The Quantitative Assay of Corticotrophin Using Rats Treated With Hydrocortisone Acetate

Abstract

An attempt has been made to modify the Sayers method for the assay of corticotrophin by substituting for hypophysectomy an injection of sufficient hydrocortisone acetate to suppress the release of endogenous pituitary corticotrophin in intact rats over the period of the assay. The results of forty such assays of subcutaneous corticotrophin seem to indicate that this procedure is about 1+ to 2 times as efficient as the original method using hypophysectomized rats. For a 2 + 2 point assay it was necessary to use a minimum of twenty-four to thirty-two rats.For some years attempts have been made to modify the technique involving adrenal ascorbic acid depletion for the assay of corticotrophin (ACTH) (Sayers, Sayers and Woodbury, 1948) by substituting hypophysectomized rats with animals in which the release of endogenous corticotrophin is sufficiently blocked by the administration of corticosteroids prior to the assay.The investigations of Ingle and Kendall (1937), Ingle, Higgins andKendall (1938), Sayers and Sayers (1947), and Gray and Munson (1951) showed that several corticosteroids inhibit the release of pituitary corticotrophin as judged by changes in ascorbic acid content and weight of the adrenal glands. At the time there was some confusion concerning the relative inhibitory effects of cortisone and deoxycortone. Sayers and Sayers (1947) were the first to suggest that several crystalline cortical steroids, including cortisone, hydrocortisone and deoxycortone, were most effective in blocking release of corticotrophin from the pituitary. Casentini, De Poli, Hukovic and Martini (1957), in a comparative test of the relative potencies of various corticosteroid preparations, found that deoxycortone had about the same activity as hydrocortisone acetate in a test on unilaterally adrenalectomized rats (Abelson and Baron, 1952;Porter and Jones, 1956).On the other hand, Fortier, Yrarrazaval and Selye (1951) showed that both cortisone and cortisone acetate were completely ineffective in preventing the release of pituitary corticotrophin in response to stress, and Moya and Selye (1948), Gershberg, Fry, Brobeck and Long (1950) and Hall, Finerty, Hall and Hess (1951) were unable to confirm that deoxycortone acetate had an D inhibitory effect under similar conditions. Hodges (1953, 1954) found that rats were far more suitable for a quantitative assay after treatment with deoxycortone acetate than after cortisone acetate, but even larger doses of deoxycortone failed to give a satisfactory preparation. Recently Hodges and Vernikos (1958) found that prednisolone and hydrocortisone were far more effective in inhibiting corticotrophin release and suggested the use of these preparations instead of deoxycortone acetate (Hodges, 1955) for a modified ascorbic acid depletion technique not involving hypophysectomy.For over two years intact rats pretreated with hydrocortisone acetate have been used in this laboratory for subcutaneous and intravenous Sayers (Munson modification) assays of various corticotrophin preparations with sati...

The Lactational Anoestrus of the Sow: The Status of the Anterior Pituitary-Ovarian System During Lactation and After Weaning

“…injection and the two ovaries were removed. The ascorbic acid content of each ovary was determined using the 2,6-dichlorophenolindophenol reaction as described by Dekanski & Harvie (1960) with minor modifications only. The ascorbic acid content of each ovary was expressed as pg.…”

Biological and Immunological Properties of Human Pituitary Follicle Stimulating Hormone Obtained by Starch Gel Electrophoresis

“…Biological assays 'Total gonadotrophin' was assayed by the uterine weight method, FSH by the ovarian augmentation method (Butt et al 1961) and luteinizing hormone (LH) by the ovarian ascorbic acid depletion method (Parlow, 1958(Parlow, , 1961. In the latter the ascorbic acid in one ovary from each rat was measured by the dichlorophenolindophenol method (Dekanski & Harvie, 1960) and the gonadotrophin injected intra¬ peritoneally in a volume of 0-2 ml. saline or buffer.…”

Circadian rhythm of the ascorbic acid content in the adrenal gland of male rats

“…The adrenal glands were quickly removed from the dorsal, weighed and homogenized in metaphosphoric acid. The determination of the ascorbic acid concentration was carried out with the dichlorphenol-indophenol method, described by Dekanski and Harvie (1960). The values obtained were calculated in jug to 100 mg adrenal wet weight.…”