Tropeines were originally identified as potent 5-HT3 receptor antagonists. Tropisetron, also known as ICS-205,930, is one of the best-known compounds of this group. It is used mainly as an anti-emetic following chemotherapy due to its ability to target 5-HT3 receptors involved in vomiting reflexes. An increasing body of evidence has shown that tropeines also allosterically modulate GlyRs of different subunit composition. Pioneering electrophysiological recordings in cultured spinal neurons have revealed that two tropeines, MDL-72222 and tropisetron, were able to potentiate GlyR chloride currents at nanomolar concentrations (Chesnoy-Marchais, 1996). In contrast, higher micromolar concentrations caused inhibition. Subsequent studies determined that potentiation only occurred in the presence of GlyR agonists, depended on the agonist concentration, and was also present in outside-out patches (Chesnoy-Marchais, 1996; Supplisson and Chesnoy-Marchais, 2000; Yang et al., 2007). The potentiation elicited by tropisetron remained unaltered in the presence of zinc, ethanol or propofol, suggesting different binding sites and mechanisms (Chesnoy-Marchais, 1999). Interestingly, tropisetron also displayed subunit-specificity. Studies in recombinant GlyRs showed that tropisetron potentiated homomeric α1 but inhibited homomeric α2-GlyRs. Furthermore the expression of β subunits significantly increased the potentiation sensitivity of α1 and switched α2-GlyR inhibition to potentiation. These results suggest that the tropeine potentiating site lies within the α-α or α-β interface (Supplisson and Chesnoy-Marchais, 2000). Other studies also found that α2-GlyR was more effectively inhibited by tropisetron than α1-GlyR, but in contrast, they did not find any potentiation even in the presence of β subunits (Maksay et al., 1999). Despite these differences, the electrophysiological data correlated well with binding studies in recombinant and native membrane preparations. For example, several tropeines have been shown to inhibit 3[H]strychnine binding to GlyRs with high nanomolar affinity. In addition, they increase the glycine potency to displace 3[H]strychnine, suggesting direct effects on glycine binding sites (Maksay, 1998; Maksay et al., 2004). In general terms, structure-activity analysis suggests that the tropeine ring itself, the tropeine nitrogen, an aromatic ring and a carbonyl group are necessary for binding and functional potentiation (Maksay, 1998; Chesnoy-Marchais et al., 2000; Maksay et al., 2004). The tropeine ring, on the other hand, appears to be a primary requirement for functional inhibition (Yang et al., 2007; Maksay et al., 2009).
Recently, several studies addressed the location of the tropeine binding sites on GlyRs. In agreement with studies performed in 5-HT3 receptors (Yan and White, 2005; Joshi et al., 2006), tropeines appear to bind to cavities within the extracellular domain located close to the ligand binding sites. Using recombinant GlyRs, Yang et al. (2007) showed that mutations to N102 in the α1, but not in the β subunit (N125), abolished tropisetron inhibition without affecting the potentiation. Subsequent work performed with a structurally related tropeine (3α-(3′-methoxy-benzoyloxy) nortropane, MBN) determined that other amino acid substitutions close to the agonist-binding domain of α1-GlyRs also alter the MBN inhibition or potentiation of GlyRs (see Figure 2, Maksay et al., 2009). In addition, homology models and molecular docking simulations also suggest that the biphasic modulation elicited by tropeines on GlyRs is likely to involve different docking modes in adjacent binding sites within the agonist-binding region (Maksay et al., 2009).
The high affinity binding and the remarkable sensitivity of GlyRs to tropeines makes this group of compounds one of the most promising candidates for the development of specific drugs targeting GlyRs. Despite the existence of some interesting differences between the chemical determinants required for tropeine binding to GlyRs and 5-HT3 receptors, most tropeines still bind and modulate 5-HT3 receptors with high affinity (Maksay et al., 2004). In addition, the biphasic nature of tropeine-GlyR modulation and the significant overlap between the requirements for potentiation and inhibition is also an important impediment to their use as enhances of GlyR function. A better understanding of the mechanisms underlying the potentiation of GlyR subtypes by tropeines will hopefully lead to new tropeine derivatives lacking glycinergic inhibition and 5-HT3 receptor binding.