Protein synthesis in vitro by mantle tissue of the land snail Otala lactea: possible insulin-like peptide function

Mantle-collar tissue from adult land snails Otala lactea continuously incorporated labelled amino acids over a 72-h period of incubation in modified culture medium. Acid-saline extract of cerebral ganglia stimulated protein synthesis by the mantle-collar tissue in vitro. This effect was dose-dependent, with the minimum and maximum doses at 0.5 and 2 cerebral ganglion equivalents, respectively. The protein synthesis-stimulating factor(s) from the cerebral ganglia appeared to be proteinaceous and hydrophobic in nature. The cerebral ganglion extract was fractionated by means of a size-exclusion HPLC column. The biological activity was induced by three fractions with estimated molecular masses of 0.82, 1.88, and 4.33 kilodaltons (kDa). Porcine insulin antiserum abolished the activity of the 4.33- and 1.88-kDa fractions but had no significant effect on the activity of the 0.82-kDa fraction. The results suggest the existence in the cerebral ganglia of more than one factor with protein synthesis-stimulating activity. One of these factors could be related to mammalian insulin. Porcine insulin, however, had no significant effect on protein synthesis by the mantle collar in vitro.

Localization and immunological characterization of insulin-like peptide(s) in the land snail Otala lactea (Mollusca: Pulmonata)

Insulin-like peptides were detected by means of immunological techniques in tissues of the land snail Otala lactea. Insulin-positive cells were detected in all the ganglia except the right parietal ganglion and visceral ganglion. In the digestive tract, insulin-positive cells were found in the muscle and connective tissue layer of the intestine. The amount of insulin-like peptide detected in acid-ethanol extract of brains and digestive tracts from active snails did not differ significantly from that in the corresponding tissues from estivating (dormant) ones. More insulin-like peptide was detected in hemolymph from active snails than in hemolymph from estivating ones. Brains from active snails released insulin-like peptide in vitro. Analysis of the cerebral ganglia or digestive tract extracts by size-exclusion chromatography and insulin RIA revealed more than one fraction with insulin immunoreactivity. Some of these fractions contained material with molecular masses close to those of mammalian insulin or its subunits. Further analysis of the extracts by reverse-phase chromatography also revealed more than one fraction with immunoreactivity. The immunoreactive material from the digestive tract was found to be less hydrophobic than insulin. Western blot analysis of the cerebral ganglia extract revealed more than one band with insulin immunoreactivity. Three of these bands had molecular masses very similar to those of vertebrate insulin, its subunits, and its precursor.