Introduction

Over the last several years evidence has emerged indicating that the 5-HT2C receptor subtype exerts a modulatory influence over the functioning of the mesocorticolimbic dopamine (DA) system originating in the ventral tegmental area. The moderately selective 5-HT2C receptor agonist Ro 60-0175 [(S)-2-(6-chloro-5-fluoroindol-1-yl)-1-methylethylamine] (Martin et al. 1998) reduces the firing rate of mesolimbic DA neurons originating in the ventral tegmental area, leading to a reduction in DA release in terminal regions of the nucleus accumbens and frontal cortex (Di Matteo et al. 2000; Gobert et al. 2000). These effects are reversed by the selective 5-HT2C receptor antagonist 6-chloro-5-methyl-1-[[2-[(2-methyl-3-pyridyl)oxy]-5-pyridyl]carbamoyl]-indoline (SB242084; Bromidge et al. 1997) (Di Matteo et al. 2000; Gobert et al. 2000). In addition, SB242084 alone increases the burst-firing of dopaminergic neurons in the ventral tegmental area leading to increased release of DA in the nucleus accumbens (Di Matteo et al. 1999). Thus, it appears that 5-HT2C receptors may exert a tonic inhibitory influence over the activity of ascending DA neurons (Di Matteo et al. 2002).

The results of behavioral studies are consistent with this modulatory role of 5-HT2C receptors on DA function. Much of this work has examined the expression of behaviors induced by cocaine, which are thought to be dopaminergically mediated, after treatment with 5-HT2C receptor ligands. Recent studies have now shown that 5-HT2C receptor activation reduces cocaine-stimulated locomotor activity, cocaine self-administration, the ability of a priming injection of cocaine to reinstate drug seeking behavior, and the discriminative stimulus properties of cocaine (Frankel and Cunningham 2004; Grottick et al. 2000). In contrast, blockade of 5-HT2C receptors increases cocaine-induced locomotion, self-administration, and cocaine-primed reinstatement of responding (Filip et al. 2004; Fletcher et al. 2002a). Mice lacking the 5-HT2C receptor also show increased responses to cocaine in tests of locomotion and self-administration (Rocha et al. 2002). The amphetamine derivative MDMA stimulates locomotor activity, and this effect is partially mediated by DA release (Gold et al. 1989; Kehne et al. 1996). In agreement with findings obtained with cocaine, 5-HT2C receptor blockade also enhanced the locomotion induced by MDMA (Fletcher et al. 2002b; McCreary and Cunningham 1999).

The results of studies involving cocaine and MDMA are consistent with the notion that blocking 5-HT neurotransmission at the level of the 5-HT2C receptor enhances DA-dependent function and behavior. However, both cocaine and MDMA also increase 5-HT neurotransmission by blocking the 5-HT transporter and by releasing 5-HT from neuronal terminals, respectively (Bradberry et al. 1993; Kankaanpaa et al. 1998; Ritz and Kuhar 1989; Teneud et al. 1996). At least in the case of MDMA, a direct role for increased 5-HT function in mediating locomotor stimulation induced by this drug has been shown (Bankson and Cunningham 2002; Callaway et al. 1990; Fletcher et al. 2002b; Kehne et al. 1996). This suggests that 5-HT2C receptor blockade may influence MDMA- and/or cocaine-mediated behavioral effects directly via a serotonergic mechanism.

The main aim of the present study was to examine the effects of pharmacological blockade of 5-HT2C receptors using SB242084 on the expression of locomotor activity elicited by a variety of manipulations that are known to have different effects on 5-HT and/or DA function. The first experiment compared the effects on locomotor activity of SB242084 given before treatment with a mixed DA and 5-HT releaser, MDMA with those resulting from treatment with the selective DA releaser amphetamine, or the selective 5-HT releaser fenfluramine. A parallel experiment examined the effects of SB242084 given in conjunction with a mixed 5-HT and DA reuptake blocker, cocaine, with those resulting from a combination of SB242084 with the selective DA reuptake blocker, methylphenidate, or the selective 5-HT reuptake blocker, citalopram. To further examine whether SB242084 modifies the expression of DA-dependent locomotor activation, we examined the interactions between SB242084 and morphine, or nicotine, both of which stimulate locomotion in part via activation of the mesolimbic DA system (Benwell and Balfour 1992; Joyce and Iversen 1979; Kelly and Iversen 1976; Reavill and Stolerman 1990). Finally, the effect of SB242084 against the mixed direct 5-HT1A/1B receptor agonist RU24969 (Barnes and Sharp 1999) was examined.

