BACKGROUND: Kisspeptin and its cognate receptor GPR54 are the central driving forces in the hypothalamus-pituitary-gonadal axis essential for sexual maturation and reproduction. Kisspeptin/GPR54 signalling stimulates gonadotropin-releasing hormone (GnRH) neurones and induces pulsatile GnRH release. The molecular signalling pathway by which kisspeptin stimulates GnRH neurones is currently under investigation. METHODS: Primary GnRH neurones were isolated from young adult rats and loaded with the calcium indicator Fura Red. Cytosolic calcium was measured while the cells were stimulated with kisspeptin. RESULTS: GnRH neurones show a maintained increase of cytosolic calcium upon stimulation with 100 nM kisspeptin-10. The calcium elevation was inhibited 30% by 1 μM tetrodotoxin, a voltage-gated sodium channel blocker, and 76% by 30 μM SKF96365, an inhibitor of receptor-mediated calcium entry. Furthermore, removal of extracellular calcium completely abolished the kisspeptin-induced calcium elevation. CONCLUSION: Our results suggest that the major part of the kisspeptin-evoked calcium signal is generated by an action potential-independent calcium influx, possibly through channels of the classical transient receptor potential type, with an additional influx through voltage-gated calcium channels activated by sodium action potentials.
BACKGROUND: Kisspeptin and its cognate receptor GPR54 are the central driving forces in the hypothalamus-pituitary-gonadal axis essential for sexual maturation and reproduction. Kisspeptin/GPR54 signalling stimulates gonadotropin-releasing hormone (GnRH) neurones and induces pulsatile GnRH release. The molecular signalling pathway by which kisspeptin stimulates GnRH neurones is currently under investigation. METHODS: Primary GnRH neurones were isolated from young adult rats and loaded with the calcium indicator Fura Red. Cytosolic calcium was measured while the cells were stimulated with kisspeptin. RESULTS:GnRH neurones show a maintained increase of cytosolic calcium upon stimulation with 100 nM kisspeptin-10. The calcium elevation was inhibited 30% by 1 μM tetrodotoxin, a voltage-gated sodium channel blocker, and 76% by 30 μM SKF96365, an inhibitor of receptor-mediated calcium entry. Furthermore, removal of extracellular calcium completely abolished the kisspeptin-induced calcium elevation. CONCLUSION: Our results suggest that the major part of the kisspeptin-evoked calcium signal is generated by an action potential-independent calcium influx, possibly through channels of the classical transient receptor potential type, with an additional influx through voltage-gated calcium channels activated by sodium action potentials.
Authors: Le Min; Kathleen Soltis; Ana Claudia S Reis; Shuyun Xu; Wendy Kuohung; Manisha Jain; Rona S Carroll; Ursula B Kaiser Journal: Mol Endocrinol Date: 2013-12-02