Elsevier

Theriogenology

Volume 141, 1 January 2020, Pages 26-34
Theriogenology

Kisspeptin induces ovulation in heifers under low plasma progesterone concentrations

https://doi.org/10.1016/j.theriogenology.2019.08.033Get rights and content

Highlights

  • Single dose of human kisspeptin-10 increased plasma LH concentration more than the murine kisspeptin-10.

  • Multiple administration of human kisspeptin-10 was more effective than the murine sequence for inducing ovulation rate.

  • Multiple human kisspeptin-10 treatments induced ovulation and follicular wave emergence at rates comparable to that of GnRH.

Abstract

The objective of the present study was to compare the effect of a single versus multiple doses of a 10-amino acid fragment of human (hKp) or murine (mKp) kisspeptin on LH secretion and the fate of the dominant follicle. In all experiments, a new wave was induced (Day 0) by ultrasound-guided ablation of >5 mm follicles, a progesterone device (CIDR) was placed in the vagina, animals given prostaglandin F analog im on Day 3.5 and 4, and hKp or mKp treatment given on Day 6. The experimental design maintained growth and ovulatory potential of the dominant follicle for 12 days and allowed hypothesis testing during the low-progesterone period (plasma progesterone ≤1.8 ng/ml on Day 6) wherein spontaneous wave emergence and ovulation did not occur between Day 6 and Day 12. In Experiment 1, heifers (n = 10/group) were given single iv dose of 45 mg hKp, 45 mg mKp, or 2 ml normal saline (control). Post-treatment plasma LH concentrations from 15 to 90 min were higher (P < 0.01) in hKp group than in the mKp and control groups. Two heifers ovulated in hKp group versus none in other groups. In Experiment 2, heifers (n = 6/group) were given 45 mg hKp over a 2 h period divided into multiple iv doses treatments or 2 ml normal saline (control). Post-treatment plasma LH concentrations were higher (P < 0.01) in all hKp treatment groups than in the control group. The ovulation rate was higher (P = 0.06) after hKp treatments (11/18) than in the control group (0/6). In Experiment 3, heifers (n = 6/group) were given 45 mg mKp over a 2 h period divided into multiple iv doses treatments or a single iv dose of gonadorelin acetate (positive control). Plasma LH concentration was higher (P < 0.01) and the ovulation rate was greater (P = 0.01) in the GnRH group (5/6) than mKp groups (1/12). In summary, hKp was more effective to induce ovulation than mKp. Human kisspeptin-10 given over a 2 h period induced ovulations at a rate similar to that of GnRH treatment in heifers under a low plasma progesterone state.

Introduction

The hypothalamo-pituitary-gonadal axis controls follicular development primarily by changes in pulsatile release of GnRH from the hypothalamus that induces downstream LH and FSH secretion from the pituitary gland. In turn, gonadotropins influence the pattern of steroid synthesis from ovarian cells [[1], [2], [3]] which provides positive and negative feedback on GnRH release and gonadotropin hormone synthesis and release. Because of a negative feedback effect, circulating concentrations of progesterone are inversely related to LH release [4,5]; hence, low concentrations of progesterone (less negative feedback) are associated with increased GnRH neuron activity and an increase in frequency of gonadotropin hormone pulses [[6], [7], [8], [9]]. However, there appears to be no direct link between sex steroid hormones and GnRH neurons in the hypothalamus since there is an absence of estradiol receptors alpha and progesterone receptors on GnRH neurons [10,11]. In 2003, the discovery that alterations of the kisspeptin/GPR54 system were associated with reproductive disturbances and idiopathic hypothalamic hypogonadism created a new understanding of steroidal control of GnRH release and its fundamental effect on reproductive physiology [12,13].