Materials and methods

Animals and housing

Adult male Sprague–Dawley rats weighing 225–250 g at the beginning of the experiments were used. The rats were housed in pairs in clear polycarbonate cages under a 12-h light/dark cycle (lights on at 8 a.m.). Food and water were freely available in the home cages.

Apparatus

All experiments were conducted in a custom-built locomotor activity monitor. This system consisted of 16 clear polycarbonate cages, measuring 25-cm-wide, 20-cm-high, and 45-cm-long. An array of six infrared photocells was attached outside the longer sides of the cages. The photocells were spaced 7.5 cm apart and 2 cm above the floor of the cage. The equipment was housed in a room different to the colony room.

Effects of SB242084 on drug-induced locomotor activity

Separate groups of rats were used to investigate the effects of pretreatment with 0.5 mg/kg of SB242084 or its vehicle on changes in locomotor activity induced by amphetamine (0, 0.25, and 0.5 mg/kg; n=8; and 0, 0.5, and 1 mg/kg; n=8), methylenedioxymethamphetamine (0, 2.5, and 5 mg/kg; n=8), d-fenfluramine (0, 1, and 5 mg/kg; n=10), cocaine (0, 5, and 10 mg/kg; n=8), methylphenidate (0, 2.5, and 5 mg/kg; n=10), citalopram (0, 10, and 20 mg/kg; n=10), and RU24969 (0, 0.5 and 1 mg/kg; n=8). In all of these experiments, before any drug testing all rats were familiarized with the activity monitors by placing them in the cages for a 2-h period on each of three consecutive days. Two additional groups of rats were used to examine the interaction between SB242084 and nicotine (0, 0.2, and 0.4 mg/kg; n=12) and morphine (0, 2.5, and 5 mg/kg; n=12). Before testing began on these animals they were exposed to nicotine or morphine to induce tolerance to the depressant effects of these drugs. In the case of the nicotine group, rats received ten daily injections of 0.4 mg/kg of nicotine in the home cage; in the case of the morphine group they received four daily injections of 10 mg/kg of morphine in the home cage. Forty-eight hours after the last injection rats were familiarized with the activity monitors by placing them in the cages for a 2-h period on each of three consecutive days.

In all experiments on the test days rats were placed in the activity cages for 30 min and then injected with 0.5 mg/kg of SB242084 or its vehicle (see below). Rats were then returned to the activity cages for a further 30 min. At the end of this period rats were injected with the appropriate dose of the test drug and were replaced in the activity cages for 2 h. The number of photocell interruptions was recorded every 10 min. For all drugs, a repeated measures design was used whereby each rat was tested under all six possible combinations of SB242084 or its vehicle plus the two doses of the test drugs and the appropriate vehicle. Treatment combinations were administered in a semirandomized fashion with each combination given to at least one animal on each day. For each compound testing was usually conducted on Mondays and Thursdays or Tuesdays and Fridays.

The dose of SB242084 was chosen based on a consideration of the literature and on in-house data showing blockade of a number of 5-HT2C-mediated responses by SB242084 (e.g., Fletcher et al. 2004; Martin et al. 2002).

Statistics

Data for all studies were analyzed using repeated measures three-way analysis of variance with SB242084 pretreatment, drug treatment, and time bin (10 min) as factors. Where appropriate, a significant three-way interaction was further analyzed by conducting separate two-way analyses of variance with SB242084 pretreatment and drug treatment as factors at each 10-min time point. Post hoc comparisons between means were conducted using the Fisher’s least significant difference test.

