Abstract

Integrated in silico chemical clustering and melatonin receptor molecular modeling combined with in vitro 2-[¹²⁵I]-iodomelatonin competition binding were used to identify carbamate insecticides with affinity for hMT₁ and hMT₂ melatonin receptors. Saturation and kinetic binding studies with 2-[¹²⁵I]-iodomelatonin revealed lead carbamates (carbaryl, fenobucarb, bendiocarb, carbofuran) to be orthosteric ligands with antagonist apparent efficacy at hMT₁ and agonist apparent efficacy at hMT₂ melatonin receptors. Further using quantitative receptor autoradiography in coronal brain slices from C3H/HeN mice, carbaryl, fenobucarb, and bendiocarb competed for 2-[¹²⁵I]-iodomelatonin binding in the suprachiasmatic nucleus (SCN), thalamic paraventricular nucleus (PVT), and pars tuberalis (PT) with affinities similar to those determined for the hMT₁ receptor. Carbaryl (10 mg/kg i.p.) administered in vivo also competed ex vivo for 2-[¹²⁵I]-iodomelatonin binding to the SCN, PVT, and PT demonstrating the ability to reach brain melatonin receptors in C3H/HeN mice. Further, the same dose of carbaryl given to C3H/HeN mice for 3 consecutive days approximately 2 hours before running wheel activity onset (CT 10) in constant dark phase-advanced circadian activity rhythms (mean = 0.91 h) similar to melatonin (mean = 1.12 h) when compared to vehicle (mean = 0.04 h). Carbaryl-mediated phase shift of overt circadian activity rhythm onset is likely mediated via interactions with SCN melatonin receptors. Based on the pharmacological actions of carbaryl and other carbamate insecticides at melatonin receptors, exposure may modulate time-of-day information conveyed to the master biological clock relevant to adverse health outcomes.

Significance Statement In silico chemical clustering and molecular modeling in conjunction with in vitro bioassays identified several carbamate insecticides (i.e., carbaryl, carbofuran, fenobucarb, bendiocarb) as pharmacologically active orthosteric MT₁ and MT₂ receptor ligands. This work further demonstrated that carbaryl competes for melatonin receptor binding in the master biological clock (suprachiasmatic nucleus) and phase advances overt circadian activity rhythms in C3H/HeN mice, supporting the relevance of circadian effects when interpreting toxicological findings related to carbamate insecticide exposure.