Mephedrone, an abused psychoactive component of 'bath salts' and methamphetamine congener, does not cause neurotoxicity to dopamine nerve endings of the striatum

Abstract

Mephedrone (4-methylmethcathinone) is a β-ketoamphetamine with close structural analogy to substituted amphetamines and cathinone derivatives. Abuse of mephedrone has increased dramatically in recent years and has become a significant public health problem in the United States and Europe. Unfortunately, very little information is available on the pharmacological and neurochemical actions of mephedrone. In light of the proven abuse potential of mephedrone and considering its similarity to methamphetamine and methcathinone, it is particularly important to know if mephedrone shares with these agents an ability to cause damage to dopamine nerve endings of the striatum. Accordingly, we treated mice with a binge-like regimen of mephedrone (4 × 20 or 40 mg/kg) and examined the striatum for evidence of neurotoxicity 2 or 7 days after treatment. While mephedrone caused hyperthermia and locomotor stimulation, it did not lower striatal levels of dopamine, tyrosine hydroxylase or the dopamine transporter under any of the treatment conditions used presently. Furthermore, mephedrone did not cause microglial activation in striatum nor did it increase glial fibrillary acidic protein levels. Taken together, these surprising results suggest that mephedrone, despite its numerous mechanistic overlaps with methamphetamine and the cathinone derivatives, does not cause neurotoxicity to dopamine nerve endings of the striatum.

Fig. 1

Effect of MEPH on DA nerve endings of the striatum 2d after treatment. Mice (n = 6 per group) were treated with MEPH in doses of 4X 20 mg/kg or 4X 40 mg/kg and the levels of DA (A), TH (B) and DAT (C) were determined at 2 days after the last injection of MEPH. Immunoblots show only 3 representative samples for each treatment group. All immunoblots were scanned and presented as means ± SEM relative to controls. The effects of either dose of MEPH on DA, TH and DAT were not significantly different from control or from each other with the exception that DAT levels after the higher dose of MEPH were significantly increased over those of the lower dose (p < 0.05, one-way ANOVA followed by Tukey’s test).

Fig. 2

Effect of MEPH on DA nerve endings of the striatum 7d after treatment. Mice (n = 6 per group) were treated with MEPH (4X 40 mg/kg) and the levels of DA (A), TH (B) and DAT (C) were determined at 7 days after the last injection of MEPH. Immunoblots show only 3 representative samples for each treatment group. All immunoblots were scanned and presented as means ± SEM relative to controls. The effects of MEPH on TH or DAT were not significantly different from control but DA levels were increased significantly (p < 0.05. one-way ANOVA followed by Tukey’s test).

Fig. 3

Effect of MEPH on striatal microglia. Mice (n = 4 per group) were treated with 4X 20 mg/kg or 4X 40 mg/kg MEPH and striatum was analyzed for microglial activation by histochemical staining of sections with ILB₄. Microglia counts were obtained as described in Materials and Methods and are presented as means ± SEM within each panel. Treatment conditions and time of sacrifice after treatments are (A) control 2d, (B) 4X 20 mg/kg MEPH 2d, (C) 4X 40 mg/kg MEPH 2d, (D) 4X 5 mg/kg METH 2d, (E) control 7d, and (F) 4X 40 MEPH 7d. None of the MEPH treatment conditions were significantly different from control but the effect of METH on microglial activation was (p < 0.0001, one-way ANOVA followed by Tukey’s test). The scale bars represent 50 µm.

Fig. 4

Effect of MEPH on striatal astrocyte expression of GFAP. Mice (n = 4 per group) were treated with 4X 20 mg/kg or 4X 40 mg/kg MEPH and striatum was analyzed for GFAP expression by immunohistochemistry. Counts of GFAP-positive astrocytes were obtained as described in the Materials and Methods and are presented as means ± SEM within each panel. Treatment conditions and time of sacrifice after treatments are (A) control 2d, (B) 4X 20 mg/kg MEPH 2d, (C) 4X 40 mg/kg MEPH 2d, (D) 4X 5 mg/kg METH 2d, (E) control 7d, and (F) 4X 40 MEPH 7d. None of the MEPH treatment conditions were significantly different from control. but the effect of METH was (p < 0.0001, one-way ANOVA followed by Tukey’s test). The scale bars represent 50 µm.

Fig. 5

Effects of MEPH on core body temperature. Mice (n = 6 per group) were treated with 4X 20 mg/kg (A) or 4X 40 mg/kg (B) and core body temperatures were recorded by telemetry at 20 min intervals for 9 h after the first injection. Results are presented as group means. SEMs are omitted for the sake of clarity and were always < 10% of the respective mean values. Injections of MEPH are indicated by arrows. Areas under the curve (AUC) for MEPH treatment groups were calculated using GraphPad Prism 5 with respect to an arbitrary baseline set to 35°C for each group and are presented in parenthesis after each treatment condition. The main effects of 4X 20 mg/kg MEPH (A) and 4X 40 mg/kg MEPH (B) and their interaction were significantly different (p < 0.0001 for each; two-way repeated measures ANOVA). Significant differences between drug and controls at individual time points (p < 0.05, Bonferroni’s test) are indicated by open red symbols () whereas those that do not differ are closed red symbols ().

Fig. 6

Effect of MEPH on locomotor activity. Mice (n= 6 per group) were treated with 4X 20 mg/kg or 4X 40 mg/mg MEPH and placed in locomotor activity monitors. Total activity counts (A), distance traveled (B), movement time (C) and stereotyped episodes (D) were recorded automatically for 60 min after each of the 4 MEPH injections. Data are presented as means ± SEM. The main effect of drug treatment was significantly different from controls for total activity, distance traveled and movement time (p < 0.0001 for all, two-way ANOVA). The main effect of drug dose was not significant for total activity, movement time and stereotyped episodes but was for distance traveled (p < 0.0001, two-way ANOVA followed by Bonferroni’s test).