB1R in schizophrenia are complex. For example, the phytocannabinoid cannabidiol (CBD) may improve some symptoms of psychosis via unknown mechanisms, whereas the CB1R partial agonist Δ9-tetrahydrocannabinol (THC) significantly worsens some symptoms of psychosis (Leweke et al., 2012; Manseau and Goff, 2015). These differences may be attributed to the pluri-dimensional signaling of CB1R, such that different ligands may elicit different responses from the same receptor (Hua et al., 2017). They may also be attributed to distinct interactions between exogenous cannabinoids and endogenous cannabinoids. For example, CBD has been found to inhibit the reuptake of AEA, and the concentration of AEA in the cerebrospinal fluid is found to be inversely related to the severity of psychosis symptoms in schizophrenia patients (Manseau and Goff, 2015; Giuffrida et al., 2004). Therefore, there is merit in studying the modulation of CB1R in the context of schizophrenia (Mielnik et al., 2020).
A positive allosteric modulator (PAM) is a ligand that binds to a site on its receptor that is distinct from the main (i.e., orthosteric) site of endogenous binding and in doing so enhances the potency of the orthosteric ligand and/or the efficacy of the receptor's signaling (Laprairie et al., 2017). True PAMs lack intrinsic efficacy and thus only modulate endogenous ligand actions on the receptor (Laprairie et al., 2017). CB1R PAMs may thus help keep the receptor's activity at a homeostatic level and normalize endogenous cannabinoid system (ECS) tone (Laprairie et al., 2017; Mielnik et al., 2020). Existing pre-clinical studies have shown that CB1R allosteric modulators are safer than traditional CB1R agonists in terms of tolerance and abuse liability (Laprairie et al., 2017; Alaverdashvili and Laprairie, 2018; Mielnik et al., 2020). GAT211 is a CB1R allosteric modulator composed of 2 enantiomers: GAT228, an allosteric agonist and PAM; and GAT229, a pure CB1R PAM without agonist actions (Laprairie et al., 2017). Therefore, GAT211 is a mixed agonist-PAM (ago-PAM). Several recent studies have demonstrated the therapeutic potential of GAT211 in various conditions including pathological pain, PTSD, ocular hypertension, and Huntington's disease (Shekhar and Thakur, 2018; Slivicki et al., 2018; Laprairie et al., 2019; Slivicki et al., 2020). However, to the best of our knowledge, no previous studies have assessed the effects of GAT211, or any CB1R PAM or ago-PAM, in rodent models of the cognitive symptoms of schizophrenia. Therefore, in the present experiment, we chose to test the effects of GAT211 on the Five-Choice Serial Reaction Time (5CSRT) task in rats.
The 5CSRT task enables the measurement of visual attention, impulsivity, and perseveration using well established, automated procedures in operant conditioning chambers (Bari et al., 2008). Our apparatus made use of touchscreens for displaying the stimuli and measuring the rats' responses during the task (Mar et al., 2013; Hvoslef-Eide et al., 2015). The effects of GAT211 on the 5CSRT task were assessed alone and in combination with acute administration with the NMDA receptor antagonist MK-801 in an effort to recapitulate some aspects of NMDA receptor hypofunction as observed in schizophrenia.
Section snippets
Subjects
Adult male Long Evans rats (Charles River Laboratories, Kingston, NY, USA) were used for this experiment. Upon arrival at the vivarium at the University of Saskatchewan animal facility, the rats were given food (Purina Rat Chow) and water ad libitum, pair-housed in plastic ventilated home cages, and left undisturbed for 1 week before handling. The vivarium was temperature- and humidity-controlled, and provided a 12:12 h light:dark cycle, with the light-on period beginning at 0700 daily. All
Results
MK-801 (0.15 mg/kg) significantly disrupted task performance on several parameters. GAT211 failed to have effects on its own or affect the MK-801-induced disruptions. Response accuracy, derived from the number of correct responses divided by the total number of responses, was significantly reduced by MK-801 (Fig. 2a; main effect of MK-801: F(1,12) = 8.71; p = 0.012). GAT211 failed to impact accuracy levels alone (main effect of GAT211: F(2,24) = 0.47; p = 0.51) and did not significantly reverse
Discussion
The present study assessed the effects of acute MK-801 and GAT211 treatments on performance of the 5CSRT task in male rats. Our results support a potent effect of subanesthetic doses of acute MK-801 in impairing accuracy, increasing omissions, and increasing premature responding in the 5CSRT task. Administration of GAT211, a CB1R ago-PAM, did not reverse these impairments or have effects on its own.
Previous studies have explored the effects of MK-801 on the 5CSRT task. Acute treatment with low
Conclusion
This experiment demonstrated that acute MK-801 treatment potently alters accuracy and premature responding in the 5CSRT task. While CB1R ago-PAMs, such as GAT211, are promising new compounds for increasing CB1R activity, GAT211 treatment did not significantly affect the effects of MK-801 on the 5CSRT task. Further characterization of the effects of altering CB1R activity on behavioral models of schizophrenia, including NMDA receptor hypofunction models, may facilitate the development of novel
Ethical statement (Onofrychuk et al. 2020)
Ethical guidelines related to the ARRIVE guidelines for the use of animals in research have been followed. This is indicated in the Methods section of the submitted paper.
Declaration of Competing Interest
None.
Acknowledgements
Funding for the project was provided by a Saskatchewan Health Research Foundation (SHRF) Establishment Grant and a Canadian Institutes of Health Research (CIHR) Partnership Grant with GlaxoSmithKline to RBL; and funds from the College of Medicine, University of Saskatchewan, and a CIHR Project Grant to JGH. This work was also supported by grant from National Institutes of Health to GAT (EY024717). TJO, SC, DLM, and AJR received scholarship support from Natural Sciences and Engineering Research
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