Immortalized Luteinizing Hormone-Releasing Hormone Neurons Show a Different Migratory Activity in Vitro*

Abstract

The development of two cell lines (GT1 and GN) of immortalized LHRH neurons has allowed an accurate study of the mechanisms controlling the synthesis and the secretion of LHRH. These cell lines, obtained in mice by genetic targeted tumorigenesis, retain many of the phenotypic characteristics of LHRH neurons. Of interest, GT1 cells derive from an hypothalamic tumor, whereas GN cells were obtained from a tumor localized in the olfactory bulb. The different origin of these cell lines lead to hypothesize that they might represent hypothalamic postmigratory neurons (GT1 cells), or LHRH neurons blocked at an early stage of their migration (GN cells). Using different experimental procedures, we found that the two cell subclones GT1-7 and GN11 express a different morphology and migratory behavior in vitro. In particular, we found that GN11 cells, but not GT1-7 cells, show the morphological shape of migrating neurons. When analyzing the spontaneous motility we found that only GN11 cells express a high capacity of migrating in a matrix of collagen gel. Moreover, in a chemomigratory assay GN11 cells did show a significant response to the chemotactic stimulus represented by the FBS. On the contrary, GT1-7 cells show very low spontaneous motility and appear insensitive to the FBS stimulus. These results suggest that the simultaneous use of the GT1-7/GN11 cells may represent an experimental tool for screening the factors possibly involved in the control of the migratory processes of LHRH neurons in normal and in pathological conditions, such as those due to their impaired migration, like it happens in Kallmann's syndrome.

Hepatocyte Growth Factor/Scatter Factor Facilitates Migration of GN-11 Immortalized LHRH Neurons

“…Previous studies have shown that GN11, but not GT1-7, cells show significant migratory activity and actively respond to the chemotactic stimulus represented by FBS (37).…”

“…This different derivation is indicative of different maturational stages of the two cell lines, as demonstrated by the fact that GT1 cells retain many characteristics of the mature hypothalamic LHRH neurons (34 -36). On the contrary, it has been found recently that GN11 cells retain the phenotypic characteristics of immature LHRH neurons and show high migratory activity in vitro, responding to fetal bovine serum (FBS) as a chemotactic stimulus (36,37).…”

“…Collagen gel was obtained as previously described (37), and 200 l were pipetted onto the bottom of a 24-well culture dish and left to set at room temperature. Cells aggregates were transferred over the cushion and then overlaid with an additional 200 l collagen.…”

Generation of kisspeptin-responsive GnRH neurons from human pluripotent stem cells

“…They not only determine the timing of puberty but ultimately integrate all reproductive inputs (Romanelli et al, 2004;Stevenson et al, 2013). In humans, the GnRHneuronal network is composed of a few hundred neurons distributed as clusters located from the preoptic area to the anterior hypothalamic regions making in vivo or ex vivo experimental studies of GnRH neurons extremely difficult (Maggi et al, 2000;Wang et al, 2015). The molecular ontogeny of GnRH neurons remains elusive.…”

The alpha-7 nicotinic acetylcholine receptor is involved in a direct inhibitory effect of nicotine on GnRH release: In vitro studies

“…In the present work we evaluated the presence of nicotinic receptors and the nature of a direct effect of nicotine on GnRH secretion in a neuronal isolated system using cell lines of immortalized hypothalamic GnRH neurons, an in vitro model widely used to study the mechanisms that control GnRH release in controlled conditions (Glidewell-Kenney et al, 2013;Gore and Roberts, 1997;Krsmanovic et al, 1998;Maggi et al, 2000;Mellon et al, 1990;Pal et al, 2007;Pimpinelli et al, 1999).…”