Kenneth T. Kirton

Measurement of prostaglandin F2α metabolites by radioimmunoassay

Abstract Antibodies against 15 keto PGF2α and 13,14 dihydro 15 keto PGF2α were produced in goats and rabbits using the appropriate prostaglandin protein conjugate. Tritium labeled 15-keto, and 13,14 dihydro 15-keto PGF2α were prepared from 3H-PGF2α. These antibodies and 3H-labeled compounds were used to develop radioimmunoassays for the respective F2α metabolites. The antibodies had relatively little cross-reactivity (≤0.1%) with the parent F2α molecule. Infusion of PGF2α in monkeys increased 15-keto-h2 levels 10–20 fold higher than PGF2α in peripheral plasma. The levels of this metabolite were not altered detectably during clotting, indicating relatively slow rates of PGF2α metabolism in vitro . These assays should be useful to follow release rates of exogenous prostaglandins from various formulations and delivery systems, and in vivo tissue synthesis of PGF2α, where low levels preclude measuring the parent compound.

Return of ovulatory cyclicity following an intramuscular injection of medroxyprogesterone acetate (Provera)

Abstract This study was carried out to determine the relationship between peripheral serum concentrations of medroxyprogesterone acetate (Provera) and progesterone following an intramuscular injection of 150 mg of depomedroxyprogesterone acetate. Medroxyprogesterone acetate concentrations were maximal at 5–20 days post-injection (10–25 ng/ml), then decreased to 5–10 ng/ml by 30 days post-injection. In general, the concentrations decreased gradually, with little fluctuation, during the remainder of the study (260 days). Medroxyprogesterone acetate concentrations were 5.1, 0.8 and

Biological activities of 17-phenyl-18,19,20-trinorprostaglandins.

In a number of assay systems, some 17-phenyl-trinor-prostaglandins were similar in activity and potency to the corresponding parent prostaglandin. In others, the 17-phenyl analogs appeared several times more potent. In the hamster antifertility assay, which is considered to measure luteolytic activity, 17-phenyl-18,19,20-trinor prostaglandin F2alpha was about 90-times PGF2alpha in potency. Rat blood pressure responses to 17-phenyl analogs were significant. The 17-phenyl-trinor PGF2alpha pressor potency was 5 times that of PGF2alpha. The 17-phenyl-trinor PGE2 blood pressure response was atypical since a pressor rebound phenomenon followed the expected depressor response. Lastly, 17-phenyl-trinor PGF2alpha was more potent than PGF2alpha in synchronizing the estrous cycle in beef cows.

Prostaglandin receptors in the hamster uterus during the estrous cycle

Abstract Uterine tissue from hamsters was incubated for 1 hr at 37°C with 3H-prostaglandin E1 (3H-PGE1) or 3H-prostaglandin F2α (3H-PGF2α) in the presence or absence of nonradioactive prostaglandin(s). Specific binding of PGE1 and PGF2α (the difference between 3H-PG in tissue incubated with and without nonradioactive PG) was detected throughout the estrous cycle. The specific binding of PGE1 varied during the estrous cycle and was at a maximum on day 3 (proestrus) and a minimum on day 1 (diestrus). The specific binding of PGF2α was similar at all stages of the estrous cycle.

Serum FSH, LH and testosterone in the male rhesus following prostaglandin injection

Adult male rhesus were treated with PGE2, PGF2 alpha or the 13,14-dihydro-15-keto metabolite of PGE2 in a randomized crossover design. Serum concentrations of FSH, LH and testosterone were determined and compared to the respective values in the same uninjected animals. No significant changes were noted in controls or following the metabolite injection. FSH increased gradually for 4 hours after metabolite treatment. In contrast, injection of PGF2 alpha was followed by an abrupt (within 15 minutes) increase in LH and testosterone. FSH increased gradually in 2 of 3 treated animals. Injection of PGE2 was followed by a similar abrupt increase in LH concentration. This was not always associated with a significant increase in testosterone or FSH. These results demonstrate that injections of PGE2 or PGF2 alpha can change serum gonadotropin and testosterone concentrations in male rhesus monkeys, and that the effects of these two prostaglandins are qualitatively different.

Contraceptive efficacy of testosterone-estradiol implants in male rhesus monkeys.

Rhesus monkeys (Macaca mulatta) were treated with testosterone (100 micrograms/kg/day) plus estradiol (0.5 micrograms/kg/day) via subcutaneous polydimethylsiloxane (PDS, Silastic) implants. This treatment caused a striking reversible sterility. No pregnancies were observed in females bred to the steroid-treated males. In contrast, there was no difference in pregnancy rate of females bred to control and steroid-treated monkeys for 14 weeks, beginning 17 weeks after removal of the steroid-filled implants.

