Imidacloprid actions in the mouse mammary gland structure and epithelial mammary cells. Role of GPER and nAChR

Abstract

The neonicotinoid imidacloprid (IMI) is used worldwide for insect control and represents a potential risk to populations, due to its potential action as an endocrine disruptor (ED). IMI binds to the postsynaptic nicotinic acetylcholine receptor (nAChR), whereas other neonicotinoids can activate the G protein-coupled estrogen receptor (GPER). Here, we examined the IMI effect on the mammary gland in a peripuberal model where female mice were exposed to IMI (0.01, 0.1 and 10 mg/kg/day). Alterations observed in the mammary morphology included an increase in transverse ductal growth at 0.1 and 10 mg/kg/day and a reduction in branch density at 0.1 mg/kg/day. Furthermore, 10 mg/kg/day IMI increased the number of terminal end buds (TEBs), induced ductal hyperplasia and epithelial cell proliferation. In mammary epithelial NMuMG cells, IMI (0.01–10 μM) showed no cytotoxic effect but boosted clonogenic capacity at 0.1 and 1 μM. Western blot findings revealed that 1 and 10 μM IMI rises GPER and aromatase expression, but declines progesterone receptor levels at all assayed doses. IMI (10 μM) also increased α7-nAChR expression at 24 h and induced c-Src phosphorylation after 1 h of treatment. Finally, 10 μM IMI promoted cell migration and the proteolytic activity of metalloproteinase (MMP)-2 and −9 through GPER, α7-nAChR, and c-Src. In summary, IMI promotes preneoplastic lesions and TEBs retention and increases mammary epithelial cell motility through GPER and α7-nAChR. Our results support the hypothesis that IMI acts as an ED, affecting mammary gland structure.