Drugs

SB242084 was synthesized in the Department of Chemistry, Vernalis Research, Wokingham, UK, prepared in 0.9% saline solution containing 8% hydroxypropyl-β-cyclodextrin and 25 mM citric acid, and was injected by the IP route. Cocaine hydrochloride (BDH, Toronto, Ontario, Canada), d-amphetamine sulfate (Bureau of Drug Surveillance, Ottawa, Ontario, Canada), fenfluramine HCl (Servier), RU24969 [5-methoxy-3-(1,2,5,6-tetrahydro-4-pyridinyl)-1H-indole; National Institute on Drug Abuse (NIDA) Drug Supply Program], and morphine sulfate (Bureau of Drug Surveillance, Ottawa, Ontario, Canada) were all dissolved in 0.9% saline and injected IP. MDMA [(±)-3,4-methylenedioxymethamphetamine HCl; NIDA Drug Supply Program] was dissolved in 0.9% saline and injected SC. Nicotine (Sigma-Aldrich, Oakville, Ontario, Canada) was dissolved in saline with the pH adjusted to 7.0 and was injected SC.

Results

Interactions between SB242084 and MDMA, amphetamine, or fenfluramine

Figure 1 shows the effects on locomotor activity of combining SB242084 with MDMA, amphetamine, or fenfluramine. MDMA significantly increased locomotor activity (F 2,14=96.5, p<0.001); and the main effect of SB242084 was also significant (F 1,7=27.38, p<0.001). The interaction between SB242084 and MDMA was significant (F 2,14=14.67, p<0001) and post hoc testing showed a significant enhancement of the effects of both doses of MDMA, measured over the 2-h test period by pretreatment with SB242084.

Fig. 1
figure 1

The effects on locomotor activity of combining 0.5 mg/kg of SB242084 (filled symbols and bars) or its vehicle (open symbols and bars) with various doses of MDMA (n=8), amphetamine (n=8), or fenfluramine (n=10). The upper panel shows the mean±SEM photocell counts over a 2-h test period. The lower panel depicts the mean number of photocell counts in each 10-min period of the 2-h test; SEMs are omitted for clarity. For each test drug, the vehicle condition is represented by triangles, the low dose of the drug by circles, and the high dose by squares. **p<0.01 compared to the Veh—same dose condition

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For the experiment involving amphetamine at doses 0.25 and 0.5 mg/kg, analysis of variance confirmed that amphetamine stimulated locomotion (F 2,14=28.36, p<0.001). The significant main effect of SB242084 (F 1,7=6.19, p<0.05) reflects the fact that overall activity levels were greater under SB242084 treatment. While both interaction terms involving SB242084 and amphetamine were not significant (p>0.08), post hoc tests indicated that the stimulant effect of 0.5 mg/kg of amphetamine measured over the 2-h testing period was enhanced under SB242084 treatment. Analysis of data for a second experiment involving amphetamine (0.5 and 1 mg/kg) showed that SB242084 potentiated the effect of 0.5 mg/kg but not 1 mg/kg (data not shown). In the experiment involving fenfluramine the main effects of fenfluramine (F 2,18=23.25, p<0.001) and SB242084 (F 1,9=60.60, p<0.001) were both significant as was the interaction between these two factors (F 2,18=11.54, p<0.001). By itself, fenfluramine did not alter locomotor activity, but the combination of SB242084 and 5 mg/kg of fenfluramine markedly increased photocell counts compared to fenfluramine or vehicle conditions alone.

Interactions between SB242084 and cocaine, methylphenidate, or citalopram

Figure 2 shows the effects on locomotor activity of combining SB242084 with cocaine, methylphenidate, or citalopram. Cocaine significantly increased locomotor activity (F 2,14=16.94, p<0.001); and the main effect of SB242084 was also significant (F 1,7=6.24, p<0.05). The two-way interaction between these factors was not significant (F 2,14=3.00, p<0.08), although the overall three-way interaction was significant (F 22,154=3.59, p<0001). Further analysis showed that the interaction between SB242084 and cocaine was significant at the 10-, 20-, and 30-min time points (p<0.05) but not at other times. SB242084 enhanced the effects of only the 10-mg/kg dose of cocaine.

