NEURAL AND GLIAL-MEDIATED EFFECTS OF GROWTH-FACTORS ACTING VIA TYROSINE KINASE RECEPTORS ON LUTEINIZING-HORMONE-RELEASING HORMONE NEURONS

It is becoming increasingly evident that the secretory activity of LHR H neurons is regulated not only by transsynaptic inputs but also by tr ophic molecules of glial and neuronal origin. The present experiments were undertaken to gain insights into the potential cell-cell mechanis ms by which basic fibroblast growth factor (bFGF) and transforming gro wth factor-alpha (TGF-alpha), two growth factors produced in the hypot halamus, may affect LHRH neuronal function. Northern blot analysis sho wed that the LHRH-producing cell line GT1-7 contains the messenger RNA (mRNA) encoding the type 1 fibroblast growth factor receptor (FGFR-1) but not that encoding the epidermal growth factor (EGF) receptors, wh ich mediates the biological actions of both TGFalpha and EGF. Ligand-i nduced receptor phosphorylation experiments demonstrated that GT1-7 ce lls possess biologically active FGFR-1s but not EGF receptors. Exposur e of the cells to bFGF resulted not only in FGFR-1 tyrosine phosphoryl ation, but also in tyrosine phosphorylation of phospholipase C-gamma, one of the initial enzymes in the intracellular signaling cascade init iated by FGFR activation. GT1-7 cells proliferated in response to this activation. Despite the presence of biologically active receptors, bF GF did not significantly stimulate release of the mature LHRH decapept ide. Instead, bFGF increased the steady-state levels of the mRNA encod ing the LHRH precursor processing endoprotease PC2, with a time course comparable to that of phorbol esters, suggesting that, as shown in th e companion paper, the actions of the growth factor on LHRH neurons in volve facilitation of the initial step in LHRH prohormone processing. The increase in PC2 gene expression was not accompanied by changes in LHRH mRNA levels. Unlike these direct actions of bFGF on GT-1 cells, T GFalpha appears to act indirectly via astroglial intermediacy. Exposur e of GT1-7 cells to TGFalpha or EGF failed to affect several parameter s of cellular activity including LHRH release, LHRH and PC2 mRNA level s, and cell proliferation. In contrast, astrocyte culture medium condi tioned by treatment with TGFalpha led to sustained stimulation of LHRH release with no changes in LHRH gene expression and a transient incre ase in PC2 mRNA levels. Although no definitive evidence for the presen ce of FGFR-1 in normal LHRH neurons could be obtained by either double immunohistochemistry or double in situ hybridization procedures, feta l LHRH neurons in primary culture responded to bFGF with neurite outgr owth. Thus, normal LHRH neurons may have an FGFR-1 content too low for detection by regular histochemical procedures, and/or detectable expr ession of the receptor may be confined to a much earlier developmental stage. The mitogenic effect of bFGF on GT1-7 cells supports this poss ibility and suggests a role for FGF in the cell proliferation events t hat precede acquisition of the LHRH neuronal phenotype. It appears tha t once this phenotype is established, bFGF may promote the differentia tion of LHRH neurons. The results also suggest that the secretory capa city of LHRH neurons develops under a dual trophic influence, one on p eptide processing exerted directly by bFGF on early neurons, and anoth er on LHRH release, exerted by TGFalpha via the intermediacy of astrog lial cells.