Endocrinology, Vol 125, 1991-1998, Copyright © 1989 by Endocrine Society

ARTICLES

Growth hormone-releasing factor production by fetal rat cerebrocortical and hypothalamic cells in primary culture

G Fernandez, L Cacicedo, MJ Lorenzo, MT de los Frailes and F Sanchez Franco
Servicio de Endocrinologia, Hospital Ramon y Cajal, Madrid, Spain.

Recent data from our laboratory and others have shown radioimmunoassayable GRF (IR-GRF) in the rat brain cortex. In the present study the ontogenesis of immunoreactive rat(r) GRF (IR-GRF) in long term dissociated fetal rat cerebrocortical and hypothalamic cell cultures and the regulation of its secretion by potassium depolarization and calcium channel-blocking agents were investigated. The chromatographic profiles of IR-rGRF from cell extracts were determined and compared with those from in vivo cerebrocortical and hypothalamic tissues. Mechanically dispersed cultured telencephalic and diencephalic cells from 17- to 21-day-old fetal rats showed a progressive increase in IR-rGRF, reaching maximum values (media plus cells) between 775-1020 (pg/mg protein) in hypothalamic cells on days 10-20 and between 450 and 950 pg/mg protein in cortical cells on days 25-30. IR-rGRF from acidic extracts of cells and adult cortical and hypothalamic tissues adsorbed onto octadecylsilyl-silica columns corresponds primarily to rGRF-(1-43)OH on HPLC. In gel filtration chromatography, almost all IR-rGRF from cultured cerebrocortical cells and fetal and adult cortical tissues coeluted with rGRF-(1-43)OH. IR- rGRF from cultured hypothalamic cells showed an additional component with a higher mol wt eluting in the void volume. To determine the influence of membrane depolarization of rGRF release, potassium (K+) concentrations in the medium were increased to 30 and 56 mM, inducing a marked increase in medium rGRF concentrations. Verapamil, a Ca2+ channel blocker (20 microns) reverses the effect of 56 mM potassium depolarization, suggesting that it is at least partially dependent upon Ca2+ transport. These data indicate that fetal rat cerebrocortical and hypothalamic cells in culture produce and release rGRF in response to depolarizing agents. The presence of rGRF in cortical tissue suggests that there are other physiological roles besides those implicated in the stimulation of GH secretion.

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