Fig. 2
figure 2

The effects on locomotor activity of combining 0.5 mg/kg of SB242084 (filled symbols and bars) or its vehicle (open symbols and bars) with various doses of cocaine (n=8), methylphenidate (n=10), or citalopram (n=10). The upper panel shows the mean±SEM photocell counts over a 2-h test period. The lower panel depicts the mean number of photocell counts in each 10-min period of the 2−h test; SEMs are omitted for clarity. For each test drug the vehicle condition is represented by triangles, the low dose of the drug by circles, and the high dose by squares. **p<0.01 compared to the Veh—same dose condition

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For the experiment involving methylphenidate, analysis of variance revealed significant main effects of methylphenidate (F 2,18=30.08, p<0.001) and SB242084 (F 1,9=10.29, p<0.01). The interaction between these two factors was also significant (F 2,18=5.11, p<0.02), although this varied as a function of time (F 22,198=3.43, p<0.001). Further analysis of this effect showed that the interactions between methylphenidate and SB242084 were significant at the 20-, 30-, 40-, and 50-min time points (smallest F 2,18=3.67, p<0.05). Post hoc tests conducted on the data for the total activity scores showed that SB242084 enhanced the locomotor activity induced by 5 mg/kg of methylphenidate; the effect for 2.5 mg/kg was not significant (p=0.06).

In the study involving citalopram, neither the main effect of SB242084 nor any of the interaction terms involving SB242084 were significant (all p>0.3). Thus, SB242084 treatment had no effect in this experiment. The main effect of citalopram was significant (F 2,16=9.26, p<0.01). Further analysis of this effect revealed that when collapsed across both the time and SB242084 factors, activity scores for both 10 and 20 mg/kg of citalopram treatment were significantly higher than for saline treatment; however, it is clear that any stimulant effect of citalopram was very minor in comparison to the other compounds.

Interactions between SB242084 and nicotine or morphine

Figure 3 shows the effects of SB242084 on nicotine-stimulated activity. Analysis of variance indicated significant main effects of nicotine (F 2,22=33.30, p<0.001) and SB242084 (F 1,11=45.13, p<0.001). In these rats, SB242084 enhanced the stimulant effects of nicotine as reflected in the significant interaction between SB242084 and nicotine (F 2,22=6.25, p<0.001) and the overall three-way interaction (F 22,242=2.32, p<0.001). Post hoc testing revealed that SB242084 potentiated the effects of both doses of nicotine.

Fig. 3
figure 3

The effects on locomotor activity of combining 0.5 mg/kg of SB242084 (filled symbols and bars) or its vehicle (open symbols and bars) with various doses of nicotine (n=12) or morphine (n=12). The upper panel shows the mean±SEM photocell counts over a 2-h test period. The lower panel depicts the mean number of photocell counts in each 10-min period of the 2-h test; SEMs are omitted for clarity. For each test drug the vehicle condition is represented by triangles, the low dose of the drug by circles, and the high dose by squares. (Note the scale difference on the Y-axis for the time course graphs). **p<0.01 compared to the Veh—same dose condition

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Figure 3 also shows the effects of SB242084 on morphine-stimulated activity. Both the main effect of morphine (F 2,22=88.3, p<0.001) and the main effect of SB242084 were significant (F 1,11=18.32, p<0.01). The SB242084 × morphine interaction was significant (F 2,22=5.52, p<0.05), indicating that SB242084 potentiated the effect of morphine. Post hoc testing on the data for the full session revealed that this effect was seen only at the 2.5-mg/kg dose of morphine.

Interactions between SB242084 and RU24969

Figure 4 shows the effects on locomotor activity of combining SB242084 with the 5-HT1A/5-HT1B receptor agonist RU24969. Analysis of variance showed a significant main effect of RU24969 only (F 1,7=0.03, p<0.001). Treatment with SB242084 did not alter the response to RU24969.

