Effects of low concentrations of paraoxon on Ca 2+ mobilization in a human parotid salivary cell-line HSY

Abstract

The salivary gland is a target organ of organophosphate pesticides (OPs). Inhibition of acetylcholinesterase (AChE) by OPs leads to a decrease in acetylcholine (ACh) breakdown that results in overstimulation of muscarinic cholinergic receptors (mChR). However, OPs may also directly interact with downstream elements of the phosphoinositide (PI) signalling pathway coupled with mChR. The present study examined the effects of exposure to low concentrations of the OP paraoxon on inositol 1,4,5-trisphosphate (IP 3) formation and Ca 2+ mobilization in response to ACh or ATP in the human parotid cell-line HSY. Exposure to 0.1 and 1 nM, but not 10 nM, paraoxon for 24 hr significantly elevated the basal cytosolic free Ca 2+ ([Ca 2+] i). This increase was abolished by atropine. Ca 2+ release from the IP 3-sensitive store in response to ACh or ATP, a P2Y-nucleotide agonist, was significantly increased in cells pre-exposed to 0.1 nM paraoxon. However, IP 3 formation was inhibited by paraoxon but mChR expression was not altered. Although IP 3 receptor expression was not changed, Ca 2+ release elicited by IP 3 in streptolysin O toxin-permeabilized cells was significantly larger in cells pre-exposed to 0.1 nM paraoxon, suggesting that paraoxon increases the sensitivity of IP 3 receptors. Paraoxon exposure also induced a concentration-dependent reduction in the total capacity of intracellular Ca 2+ stores, whereas the capacity of the IP 3-sensitive Ca 2+ store was not altered by paraoxon, as judged by discharging of the IP 3-sensitive Ca 2+ store with thapsigargin (TG). Ca 2+ influx stimulated by ACh or ATP was also enhanced by 0.1 nM, but not 1 and 10 nM, paraoxon. On the other hand, Ca 2+ influx activated by TG was enhanced by exposure to all concentrations of paraoxon, indicating that paraoxon modulates the Ca 2+ entry pathway. These results suggest that low concentrations of paraoxon interact with elements of the PI pathway, enhancing Ca 2+ release and influx mechanisms.