PLGamide characterization and role in osmoregulation in leech brain.

ABSTRACT

Neurons immunoreactive to an antiserum specifically directed against the Prolyl-Leucyl-Glycinamide peptide (PLGamide Melanocyte Inhibiting Factor MIF) were detected in the brain of the leech Theromyzon tessulatum. Radioimmunoassay titrations of the PLGamide-like material at different physiological stages of the life cycle indicated a maximal amount at stage 3B, which is correlated to phase of both maximal water uptake and coelomic vitellogenin accumulation. In vivo experiments demonstrate that this cerebral PLGamide-like material is an anti-diuretic factor that would act at stage 3B in order to permit a water uptake leading to water retention allowing coelomic yolk protein accumulation. In brains of the Gnatobdellid leech Hirudo medicinalis and the Pharyngobdellid leech Erpobdella octoculata, anti-PLGamide material was also detected with an amount not differing with the degree of sex maturation of the animals, confirming the link between osmoregulation and ovogenesis in rhynchobdellid leeches. Using a combination of biochemical techniques including high-pressure gel permeation chromatography followed by reversed-phase HPLC on brain extracts and Edman degradation, we demonstrated the presence of an authentic MIF-1 peptide in leech brain. Finally, since in vertebrates MIF-1 belongs to the non-classical opioid peptide family, we studied its binding displacement, in contrast to morphine, on mu-receptors and on nitric oxide (NO) release experiments in leech brain. PLGamide did not bind to mu-alkaloid opioid receptors and did not stimulate NO release.