Fig. 4
figure 4

The effects on locomotor activity of combining 0.5 mg/kg of SB242084 (filled symbols and bars) or its vehicle (open symbols and bars) with various doses of RU24969 (n=8) The upper panel shows the mean±SEM photocell counts over a 2-h test period. The lower panel depicts the mean number of photocell counts in each 10-min period of the 2-h test; SEMs are omitted for clarity. For each test drug the vehicle condition is represented by triangles, the low dose of the drug by circles, and the high dose by squares

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Discussion

The selective 5-HT2C receptor antagonist SB242084 potentiated the locomotor activation induced by several drugs from distinct pharmacological classes and with diverse neuropharmacological actions, including cocaine, MDMA, amphetamine, methylphenidate, nicotine, and morphine. This list can also be expanded to include phencyclidine (Hutson et al. 2000). One possible explanation for the ability of SB242084 to potentiate the effects of these drugs is that this effect results from pharmacokinetic interactions. Cytochrome P450 enzymes are responsible for metabolism of many drugs, including those tested in the present experiments, and so interference with the activity of these enzymes could alter the behavioral response to these drugs simply through changes in plasma exposure level. Several lines of evidence suggest that pharmacokinetic interactions are unlikely to account for the effects of SB242084. Firstly, in vitro SB242084 has negligible affinity for several major cytochrome P450 enzymes responsible for metabolism of cocaine, amphetamine, nicotine, morphine, and fenfluramine (Bromidge et al. 1997). Secondly, SB242084 does not alter brain levels of MDMA (Fletcher et al. 2002b), amphetamine, or cocaine (unpublished observations) at the doses used in the present studies. Thirdly, mice lacking the 5-HT2C receptor gene also show enhanced stimulant and reinforcing effects of cocaine (Rocha et al. 2002). These converging findings make it likely that the effect of SB242084 to increase the stimulant effects of a variety of drugs reflects its neuropharmacological action to block 5-HT2C receptors.

Consistent with earlier reports, 5-HT2C receptor blockade enhanced the locomotor stimulatory effects of cocaine (Fletcher et al. 2002a; McCreary and Cunningham 1999) and MDMA (Bankson and Cunningham 2002; Fletcher et al. 2002b). Locomotor activity stimulated by both drugs is blocked or attenuated by destruction of DA terminals in the nucleus accumbens (Kelly and Iversen 1976; Gold et al. 1989), pointing to a role for elevated DA function in mediating the motor stimulant effects of cocaine and MDMA. In the case of cocaine this results from blockade of the DA transporter (Ritz and Kuhar 1989), and in the case of MDMA from increased release of DA (Gold et al. 1989; Kankaanpaa et al. 1998; Nash and Nichols 1991). Given that 5-HT2C receptor blockade increases the firing rate of DA neurons in the ventral tegmental area (VTA) (Di Giovanni et al. 1999; Di Matteo et al. 1999), it is possible that SB242084 potentiates the effects of cocaine and MDMA by enhancing the ability of these drugs to elevate extracellular levels of DA. In keeping with this explanation, SB242084 increased the motor stimulant effects of the DA reuptake inhibitor methylphenidate and the DA releaser amphetamine. At the doses used, methylphenidate and amphetamine increase extracellular DA levels with no effect on 5-HT (Kuczenski and Segal 1989, 1997, 2001). Thus, 5-HT2C receptor blockade enhances the motor stimulant effects of drugs that selectively increase extracellular levels of DA.

Recently, SB242084 was shown to potentiate the increase in extracellular levels of DA in the nucleus accumbens and the striatum elicited by 15 mg/kg of cocaine (Navailles et al. 2004); this provides direct evidence for the postulated dopaminergic mechanism by which 5-HT2C receptor blockade increases the effect of cocaine. In contrast, the mixed 5-HT2B/2C antagonist SB206553, which enhances the motor stimulant effect of 0.5 mg/kg of amphetamine (Bankson and Cunningham 2002), did not alter the DA-elevating effects of 2 mg/kg of amphetamine (Porras et al. 2002). The studies reported here show that the ability of SB242084 to enhance the stimulant effect of amphetamine depends on the dose of amphetamine; the response to 0.5 mg/kg was enhanced in two separate experiments, but the response to 1 mg/kg of amphetamine was unaffected. These data suggest that any modulatory effect of 5-HT2C receptor blockade on the behavioral effects of amphetamine may be overwhelmed in the face of a high degree of DA release.

