Distribution and characterization of immunoreactive growth hormone (GH) in the pituitary of the frog Rana ridibunda using an antiserum against purified bullfrog GH

Abstract

The presence of growth hormone (GH) in the pituitary of the frog Rana ridibunda was investigated using an antiserum raised against purified bullfrog GH. The immunofluorescence technique revealed that GH-containing cells are exclusively located in the dorsal area of the distal lobe of the pituitary. The relative abundance of these GH-positive cells, which correspond to acidophilic type 2 cells, was 1ζ ± 1% of the total population of endocrine cells of the pars distalis. Frontal sections of the distal lobe indicated that GH-producing cells are distributed in an arc of a circle occupying all of the dorsal part of the lobe. At the electron microscopic level, GH-immunoreactive material was sequestered in large polymorphic granules (200–700 nm). GH was quantified in R. ridibunda pituitary extracts using a radioimmunoassay for bullfrog GH. The displacement curves obtained with serial dilutions of pars distalis extracts were not strictly parallel to the standard curve made with purified bullfrog GH. In contrast, Western blot analysis revealed that GH from R. ridibunda had a molecular weight (22 kDa) similar to that of bullfrog GH. In the pars distalis, the apparent amount of GH was 0.61 ± 0.14 μg per lobe, corresponding to 0.92 ± 0.17% of total proteins in the extracts. In contrast, frog neurointermediate lobe or hypothalamus did not contain significant concentrations of immunoreactive GH (less than 0.006% of total proteins in the extracts). Taken together, these results validate the use of an antiserum to bullfrog GH to investigate the regulation of GH secretion in R. ridibunda.