Expression of Vesicular Glutamate Transporter 2 (vGluT2) on Large Dense-Core Vesicles within GnRH Neuroterminals of Aging Female Rats

• Published in: PloS one Vol. 10; no. 6; p. e0129633
• Main Authors: Yin, Weiling, Sun, Zengrong, Mendenhall, John M., Walker, Deena M., Riha, Penny D., Bezner, Kelsey S., Gore, Andrea C.
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 08.06.2015; Public Library of Science (PLoS)
• Subjects: 17β-Estradiol; Age Factors; Aging; Amino acids; Animals; Brain; Confocal microscopy; Dense core vesicles; Electron microscopy; Embedding; Endocrinology; Estrogens; Female; Females; Gene Expression; Glutamate; Glutamatergic transmission; Glutamic acid transporter; Gonadotropin-releasing hormone; Gonadotropin-Releasing Hormone - blood; Gonadotropin-Releasing Hormone - secretion; Hormones; Hypothalamus; Immunofluorescence; Immunoreactivity; Laboratory animals; Life cycle engineering; Life cycles; Median Eminence - metabolism; Menopause; Microscopy; Middle age; Morphology; Nerve endings; Neurons; Neuropeptides; Neurophysiology; Neurosciences; Neurotransmitters; Ovariectomy; Pharmacology; Pharmacy; Phenotypes; Physiological aspects; Presynaptic Terminals - metabolism; Presynaptic Terminals - ultrastructure; Progesterone; Protein Transport; Rats; Rodents; Secretory Vesicles - metabolism; Senescence; Sex hormones; Signal transduction; Signaling; Steroid hormones; Toxicology; Transport buildings, stations and terminals; Vesicles; Vesicular Glutamate Transport Protein 2 - genetics; Vesicular Glutamate Transport Protein 2 - metabolism; United States > US; Texas
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0129633

The pulsatile release of GnRH is crucial for normal reproductive physiology across the life cycle, a process that is regulated by hypothalamic neurotransmitters. GnRH terminals co-express the vesicular glutamate transporter 2 (vGluT2) as a marker of a glutamatergic phenotype. The current study sought to elucidate the relationship between glutamate and GnRH nerve terminals in the median eminence--the site of GnRH release into the portal capillary vasculature. We also determined whether this co-expression may change during reproductive senescence, and if steroid hormones, which affect responsiveness of GnRH neurons to glutamate, may alter the co-expression pattern. Female Sprague-Dawley rats were ovariectomized at young adult, middle-aged and old ages (~4, 11, and 22 months, respectively) and treated four weeks later with sequential vehicle + vehicle (VEH + VEH), estradiol + vehicle (E2 + VEH), or estradiol + progesterone (E2+P4). Rats were perfused 24 hours after the second hormone treatment. Confocal microscopy was used to determine colocalization of GnRH and vGluT2 immunofluorescence in the median eminence. Post-embedding immunogold labeling of GnRH and vGluT2, and a serial electron microscopy (EM) technique were used to determine the cellular interaction between GnRH terminals and glutamate signaling. Confocal analysis showed that GnRH and vGluT2 immunofluorescent puncta were extensively colocalized in the median eminence and that their density declined with age but was unaffected by short-term hormone treatment. EM results showed that vGluT2 immunoreactivity was extensively associated with large dense-core vesicles, suggesting a unique glutamatergic signaling pathway in GnRH terminals. Our results provide novel subcellular information about the intimate relationship between GnRH terminals and glutamate in the median eminence.