146 Evidence for a role of a noradrenergic influence on nerve growth factor–induced luteinising hormone secretion in llamas

Nerve growth factor (NGF) has been established as the main factor triggering induced ovulation in South American camelids. Although the mechanism of action of NGF has not been fully characterised, recent data suggest that an inter-neuronal transduction pathway within the hypothalamus is the functional target of NGF (Carrasco et al. 2021 Biol. Reprod. 104, 578–588). One strong candidate is the noradrenergic system, which in rabbits (another induced ovulator) is a clear mediator of mating-induced ovulation at the level of the brain. The objective of this study was to test the hypothesis that a hypothalamic noradrenergic neuronal network plays a role in mediating the NGF-induced preovulatory luteinising hormone (LH) surge. In Experiment 1, we compared the LH-releasing response to NGF (500 µg IM) in llamas that were pre-treated intravenously with an α-1 and -2 noradrenergic receptor antagonist (0.8 mg per kg; phenoxybenzamine) or vehicle (n = 5 per group). In Experiment 2, we compared the LH response to NGF in llamas that were pre-treated in the lateral cerebral ventricle with an α-1 noradrenergic receptor antagonist (doxazosin; infusion of 300 nmoles per hour for 3 h with a syringe pump; n = 3) or left untreated (n = 4). The LH response in Experiments 1 and 2 was detected by measurement of plasma LH concentration in blood samples taken every 30 minutes for 7 and 3 h, respectively. In Experiment 3, we used double immunohistochemistry and immunofluorescence to compare the neuroanatomic association between neurons that contain tyrosine hydroxylase (TH), an enzyme that is part of the noradrenergic biosynthetic pathway, and GnRH neurons in the hypothalamus of llamas (n = 5). Serial data were compared by one-way analysis of variance for repeated-measures, and differences were considered significant when P < 0.05. In Experiment 1, no differences were detected in the LH response to NGF in llamas pre-treated with either phenoxybenzamine or vehicle; LH concentrations increased immediately after NGF treatment, reaching a maximum 2 h later, and remaining elevated 6 h after treatment. In Experiment 2, intra-cerebroventricular infusion of doxazosin reduced the magnitude of the LH surge at 30 and 90 to 150 minutes of NGF treatment (P < 0.05). In Experiment 3, we found that GnRH neurons did not express TH immunoreactivity, suggesting that these two neuronal populations were neuroanatomically distinct. However, we found abundant TH immunoreactive appositions onto GnRH neurons and their axons. Results support the hypothesis that a noradrenergic network within the hypothalamus plays a role in mediating the NGF-induced ovulatory pathway in llamas, and it could lead to understanding the biology of induced ovulation. Although detection of the noradrenergic enzyme tyrosine hydroxylase in potential synapses with GnRH nerve cell bodies and axons within the hypothalamus indicates a direct effect, the neuroanatomic source of such noradrenergic inputs remains unknown.

This research was supported by the Natural Science and Engineering Research Council of Canada.