Nicotine and morphine increase locomotion by indirectly activating mesolimbic DA neurons originating in the VTA (Benwell and Balfour 1992; Joyce and Iversen 1979; Reavill and Stolerman 1990). SB242084 increased the stimulant effects of both drugs, although the interaction with nicotine seemed more pronounced than that with morphine. These findings are in broad keeping with demonstrations that the motor stimulant effect of nicotine and the ability of morphine and nicotine to elevate DA levels in the nucleus accumbens are reduced by 5-HT2C receptor agonists (Di Matteo et al. 2004; Grottick et al. 2001; Willins and Meltzer 1998). Again, given that SB242084 modulates the activity of DA neurons in the VTA, a plausible explanation for the increased response to nicotine and morphine under 5-HT2C receptor blockade is that this involves an increased capacity of morphine and nicotine to elevate DA levels in the nucleus accumbens. This possibility is supported, at least in the case of morphine, by the finding that SB206553 enhanced the ability of morphine to increase DA release (Porras et al. 2002).

In addition to releasing DA, MDMA also releases 5-HT (e.g., Kankaanpaa et al. 1998). The more selective 5-HT releaser fenfluramine, when given alone, had no effect on locomotion, but pretreatment with SB242084 led to strong locomotor activation. These results confirm an earlier report using d-fenfluramine (Higgins et al. 2001). At the doses used, fenfluramine has little impact on DA activity (Balcioglu and Wurtman 1998; Shoaib et al. 1997), suggesting that released 5-HT, acting presumably via non-5-HT2C receptors, elicits motoric activation. One candidate receptor is the 5-HT2A receptor, as implied from findings that the selective 5-HT2A receptor antagonist M100907 blunts the locomotor response to MDMA (Fletcher et al. 2002b; Herin et al. 2005; Kehne et al. 1996) and a d-fenfluramine/SB242084 combination (Higgins et al. 2001). A second candidate 5-HT receptor subtype for mediating the enhanced stimulant effects of MDMA/SB242084 combination and fenfluramine/SB242084 combination is the 5-HT1B receptor. This is supported by the observation that mixed 5-HT1A/1B receptor agonists such as RU24969 and CGS12066B increase locomotor activity in rodents (Cheetham and Heal 1993; Green et al. 1984), and by the fact that the 5-HT1B/1D receptor antagonist GR127935 attenuates the stimulant effect of MDMA (Bankson and Cunningham 2002; Fletcher et al. 2002b; McCreary et al. 1999).

Cocaine elevates 5-HT function by inhibiting 5-HT reuptake. Like MDMA, the stimulant effects of cocaine are attenuated by 5-HT1B and 5-HT2A receptor antagonists (Castanon et al. 2000; Filip et al. 2004; Fletcher et al. 2002a; McMahon and Cunningham 2001), suggesting that activation of these receptors by endogenous 5-HT, subsequent to reuptake inhibition, is part of the mechanism by which cocaine increases locomotion. In the present experiments a selective 5-HT reuptake inhibitor citalopram (SSRI) did not alter locomotion even in the presence of 5-HT2C receptor blockade. Both fenfluramine and citalopram selectively increase synaptic 5-HT levels (Boothman et al. 2006; Bymaster et al. 2002; Millan et al. 2000), as opposed to the more generalized monoamine changes induced by cocaine and MDMA. Given the significant fenfluramine/SB242084 interaction (see also Higgins et al. 2001), the lack of an equivalent interaction between citalopram and SB242084 may relate to qualitative or quantitative differences in synaptic 5-HT changes produced by selective 5-HT reuptake inhibition, compared to the 5-HT-releasing property of fenfluramine (Fuller et al. 1988). Thus, despite its capacity to elevate extracellular 5-HT (Boothman et al. 2006; Millan et al. 2000) the magnitude of this effect induced by citalopram may have been below a threshold for induction of locomotor activation via 5-HT2A and/or 5-HT1B receptor activation, even in the presence of concomitant 5-HT2C receptor block. It is interesting to note that in the mouse citalopram elicits a mild hyperactivity that is potentiated by SB242084 (Millan et al. 2003). Combined with reports of a synergistic effect between citalopram and SB242084 on extracellular 5-HT levels in the hippocampus and immobility time in the mouse tail suspension test (Boothman et al. 2006; Cremers et al. 2004), it is clear that 5-HT2C receptor blockade can potentiate certain SSRI-mediated effects.

