Increased Vesicular Monoamine Transporter 2 (VMAT2; Slc18a2) Protects against Methamphetamine Toxicity

Abstract

The psychostimulant methamphetamine (METH) is highly addictive and neurotoxic to dopamine terminals. METH toxicity has been suggested to be due to the release and accumulation of dopamine in the cytosol of these terminals. The vesicular monoamine transporter 2 (VMAT2; SLC18A2) is a critical mediator of dopamine handling. Mice overexpressing VMAT2 (VMAT2-HI) have an increased vesicular capacity to store dopamine, thus augmenting striatal dopamine levels and dopamine release in the striatum. Based on the altered compartmentalization of intracellular dopamine in the VMAT2-HI mice, we assessed whether enhanced vesicular function was capable of reducing METH-induced damage to the striatal dopamine system. While wildtype mice show significant losses in striatal levels of the dopamine transporter (65% loss) and tyrosine hydroxylase (46% loss) following a 4 × 10 mg/kg METH dosing regimen, VMAT2-HI mice were protected from this damage. VMAT2-HI mice were also spared from the inflammatory response that follows METH treatment, showing an increase in astroglial markers that was approximately one-third of that of wildtype animals (117% vs 36% increase in GFAP, wildtype vs VMAT2-HI). Further analysis also showed that elevated VMAT2 level does not alter the ability of METH to increase core body temperature, a mechanism integral to the toxicity of the drug. Finally, the VMAT2-HI mice showed no difference from wildtype littermates on both METH-induced conditioned place preference and in METH-induced locomotor activity (1 mg/kg METH). These results demonstrate that elevated VMAT2 protects against METH toxicity without enhancing the rewarding effects of the drug. Since the VMAT2-HI mice are protected from METH despite higher basal dopamine levels, this study suggests that METH toxicity depends more on the proper compartmentalization of synaptic dopamine than on the absolute amount of dopamine in the brain.

Keywords: Methamphetamine; VMAT2; dopamine; inflammation; neurodegeneration; vesicle.

Figure 1

Increased VMAT2 protects against TH and DAT loss in the striatum. Male wildtype and VMAT2-HI mice were treated with four injections of either saline or 10 mg/kg methamphetamine every 2 h and sacrificed 48 h after the final injection. (A–D) Wildtype animals show significant losses in DAT and TH levels. VMAT2-HI mice are significantly protected from losses in both DAT (p < 0.01) and TH (p < 0.05) following METH. (E,F) Wildtype mice also show a trend toward VMAT2 protein loss following METH, while VMAT2-HI mice maintain a VMAT2 level greater than wildtype even following METH. Data are presented as percent of saline-treated wildtype mice. Different letters at the tops of bars indicate differences of at least p < 0.05 as determined by a two-way ANOVA with Bonferroni posthoc tests (n = 6).

Figure 2

Increased VMAT2 protects against TH+ fiber denervation in the striatum. VMAT2-HI mice are protected from the loss of TH+ fibers in the striatum. Representative images of dorsolateral striatum pictured with cortex on the right side of each image. Scale bar = 200 μm.

Figure 3

METH treatment did not induce SNpc cell body loss. (A,B) There was no difference in TH+ cells (p > 0.05) or Nissl+ cells (p > 0.05) between the genotypes following a 4 × 10 mg/kg METH dose (n = 6). (C) Representative images of TH staining of the midbrain with and without METH treatment.

Figure 4

Increased VMAT2 protects against gliosis in the striatum. (A,B) VMAT2-HI mice show a significantly smaller increase in astrogliosis as indicated by GFAP expression (n = 6). Different letters above the bars indicate difference of p < 0.05. Data are presented as percent of saline-treated wildtype mice. (C,D) VMAT2-HI mice show less ramified microglia as shown by IB4 staining. Representative images of dorsolateral striatum pictured with corpus callosum in the upper right corner of each image. Scale bar = 200 μm.

Figure 5

Preferential targeting of the striosomes following METH treatment. (A,B) Selective loss of striosomal DAT was shown in both wildtype and VMAT2-HI mice. However, this loss was not seen until a higher 4 × 10 mg/kg METH dose in the VMAT2-HI animals (A). (C) Representative tracing of DAT loss in striosomes of wildtype METH-treated striatum following a 4 × 5 mg/kg METH dose.

Figure 6

Increased VMAT2 level does not alter the hyperthermic response following METH treatment. Core temperatures taken 1 h post each METH injection (injections indicated by arrows). While there was a significant increase in core temperature in both genotypes, there was no difference between the hyperthermic profiles between wildtype and VMAT2-HI mice (p > 0.05) (n = 12).

Figure 7

Increased VMAT2 does not change METH-induced conditioned place preference or METH-stimulated locomotor activity. Both genotypes show a preference at 1 mg/kg METH (n = 9). However, there was no difference between genotypes on time spent in the METH-paired side of the chamber on test day. Similarly, wildtype and VMAT2-HI mice show no difference in locomotor activity when the genotypes were treated with METH. Different letters at the tops of the bars indicate difference of p < 0.01.