SAT-405 Activation of Kiss1 Neurons in the Posterodorsal Medial Amygdala Stimulates Gonadotropin Release via the Release of Kisspeptin in Female Mice

Abstract

Reproduction is regulated by a complex neuronal network the precise components of which are still unknown. Kisspeptins, secreted from Kiss1 neurons in the arcuate nucleus (Kiss1 ^ARC) and anteroventral periventricular/periventricular (Kiss1 ^AVPV/PeN) nuclei, have been directly linked to the release of GnRH from the hypothalamus and hence, LH from the anterior pituitary. However, there is a population of Kiss1 neurons in the posterodorsal medial amygdala (Kiss1 ^MePD) whose contribution to GnRH/LH stimulation has not yet been described. Here, we aimed to determine whether the release of kisspeptin and/or other components, besides kisspeptin, within the Kiss1 ^MePD neuron, can stimulate LH release into the peripheral circulation. In this regard, we used a chemogenetic approach to specifically activate Kiss1 ^MePD neurons in adult Kiss1 ^Cre/+ (heterozygous state) females which we studied in parallel to Kiss1 ^Cre/Cre (Kiss1 knock-out state) littermates (n=5/group). Females received bilateral stereotaxic injections of an adeno-associated virus (pAAV) encoding a Cre-driven Gq-coupled excitatory DREADD (pAAV5/hSyn-DIO-hm3Dq:mCherry; titer 3x10 ¹² genome copies per ml; 1 µl per hemisphere). Following infection, mice were given 3 weeks for recovery and maximum expression of the AAV vector. On day 1 of the experiment, animals were administered an ip bolus injection of vehicle saline (0.9% NaCl; day 1) and then hM3D receptors were activated by ip injection of its agonist, clozapine N-oxide (CNO; 10 mg/kg dissolved in saline; day 2). Blood samples were collected just before saline or CNO treatment (0) and then every 15 min for 90 min. Kiss1 ^Cre/+ mice expressing hM3Dq:mCherry in the MePD and treated with CNO to activate the Kiss1 ^MePD neurons, showed an increase in LH within 30 min after the injection (P=0.0107) compared to animals receiving saline treatment, which was sustained for a further 30 min before returning to basal levels. Furthermore, no alteration in LH was observed in Kiss1 ^Cre/Cre (i.e., Kiss1 KO) animals treated with either saline or CNO indicating that kisspeptin is the only component within the Kiss1 ^MePD neuron that has the ability to stimulate LH release. Analysis following the completion of pharmacological studies demonstrated that mCherry expression was evident in 89% of the targeted Kiss1 ^MePD neurons. Interestingly, mCherry labeled projections were observed in the ARC and specifically in close contact with Kiss1 ^ARC neurons. Thus, in the female mouse, Kiss1 ^MePD neurons have the ability to stimulate the gonadotropic axis but only when kisspeptin is present. Furthermore, this study suggests that this action is, at least in part, via the activation of Kiss1 ^ARC neurons which, in turn, stimulate GnRH/LH release. Overall, our data provide insight into the potential mechanism via which amygdala regulated social cues can enhance reproductive function.