In the present study we blocked the adrenal response to MA treatment to determine if the learning deficits observed previously would be blocked or attenuated (
Herring et al., 2008). We also determined whether ADX altered the neurotoxic and hyperthermic effects of binge MA treatment. ADX did not change the magnitude or extent of water maze learning deficits (CWM performance in the absence of distal cues), the decreases in DA at 72 h, or the increases in GFAP at 72 h, even though the ADX-MA-treated animals showed no hyperthermia during treatment. It was noted too that ADX, regardless of treatment, had slightly higher 5-HT in some regions and whether this was the result of the absence of hyperthermia in the ADX groups is not known.
Hyperthermia is thought to be permissive for neurotoxicity to occur (
Ali et al., 1994;
Bowyer et al., 1992;
Bowyer et al., 1994;
Cappon et al., 1997) and the lack of protection on dopaminergic and GFAP markers of neurotoxicity at 72 h is noteworthy given that ADX-MA animals demonstrated essentially normal body temperatures while SHAM-MA animals displayed the typical pattern of hyperthermia during MA treatment. Previous studies have indicated that hyperthermia during MA exposure plays a role in the neurotoxic effects. For example, rats treated at an ambient temperature of 23°C with 4 × 5 mg/kg MA showed 60% decreased striatal DA 3 days after treatment, but only 30% decreased levels 14 days after treatment (
Bowyer et al., 1992), however rats with core body temperatures >41°C had greater DA depletions than rats with core body temperatures <41°C (
Bowyer et al., 1994). In the present experiment we intervened before body temperatures exceeded 41°C because past experience shows that this reduces mortality. Contrary to hyperthermic conditions, when MA is administered in a cold environment (4°C) and hypothermia is induced, only a transient 30% decrease in striatal DA levels are observed 3 days following MA treatment with levels similar to control animals 14 days post-treatment (
Bowyer et al., 1992). Similarly, mice administered MA (4 × 10 mg/kg) show 80% depletions of striatal DA when dosed at 23°C, but only 20% reductions when dosed at 4°C (
Ali et al., 1994). However, under certain conditions hyperthermia is not necessary to induce neuronal damage. For instance, MA-induced neurotoxicity was not prevented in mice given reserpine, a vesicular monoamine transporter (VMAT2) inhibitor and hypothermia-inducing agent (
Albers and Sonsalla, 1995;
Thomas et al., 2008). ADX has previously been shown to suppress the hyperthermic response to 1 mg/kg MA (
Makisumi et al., 1998) and in the present experiment we show that ADX suppresses hyperthermia following a higher dose of MA (10 mg/kg × 4). Unique to the present experiment is the fact that ADX-MA animals showed only a small and transient decrease in body temperature but still showed dopaminergic reductions and reactive astrogliosis 72 h later, essentially the same as those seen in the SHAM-MA animals.
Makisumi et al. (1998) suggest that glucocorticoids are permissive for the hyperthermic response and MA (4 × 10 mg/kg) induces a marked increase in corticosterone that lasts for at least 3 days (
Herring et al., 2008). Interestingly, we also noted slightly higher 5-HT levels in the ADX groups an observation that may benefit from further investigation since it is known that the serotonergic system and corticosterone are linked (
Fuller, 1992;
Maines et al., 1998;
Meijer et al., 1997). Taken together, the data suggest that hyperthermia and/or increased corticosterone levels may have a larger role in MA-induced serotonergic reductions than in dopaminergic reductions.
We have previously demonstrated that MA binge-treated animals exhibit greater than four-fold increases in CWM latency and errors compared to SAL-treated animals and show no evidence of performance catch-up after 15 days of testing (
Herring et al., 2008). In the present experiment, we extended testing to 18 days and demonstrated that MA-treated animals did not catch-up to the SAL-treated groups’ levels of performance for latencies, errors, and start returns even with three additional days of testing. Therefore, the magnitude and extent of the deficits observed in maze learning after MA treatment are unique compared to learning deficits utilizing other tasks (
cf., novel object recognition or MWM). While ADX-MA animals appeared to perform slightly better than the SHAM-MA animals, direct comparison between the groups showed no differences. Other psychostimulants, such as fenfluramine, 5-methoxy-diisopropyltryptamine (5-MEO-DIPT), and ±3,4-methylenedioxymethamphetamine (MDMA), have also been shown to disrupt learning in the CWM but to a much lesser extent than seen with MA (
Able et al., 2006;
Skelton et al., 2004;
Williams et al., 2002;
Williams et al., 2007). However, no direct comparison is possible since those earlier experiments tested animals in the CWM under lighted conditions where distal cues could be used to help navigate through the maze, whereas in the previous binge MA experiment (
Herring et al., 2008) and the present one, distal cues were eliminated by testing under infrared lighting, which we have shown makes the task much more difficult (
Skelton et al., 2004;
Williams et al., 2002;
Williams et al., 2007). In a previous examination of the relationship between corticosterone and CWM deficits, we demonstrated that the smaller CWM deficits observed with fenfluramine were prevented if the increase in corticosterone produced by the drug was blocked pharmacologically (
Skelton et al., 2004) or surgically (ADX) (
Williams et al., 2002).
