NEUROPEPTIDES AND RELATED NUCLEIC-ACID SEQUENCES DETECTED IN PENEID SHRIMPS BY IMMUNOHISTOCHEMISTRY AND MOLECULAR HYBRIDIZATIONS

The neurosecretory cells in the eyestalks of Penaeus indicus and P. vannamei were studied by immunocytochemistry using polyclonal antisera raised against purified Homarus americanus neuropeptides. Cross-reactions between two H. americanus anti-crustacean hyperglycemic hormone antisera and Penaeus neurosecretory material were observed. The specific anti-vitellogenesis inhibiting hormone antiserum only showed an immunological reaction in the nervous tract and the sinus gland of Penaeus, suggesting a progressive exposure of a characteristic epitope which was amenable to immunological detection. Molecular hybridizations were performed in P. indicus and P. vannamei with a digoxigenin tailed 23mer oligonucleotide probe deduced from two partial sequences of uncharacterized, purified P. duorarum neuropeptides. Two distinct clusters of positive cells were observed by in situ hybridization experiments in the medulla terminalis ganglionic X-organ. Classical control tests gave negative results. Northern and Southern blot analyses were performed with the same tailed probe and allowed the determination of molecular weights for the mRNA and for a DNA restriction fragment (Pst 1), 1.7 kb and 200 bp, respectively. These observations show the existence of a strong homology between the P. duorarum sequence (selected for synthesizing the probe), and some P. indicus and P. vannamei neuropeptide sequence(s). Heterologous antisera were tested in other Arthropods to complete our analyses. In the centipede Lithobius forficatus and the scorpion Euscorpius carpathicus, the anti-crustacean hyperglycemic hormone antiserum induced a strong cross-reaction. A monoclonal anti-bombyxin-1 antiserum showed an immunoreaction in the neurosecretory system of the insects Tenebrio molitor and Labidura riparia. In contrast, the anti-bombyxin-1 antiserum did not react either in Penaeus or in Lithobius, and the Homarus hyperglycemic hormone antiserum did not react in the insects that were tested. A comprehensive view of these observations is discussed in relation to a divergence in Arthropod evolution.