STEROID SEX HORMONES 



651 



tions constant and to minimize the effects 

 of the injected hormone. In addition, there 

 is a maze of interactions, some synergistic 

 and some antagonistic, between the different 

 hormones both in the endocrine gland and 

 in the target organs, so that the true effect 

 of the substance injected may be veiled. Our 

 growing understanding of the interconver- 

 sions of the steroid hormones warns us that 

 an androgen, for example, may be rapidly 

 converted into an estrogen, and the meta- 

 bolic effects observed on the administration 

 of an androgen may, at least in part, result 

 from the estrogens produced from the in- 

 jected androgen. 



To eliminate some of the confusing effects 

 of these homeostatic mechanisms some in- 

 vestigators remove the liver, kidneys, and 

 other viscera before injecting the hormone 

 under investigation. Such eviscerated prep- 

 arations have been used by Levine and his 

 colleagues in their investigations of the 

 mode of action of insulin (c/. Levine and 

 Goldstein, 1955). 



Other investigators have incubated slices 

 of liver, kidney, muscle, endocrine glands, 

 or other tissues in glass vessels in a chem- 

 ically defined medium and at constant tem- 

 perature. Such experiments have the ad- 

 vantage that metabolism can be studied 

 more directly, oxygen consumption and car- 

 bon dioxide production can be measured 

 manonietrically, and aliquots of the incuba- 

 tion medium can be withdrawn for chemical 

 and radiochemical analyses. The amounts of 

 substrate, cofactors, and hormone present 

 can be regulated and the interfering effects 

 of other hormones and of other tissues are 

 eliminated. Theoretically, working with a 

 simpler system such as this should lead to 

 greater insight into the physiologic and 

 chemical events that occur when a hormone 

 is added or deleted. The chief disadvantage 

 of this experimental system is that it is 

 difficult to prove that the conditions of the 

 experiment are "physiologic." With tissue 

 slices there is the possibility that the cut 

 edges of the cells may introduce a sizeable 

 artifact. Kipnis and Cori (1957) found that 

 the rat diaphragm, as it is usually prepared 

 for experiments in vitro, has an abnormally 

 large extracellular space and is more per- 

 meable to certain pentoses than is the intact 

 diaphragm. 



It has been postulated that a hormone 

 may influence the metabolism of a particu- 

 lar cell by altering the permeability of the 

 cell membrane or of the membrane around 

 one of the subcellular particles. Experiments 

 with tissue homogenates, in which the cell 

 membrane has been ruptured and removed, 

 provide evidence bearing on such theories. 

 If an identical hormone effect can be ob- 

 tained in a cell-free system, and if suitable 

 microscopic controls show that the sys- 

 tem is indeed cell-free, the permeability 

 theory may be ruled out. 



Ideally the hormone effect should be 

 studied in a completely defined system, 

 with a single crystalline enzyme, known 

 concentration of substrates and cofactors, 

 and with known concentration of the pure 

 hormone. Colowick, Cori and Slein (1947) 

 reported that hexokinase extracted from 

 diabetic muscle has a lower rate of activity 

 than hexokinase from normal muscle and 

 that it could be raised to the normal rate 

 by the addition of insulin in vitro. The 

 reality of this effect has been confirmed by 

 some investigators and denied by others 

 who were unable to repeat the observations. 

 Cori has suggested that the decreased rate 

 of hexokinase activity in the diabetic re- 

 sults from a labile inhibitor substance pro- 

 duced by the pituitary. Krahl and Bornstein 

 (1954) have evidence that this inhibitor is 

 a lipoprotein which is readily inactivated 

 by oxidation. 



The two hormones whose effects can be 

 demonstrated reproducibly in an in vitro 

 system at concentrations in the range which 

 obtains in the tissues are epinephrine (or 

 glucagon) and estradiol (and other estro- 

 gens) . Epinephrine or glucagon stimulates 

 the reactivation of liver phosphorylase by 

 increasing the concentration of adenosine- 

 3'-5'-monophosphate (Haynes, Sutherland 

 and Rail, 1960), and estrogens stimulate an 

 enzyme system found in endometrium, 

 placenta, ventral i)rostate of the rat, and 

 mammary gland. The estrogen-stimulable 

 enzyme was originally described as a DPN- 

 linked isocitric dehydrogenase, but the es- 

 trogen-sensitive enzyme now appears to be 

 a transhydrogenase which transfers hydro- 

 gens from TPN to DPN (Talalay and Wil- 

 liams-x\shman, 1958; Yillee and Hngerman, 

 1958). 



