S9 LIVER FRACTION IS CYTOTOXIC TO NEURONS IN DISSOCIATED CULTURE

Methods to evaluate the neurotoxicity of chemicals in vitro must take into account that many xenobiotics exert their toxic effects through m etabolites. S9 fraction of liver homogenate has been used in cultures of bacterial and mammalian cells as a system for metabolic activation. The suitability of the S9 activation system for long-term neurotoxici ty studies in vitro was investigated in dissociated cultures of murine spinal cord-dorsal root ganglia. Exposure to amounts of S9 greater th an 0.07 mg S9 protein/ml of culture medium for 4 days or longer was cy totoxic to all types of neurons in the cultures. Nonneuronal cells tol erated higher exposures, but contained numerous cytoplasmic inclusions when 0.35 mg S9 protein was included in the medium. It was demonstrat ed that cytotoxicity was caused by the particulate, microsomal fractio n of S9. It is concluded that direct incorporation of S9 fraction in t he growth medium (0.07 mg S9 protein/ml or greater) is not a suitable method of generating metabolites in dissociated cultures of central ne rvous system when several days are required to elicit a biological res ponse. Cytotoxicity can be prevented by using tissue culture inserts t o separate cultured cells from S9 particulate fraction by a microporou s membrane. (C) 1993 Intox Press, Inc.