Neurochemical Research

A modification of the tyrosine hydroxylase assay is described in which ascorbate, rather than 2-mercaptoethanol or dihydropteridine reductase with NADPH, is used as the reductant. Enzyme activity is 3–4 times higher with ascorbate than with the other reducing agents. Low blanks are obtained with the ascorbate system provided that catalase is also included. The tissue distribution and kinetic activation of the enzyme have been studied with the ascorbate assay. The results obtained are consistent with the biological and regulatory properties of the enzyme which have been determined with the other reducing systems.

C-Jun and c-Fos belong to the family of immediate early genes. Apart from their role as transcription factors, a basal expression was shown for them in central nervous system tissues. The expression of c-Jun and c-Fos in sensory ganglia of guinea pig, rat and murine sensory ganglia was examined under normal, unstimulated conditions by quantitative double-immunohistochemistry. 4.6 ± 2.8% of neuron-specific protein gene-product 9.5 -positive cells in nodose ganglia, 51.6 ± 2.1% in jugular ganglia, 46.4 ± 3.0% in trigeminal ganglia and 42.5 ± 1.3% of cervical dorsal root ganglia neurons were positive for c-Jun in the guinea pig (less than 1% for c-Fos). In rat and mouse, less than 1% of the sensory neurons exhibited c-Jun and c-Fos-immunoreactivity. The high basal expression of c-Jun in guinea pig sensory neurons suggests that in this species the presence of c-Jun does not only depend on specific stimulation and is not exclusively associated with neuronal plasticity of gene expression and functional changes.

Microglia, innate immune cells of the brain, constantly monitor the dynamic changes of the brain microenvironment under physiological conditions and respond in time. Growing evidence suggests that microglia-mediated neuroinflammation plays an important role in the pathogenesis of Alzheimer’s disease. In this study, we investigated that the expression of IFITM3 was significantly upregulated in microglia under the Aβ treatment, and knockdown of IFITM3 in vitro suppressed the M1-like polarization of microglia. Moreover, IFITM3 was regulated by cGAS-STING signaling in activated microglia, and inhibition of cGAS-STING signaling reduces IFITM3 expression. Taken together, our findings suggested that the cGAS-STING-IFITM3 axis may be involved in Aβ-induced neuroinflammation in microglia.

The basic mechanisms of nerve protection by estrogen remain to be clarified. This study was undertaken to confirm estrogen-induced insulin-like growth factor 1 (IGF-1) mRNA expression in the immortalized rat hippocampal cell H19-7 using a real-time quantitative polymerase chain reaction (PCR) assay, which has considerably increased accuracy and rapidity over other current methods. Upon stimulation by estradiol, the copy number of ERα mRNA showed a 1.4-fold increase, and that of IGF-1 mRNA showed a 38.5-fold increase when compared with the control value. ICI182,780 inhibited the estradiol-induced upregulation of ERα RNA completely, while it inhibited estradiol-stimulated IGF-1 mRNA expression partially. The increase of the copy number of IGF-1 mRNA was accomanied by enhancement of IGF-1 protein as observed by Western blot analysis.

Studies have demonstrated that berberine exhibits the antineoplastic action in rat model. Rat glial tumor cells also have been shown to have N-acetyltransferase activity. In this study, we reported the effects of berberine on arylamine N-acetyltransferase (NAT) activity, gene expression, and DNA adduct formation in human brain tumor cell lines (G95/VGH and GBM 8401). The activity of NAT (N-acetylation of substrate) was measured and determined by high-performance liquid chromatography (HPLC) assaying for the amounts of acetylated 2-aminofluorene (AF) and nonacetylated AF. Human brain tumor cells (G9T/VGH and GBM 8401) were used for examining NAT activity and gene expression and AF-DNA adduct formation. NAT gene expression was determined by polymerase chain reaction (PCR) for the levels of mRNA NAT in both examined cells lines. The amounts of AF-DNA adducts were also determined and quantities by HPLC. The results demonstrated that NAT activity, levels of mRNA NAT1 and AF-DNA adduct formation in both examined cell were inhibited and decreased by berberine in a dose-dependent manner. The apparent values of Km and Vmax from NAT of both examined cells were also determined with or without berberine cotreatment. The data also indicated that berberine decreased the apparent values of Km and Vmax. These effects also indicate that berberine is a uncompetitive inhibitor.

Repeated morphine administration results in analgesic tolerance. However, the underlying mechanism of morphine analgesic tolerance remains unclear. NADPH-oxidase 2 (NOX2) is the first discovered NADPH oxidase, which mainly functions to produce reactive oxygen species. Its specific role in morphine tolerance has not been fully investigated. In this work, we found that chronic morphine administration significantly increased the expression of NOX2 in spinal cord. Pretreatment of NOX2 inhibitor blocked the upregulation of NOX2 and autophagy markers, including LC3B and P62, and consequently the development of morphine tolerance. NOX2 and LC3B were both colocalized with NeuN in spinal dorsal horn in morphine-tolerant rats. Our results suggest that the increased autophagy activity in spinal neurons promoted by NOX2 activation contributes to the development of morphine tolerance. NOX2 may be considered as a new therapeutic target for morphine tolerance.