Alteration of catecholamines in pheochromocytoma (PC12) cells in vitro by the metabolites of chlorotriazine herbicide

The effects of four major chlorotriazine metabolites on the constitutive sy nthesis of the catecholamines dopamine (DA) and norepinephrine (NE) were ex amined, using undifferentiated PC12 cells. NE release and intracellular DA and NE concentrations were quantified, for up to 24 h after initiation of t reatment with different concentrations, ranging from 0 to 400 muM, of the m etabolites hydroxyatrazine (HA), 2-amino-4-chloro-6-isopropylamino-s-triazi ne (deethylchlorotriazine), 2-amino-4-chloro-6-ethylamino-striazine (deisop ropylchlorotriazine), and diaminochlorotriazine (DACT), Hydroxyatrazine sig nificantly decreased intracellular DA and NE concentrations in a dose- and time-dependent manner. This metabolite also caused a significant inhibition of NE release from the cells. In contrast, deethylchlorotriazine and deiso propylchlorotriazine significantly increased intracellular DA concentration following exposure to 50-200 muM from 12 to 24 h. Intracellular NE was sig nificantly reduced at these same concentrations of deethylchlorotriazine at 24 h while the concentration of NE in PC12 cells exposed to deisopropylchl orotriazine was not altered at any dosage or time point measured, NE releas e was decreased at 18 (200 muM) and 24 h (100 and 200 muM) following exposu re to deethylchlorotriazine and at 24 h (100 and 200 muM) following deisopr opylchlorotriazine. DACT, at the highest concentration (160 muM), significa ntly increased intracellular DA and NE concentrations at 18 and 24 h. NE re lease was also increased at 40-160 muM at 24 h. The viability of the PC12 c ells was tested using the trypan blue exclusion method. Following 18 to 24 h of treatments with HAI cell viability was reduced 10-12% at the two highe r concentrations (200 and 400 muM), but, with other metabolites, the viabil ity was reduced by only 2 to 5% at the highest concentrations. These data i ndicate that HA affects catecholamine synthesis and release in PC12 cells i n a manner that is similar to synthesis of atrazine and simazine. On the ot her hand, deethylchlorotriazine and deisopropylchlorotriazine altered catec holamine synthesis in a manner similar to that observed in the rat brain fo llowing in vivo exposure (i.e., increased DA and decreased NE concentration ), whereas DACT appeared to produce an increase in NE release as well as in the intracellular DA and NE concentrations. Overall, these findings sugges t that the catecholamine neurons may be a target for the chlorotriazines an d/or their metabolites, that the metabolites produce a unique pattern of ca techolamine response, and that all of the changes mere seen within the same range of doses.