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Compounds known as chemical mediators, including acetylcholine, have been found not only in humans and animals, but also in living organisms, like protozoa, which lack nervous system. In Paramecium primaurelia has been described a cholinergic system, which is proven to play an important role in cell-cell interactions during its developmental cycle. In our work we investigated the effects of exposure to simulated microgravity (3D Random Positioning Machine, 56 rpm, 10-6 g) on the cholinesterase activity of the eukaryote unicellular-organism alternative-model P. primaurelia. Our results show that the exposure of P. primaurelia to microgravity for 6 h, 24 h, 48 h affects the localization and the amount of cholinesterase activity compared to cells grown under Earth gravity conditions (1 g). However, these effects are transient since P. primaurelia restores its normal cholinesterase activity after 72 h under microgravity conditions, as well as cells exposed up to 72 h to microgravity and then placed under terrestrial gravity for 48 h.