| Summary: | 碩士 === 國立成功大學 === 藥理學研究所 === 93 === Pituitary adenylate cyclase-activating polypeptide (PACAP), a neuropeptide, is observed in the adrenal gland and sympathetic ganglia to regulate catecholamine (CA) synthesis and release. Both of PACAP and glucocorticoid have been reported to elevate CA content in the line from stimulating biosynthesis. We assumed that glucocorticoid could modulate PACAP gene expression to increase CA content in adrenal chromaffin cell. In this study, we found that PACAP mRNA in rat pheochromocytoma cell (PC12 cell) examined by RT-PCR was stimulated after treatment of dexamethasone (DEXA). This effect was blocked by pre-incubation with RU486(Mifepristone), a glucocorticoid receptor(GR) antagonist. In addition, transcriptional inhibitor actinomycin D was used to confirm that DEXA could elevate PACAP gene expression through activating its transcriptional activity.
Poly (ADP -ribose ) polymerase (PARP), an enzyme that exists in the nucleus, has been reported to play an important role in regulating the transcription of particular sets of genes. Thus, we investigated the role of PARP on GR modulating PACAP gene. PC12 cells were treated with two different PARP inhibitors, 3-aminobenzamide (3-AB) and 1,5- isoquinolinediol (ISO). The effect of DEXA on PACAP gene were inhibited after treatment of 3-AB or ISO, respectively.
Dopamine (DA) content of PC12 cells was measured with HPLC after incubation for 24h with DEXA. DEXA increases DA content of PC12 cells and this effect could be blocked by pre-incubation with PACAP type 1 (PAC1) receptor antagonist PACAP(6-38). This result indicates that DEXA could increase CA content in PC12 cells via modulating PACAP gene. Therefore, we proved that DEXA has the ability to stimulate PACAP gene transcription via activating GR in PC12 cells, resulted in the increase of CA content.
β-amyloid(Aβ) is a neurotoxic peptide, deposits of Aβ peptide are found in plaques in the brains with Alzheimer’s disease (AD). It has been reported that the intracellular and extracellular deposits of Aβ may lead to neuronal cell death and apoptosis. In neuron cell, PACAP is a neurotropic factor, which could promote cell survival. In this study, we found that incubation of PC12 or rat brain astrocyte type 1 (RBA1) with Aβ (1μM) could inhibit the endogenous PACAP expression. The inhibition effect of Aβ could be rescued by DEXA, and the effect of DEXA was blocked by RU486.
It has been reported that both 17β-estradiol (E2) and ginseng extract – gensenoside Rh2 have neuroprotective effect. In this experiment, treatment of RBA1 cells with E2 or Rh2 could stimulate PACAP gene expression, respectively. And both of them could reverse Aβ reduced PACAP gene. Thus, our results revealed that, in PC12 cells, DEXA could reverse Aβ reduced PACAP gene via activating GR. In RBA1 cells, E2 and Rh2 could attenuate Aβ induced toxicity via reversing Aβ reduced PACAP gene expression.
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