Characterization of Luteinizing Hormone and Luteinizing Hormone Receptor and Their Indispensable Role in the Ovulatory Process of the Medaka

Abstract

The molecular properties and roles of luteinizing hormone (Lh) and its receptor (Lhcgrbb) have not been studied for the medaka (Oryzias latipes), which is an excellent animal model for ovulation studies. Here, we characterized the medaka Lh/Lhcgrbb system, with attention to its involvement in the ovulatory process of this teleost fish. In the medaka ovary, follicle-stimulating hormone receptor mRNA was expressed in small and medium-sized follicles, while lhcgrbb mRNA was expressed in the follicle layers of all growing follicles. Experiments using HEK 293T cells expressing medaka Lhcgrbb in vitro revealed that gonadotropin from pregnant mare’s serum and medaka recombinant Lh (rLh) bound to the fish Lhcgrbb. The fish gonadotropin subunits Gtha, Fshb, and Lhb were essentially expressed at fairly constant levels in the pituitary of the fish during a 24-h spawning cycle. Using medaka rLh, we developed a follicle culture system that allowed us to follow the whole process of oocyte maturation and ovulation in vitro. This follicle culture method enabled us to determine that the Lh surge for the preovulatory follicle occurred in vivo between 19 and 15 h before ovulation. The present study also showed that oocyte maturation and ovulation were delayed several hours in vitro compared with in vivo. Treatment of large follicles with medaka rLh in vitro significantly increased the expression of Mmp15, which was previously demonstrated to be crucial for ovulation in the fish. These findings demonstrate that Lh/Lhcgrbb is critically involved in the induction of oocyte maturation and ovulation.

Leporinus elongatus induced spawning using carp pituitary extract or mammalian GnRH analogue combined with dopamine receptor antagonists

“…The induced ovulation failed in Rhamdia quelen (two doses: 2 µg mGnRHa kg -1 + 1 mg metoclopramide kg -1 and 20 µg mGnRHa kg -1 + 10 mg metoclopramide kg -1 metoclopramide) (Carneiro and Mikos, 2008). When applied to Piaractus mesopotamicus, Brycon orbygnianus and Prochilodus lineatus, busserelin acetate led to ovulation, but no viable embryos were obtained after fertilization (Paulino et al, 2011). In addition, different from most South American rheophilic species, viable embryos and successful ovulation can be unusually obtained with very low doses of CPE (0.5mg kg -1 (with successive doses every six hours, if necessary)) in Schizodon fasciatus (Lopes and Leal, 2010).…”

“…However, the results obtained here reinforced the existing hypothesis of an ovarian hyperstimulation with consequent egg toxicity due to the hormonal treatment in this rheophilic species and other South American ones, that must be addressed in futures studies. Recent reports on embryonic mortality obtained in spawning induced South American migratory species (Acuña and Rangel, 2009;Carneiro and Mikos, 2008;Paulino et al, 2011;Pereira et al, 2017), coupled with the need for hormone-inducing such species in order to obtain embryos, point out to an immediate need for speciesspecific studies (probably in vitro) in order to confirm which substances used in hormone therapies (and their derivatives), as well as which levels are indeed toxic to the eggs. This aspect needs to be addressed for the development of aquaculture in countries which depend on these species, since rheophilic fish are known to be among the most important fish for commercial purposes in South American countries (MPA 2013;FAO 2014) and the constant and predictable supply of fingerlings is imperative for the consolidation of the use of any species for aquacultural purposes.…”

“…Moreover, due to their synthetic nature, they pose no risk of transmitting diseases, such as carp pituitary extracts (CPE) do, and since they act at higher levels of the hypothalamic-pituitary gonad axis, they stimulate the release of LH and FSH, as well as other pituitary hormones, which may serve important reproductive functions (review in Mylonas et al, 2010). However, although the efficiencies of the use of synthetic products in some South American rheophilic species (Ittze's et al, 2015;Viveiros et al, 2015) have already been shown -such as obtaining ovulation and viable embryos -and even if they sometimes have a higher potential for inducing ovulation compared to that of CPE (Pereira et al, 2017), the use of mGnRH in these species is frequently associated with ovulation failure (Carneiro and Mikos, 2008) and / or failure in obtaining viable embryos (Acuña and Rangel 2009;Paulino et al, 2011;Pereira et al, 2017).…”

Disruption of Zebrafish Follicle-Stimulating Hormone Receptor (fshr) But Not Luteinizing Hormone Receptor (lhcgr) Gene by TALEN Leads to Failed Follicle Activation in Females Followed by Sexual Reversal to Males

“…Most studies on Fshr and Lhcgr in fish have focused on their expression profiles during reproductive cycle (25)(26)(27)(28). In the zebrafish, both fshr and lhcgr are exclusively expressed in the somatic follicle cells in the ovary (29).…”

Targeted Gene Disruption in Zebrafish Reveals Noncanonical Functions of LH Signaling in Reproduction

“…In addition, marked differences exist between mammals and fish with regard to the specificity of the gonadotropins toward their receptors (1,23,24). It has been demonstrated in several fish species including zebrafish that FSH stimulates only FSHR, whereas LH stimulates both FSHR and LHR (11,(25)(26)(27)(28). This difference implies functional differentiation of LH signaling from fish to mammals.…”