Gonadotropin-releasing hormone (GnRH) is a pivotal neuroendocrine regulator controlling reproductive functions. However, the scattered distribution of GnRH neurones in the mammalian brain has hindered studies on the development and differentiation of GnRH neurones. In the present study, we used the immortalized GnRH-producing GT1-1 cells to examine whether activation of protein kinase C (PKC) pathway with 12-O-tetradecanoyl-13-acetate (TPA) induces morphological and functional differentiation of GnRH neurones. TPA induced neurite outgrowth and inhibited proliferation of GT1-1 cells that were specifically antagonized by cotreatment of PKC inhibitor, calphostin C. The functional significance of TPA-induced differentiation of GT1-1 cells was manifested in part by the changes in the effects of gamma-aminobutyric acid (GABA) on intracellular Ca2+ levels. In untreated GT1-1 cells, activation of GABA-A receptor with 10 microM muscimol increased intracellular Ca2+ levels, whereas such stimulatory effects disappeared in GT1-1 cells bearing neurites. Accordingly, muscimol could not stimulate GnRH release in TPA-treated GT1-1 cells. To elucidate the molecular mechanism underlying TPA-induced neurite outgrowth, we performed differential display reverse transcription-polymerase chain reaction. Among several genes that are affected by TPA treatment, we found a significant induction of beta-catenin mRNA expression. Along with the rapid induction of beta-catenin protein levels, we observed that beta-catenin was reallocated from cell-cell adhesion sites to the growth cones within 3 h of TPA treatment. Transient transfection studies with green fluorescent protein as a reporter gene demonstrated that beta-catenin overexpression alone can promote neurite outgrowth in GT1-1 cells. Moreover, TPA was found to increase the transcription-activational roles of beta-catenin. Together, these data provide evidence that beta-catenin is involved in the TPA-induced functional differentiation of immortalized GnRH neurones.

The growth kinetics of passive films on iron and stainless steel (Fe−16.31%Cr) substrates in pH 8.4 borate buffer solution was investigated by using in situ specular X-ray reflectivity. The oxide growth rate decays exponentially with increasing oxide thickness consistent with the point defect model in which the electric field in the oxide is maintained constant during growth. In stainless steel, however, the electric field depends strongly on the applied potential, indicating that the oxide properties change as the applied potential varies. Using the electric field in the oxide and the observed saturation oxide thickness in a quasi steady state, we estimate the potential drop at the metal/oxide interface, in the oxide, and at the oxide/solution interface.

The chemical depth profile of the passive oxides formed on a stainless-steel surface (type 430, Fe–16.3%Cr) in pH 8.4 borate solution was obtained quantitatively by in situ anomalous x-ray reflectivity. The passive film consists of a Cr oxide inner layer and a Fe∕Cr oxide outer layer. The absence of Fe oxide underneath Cr oxide indicates that Cr oxide provides the passivity in stainless steel by inhibiting oxygen anion diffusion. The oxide grows by limited metallic cation transfer. The passivity breaks down in transpassive regime due to the oxidation of the Cr oxide to a soluble oxide.

In situ specular x-ray reflectivity was applied to study the growth kinetics of passive oxide films on iron and stainless steel substrates in pH 8.4 borate buffer solution. Under electrical potential from 0 to 800 mV, the growth rate of oxide films decreases exponentially in thickness following the direct logarithmic growth law predicted in the point defect model. The electric field in the oxide on iron is independent of the applied potentials consistent with the point defect model. In stainless steel, however, the electric field depends strongly on the applied potential indicating that the oxide properties change as the applied potential varies.