Prostaglandin E1 specific binding in rhesus myometrium

Myometrial low speed supernatant prepared from non-pregnant rhesus uteri was incubated with 3H-Prostaglandin (PG)E1 with or without addition of unlabelled prostaglandins. The uptake of 3H-PGE1 was inhibited in a dose dependent fashion by PGE2greater thanPGE1greater thanPGA1greater thanpgf2alpha=PGA1greater thanPGB1=PGB2greater than or equal toPGD2. PGE1 metabolites inhibited 3H-PGE1 binding in the following order: 13, 14-dihydro-PGE1greater than13,14-dihydro-15-keto-PGE1=15-keto-PGE1. The specific binding of 3H-PGE1 and 3H-PGF2alpha was similarly affected by the temperature and time of incubation. Equilibrium binding constants determined using rhesus uteri obtained during the luteal phase of the menstrual cycle indicate the presence of high affinity PGE1 binding sites with an average (n=3) apparent dissociation constant of 2.2 X 10(-9)M and a lower affinity PGE1 binding site with a Kd approximately equal to 1 X 10(-8)M. No high affinity - low capacity 3H-PGF2alpha sites could be demonstrated. Relative uterine stimulating potencies of some natural prostaglandins and prostaglandin analogs tested after acute intravenous administration in mid-pregnant rhesus monkeys corresponded with the PGE1 binding inhibition of the respective compound. The uterine stimulating potencies of the prostaglandin analogs tested were: (15S)-15-methyl-PGE2=16,16-dimethyl-PGE2greater than17-phenyl-18,19,20-trinor-PGE2greater than16 phenoxy-17,18,19,20-tetranor-PGF2alpha=PGE2=PGE1=(15S)-15-methyl-PGF2alpha greater than PGF2alpha.

Evaluation of progesterone-containing silicone vaginal devices in rhesus monkeys

Abstract The biological activity of progesterone-containing silicone vaginal devices was evaluated in rhesus monkeys. Eighteen animals (six in each concentration group) were utilized to determine the release rate for the three drug concentrations evaluated, 2, 10 or 30%. Devices were inserted on day 5–7 of the cycle, and removed 20–21 days later. One control (placebo device) and one treatment cycle were evaluated for each animal. Sixteen of 18 control cycles were ovulatory, while there was only one in 18 treatment cycles, that occurring during a 2% cycle. Blood progesterone concentrations in the 10 and 30% groups were similar to concentrations during the luteal phase of the placebo cycles. Concentrations were lower, and dropped off sooner, in animals with 2% devices. Initiation of menses was closely associated with the day of ring removal in the 10 and 30% groups. Residual drug analysis illustrated that the 2% device was depleted within the 21-day period; the 10 and 30% devices lost 28.7 and 26.0 mg of progesterone, respectively. These release rates were more rapid than rates noted for other progestins, were independent of drug concentration, and were predictive based on previous in vitro studies.

Prostacyclin (PGI2) effets on anterior pituitary hormones in the rat in vivo

Intravenous injection of 600 microgram PGE2 or PGI2 significantly increased serum LH and prolactin levels in estradiol treated ovariectomized rats. There was no effect on serum FSH concentration. PGE2 and PGI2 stimulated LH release in a non-dose dependent manner, while prolactin levels were positively correlated with the dose administered following PGI2 treatment. 6-keto-PGF1 alpha at a comparable dose had no effect on pituitary hormone levels. Subcutaneous administration of 1 mg/kg or 60 mg/kg PGI2 for seven days significantly depressed serum LH level both in male and female rats. These doses had no effect on serum FSH or prolactin levels.

Male antifertility: An approach

Abstract Experimental data in the rat is presented for an approach to male fertility regulation. The concept involves administration of an anti-gonadotropic steroid plus a replacement androgen to inhibit spermatogenesis and to maintain normal secondary sex characteristics. A model system utilizing the adult male rat is described. FSH and LH concentrations in the plasma or serum increase 8–10 fold in response to castration. Secondary accessory organs atrophy significantly within 10 days following surgery. The near optimal replacement dosage of testosterone propionate (TP) in castrated male rats was 40 μg100g body weight day. Provera® (300 μg100g BW day), an antigonadotropic steroid, when administered to castrate males for 10 days decreased plasma LH and FSH concentrations to values observed in intact rats. TP (40 μg100g BW day) in intact males exerted an effect on accessory organs by the fifth day of treatment. The effect was still increasing after 10 days of treatment. Provera, in intact rats, inhibited accessory organ weights by the fifth day of treatment and no additional inhibition was detected after 10 days. A combination of Provera and TP in the intact male did not significantly affect accessory organ weights after 10 days treatment; however, testicular weights were significantly inhibited. In all animals treated with Provera, significant adrenal atrophy was evident.