Immunohistochemical Demonstration of proGnRH and GnRH in the Preoptic-Basal Hypothalamus of the Primate

Abstract

An antiserum (ARK-1) specific to the gonadotropin-releasing hormone precursor (proGnRH) was produced by immunizing with a synthetic peptide (proGnRH 6–16; Gly-Leu-Arg-Pro-Gly-Gly-Lys-Arg-Asp-Ala-Glu) which bridges the proteolytic cleavage site of proGnRH. When used in the radioimmunoassay, ARK-1 bound 25% of the iodinated 5–16 fragment at a 1:30,000 dilution with a sensitivity of 1 pg/tube. Using immunohistochemical techniques, we observed that in serial and the same sections through the preoptic-basal hypothalamus (POA-BH), the precursor molecule was primarily present in the cell soma, whereas GnRH was found in the cell soma, nerve fibers, and terminals of the same neurons. These data indicate that the processing of proGnRH to biologically active peptides (e.g., GnRH) in the rhesus macaque and the baboon POA-BH primarily occurs in the cell soma.

Inward Rectification (Ih) in Immunocytochemically‐ldentified Vasopressin and Oxytocin Neurons of Guinea‐Pig Supraoptic Nucleus

“…The slice was sectioned at 16 pm on a cryostat, mounted on Po1)-l-lysine coated slides and stored frozen at a temperature of -20 "C. All of the sections were rinsed in PB and alternate sections were incubated with OT (14) or AVP (15) antisera at 1:1,000 and 1:2,000 dilutions, respectively. Next, the sections were rinsed in PB and reacted for 1 h with a mixture of StWptavidin-FITC diluted 1:600 and IgG conjugated to Texas Red diluted 1:50 (16). The last step was repeated once following a 7-min wash in PB.…”

Effects of Ovariectomy on GnRH mRNA, proGnRH and GnRH Levels in the Preoptic Hypothalamus of the Female Rat

“…extracts were resuspended by sonication in phos phate-buffered saline and aliquoted for duplicate determinations or serial dilutions. We measured GnRH with the El-14 antisera, a conformational antisera which does not recognize any form of preproGnRH [36]. The assay procedure and the specificity of the El-14 antiserum against GnRH have been published [9].…”

“…The ARK-2 antiserum was generated against proGnRH 6-16 (human) as described previously [36], but differs from ARK-1 in that the sera was obtained from a different rabbit and was found to stain the cell soma of GnRH neurons in the rat ( fig. 3 and [35]).…”

“…We developed a specific antisera to proGnRH, designated as ARK-I, by using a peptide sequence (amino acids 6-16 of proGnRH) which spans the bridging site between GnRH and GAP. Using the ARK-1 antiserum in concert with El-14, which is a conformational antiserum specific for GnRH [9], we demonstrated that (I) proGnRH is colocalized with GnRH in the soma of neurons throughout the POA and BH of the monkey, and (2) processing of proGnRH to GnRFI begins in the cell soma before transport down to the nerve terminal [36],…”

Effect of maternal deprivation on the gonadotrophin-releasing hormone (GnRH) and GnRH-associated peptide neurobiology in lambs during the transition from infancy to prepuberty.

“…The processing of proGnRH to biologically active peptides, both the GnRH and GAP, begins in the cell soma and then progresses in secretory granules during axonal transport to the nerve terminals, where the proteolytic cleavage products are accumulated and released. Thus, in the perikarya we can find preproGnRH, proGnRH, GnRH and/or GAP, whereas in the nerve terminals we can recognize GnRH and GAP (the storage forms within the secretory vesicles) and some lower molecular weight GAP-like peptides according to the current neurosecretory activity of GnRH neurons [11,[24][25][26][27][28][29].…”