Acetylcholinesterase activity is affected by stress conditions in Paracentrotus lividus coelomocytes

The presence and function of acetylcholinesterase (AChE) in blood cells and plasma is well known. The enzyme activity is associated to red blood cell membranes, as well as to the perinuclear region of leucocytes, suggesting a role in the function of the latter. Recently coelomocytes, acknowledged progenitors of immune cells in vertebrate systems, have been shown to respond to stress conditions, both in laboratory experiments and field studies. Bearing in mind the similarity between coelomocytes and vertebrate blood cells, we investigated the presence of AChE activity in coelomocytes of the sea urchin Paracentrotus lividus characterized by specific inhibitors, in relation to cold stress induced in the laboratory. Exposure to stress was followed by electrophoretic analysis of AChE activity and Western blot analysis of AChE-like immunoreactivity. Moreover, quantitative analysis of the different molecular forms of AChE was carried out with the aid of a spectrophotometric method. The effects of stress on AChE activity were time dependent. In fact, the enzyme activity was detected by the catalytic activity in cleaving the substrate acetyl thiocholine iodide, which appears to be formed by different isoforms of the enzyme, compared to that of Electrophorus, which was used as a control. Western blot analyses showed the presence of different immunoreactive molecules, with molecular mass ranging from 265 kDa (corresponding to Electrophorus molecules) to about 180 kDa. According to quantitative analysis, the isoforms that were differentiated corresponded to an AChE with the capacity to cleave acetyl-beta-methyl thiocholine iodide (the so-called true AChE, E.C. 3.1.1.7, corresponding to the 265 kDa band) and to a ChE with the capacity to cleave propionyl thiocholine iodide as well. The latter is an ancestral form of AChE, mainly present in invertebrates. Each of these forms was affected differently by varying time exposures