Differentiation of growth hormone- and prolactin-containing acidophils with peroxidase-labeled antibody.

In order to verify the concept that growth hormone and prolactin are contained in two different populations of acidophils, sections of Bouin-fixed rat hypophyses were stained immunochemically. For this purpose the histochemical demonstration of peroxidase was utilized after sequential application to the tissue section of rabbit antiserum to human growth hormone (or antiserum to rat prolactin) followed by application of peroxidase-labeled sheep antiserum to rabbit gamma-globulin. It was found that growth hormone cells and prolactin cells, when revealed immunochemically, corresponded structurally to cell types that could be differentiated with reasonable certainty in sections stained with the Masson trichrome procedure. When delineated immunochemically, growth hormone cells were larger and more densely arranged in the adult male than in the intact female; thcy exhibited little change in the female after ovariectomy. In contrast, prolactin cells were large and frequent in the female hypophysis but were small and less frequent in the male and in the female after ovariectomy. By double-staining, growth hormone and prolactin cells were differentiated in the same section. It was concluded that ( a ) growth hormone and prolactin are contained in, and presumably secreted by, two different populations of acidophils; and ( b ) the Masson procedure permits a reasonably accurate differentiation of the two cell types. By means of chemical staining procedures (Dawson and Friedgood, '65; Sanders and Rennels, '59; Purves, '61) and electron microscopy (Hedinger and Farquhar, '57), acidophils in the hypophyseal pars distalis of the rat may be subdivided into two types. The cells of one type tend to be arranged in rows alongside sinusoids, contain granules measuring not more than 350 mcl in diameter, and in adult rats have a small Golgi apparatus. These cells are believed to secrete growth hormone. Acidophils of the second type usually occur singly, possess cyto lasmic processes endapparatus, and contain sparsely distributed granules measuring up to 700 mu in diameter. This cell type is the probable source of prolactin. Of still less certain identity is a third form of acidophil that according to Herlant ('64) secretes corticotropin in the rat. The development of immunochemical staining methods has given hope that the functional roles of the pituitary cell types can be demonstrated conclusively. Using ing on sinusoids, o P ten have a large Golgi ANAT. REC., 164: 163-172. fluorescein-labeled antibody to human growth hormone, Leznoff et al. ('60) localized this hormone in acidophils of the human hypophysis; a similar result was obtained in the rat by Parker et al. ('65) with antibody to porcine growth hormone. Similarly, Emmart et al. implicated acidophils as being the source of prolactin in the cat ('63) and rat ( '65). After Riimke and Ladiges ('65) had found that growth hormone and prolactin are contained in different acidophilic cells in hypophyses of several species, Shiino and Rennels ('66) demonstrated that growth hormoneand prolactin-containing acidophils could be differentiated in a single section by use of antibodies conjugated with different fluorescent dyes; Nayak et al. ('68) confirmed this observation in beef. Nevertheless in spite of these provocative observations, limitations inherent in methods for immunochemical staining based on fluoresReceived Dec. 12, '68. Accepted Jan. 21, '69. 1 Supported in part by USPHS research grants HD 03159-01Al and HD 02193, and a grant from the Universitv of Michiean Horace H. Rackham School of Graduite Studies-