Mechanism of Action of Antipsychotics, Haloperidol and Olanzapine in vitro

Schizophrenia affects 1-1.5% of people in the United States alone. Haloperidol (HP), a butyrophenone and a typical antipsychotic, has been used as an antipsychotic drug in human. Unfortunately, the therapeutic effects of HP also come with severe extrapyramidal side effects, resulting in movement disorders in patients. Olanzapine, a new atypical neuroleptic, seems to have better efficacy, with less severe adverse effects. There has been increasing evidence of the role of reactive oxygen species (ROS) and oxidative stress in the pathogenesis of Schizophrenia. We therefore hypothesized that the differences between HP and Olz could be partly because of the differences in the oxidative stress they cause. We studied the pro-oxidant and antioxidant effects of these two drugs in vitro and examined the mechanism of their cytotoxicity in a neuronal cell model using PC-12 cells. HP was found to be ineffective as a superoxide radical scavenger but appeared to be a potent scavenger of hydroxyl radicals with a rate constant of ~6.78 X 109 M-1s-1. Olz on the other hand was found to scavenge hydroxyl radical at a rate of 34.1 X 109 M-1s-1. This was shown using the hydroxyl radical dependent deoxyribose degradation assay and EPR spin trapping methods. HP was also found to quench singlet oxygen in a dose-dependent manner. HP was found to enhance the microsomal lipid peroxidation in a dose-dependent manner and at 10 µM it augmented the lipid peroxide accumulation by 100% whereas Olz, at the same concentrations had trivial effects. Light microscopy and two cytometric apoptotic/viability probes (7-aminoactinomycin D and Annexin-V) were employed to evaluate mechanisms of drug-induced cell death in PC-12 pheochromocytoma cells exposed to HP or Olz. At low dose (50 µM), HP was more cytotoxic than Olz. At high concentrations (150 mM) each of these antipsychotic drugs caused a significant increase in cell death that was readily detectable by all the techniques. Light microscopy with trypan blue staining indicated that necrosis was the predominate form of cell death with both drugs. Apoptotic cells were rarely observed by microscopy in vehicle or drug-exposed cells. Further, no increase in early cellular apoptosis was observed using the Annexin-V probe. 7AAD and Annexin-V both showed drug-related increases in the late apoptotic/necrotic cell death window. These data, along with the cytologic evaluations suggest that cell death in PC-12 pheochromocytoma cells exposed to HP or Olz may primarily be necrotic in nature, rather than apoptotic. Because Olz at a low dose was less cytotoxic and was found to have lower pro-oxidant action than HP the secondary effects manifested in patients with chronic treatment with HP may, at least in part, be attributed to the pro-oxidant effects of the drug.