title: Colocalization of FM1-43, Bassoon, and GnRH-1: GnRH-1 release from cell bodies and their neuroprocesses.
abstract: Pulsatile release of GnRH-1 is critical for reproductive function. However, the cellular mechanism of GnRH-1 neurosecretion is still elusive. In this study, we examined the neurosecretory process of GnRH-1 neurons using time-lapse image acquisition followed by immunocytochemistry with confocal microscopy. To monitor exocytotic processes, cultured GnRH-1 neurons derived from monkey embryos were labeled with the lipophilic dye, FM1-43, or its fixable form FM1-43Fx, in the presence or absence of depolarization signals, and changes in vesicles labeled with FM1-43 were analyzed. The results show FM1-43 was taken up into the cell and labeled puncta in the soma and neuroprocesses in the absence of depolarization signals, indicating that GnRH-1 neurons were spontaneously active. Depolarization of GnRH-1 neurons with high K+ or veratridine challenge increased the intensity and size of puncta in both soma and neuroprocesses, and the veratridine-induced changes in puncta were blocked by tetrodotoxin, indicating that changes in the puncta intensity and size reflect neurosecretory activity. Subsequent double immunocytochemistry for GnRH-1 and the synaptic vesicle marker, vesicle-associated membrane protein, demonstrated that the FM1-43Fx-labeled puncta were synaptic vesicles with the GnRH-1 peptide. Additional double immunocytochemistry for GnRH-1 and the marker of the neurosecretory active zone, Bassoon, indicated that the FM1-43Fx-labeled puncta were located at the sites of neurosecretory active zones in GnRH-1 neurons. These results suggest that GnRH-1 neurons have the capacity to release the peptide from the soma and dendrites. Collectively, we hypothesize that soma-dendritic release of the peptide may be a mechanism of synchronized activity among GnRH-1 neurons.
authors: Department of Pediatrics and Wisconsin National Primate Research Center, 1223 Capitol Court, Madison, Wisconsin 53715-1299, USA.
authors: Lidia C
authors: LC
authors: Fuenzalida
authors: Kim L
authors: KL
authors: Keen
authors: Ei
authors: E
authors: Terasawa
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D005456:Fluorescent Dyes / Q000494:pharmacology; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism*; D008253:Macaca mulatta; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D009489:Neurosecretion / Q000502:physiology*; D011726:Pyridinium Compounds / Q000494:pharmacology; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology; D050683:R-SNARE Proteins / Q000378:metabolism; D013572:Synaptic Vesicles / Q000378:metabolism
chemical_list: C073804:FM1 43; D005456:Fluorescent Dyes; D009419:Nerve Tissue Proteins; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D050683:R-SNARE Proteins; D007987:Gonadotropin-Releasing Hormone