Abstract
Gonadotropin-releasing hormone (GnRH) neurons are the final output neurons of a hypothalamic network controlling fertility in mammals. They release GnRH from axon terminals into the portal blood circulation of the pituitary, where GnRH triggers the secretion of luteinizing and follicle stimulating hormones. GnRH neuron anatomy, morphology and physiology have been studied extensively at the levels of the cell bodies and dendrites in the basal forebrain and axons in the median eminence. However little is known about the subcellular origin of the axon and the physiological role of the dense GnRH fiber innervation of the organum vasculosum of the lamina terminalis (OVLT).
Using a cell-filling approach in acute brain slices from GnRH-green fluorescent protein mice, GnRH neuron morphology was analyzed with a confocal microscope. The vast majority (83%) of GnRH neurons were found to extend long dendrites in the direction of the median eminence. When these dendrites were analyzed in a novel brain slice preparation that keeps GnRH neuron projections to the median eminence intact, dendrites were observed to innervate the median eminence and elaborate hypophysiotropic terminals. This projection received continuous synaptic input as indicated by the presence of spines and VAMP2-positive appositions. Moreover, analysis of the position of the action potential-generating initial segment using the marker Ankyrin G, revealed that GnRH neurons exhibit initial segments in one of their dendrites in >80% of cases. This suggests that the long GnRH neuron dendrite has both dendritic and axonal functions and that this unique structure, termed the “dendron”, necessitates a considerable revision of how GnRH neurons integrate inputs and regulate GnRH release at the median eminence.
Cell-filling of GnRH neurons with cell bodies located near the OVLT revealed that, unlike other GnRH neurons in the preoptic area, these neurons elaborate complex, highly branched dendritic trees which specifically target the OVLT. Dendritic identity was confirmed morphologically at the light and electron microscopic level and functionally by electrophysiological recordings of the response to glutamate. Further, these dendrites were shown to reside outside the blood-brain barrier and belong to GnRH neurons involved in the generation of the GnRH/LH surge. This suggests that GnRH neuron activity, and hence fertility, may be directly modulated by blood-borne factors. While kisspeptin puffed onto dendrites in the OVLT was found to activate GnRH neurons, luteinizing hormone did not exert feedback actions at this site and no evidence was found for a role of OVLT-GnRH neurons in the suppression of fertility during systemic infection. In summary, work in this thesis has re-defined the morphology of the GnRH neuron by showing that the dendrites of GnRH neurons target highly vascularized regions outside the blood-brain barrier. Most importantly, the work demonstrates that the long hypophysiotropic projections of GnRH neurons to the median eminence are, in fact, dendritic in nature.