Maurice H. Friedman

Effect of Progestin and Progesterone on Ovulation in the Rabbit

Although the post partum rabbit (oestrus) uniformly ovulates after coitus, the pregnant or pseudopregnant rabbit does not ovulate after coitus. This failure of post-coital ovulation in the pregnant or pseudopregnant rabbit, as well as the absence of spontaneous ovulation during pregnancy in other species, has been attributed to some inhibitory influence of the corpus luteum. It is obvious that this inhibition could be effected either by rendering the ovarian follicles refractory to the normal concentrations of the gonadotropic hormone, or by interfering with the normal supply of this hormone to the ovarian follicles. To determine the mechanism of this inhibition, post partum rabbits were injected daily with varying doses of progestin∗ or progesterone† for 5 days. At the end of the fifth day an attempt was made to mate the treated animals. Those animals which refused the male were immediately injected with one minimal ovulating dose of pregnancy urine extract‡ which had just been closely assayed by the rabbit method. 1 , 2 Laparotomy was performed 18–24 hours later to determine whether or not ovulation had occurred. Of the 24 females which had been injected with corpus luteum preparations, 9 accepted the male. In not a single instance did coitus provoke ovulation. The remaining 15 females were injected with the P. U. extract, and this was followed by ovulation in 10 animals. From these results it is clear that post-coital ovulation in the oestrous rabbit is prevented by the daily injection of corpus luteum preparations in amounts equal to, or greater than, the quantity necessary to sustain pregnancy in the castrate rabbit. 3 It is equally clear that this inhibition of ovulation is not effected by altering the sensitivity of the ovarian follicle as measured by the response to P.U.

Gonad-Stimulating Extract from Alfalfa Meal.

For reasons which can better be stated in a more complete report, it was thought highly probable that the pituitary hormone was not elaborated in the animal body from simple chemical substances, but on the contrary, was built up from complex precursors, or, taken in from the environment in a form not far removed from the hormone itself. If such hypothesis were correct, one should be able to isolate from foods these complex substances, or precursors, which when given to an animal in sufficient amounts, would prove to be gonad-stimulating. Of the foodstuffs considered, green plants seemed to be the most likely source of such material. Consequently, alfalfa was investigated from this standpoint and found to yield an extract capable of affecting the rabbit ovary in a fashion similar to that of the material found in pregnancy urine; i. e., the production of corpora lutea and corpora hemorrhagica in unmated rabbits. The extract is prepared by mixing 1 kilo of alfalfa meal with 10 liters of water and sufficient 3N alkali to bring the reaction to pH 9-10. After 4 hours the mixture is filtered by suction through a mat of moistened alfalfa meal, and the filtrate then subjected to the Katzman-Doisy benzoic acid method for the extraction of the active material from pregnancy urine. 1 The extracts prepared so far are highly toxic so that they can not be injected intravenously, and therefore cannot be satisfactorily assayed by the rabbit method. The material must be injected either intraperitoneally or subcutaneously, and in order to obtain a positive response it has been necessary to inject the equivalent of 300 to 500 gm. of dry alfalfa in 2 doses not more than 12 hours apart. From previous experience with injections of pregnancy urine by the intraperitoneal route, it is estimated that the yield from 1 kilo of dry material is probably more than 30 rat units and probably less than 100 rat units. The negative results obtained with the injections of inactive extracts, prepared in an identical manner, except for 24 hours (or more) initial exposure to the dilute alkali instead of 4 hours, together with the intraperitoneal injections of a variety of materials, including male urine and sterol suspensions, serve adequately as controls.

The development of the rabbit test for pregnancy

The development of the Friedman rabbit test for pregnancy at the end of the 1920s is presented. Based on the reports of humoral substances in the urine of pregnant women by Aschheim and Zondek rabbits were injected with pregnant urine over a short period and their ovaries inspected 48 hours after the initial injection. This test was well received because of the regular response of the rabbit to this stimulation however it is only a modification of the Aschheim-Zondek test.