Role of Locally Produced Growth Hormone-Releasing Factor in Somatostatin Regulation by Fetal Rat Brain Cells in Culture

Abstract

To determine the possible physiological role of endogenous growth hormone-releasing factor (GRF) in the neuronal content and release of cerebral somatostatin (SS), we studied the effect of endogenous GRF blockade on the immunoreactive SS (IR-SS) content of cells and media in fetal rat cerebral cortical and hypothalamic cells in culture. Cells were cultured in minimum essential medium (MEM) with 10% fetal calf serum and 10% horse serum. After 7–10 days in vitro, media were replaced with MEM without sera containing anti-GRF immunoglobulins G (IgG) for 1, 5 or 24 h. Controls were incubated with equal amounts of IgG from normal rabbit serum (NRS). In another group of experiments, cells were incubated with GRF (10^–11 to 10^–7M) for 1 or 24 h. Long-term exposure (24 h) to anti-GRF IgG resulted in decreased media and intracellular IR-SS content, in both cerebral cortical and hypothalamic cells. 24 h treatment with GRF caused a dose-dependent increase in the IR-SS content of cells and media, the stimulatory action being abolished by the addition of anti-GRF to plates containing 10^–7M GRF. On the contrary, when cells were exposed to anti-GRF IgG for 1 h, IR-SS increased in the media as compared to the control group. Short-term incubation (1 h) with GRF (10^–9 to 10^–7M) resulted in a dose-dependent inhibition of IR-SS content in the cells and media. This inhibitory action was partially prevented by the addition of anti-GRF to plates containing 10^–7M GRF. Patterns of IR-SS cell and media content were qualitatively similar in both cerebral cortical and hypothalamic cells. In plates containing anti-GRF, the accumulation of IR-SS in the cells and media was lower than in the control groups (IgG from NRS) throughout the incubation time.

Growth Hormone-Releasing Factor Regulation by Somatostatin, Growth Hormone and Insulin-Like Growth Factor I in Fetal Rat Hypothalamic-Brain Stem Cell Cocultures

“…IR-SS was quantitated by RIA [23] using an antiserum against SSI4, raised in our laboratory by immunizing rabbits with SSI4 bound to cBSA with carbodiimide. Sensitivity was 2 pg/tube and cross-reactivity with SS28 was 20%.…”

Joint pituitary-hypothalamic and intrahypothalamic autofeedback construct of pulsatile growth hormone secretion

“…Unrestricted access of GHRH to SRIF-releasing neurons in this construct represses GH pulse amplitude by delayed feedforward on SRIF. The latter phenomenon is observed after in vivo intracerebroventricular infusion of biosynthetic GHRH in the mature male rat and after in vitro incubation of GHRH with fetal hypothalamic neurons or adult median-eminence neural tissue (1,2,11,27,29,32,33). In further simulations, we observed that the onset of continuous GHRH infusion initially induces high-amplitude GH pulses.…”

“…Distinguishing between the foregoing formulations (and possible hybrid models that include each) is not facile. Indeed, interconnectivity may be redundant or complementary, inasmuch as immunoreactive peptidergic neuronal terminals and cognate receptors for GHRH and SRIF are each detected in both the periventricular and the arcuate nucleus (1,2,8,11,24,27,29,32,33,42,47). In light of the foregoing complex issues, the present analyses underscore the expected complementarity of experimental data and model-assisted predictions to probe physiological regulation.…”

“…On the other hand, according to the simulations described here, failure to inhibit rebound GH release in the human administered high concentrations of a GHRH antagonist could denote central stimulatory effect directly on the hypothalamic GHRH-SRIF oscillator. The latter notion follows from extensive experimental data documenting (paradoxical) inhibition of GH secretion via GHRHspecific central neural stimulation of SRIF release (1,2,5,11,16,27,30,34) and structural evidence of bidirectional GHRH-SRIF connectivity (24,26). Simulated central (SRIF) access of infused GHRH also paradoxically suppresses pulsatile GH secretion (Fig.…”

“…Thus we compare the impact of constant systemic GHRH stimulation under the assumption of either no or variable access of infused GHRH to SRIF neurons. A truncated transcript of the GHRH gene is expressed in the foregoing two hypothalamic nuclei, as quantitated by RT-PCR, sequencing of the cDNA product, and Northern blot hybridization (47); and, central delivery of synthetic GHRH stimulates SRIF release acutely in vivo in the adult male rat and in vitro from incubated fragments of the median eminence (1,2,11,24,27,29,(32)(33)(34)37).…”