The purpose of this study was to evaluate intentional replantation of vertically fractured roots reconstructed with dentin-bonded resin. Twenty vertically fractured teeth were extracted intentionally and reconstructed with 4-META/MMA-TBB dentin-bonded resin. After reconstruction, the teeth were replanted into the original sockets. The replanted teeth were evaluated by clinical criteria and radiographic examination. The observation periods ranged between 4 and 45 months. Longevity was calculated by the Kaplan-Meier method, and factors that had significant influence on the longevity were analyzed with the Log rank test at a 95% level of confidence. From this short-term observation, 14 of 20 cases were functioned in the oral environment. Of these 14 cases, 6 could be considered truly successful and 8 cases needed further observation. The other six cases were total failures and were extracted. The longevity was calculated as 83.3% at 12 months and 36.3% at 24 months. Teeth with thin roots or with longitudinal fractures extending over 2/3 of the root from the cervical portion toward the apex showed significantly lower longevity. It was concluded that this method had the potential to preserve vertically fractured teeth.

Recent studies suggest that the AMP-activated protein kinase (AMPK) signaling in the hypothalamus is the master regulator of energy balance. We reported in previous studies that glucocorticoids play a permissive role in the regulation of orexigenic neuropeptide Y (Npy) gene expression in the arcuate nucleus. In this study, we examined whether any cross talk occurs between glucocorticoids and AMPK signaling in the hypothalamus to regulate Npy as well as agouti-related peptide (Agrp) gene expression in the arcuate nucleus. In the hypothalamic organotypic cultures, the addition to the medium of the AMPK activator, 5-aminoimidazole-4-carboxamide-1-b-d-ribofuranoside, increased phosphorylated AMPK (p-AMPK) as well as phosphorylated acetyl-coenzyme A carboxylase (p-ACC) in the explants, accompanied by significant increases in Npy and Agrp gene expression in the arcuate nucleus. The incubation with dexamethasone (DEX) also activated AMPK signaling in the explants, accompanied by significant increases in Npy and Agrp gene expression in the arcuate nucleus. The addition of the AMPK inhibitor compound C to the medium, which blocked increases of p-AMPK and p-ACC by DEX, significantly attenuated Npy and Agrp gene expression stimulated by DEX. Furthermore, p-AMPK and p-ACC levels in the arcuate nucleus were significantly decreased in adrenalectomized rats compared with sham-operated rats, and a replacement of glucocorticoids reversed the AMPK signaling in adrenalectomized rats. Thus, our data demonstrated that glucocorticoids up-regulate the Npy and Agrp gene expression in the arcuate nucleus through AMPK signaling, suggesting that the activation of the hypothalamic APMK signaling by glucocorticoids might be essential to the energy homeostasis.

No abstract

Neuropeptide Y (NPY) in the arcuate nucleus is an orexigenic hormone of which levels are regulated by humoral as well as neural signals. In this study, we examined the regulation of NPY gene expression in the arcuate nucleus in hypothalamic organotypic cultures. Dexamethasone (DEX) (10 Ϫ9 to 10 Ϫ7 M) significantly increased NPY mRNA expression, and the effects were not influenced by coincubation with the sodium channel blocker tetrodotoxin (TTX), indicating that the action of DEX is independent of action potentials. Conversely, insulin (10 Ϫ11 to 10 Ϫ9 M) significantly inhibited NPY expression stimulated by DEX, and the inhibitory action of insulin was abolished in the presence of TTX. Because GABA and its receptors are expressed in the arcuate nucleus in vivo, we examined whether GABAergic systems were involved in the insulin action. The GABA B agonist baclofen significantly inhibited NPY expression stimulated by DEX, and the inhibitory action of insulin was completely abolished in the presence of either the GABA A antagonist bicuculline or the GABA B antagonist CGP35348 ( p-3-aminopropyl-p-diethoxymethyl phosphoric acid). Furthermore, increases in the GABA-synthesizing enzyme glutamic acid decarboxylase 65 (GAD65) mRNA expression preceded decreases in NPY mRNA expression in the arcuate nucleus in the cultures. Experiments in vivo also demonstrated that increases in GAD65 mRNA expression in the arcuate nucleus preceded decreases in the NPY mRNA expression in a fasting-refeeding paradigm and that intracerebroventricular injection of insulin increased GAD65 mRNA expression in the arcuate nucleus in fasted rats. These data suggest that insulin inhibits NPY gene expression in the arcuate nucleus through GABAergic systems.

