Exploring the Aversive and Anxiogenic Effects of Novel Kappa Opioid Receptor Agonists in Rats

Drug addiction is characterised by uncontrolled, compulsive drug use despite negative consequences. As this disease has a high social and economic cost, greater attention is required in finding an effective treatment for individuals suffering addiction. Kappa opioid receptor (KOPr) agonists demonstrate anti-addiction effects in the rodent cocaine drug-prime model of reinstatement. Salvinorin A (Sal A), a novel non-nitrogenous KOPr agonist, has demonstrated reduced side-effects compared to traditional agonists. However, its short half-life and duration of action limit clinical development. The design of novel Sal A analogues with improved pharmacokinetics, anti-addiction effects, and reduced side-effects is an important step towards the pharmaceutical development of KOPr agonists. β-Tetrahydropyran Sal B (β-THP Sal B), Mesyl Sal B, ethoxymethyl salvinorin B ether (EOM Sal B), and Ethynyl Sal A (Ethy Sal A) have demonstrated anti-addiction effects by reducing cocaine-seeking behaviour in rats, but their aversive and anxiogenic properties have yet to be examined. Here the conditioned place aversion (CPA) paradigm is used to evaluate aversion and the elevated plus maze (EPM), light/dark test, and open field are utilised to measure anxiety in male Sprague-Dawley rats.  EOM Sal B (0.1 mg/kg, i.p) and Ethy Sal A (0.3 mg/kg, i.p) did not produce aversive effects, whereas the traditional KOPr agonist U50,488 (10 mg/kg, i.p), Sal A (0.3 mg/kg, i.p), and the novel analogue β-THP Sal B (1 mg/kg, i.p) produced significant aversion using the CPA protocol.  In the EPM all the novel analogues, β-THP Sal B, EOM Sal B, Mesyl Sal B, and Ethy Sal, A did not show a reduction in time spent on the open arm. In addition, EOM Sal B showed a significant increase in time spent on the open arm compared with Sal A (0.3 mg/kg, i.p). Sal A (0.3 and 1 mg/kg, i.p) showed significant anxiogenic effects, but the traditional agonist U50,488 did not. In the light/dark test Sal A (1 mg/kg, i.p) showed significant dose dependent anxiogenic effects with significant effects observed at 1 but not 0.3 mg/kg dose. This is in contrast to results observed in the EPM. The novel analogues EOM Sal B and β-THP Sal B demonstrated a non-significant trend toward anxiogenic behaviour in the light/dark test, but U50,488, Mesyl Sal B, and Ethy Sal A did not show significant reductions in time spent in the light box.  KOPr stimulation activates its associated G-proteins, allowing them to interact with several intracellular effectors. Activation of cAMP response element binding protein (CREB) can occur downstream of the KOPr signalling cascade. The phosphorylation of CREB is associated with dysphoria and stress-induced reinstatement of drug-seeking behaviour. An initial attempt to validate CREB assays was made.  The lack of behavioural anxiogenic and aversive side-effects with EOM Sal B, Mesyl Sal B and Ethy Sal A treatment demonstrates that the development of KOPr agonists with desirable effects and reduced side-effects is possible. These novel Sal A agonists provide promising candidates for pharmacotherapy development.