RNA synthesis and turnover in the molluscan nervous system studied by Click-iT method

●5-ethynyluridine labeled neurons in snail CNS after 4 h of incubation in EU solution.●Maximum staining was observed after 12 h of incubation, and varied between neurons.●In juveniles, 1 h incubation labeled apical part of the procerebrum.●EU readily penetrates into the bodies of juvenile snails.

RNA synthesis can be detected by means of the in vivo incorporation of 5-ethynyluridine (EU) in newly-synthesized RNA with the relatively simple Click-iT method. We used this method to study the RNA synthesis in the CNS tissue of adult and juvenile terrestrial snails Helix lucorum L. Temporally, first labeled neurons were detected in the adult CNS after 4-h of isolated CNS incubation in EU solution, while 12-h of incubation led to extensive labeling of most CNS neurons. The EU labeling was present as the nuclear and nucleolar staining. The cytoplasm staining was observed after 2–3 days of CNS washout following the EU exposure for 16 h. In juvenile CNS, the first staining reaction was apparent as the staining of apical region in the procerebral lobe of cerebral ganglia after 1 h of CNS incubation in EU, while the maximum pattern of staining was obtained after 4 h of CNS incubation. Thus, age-related differences in RNA synthesis are present. Activation of neurons elicited by serotonin and caffeine applications noticeably increased the intensity of staining. EU readily penetrates into the bodies of juvenile snails immersed in the EU solution. When the intact juvenile animals were immersed in the EU solution for 1 h, the procerebrum staining, similar to the one detected in the incubated juvenile CNS, was observed.