In rats, kisspeptin cells were closely associated with GnRH neurons that expressed the kisspeptin receptor GPR-54 [14,15]. In search of the bioactive portion of the 145-amino acid peptide, kisspeptin has been proteolytically cleaved into shorter peptides of 54-amino acids (kisspeptin-54) and 14-, 13- and 10-amino acids (kisspeptin-14, kisspeptin-13, kisspeptin-10) [16]. Bioactivity appears to be confined primarily to kisspeptin-10 since, in rats and sheep, administration of this short segment induced GnRH neuron activation, release of GnRH in the portal circulation, and a rise in plasma gonadotropin concentrations [15,17]. Administration (intraperitoneal, intravenous and subcutaneous) of kisspeptin 10 induced a marked increase in plasma concentrations of LH and FSH in several species including goats, sheep, mice and primates [[18], [19], [20], [21], [22]]. Continuous intravenous administration of kisspeptin over a period of 30 or 48 h induced ovulation in sheep during the anovulatory season [19]. The effectiveness of prolonged treatment, or the lack of effectiveness of short treatments, has been attributed to the short half-life of kisspeptin in circulation [23,24]. Predicted bovine kisspeptin-10 amino acid sequence is identical to murine C-terminal decapeptide and differs from human sequence by one amino acid [16]. While there is only slight variation among mammalian species in the sequence of the final 10 amino acids (i.e., the biologically-active fragment) at the C-terminal part of kisspeptin [16], the biological effect of heterospecific kisspeptin-10 sequences has not been critically examined in cattle [25,26].

The objective of this study was to compare the effect of a single iv bolus versus multiple doses of a 10-amino acid fragment of human or murine kisspeptin on LH secretion and the fate of the dominant follicle (ovulation, growth rate, regression and time to next wave emergence) during development in a low-progesterone environment. We predicted that 45 mg of kisspeptin-10 (administered as a single iv dose or given by multiple doses over a 2 h period) would elicit a surge in plasma LH leading to ovulation of the dominant follicle. We tested the hypotheses that (1) a single dose of murine kisspeptin-10 (homolog to the predicted bovine kisspeptin-10 sequence) will induce a greater response than human kisspeptin-10, (2) multiple-dose treatment with kisspeptin-10 (total dose of 45 mg) will induce a greater rise in plasma LH concentration and more ovulations than a single dose, and (3) multiple-dose murine kisspeptin-10 will induce ovulations at a rate comparable to that of GnRH treatment.

Section snippets

Animals

Three experiments were conducted on sexually mature Hereford cross-bred heifers. Experiment 1 was done on 30 heifers (455 ± 12 Kg body weight, 14–16 months of age) during the Spring (May–June). Experiment 2 was done on 24 heifers (426 ± 8 Kg body weight, 17–18 months of age) in the Summer (July–August). Experiment 3 was done on 18 heifers (426 ± 10 Kg body weight, 19–20 months of age) in the Fall (September–October). The animals were maintained in outdoors pens at the University of Saskatchewan

Experimental model

To validate the experimental protocol of dominant follicle development, data from the control groups in Experiment 1 (n = 10) and Experiment 2 (n = 6) were combined. The mean dominant follicle profile from wave emergence to ovulation after CIDR removal is shown in Fig. 2. The dominant follicle continued to grow from the day of wave emergence (Day 0) to Day 14. Ovulation did not occur in any of the heifers until after removal of the CIDR. The maximum diameter of the dominant follicle was

Discussion

Estrus synchronization, fixed-time artificial insemination, and management of embryo donors and recipients require exogenous control of ovarian follicular wave emergence and ovulation. Both single and multiple iv doses of human and murine kisspeptin-10 increased plasma LH concentrations, but the ovulation rates differed depending on the frequency of treatment. We found that: 1) a single dose of human kisspeptin-10 increased plasma LH concentration more than the bovine predictor sequence (i.e.,

Conflicts of interest

The authors declare that they have no competing interests.

Authors’ contributions

Carlos E.P. Leonardi participated in designed of study, collection, analysis and interpretation of data, and in writing and revising the manuscript. Fernanda C.F. Dias participated in design of study, interpretation of data and revising the manuscript. Estela R. Araujo participated analysis and interpretation of data, as well as, in writing and revising the manuscript. Gregg P. Adams participated in designing the study, interpretation of data and revising the manuscript. As Principal

Acknowledgements

The research was supported by grant from the Natural Sciences and Engineering Research Council of Canada. Carlos Leonardi was financially supported by CAPES scholarship from the Ministry of Education of Brazil. We thank Taryn Roberts, Dr. Rodrigo Carrasco and Eric Zwiefehofer for assistance with data collection, and Dr. OJ Ginther for all support to analyse LH and Progesterone concentrations.

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