In addition to their effects on DA and 5-HT systems, amphetamine, methylphenidate, cocaine, and MDMA increase noradrenaline (NA) neurotransmission (Florin et al. 1994; Kuczenski and Segal 1997; Rothman et al. 2001). SB242084 elevates extracellular levels of NA (Millan et al. 1998) and so it is possible that some of the effects of SB242084 involve interactions between 5-HT and NA. Selective NA reuptake inhibitors do not stimulate activity (Brocco et al. 2002; Davids et al. 2002), although the motor-stimulant effect of amphetamine is blocked by prazosin and clonidine, and is enhanced by propranolol (Vanderschuren et al. 2003). In contrast, none of these NA receptor antagonists alter cocaine-stimulated activity (Vanderschuren et al. 2003). Thus, the role of NA in mediating the effects of psychomotor stimulants is not precisely known, although it is likely to be complex. Therefore, the most parsimonious explanation for the effect of SB242084 on drug-induced locomotor activation is that it involves dopaminergic and/or serotonergic mechanisms.

As a final part to these studies, we investigated the interaction between SB242084 and the direct 5-HT1A/1B receptor agonist RU24969. The failure of SB242084 to potentiate the RU24969-induced hyperactivity suggests a certain pharmacological specificity to the effect of SB242084 on the other drugs tested in this report. This lack of effect of SB242084 on the stimulant effect of RU24969 fails to support an interesting interaction reported by Dalton et al. (2004). These authors found that in mice the hyperlocomotion elicited by joint activation of 5-HT1A and 5-HT1B receptors through 8-OH-DPAT and CP-94,253 cotreatment is dependent on intact 5-HT2C receptor function. The different results between that study and the present one may reflect either species differences or differences among the pharmacological approaches used to influence these 5-HT receptor subtypes.

Overall, these experiments suggest that 5-HT2C receptor blockade alters drug-induced locomotion by two distinct mechanisms. The first is a modulation of DA neural activity. Evidence for this mechanism is shown by the ability of SB242084 to potentiate the effects of amphetamine and methylphenidate, two drugs that selectively and directly increase DA neurotransmission, as well as the effects of nicotine and morphine, two drugs that indirectly activate mesolimbic DA neurons. The second may involve an unmasking of a serotonergic mechanism, as evidenced by the potentiation of the selective 5-HT releaser fenfluramine. The failure to see a similar interaction with citalopram may suggest that the degree of increased synaptic 5-HT tone is a critical factor for hyperlocomotion. The interaction between MDMA and SB242084 could reflect the recruitment of both mechanisms, which may explain why the greatest magnitude of response noted across all experiments was apparent after the MDMA/SB242084 combination. On the basis of the present data, showing a lack of interaction between SB242084 and the SSRI citalopram, it seems that the ability of SB242084 to enhance the stimulant effect of cocaine, at least in the rat, is likely due to the dopaminergic mechanism, rather than unmasking of a serotonergic mechanism.

The fact that blockade of 5-HT2C receptors enhances the stimulant effects of a number of drugs of abuse indicates a possible endogenous modulation by 5-HT2C receptors of the effects of these drugs. The 5-HT2C receptor is the only known G protein-coupled receptor that undergoes RNA editing (Burns et al. 1997). This posttranscriptional modification gives rise to functionally distinct receptor isoforms (Burns et al. 1997). For some of these isoforms basal receptor activity is reduced resulting from less efficient G protein coupling (Fitzgerald et al. 1999; Herrick-Davis et al. 1999; Niswender et al. 1998, 1999). Distinct patterns of 5-HT2C RNA editing have been linked to depression, suicide (Gurevich et al. 2002; Niswender et al. 2001), and schizophrenia (Sodhi et al. 2001). Perhaps distinct patterns of 5-HT2C receptor isoforms resulting from altered RNA editing may contribute to individual differences in responsivity to drugs of abuse and vulnerability to addiction.