LATERAL INTERFACIAL CHANGES CORRELATED TO MONOTERPENE MODULATION OF ACETYLCHOLINESTERASE ACTIVITY

Acetylcholinesterase (AChE) is a key enzyme in neurotransmission that hydrolyzes acetylcholine and terminates nerve impulses. In many cells, AChE is anchored to the extracellular face of the plasma membrane, where its activity may be influenced by the surrounding membrane environment. This study investigated whether monoterpenes (MTs) membrane association inhibits AChE and whether these effects are associated with changes in membrane lateral organization. 

AChE activity was characterized in bovine erythrocyte membranes (AChE_BEM) and after release from membrane  (AChE_R) in the presence of the MTs: geraniol, 1,8-cineole, menthol, and camphor. For each MT, the half-maximal effective concentration (EC₅₀) was determined, and the kinetic parameters K_M and V_max were subsequently evaluated in the presence of the corresponding EC₅₀. MT–membrane interactions were further assessed through penetration experiments in bovine erythrocyte membrane (BEM) monolayers and Brewster angle microscopy (BAM).

All four MTs inhibited AChE in a concentration-dependent manner, with geraniol and 1,8-cineole showing the greatest effects and lower apparent potency in AChE_BEM compared with AChE_R. Geraniol and 1,8-cineole also induced differential changes in kinetic parameters depending on membrane association, whereas menthol and camphor produced minor kinetic effects at the studied MTs concentration (EC₅₀). Instead of the previous results, this pattern is not reflected on the MTs π_cut-of (geraniol>menthol>>camphor>1,8-cineole), indicating distinct membrane penetration behaviors in two groups, according to their molecular structural features. In contrast, BAM revealed that geraniol and 1,8-cineole produced the most pronounced changes in lateral organization, including low-reflectivity domains occupying up to ~30% and ~4% of the analyzed area, respectively; grouping the MTs in a different pattern. Overall, the inhibitory effects of MTs were more closely associated with the extent of lateral membrane perturbation, as seen by BAM, than with their penetration capacity, as seen by π_cut-of, suggesting that membrane reorganization may contribute to the modulation of AChE activity.

The present work was partially financed by SeCyT-Universidad Nacional de Córdoba, Foncyt and CONICET from Argentina.