Effects of the chlorotriazine herbicide, cyanazine, on GABA A receptors in cortical tissue from rat brain

Abstract

Chlorotriazine herbicides disrupt luteinizing hormone (LH) release in female rats following in vivo exposure. Although the mechanism of action is unknown, significant evidence suggests that inhibition of LH release by chlorotriazines may be mediated by effects in the central nervous system. GABA A receptors are important for neuronal regulation of gonadotropin releasing hormone and LH release. The ability of chlorotriazine herbicides to interact with GABA A receptors was examined by measuring their effects on [ 3H]muscimol, [ 3H]Ro15-4513 and [ 35S] tert-butylbicyclophosphorothionate (TBPS) binding to rat cortical membranes. Cyanazine (1–400 μM) inhibited [ 3H]Ro15-4513 binding with an IC 50 of approximately 105 μM ( n=4). Atrazine (1–400 μM) also inhibited [ 3H]Ro15-4513 binding, but was less potent than cyanazine (IC 50=305 μM). However, the chlorotriazine metabolites diaminochlorotriazine, 2-amino-4-chloro-6-ethylamino- s-triazine and 2-amino-4-chloro-6-isopropylamino- s-triazine were without significant effect on [ 3H]Ro15-4513 binding. Cyanazine and the other chlorotriazines were without effect on [ 3H]muscimol or [ 35S]TBPS binding. To examine whether cyanazine altered GABA A receptor function, GABA-stimulated 36Cl − flux into synaptoneurosomes was examined. Cyanazine (50–100 μM) alone did not significantly decrease GABA-stimulated 36Cl − flux. Diazepam (10 μM) and pentobarbital (100 μM) potentiated GABA-stimulated 36Cl − flux to 126 and 166% of control, respectively. At concentrations of 50 and 100 μM, cyanazine decreased potentiation by diazepam to 112 and 97% of control, respectively, and decreased potentiation by pentobarbital to 158 and 137% of control ( n=6). Interestingly, at lower concentrations (5 μM), cyanazine shifted the EC 50 for GABA-stimulated 36Cl − flux into synaptoneurosomes from 28.9 to 19.4 μM, respectively ( n=5). These results suggest that cyanazine modulates benzodiazepine, but not the muscimol (GABA receptor site) or TBPS (Cl − channel), binding sites on GABA A receptors. Furthermore, at low concentrations, cyanazine may slightly enhance function of GABA A receptors, but at higher concentrations, cyanazine antagonizes GABA A receptor function and in particular antagonizes the positive modulatory effects of diazepam and pentobarbital.