Expression and activation of N-methyl-D-aspartate receptor subunit-1 receptor subunits in gonadotrophin-releasing hormone neurones of young and middle-aged mice during the luteinising hormone surge

Abstract

Glutamate is an important excitatory neurotransmitter that stimulates the release of gonadotrophin-releasing hormone (GnRH) and participates in the generation of the luteinising hormone (LH) surge. To determine the mechanisms of action of glutamate and possible changes in the glutamatergic input to GnRH neurones during reproductive ageing, we measured the expression and activation of the mandatory N-methyl-D-aspartate receptor subunit-1 (NMDAR1) in GnRH neurones of young and middle-aged mice prior to and during a steroid-induced LH surge. The results show that, in young animals, approximately 55% of all GnRH neurones contain immunoreactive NMDAR1 protein and this percentage does not change during the day of the LH surge. In approximately 10% of the GnRH neurones, NMDA receptor protein is phosphorylated at Ser 890 prior to the surge, whereas, in approximately 55% of the GnRH neurones, NMDAR1 subunits are phosphorylated during the LH surge. Activation of NMDAR1 receptor subunits stimulates the calcium-calmodulin-kinase IV (CaMK IV). pathway, which leads to the translocation of CaMK IV into the nucleus where this enzyme can phosphorylate the cAMP response element-binding protein (CREB) and CREB-binding protein. We show that, in young animals, approximately 20% of the GnRH neurones contain CaMK IV in their nuclei 7 h prior to the LH surge; this percentage increases to 60% at the beginning of the surge and decreases to approximately 40% some 2 h into the LH surge. In middle-aged animals, approximately 25% of the GnRH neurones contain NMDAR1 protein and only 10% of the GnRH neurones contain phosphorylated NMDAR1 protein prior to and during the surge; however 2 h after the peak of the surge, 20% of the GnRH neurones contain phosphorylated NMDAR1 subunits. Similarly, 20% of GnRH neurones contain nuclear CaMK IV and this percentage does not change during the day of the LH surge. The results suggest that, in the young animal, glutamatergic innervation of GnRH neurones during the LH surge causes the activation and phosphorylation of NMDAR1 receptor subunits which results in the translocation of CaMK IV into the nucleus. However, both, the expression and activation of NMDAR1 receptor subunits are greatly reduced in the middle-aged animals, which could result in the absence of LH surges.