Development of a non-steroidal aromatase inhibitor-based protocol for the control of ovarian function using a bovine model
Five studies were designed to characterize the effects of a non-steroidal aromatase inhibitor, letrozole, on ovarian function in cattle. The general hypothesis was that non-steroidal aromatase inhibitors have potential as a steroid-free option for the control of ovarian function for the purposes of fixed-time artificial insemination and embryo production. The specific objectives were to determine the effect of route and vehicle, type of aromatase inhibitor, and duration of aromatase inhibitor treatment (short vs prolonged) on ovarian follicles in cattle, and to test the efficacy of an aromatase inhibitor-based protocol to synchronize ovulation in cattle. In the first experiment, heifers were treated with letrozole intravenously (n=10) or intramuscularly (n=10) or allocated in iv and im control groups (n=5/group). During the second experiment, heifers were divided randomly into two groups (n=15/group) and an intravaginal device containing 1 g of letrozole or a blank device (control) was inserted. The third experiment was designed with the goal of formulating and testing an intravaginal device that provides biologically active circulating concentrations of an aromatase inhibitor for a minimum of 4 days. The biological significance of the pharmacokinetic differences between the letrozole intravaginal devices resulting from the third study was evaluated during the fourth study. A final study was designed to determine the effect of stage of the estrous cycle on the proportion of animals that ovulated and the synchrony of ovulation of heifers treated with an aromatase inhibitor-based ovulation-synchronization protocol and to determine subsequent pregnancy outcomes. In all the studies, the effects of aromatase inhibitor on ovarian function were assessed by transrectal ultrasound examination of the ovaries, and blood samples were collected for hormone concentration determination. Results demonstrated that route of administration, or more precisely, the nature of iii the vehicle used for the administration of letrozole (intravenous, intramuscular depot, short release intravaginal or prolonged release intravaginal) has an impact on the effects of letrozole on hormonal profiles and ovarian dynamics. The intramuscular route appeared to provide a prolonged release of letrozole from the injection site which had a marked effect on estradiol production, dominant follicle lifespan, and CL form and function. Letrozole treatment during the ovulatory follicle wave by means of a gel-based intravaginal releasing device during the second study resulted in more rapidly growing dominant follicles and larger ovulatory follicles, delayed ovulation (by 24 h) of a single follicle and formation of a CL that secreted higher levels of progesterone. A wax-based vehicle allowed for a steady and continuous delivery of the active compound over the treatment period. During the third study, the addition of a letrozole-containing gel coating increased the rate of initial absorption and hastened the increase on plasma concentrations of the active ingredient, while the letrozole-containing wax-based vehicle prolonged drug-delivery from the intravaginal device. When tested in vivo during the fourth study, we confirmed that letrozole-impregnated intravaginal devices formulated with a wax base plus a gel coat vehicle was most suitable for the application of a letrozole-based protocol for the synchronization of ovulation in cattle, since it effectively delivered elevated concentrations of letrozole, and reduced estradiol production resulting in increased follicular growth and lifespan, without adversely affecting progesterone production. The application of a letrozole-impregnated intravaginal device for 4 days, combined with PGF treatment at device removal and GnRH 24 h post-device removal increased the percentage of ovulations and synchrony of ovulation in cattle, regardless the stage of the estrous cycle at initiation of treatment. As observed in previous studies, the effects observed could be associated with an increase in circulating LH iv concentrations. However, the effects of treatment on gonadotropin concentrations are inconclusive, possibly due to inadequate sampling frequency. The impact of letrozole treatment of oocyte fertility remains unknown. The results of the five experiments support our general hypothesis that non-steroidal aromatase inhibitors have potential as a steroid-free option for the control of ovarian function in cattle. However, further research is needed in order to elucidate the effects of letrozole treatment during the proestrous on oocyte competence and fertility of the resulting ovulations in cattle.
DegreeDoctor of Philosophy (Ph.D.)
DepartmentVeterinary Biomedical Sciences
ProgramVeterinary Biomedical Sciences
SupervisorAdams, Gregg P.
CommitteeMapletoft, Reuben J.; Singh, Jaswant; Pierson, Roger; Dragan, Rogan
Copyright DateJune 2013
aromatase inhibitor, letrozole, synchronization