Antagonism by haloperidol and its metabolites of mechanical hypersensitivity induced by intraplantar capsaicin in mice: role of sigma-1 receptors

Jos M. Entrena Enrique J. Cobos Francisco R. Nieto Cruz M. Cendn Jos M. Baeyens Esperanza Del Pozo 0 ) Department of Pharmacology and Institute of Neuroscience, Faculty of Medicine, University of Granada , Avenida de Madrid 11, 18012 Granada, Spain Rationale We evaluated the effects of haloperidol and its metabolites on capsaicin-induced mechanical hypersensitivity (allodynia) and on nociceptive pain induced by punctate mechanical stimuli in mice. Results Subcutaneous administration of haloperidol or its metabolites I or II (reduced haloperidol) dose-dependently reversed capsaicin-induced (1 g, intraplantar) mechanical hypersensitivity of the hind paw (stimulated with a nonpainful, 0.5-g force, punctate stimulus). The order of potency of these drugs to induce antiallodynic effects was the order of their affinity for brain sigma-1 (1) receptor ([3H](+)-pentazocine-labeled). Antiallodynic activity of haloperidol and its metabolites was dose-dependently prevented by the selective 1 receptor agonist PRE-084, but not by naloxone. These results suggest the involvement of 1 receptors, but discard any role of the endogenous opioid system, on the antiallodynic effects. Dopamine receptor antagonism also appears unlikely to be involved in these effects, since the D2/D3 receptor antagonist ()-sulpiride, which had no affinity for 1 receptors, showed no antiallodynic effect. None of these drugs modified hind-paw withdrawal after a painful (4 g force) punctate mechanical stimulus in noncapsaicin-sensitized animals. As expected, the control drug gabapentin showed antiallodynic but not antinociceptive activity, whereas clonidine exhibited both activities and rofecoxib, used as negative control, showed neither. Conclusion These results show that haloperidol and its metabolites I and II produce antiallodynic but not antinociceptive effects against punctate mechanical stimuli and suggest that their antiallodynic effect may be due to blockade of 1 receptors but not to dopamine receptor antagonism. - Sigma () receptors, initially considered a subtype of opioid receptors and later confused with phencyclidine binding sites in N-methyl-D-aspartate (NMDA) receptors, are now described as a distinct pharmacological entity (for reviews, see Guitart et al. 2004; Monnet and Maurice 2006). Two subtypes (1 and 2) have been pharmacologically characterized. The 1 receptor, cloned in several animal species and humans, has been described as a unique protein with no homology with known mammalian proteins (Guitart et al. 2004; Monnet and Maurice 2006). Drug binding to 1 receptors is allosterically modulated by phenytoin (Quirion et al. 1992), and testing for this modulation has been proposed as a method to discriminate between 1 receptor agonists and antagonists in vitro (Cobos et al. 2005, 2006). The pharmacology of 1 receptors is now well-characterized, and selective agonists, such as (+)-pentazocine and PRE-084 [2-(4-morpholinethyl) 1-phenylcyclohexanecarboxylate) hydrochloride], and antagonists, such as BD-1063 (1-[2-(3,4-dichlorophenyl) ethyl]-4-methylpiperazine dihydrochloride) and NE-100 (N, N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]ethylamine hydrochloride), are both available (Guitart et al. 2004; Hayashi and Su 2004). Some neurosteroids, psychostimulants, and antipsychotics also bind to 1 receptors (Maurice et al. 2001; Monnet and Maurice 2006; Cobos et al. 2008). Among the antipsychotics, haloperidol (HP) is mainly known as a D2 receptor antagonist, although it shows the same affinity for D2 and 1 receptors (Bowen et al. 1990; Matsumoto and Pouw 2000) and exhibits 1 receptor antagonistic activity (Maurice et al. 2001; Hayashi and Su 2004). Two major metabolic pathways for HP have been identified in experimental animals and humans (for references, see Cobos et al. 2007). One is a reversible reductive pathway that produces HP metabolite II (HP-Met-II), also called reduced HP [4-(4-chlorophenyl)--(4-fluorophenyl)4-hydroxy-1-piperidinebutanol]. The other is an oxidative N-dealkylation pathway that leads to HP metabolites I (HP-Met-I, 4-(4-chlorophenyl)-4-hydroxypiperidine) and III (HP-Met-III, p-fluorobenzoylpropionic acid). Studies performed in rodent brain membranes and human neuroblastoma cells showed that metabolites I and II of HP bind to 1 receptors with less affinity than HP, but show much lower (HP-Met-II) or no affinity (metabolite I) for D2 receptors, whereas metabolite III has no affinity for either 1 or D2 receptors (Bowen et al. 1990; Matsumoto and Pouw 2000; Cobos et al. 2007). Sigma-1 receptors are involved in nociception, among other processes. They are distributed in the central nervous system in areas of great importance for pain control, such as the superficial layers of the spinal cord dorsal horn, the periaqueductal gray matter, the locus coeruleus, and rostroventral medulla (Alonso et al. 2000; Kitaichi et al. 2000). Functional studies have postulated that an endogenous 1 system tonically modulates the opioid system. The antinociception induced by agonists of opioid receptors in the tail flick test is antagonized by systemic administration of the selective 1 agonist (+)-pentazocine, whereas it is enhanced by the 1 antagonist HP (Chien and Pasternak 1993, 1994; Mei and Pasternak 2002, 2007). New 1 ligands such as the 1 antagonist (+)-MR 200 [(+)-methyl (1R,2S)-2-{[4-(4-chlorophenyl)-4-hydroxypiperidin-1-yl] methyl}-1-phenylcyclopropanecarboxylate] and the proposed 1 agonist ()-PPCC [(1R,2S/1S,2R)-2-[4-hy droxy-4-phenylpiperidin-1-yl)methyl]-1-(4-methylphenyl) cyclopropanecarboxylate] also modulate opioid receptor agonist-induced antinociception (Marrazzo et al. 2006; Prezzavento et al. 2008). Sigma ligands are also able to modulate nociception per se (i.e., not associated to opioid agonists). Selective 1 agonists induce nociception when used alone in the nociceptive flexor response test and the effects of (+)-pentazocine are reversed by selective 1 receptor antagonists (Ueda et al. 2001). Moreover, both phases of pain behavior in the formalin test are diminished in 1 receptor knockout mice (Cendn et al. 2005a) and after the systemic administration of the 1 receptor antagonists HP and reduced HP (Cendn et al. 2005b). Pain behavior in the second phase of the formalin test is also reduced after intrathecal administration of the 1 receptor antagonists BD-1047 (N-[2-(3,4-dichlorophenyl)ethyl]-Nmethyl-2-(dimethylamino ethylamine dihydrobromide) and BMY-14802 (-(4-fluorophenyl)-4-(5-fluoro-2-pyrimidinyl)1-piperazinebutanol) (Kim et al. 2006). However, the possible role of 1 receptors in mechanical stimulusinduced pain is unknown. The intradermal injection of capsaicin induces an immediate pain behavior response followed by longerlasting secondary mechanical hypersensitivity (Gilchrist et al. 1996; Joshi et al. 2006). The mechanisms underlying the mechanical hypersensitivity (allodynia) produced by the intradermal injection of capsaicin and the second phase of the formalin (...truncated)