Expression of gonadotropin-releasing hormone receptors and autocrine regulation of neuropeptide release in immortalized hypothalamic neurons.

Abstract

The hypothalamic control of gonadotropin secretion is mediated by episodic basal secretion and midcycle ovulatory surges of gonadotropin-releasing hormone (GnRH), which interacts with specific plasma membrane receptors in pituitary gonadotrophs. Similar GnRH receptors and their mRNA transcripts were found to be expressed in immortalized hypothalamic neurons, which release GnRH in a pulsatile manner in vitro. Activation of these neuronal GnRH receptors elicited dose-related intraceDlular Ca2+ concentration responses that were dependent on calcium mobilization and entry and were inhibited by GnRH antagonists. Exposure of perifused neurons to a GnRH agonist analog caused a transient elevation of GnRH release and subsequent suppression of the basal pulsatile secretion. This was followed by dose-dependent induction of less frequent but larger GnRH pulses and ultimately by single massive episodes of GnRH release. The ability of GnRH to exert autocrine actions on its secretory neurons, and to promote episodic release and synchronized discharge ofthe neuropeptide, could reflect the operation of the endogenous pulse generator and the genesis of the preovulatory GnRH surge in vivo.The major regulator of reproduction in mammals, gonadotropin-releasing hormone (GnRH), is produced by neuronal cells located in the preoptic area and adjacent sites in the rostral portion of the hypothalamus and secreted into the hypophyseal portal vessels at the median eminence (1, 2). The secretion of GnRH occurs in an episodic manner due to the activity of a hypothalamic GnRH pulse generator (3). Recently, pulsatile neuropeptide secretion was found to be an intrinsic property of GnRH neuronal networks and to depend on voltage-sensitive Ca2+ influx for its maintenance (4-7). The activity of the GnRH pulse generator is influenced by several factors, including endothelin, N-methyl-D-aspartate, opiates, -aminobutyrate, and a-adrenergic input, as well as estrogens and androgens (8)(9)(10)(11)(12)(13)(14)(15)(16)). In addition, GnRH has been proposed to exert an inhibitory action on its own secretion (17-19); however, the mechanism and circuitry of such an ultrashort loop feedback effect have not yet been defined. We now report that GnRH acts directly on immortalized GnRH neuronal cells to regulate its own secretion and that this autocrine action of the neuropeptide is associated with activation of calcium-mobilizing GnRH receptors expressed in its cells of origin. MATERIALS AND METHODSBinding Studies. Plasma membrane receptors for GnRH were analyzed by binding studies with 125I-labeled des- Glyl0[D-Ala6]GnRH N-ethylamide (Hazleton LaboratoriesAmerica, Vienna, VA). The radioligand (150 pM) and nonradioactive peptides were added in 100-,ul aliquots to monolayers of GT1-7 cells (generously provided by Richard Weiner, University of California, San Francisco) cultured in 12-well Falcon plates at 24°C. After incubation to equilibrium for 90 min at room temperature, the cells were washed three times with ice-cold phosphate-buffered saline/0.1% bo...

Expression and Function of Gonadotropin-releasing Hormone (GnRH) Receptor in Human Olfactory GnRH-secreting Neurons

“…be important for its pulsatile release (39,40). Our results in human olfactory FNC-B4 cells are in agreement with those of the aforementioned studies.…”

Dependence of Gonadotropin-releasing Hormone-induced Neuronal MAPK Signaling on Epidermal Growth Factor Receptor Transactivation

“…These cells retain many of the characteristics of the native GnRH neurons, including the ability to maintain pulsatile GnRH release (1,3). Recent evidence suggests that the autocrine action of GnRH on hypothalamic GnRH neurons is involved in the mechanism of pulsatile GnRH secretion (3).…”

“…The major signal transduction pathways in cells expressing GnRH receptors are initiated by activation of phospholipase C. The consequent calcium (Ca 2ϩ ) mobilization and activation of protein kinase C (PKC) by GnRH are key elements in the hypothalamic control of gonadotropin secretion from the anterior pituitary gland (1,3,5). Activation of PKC and Ca 2ϩ mobilization during GnRH receptor stimulation are also responsible for mediating downstream signals leading to activation of extracellularly regulated mitogen-activated protein kinases (ERK1/2 MAPKs) that transmit signals from the cell surface to the nucleus to regulate transcriptional and other processes (7)(8)(9)(10)(11)(12)(13).…”

“…In addition to regulating pituitary gonadotropin release, GnRH has extrapituitary actions in neural and nonneural tissues and in several types of tumor cells (1). Immortalized GnRH-producing neurons (GT1-7 neurons) express several G protein-coupled receptors (GPCRs), including those for GnRH and luteinizing hormone/ human chorionic gonadotropin (2,3), as well as ␣-and ␤-adrenergic (4), muscarinic (5), and serotonergic receptors (6). These cells retain many of the characteristics of the native GnRH neurons, including the ability to maintain pulsatile GnRH release (1,3).…”

“…Immortalized GnRH-producing neurons (GT1-7 neurons) express several G protein-coupled receptors (GPCRs), including those for GnRH and luteinizing hormone/ human chorionic gonadotropin (2,3), as well as ␣-and ␤-adrenergic (4), muscarinic (5), and serotonergic receptors (6). These cells retain many of the characteristics of the native GnRH neurons, including the ability to maintain pulsatile GnRH release (1,3). Recent evidence suggests that the autocrine action of GnRH on hypothalamic GnRH neurons is involved in the mechanism of pulsatile GnRH secretion (3).…”