Ghrelin, which was identified from the rat stomach, is a potent stimulant for food intake. Several lines of evidence suggest that the orexigenic action of ghrelin is mediated via the neuropeptide Y (NPY) neurons in the arcuate nucleus, although the detailed mechanisms by which ghrelin stimulates NPY neurons are not clear. In this study, we examined the gene regulation of NPY and agouti-related peptide (AGRP), another orexigenic peptide synthesized in the NPY neurons, in the arcuate nucleus by ghrelin in hypothalamic organotypic cultures. Incubation of the hypothalamic explants with ghrelin significantly increased NPY and AGRP mRNA expression in the presence, but not absence, of dexamethasone. Glucocorticoids were also necessary for ghrelin action in vivo because an intracerebroventricular injection of ghrelin significantly increased NPY and AGRP mRNA expression in the arcuate nucleus only in sham-operated, but not in adrenalectomized rats. The stimulatory effects of ghrelin on gene expression were not blocked by a sodium channel blocker tetrodotoxin in the organotypic cultures. Ghrelin also increased NPY heteronuclear (hn) RNA expression, the first transcript that has been used as an indicator for gene transcription. The stimulatory effects of ghrelin on NPY gene expression were abolished in the presence of cycloheximide, which blocks translation, suggesting that de novo protein synthesis is required for ghrelin action. These data suggest that ghrelin stimulates NPY and AGRP gene expression independently of action potentials only in the presence of glucocorticoids. Furthermore, our data demonstrate stimulatory action of ghrelin on NPY gene transcription, which requires de novo protein synthesis.

No abstract

Although acute decreases in plasma volume are known to enhance the osmotically induced arginine vasopressin (AVP) release, it is unclear whether there is also such interaction at the level of gene transcription. It also remains to be established how sustained changes in plasma volume affect the osmoregulation. In this study, we examined how acute and chronic decreases in blood volume affected the osmoregulation of AVP release and gene transcription in rats. Acute hypovolemia was induced by intraperitoneal injection of polyethylene glycol (PEG), and chronic hypovolemia was induced by 3 days of water deprivation (WD) or 12 days of salt loading (SL). Rats were injected with isotonic or hypertonic saline, and plasma AVP levels and AVP heteronuclear (hn)RNA expression in the supraoptic and paraventricular nuclei, an indicator of gene transcription, were examined in relation to plasma osmolality in each group. Plasma AVP levels were correlated with plasma Na levels in all groups. Whereas the regression lines relating plasma AVP to Na were almost identical among control, WD, and SL groups, the thresholds of plasma Na for AVP release were significantly decreased only in the PEG group. AVP hnRNA levels were also correlated with plasma Na levels in control and PEG groups, and the thresholds were significantly decreased in the PEG group. In contrast, there was no significant correlation of AVP hnRNA and plasma Na levels in WD and SL groups. Thus it was demonstrated that acute and chronic reduction in plasma volume affected the osmoregulation of AVP release and gene transcription in different ways.

Arginine vasopressin (AVP) in the parvocellular neurons of the paraventricular nucleus (PVN) is known to play an important role in the hypothalamo-pituitary-adrenal axis. In the present study, we examined how cAMP and glucocorticoids regulate AVP gene expression in the parvocellular neurons of the PVN in rat hypothalamic organotypic cultures with in situ hybridization. AVP heteronuclear (hn) RNA, an indicator for gene transcription, was induced in the PVN with incubation of forskolin as reported previously, and AVP mRNA was increased by forskolin in the presence of the gene transcription inhibitor 5,6-dichloro-1-D-ribofuranosylbenzimidazole (DRB). These data indicate that cAMP could increase not only gene transcription but also mRNA stability. Dexamethasone treatment, in contrast, significantly decreased AVP mRNA expression levels in the PVN, but this inhibitory action was abolished in the presence of DRB or the sodium channel blocker tetrodotoxin (TTX). However, when the hypothalamic slices were treated with forskolin, dexamethasone decreased AVP mRNA expression even in the presence of DRB and/or TTX. Furthermore, AVP hnRNA expression induced by forskolin was attenuated by dexamethasone treatment in the presence of TTX. These data indicate that dexamethasone could act on AVP cells independently of action potentials to decrease mRNA stability and to suppress AVP gene transcription during stimulation by cAMP. Thus, it was demonstrated that: (1) cAMP upregulates AVP gene transcriptionally and post-transcriptionally, (2) the mode of action of glucocorticoids was dependent on whether the cells were stimulated by cAMP, and (3) the interactions between cAMP and glucocorticoids encompass both gene transcription and mRNA stability.