ONTOGENY OF THE LUTEINIZING HORMONE RELEASING HORMONE SYSTEM

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Luteinizing hormone releasing hormone (LHRH) neurons are derived from the olfactory placode and migrate into the brain, where they become integral members of the hypothalamic-pituitary-gonadal axis. To study the migratory mechanism(s) involved in LHRH neuronal movement into the CNS, we use normal and transgenic animals, as well as olfactory explants. In addition, long-term organotypic slice cultures are used to study mechanisms underlying intrinsic and trans-synaptic regulation of LHRH gene expression, peptide synthesis and secretion in postnatal differentiated LHRH neurons. Working on the hypothesis that LHRH neurons migrate on peripherin positive (+) olfactory axons from the olfactory pit to diencephalon, we found that: (1) LHRH neurons do not express peripherin mRNA; (2) LHRH neurons do not express N-CAM mRNA, although olfactory axons are N-CAM+; and (3) olfactory pit cells differentially express peripherin mRNA and N-CAM mRNA, suggesting distinct populations. In embryonic explants, we distinguished N-CAM+ and peripherin+ axons, and verified that LHRH neurons moved via peripherin+ but not N-CAM+ axons. Examination of voltage- and ligand-gated channels on embryonic LHRH neurons revealed membrane characteristics of highly differentiated neurons. In addition, we have generated embryonic explants from transgenic mice expressing luciferase in LHRH neurons. When given luciferin, a detectable signal is measurable in lysed cells from these explants. We examined second messengers active in LHRH cells and oxytocin (OT) cells maintained in organotypic slice explants. Forskolin and/or phorbol 12-myristate 13-acetate (PMA) treatment significantly decreased LHRH mRNA levels at 4 hr. In contrast, forskolin treatment significantly increased OT mRNA levels by 8 hr. Using actinomycin D (a transcription inhibitor), we determined neuropeptide mRNA turnover rates: LHRH mRNA has a very fast turnover rate (approximately 4 hr), while OT mRNA is much slower (approximately 40 hr). We propose that second messengers act primarily to increase transcription of OT mRNA in OT cells, but decrease LHRH mRNA transcription and/or increase LHRH mRNA degradation in LHRH neurons. Currently, we are determining: (1) cell surface glycoproteins expressed on LHRH neurons and/or the peripherin+ axons with which they associate; (2) the identity of cells expressing N-CAM vs those expressing peripherin in nasal regions; (3) whether LHRH neurons maintained in cultures release LHRH in a pulsatile manner; and (4) whether tagged-LHRH neurons can be visualized in situ to monitor movement in embryonic explants and~or determine the membrane properties of postnatal LHRH neurons in organotypic slices.