Microglial activation precedes dopamine terminal pathology in methamphetamine-induced neurotoxicity

Abstract

Previous studies have demonstrated methamphetamine (METH)-induced toxicity to dopaminergic and serotonergic axons in rat striatum. Although several studies have identified the nature of reactive astrogliosis in this lesion model, the response of microglia has not been examined in detail. In this investigation, we characterized the temporal relationship of reactive microgliosis to neuropathological alterations of dopaminergic axons in striatum following exposure to methamphetamine. Adult male Sprague–Dawley rats were administered a neurotoxic regimen of methamphetamine and survived 12 h, or 1, 2, 4, and 6 days after treatment. Immunohistochemical methods were used to evaluate reactive changes in microglia throughout the brain of methamphetamine-treated rats, with a particular focus upon striatum. Pronounced morphological changes, indicative of reactive microgliosis, were evident in the brains of all methamphetamine-treated animals and were absent in saline-treated control animals. These included hyperplastic changes in cell morphology that substantially increased the size and staining intensity of reactive microglia. Quantitative analysis of reactive microglial changes in striatum demonstrated that these changes were most robust within the ventrolateral region and were maximal 2 days after methamphetamine administration. Analysis of tissue also revealed that microglial activation preceded the appearance of pathological changes in striatal dopamine fibers. Reactive microgliosis was also observed in extra-striatal regions (somatosensory and piriform cortices, and periaqueductal gray). These data demonstrate a consistent, robust, and selective activation of microglia in response to methamphetamine administration that, at least in striatum, precedes the appearance of morphological indicators of axon pathology. These observations raise the possibility that activated microglia may contribute to methamphetamine-induced neurotoxicity.

Introduction

Repeated administration of methamphetamine (METH) to rodents has been shown to induce the degeneration of dopamine and serotonin terminals of the striatum, serotonin terminals of the hippocampus Hotchkiss and Gibb, 1980, Ricaurte et al., 1982, Seiden et al., 1976, Wagner et al., 1980 as well as cortical and striatal neurons Deng et al., 2001, Eisch et al., 1998, Ryan et al., 1990. Although the mechanism of METH-induced toxicity is unknown, several factors appear to be involved (Cadet et al., 2003). For example, it is known that the neurotransmitter dopamine plays a significant role in METH-induced toxicity Gibb and Kogan, 1979, LaVoie and Hastings, 1999, Schmidt et al., 1985. There is also considerable evidence of a role for oxidative stress De Vito and Wagner, 1989, Gibb and Kogan, 1979, Giovanni et al., 1995, Hirata et al., 1995, LaVoie and Hastings, 1999, Yamamoto and Zhu, 1998, although METH-induced increases in extracellular levels of glutamate suggest that excitotoxicity may also contribute to the resulting pathology Abekawa et al., 1994, Nash and Yamamoto, 1992. Finally, it has been established that the toxicity of many substituted amphetamine analogs is dependent upon an acute drug-induced hyperthermia Albers and Sonsalla, 1995, Ali et al., 1994, Colado et al., 1995, Malberg and Seiden, 1998, Schmidt et al., 1990. How these many factors interact to produce the selective degeneration of monoaminergic terminals of the rodent striatum is unknown.

Many of the hypotheses regarding the mechanism of METH-induced toxicity have focused on intra-neuronal events such as dopamine oxidation, oxidative stress, and excitotoxicity. However, available evidence suggests that METH-induced neuropathology may result from a multicellular response in which glial cells may play a prominent role. Several previous studies have demonstrated the appearance of astrocyte activation in METH-induced toxicity Cappon et al., 1997, Fukumura et al., 1998, O'Callaghan and Miller, 1994, Pu and Vorhees, 1993. For example, Pu and Vorhees (1993) demonstrated that METH-induced toxicity was associated with dramatic increases in the astrocyte marker glial fibrillar acidic protein (GFAP) that was most pronounced in the ventrolateral striatum. This striatal subregion has been shown to be most vulnerable to the toxic effects of METH and exhibits the greatest loss of tyrosine-hydroxylase immunoreactivity (Pu and Vorhees, 1993) and dopamine-transporter binding sites (Eisch et al., 1992) following neurotoxic administration of METH. A temporal analysis of astrogliosis in METH-treated mice demonstrated that the astroglial response peaked 2 days after administration and remained elevated for at least 7 days following METH treatment (O'Callaghan and Miller, 1994). In addition, the activation of astrocytes in response to METH was correlated with toxicity (Pu and Vorhees, 1993). Therefore, the astrocyte response occurs rapidly following METH treatment and is relatively prolonged. Nevertheless, the extent to which this response contributes to pathological changes in the striatum is unknown.

Although several studies have investigated changes in GFAP immunoreactivity or protein levels in METH-induced toxicity, the microglial response to METH administration has not been reported. Microglia are classically recognized for their phagocytic capabilities and are considered the resident immune cells of the brain (Streit et al., 1999). However, they also exhibit dynamic response properties and phenotypic changes that can exert an important influence upon the outcome of CNS disease or injury (Aschner et al., 1999). This raises the possibility that METH-induced microglial responses may contribute to the neuropathological changes that result from neurotoxic levels of this amphetamine. As a first step in testing this hypothesis, we defined the spatial and temporal relations of reactive microglia responses to METH-induced pathology.