FALLAS DEL SISTEMA

___________________ Chapter Three__________Neurochemical basis o f the 3-NPM model 3.1 Introduction

Methamphetamine is a common drug o f abuse known to release DA and glutamate from the respective neurones (Sonsalla et al., 1989). The release o f DA from the cytosolic pool

(Wagner et al., 1983) or the vesicular stores (Anderson et a l, 1998; Sabol and Seiden, 1998) occurs via the reversal of the DA transporter (Albers et a l, 1996; Fleckenstein et a l, 1997; Wagner et al, 1983). In energy-deficiency states, this ATP-dependent mechanism has been shown to be sufficient to trigger mechanisms that result in neuronal death via metabolic stress (Albers et al 1996; Chan, et al, 1994), and to render the neurones susceptible to the

excitotoxic insults of endogenous glutamate (Albers et al, 1996; Nash et a l, 1992; Sonsalla et al, 1998). Simultaneously, the release of DA in such large amounts accelerates the build-up of free radicals associated with DA auto-oxidation (O’Dell et a l, 1993). This overwhelms the neutralizing mechanisms, producing a state o f oxidative stress (Jenner, 1992; Zeevalk et al,

1998). Furthermore, the DA build-up has been shown to facilitate the condensation reaction of DA with acetaldehyde to yield the potent, endogenous neurotoxin, salsolinol (Maruyama et al, 1998). The ultimate effect is the death of the terminals (Albers et a l, 1996) and/or cell bodies (Sonsalla et a l, 1996, 1997) of DA, 5-HT and other neurones. The susceptibility of the neurones to energy impairment or METH toxicity varies from one region o f the CNS to another, and from one transmitter system to the next, with DA neurones showing greater vulnerability than GAB A and 5-HT neurones to lesioning (Araujo and Hilt, 1998; Hotchkiss et al, 1979; Zeevalk et a l, 1998). Furthermore, the direct stratonigral output pathway is more susceptible than the indirect striatopallidal output pathway to 3-NP toxicity (Araujo and Hilt,

1998).

The ability of METH to incorporate both these fundamental theories - Metabolic and

Oxidative Stress - sets the stage for its potentiation by a variety o f neuroactive substances. PD is known to occur more commonly in the later stages o f life, stages that are synonymous with impairment in energy metabolism or energy deficiency. An allusion to a causal role for energy impairment in the pathogenesis of PD can be seen in the deficiency of Complexes I, II and III of the mitochondrial energy cycle in patients with the early, untreated form of the disorder (Haas et al, 1995; Jenner, 1992). Examination of the substantia nigra o f these patients reveals decreased levels of NADH cytochrome c reductase, a marker of Complexes I, II and III

________________________ Chapter Three_____Neurochemical basis o f the 3-NPM model

(Schapira et al, 1990). The leading primate model of the disease, the MPTP model, is based on the ability of its active ion (MPP^) to also inhibit the said Complex I (Adams and Odunze,

1991).

3-NP is an irreversible inhibitor of succinate dehydrogenase (Complex II) o f the mitochondrial electron transport chain (Zeevalk et al., 1995). Our preference for 3-NP over malonate

(another Complex II inhibitor) stemmed from the irreversible inhibition o f Complex II with the former (Ludolph et al., 1991), its being ten times more potent than malonate (Zeevalk et al.,

1995), its efficacy via parenteral administration, and its natural occurrence, being present as a contaminant in sugar-cane (Gould and Gustine, 1987). Furthermore, 3-NP is not known to cause release of DA or glutamate (Zeevalk and Nicklas, 1991; Zeevalk et al., 1995).

In establishing a dose for 3-NP, care was taken to achieve a balance between maximal inhibition or neuronal damage (as was demonstrated by the stage 3 phenomenon (Ludolph et al., 1991)) and minimum paralysis or death. Although the higher doses tended to produce the said phenomenon rapidly, they were more likely to cause paralysis, or even death. The lower dose o f 20 mg/kg produced the phenomenon by the second or third day of injection, and yielded no hind-limb paralysis or fatality until combined with METH.

The rationale behind the different dosing schedules in the combination of both agents stemmed from the keenness to produce a simple and effective means of lesioning the nigrostriatal pathway in the rat, as determined by selective reductions in nigrostriatal DA synthesis and content, and from corresponding alterations in the spontaneous and drug-induced motor behaviour of the animals.

3.2 Results

3.2.1 Preliminary investigations

The preliminary experiments were geared towards the determination of optimum doses and treatment schedules for combining METH and 3-NP, so as to yield maximum nigrostriatal DA depletion and behavioural signs of increased sensitivity to DA receptor agonists, while limiting the toxic effects of both agents. The findings formed the basis for more elaborate investigation in the “3-NPM” treatment schedule.

____________________________ Chapter Three______N eurochem ical basis o f the 3-N PM m odel 3.2.1.1 Effects of METH on tissue DA levels

The brains o f rats treated with four, two-hourly doses o f M ETH (4, 8 and 12.5 m g/kg i.p.) were assayed for D A seven days after treatment. N one o f the doses significantly (p >0.05) altered tissue dopamine levels in the N A cc, PFC and SN (Fig. 3.1). H owever, treatment with 4x8 and 4x12.5 mg/kg M ETH gave rise to significant decreases in striatal D A (p<0.01). A dose o f 4x8 mg/kg, q2 h METH w as selected for future combination with 3-NP, as this gave a significant depletion o f D A in the ST (43 % o f control levels). Although a dose o f 4x12.5 m g/kg produced a greater percentage o f depletion, its high fatality rate o f 80 % (compared to the 33% fatality for the 4x8 mg/kg dose (Fig. 3.27)) made it a second-choice dose. The 50 % striatal D A depletion observed with 4 x 12.5 mg/kg M ETH differs from the 82 % depletion reported for mice (Johnson et al., 1992).

Tissue dopamine levels following treatment