We demonstrated increased GFAP in the neostriatum and decreased monoamines at 72 h after MA treatment in both the neostriatum and hippocampus (
Herring et al., 2008) as have others (
Bowyer et al., 1994;
Cappon et al., 1997;
O’Callaghan and Miller, 2002;
O’Dell and Marshall, 2002). The neostriatum has been implicated in sequence learning (
Cook and Kesner, 1988;
Potegal, 1972) and the CWM involves learning a sequence of turns to find the escape. Others have shown that binge MA exposure disrupts learning in a test of route-based motor learning (in which animals learned a specific path through corridors without choices) by showing longer latencies to complete the task or reductions in ‘directness’ to the goal (
Chapman et al., 2001;
Daberkow et al., 2005), but the exact relationship between sequence and egocentric learning is unknown. It is known that some types of egocentric learning depend on head-direction cells in the presubiculum, grid cells in the entorhinal cortex, and other regions (
Fuhs and Touretzky, 2006;
McNaughton et al., 2006;
Rondi-Reig et al., 2006;
Sargolini et al., 2006;
Whishaw et al., 1997;
Witter and Moser, 2006). Loss of 5-HT has been implicated in a variety of specific human and animal cognitive deficits (
Chudasama and Robbins, 2006;
Schmitt et al., 2006), thus the depletions in 5-HT and 5-HIAA in the entorhinal cortex may be involved in MA-induced CWM deficits. Taken together, the serotonergic system should be an area of focus in future research on MA-induced learning and memory deficits.
Acutely, the binge dosing regimen used here increased stereotypic behavior in the MA groups during the first 24 h post-treatment an effect that was gone by 30 h. The ADX-MA group showed a small but significantly higher stereotypy score over this period than did the SHAM-MA group, however given how small the difference was it does not suggest a meaningful difference as a result of the ADX surgery.
One concern with using ADX is that the learning ability of the animals may be affected, however examination of shows that ADX-SAL animals performed as well as SHAM-SAL animals, a finding we have previously observed in both the MWM and CWM (
Williams et al., 2002). This is an interesting finding in that intermediate levels of corticosterone have been shown in other studies to augment spatial learning and memory, but very low or high levels have been shown to be detrimental (
Lupien and McEwen, 1997). It is possible that the short interval between ADX and behavioral testing and/or the different cognitive tests utilized here resulted in the lack of learning deficits observed between intact and ADX animals.
In this experiment, we did not find evidence of novel object recognition deficits, although these have been reported previously after binge MA treatment (
Belcher et al., 2005;
Belcher et al., 2006;
Belcher et al., 2008;
Bisagno et al., 2002;
He et al., 2006;
Herring et al., 2008;
Schroder et al., 2003). Finding no novel object recognition effects is similar to recently reported data (
Clark et al., 2007). Another recent study showed that if MA-treated animals are compared to SAL-treated animals for time spent with the novel object, no differences were obtained, but if the amount of time spent with the familiar object was used to adjust retention performance then differences were seen (
Belcher et al., 2008). This suggests that MA-related novel object recognition differences may be sensitive to time spent with the familiar object when familiarization times are not equal.
We also found that MA had no effects on elevated zero maze performance. Previous studies examining anxiety-related behavior after MA have reported mixed results. Low dose MA appears to increase the amount of time spent in the open arms of the elevated plus maze 20 min following injection (
Szumlinski et al., 2001). Chronic (20 mg/kg, 5 days) MA treatment has been reported to increase the time spent in open arms ~2 weeks after the last treatment (
He et al., 2005); however, another study comparing chronic (2 mg/kg, 7 days) and single (4 mg/kg) MA treatment revealed a preference for closed arms 3 and 5 days following the last dose (
Hayase et al., 2005). In addition, no effects of MA on sensory gating (prepulse inhibition of the acoustic startle response) were observed in the present experiment. To the extent that sensory gating differences might interfere with learning, no evidence for such effects was obtained.
The present results indicate that the deficits in egocentric learning are not attributable to the increase in adrenal output during or after MA exposure. In addition, hyperthermia does not appear to be required for egocentric water maze learning impairments or for dopaminergic or serotonergic reductions after MA exposure. Nor did the significantly greater DA or 5-HT neostriatal or hippocampal 5-HT recovery seen in the ADX-MA vs. SHAM-MA groups make any significant difference in CWM performance. Given this, the robust deficits observed in MA-treated animals in egocentric water maze learning may provide a useful model for investigations of the mechanisms underlying the cognitive deficits reported in chronic and/or abstinent MA users (
Barr et al., 2006;
Meredith et al., 2005).
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