Abstract-Spontaneous beating of rabbit sinoatrial node cells (SANCs) is controlled by cAMP-mediated, protein kinase A-dependent local subsarcolemmal ryanodine receptor Ca 2ϩ releases (LCRs). LCRs activated an inward Na ϩ /Ca 2ϩ exchange current that increases the terminal diastolic depolarization rate and, therefore, the spontaneous SANC beating rate. Basal cAMP in SANCs is elevated, suggesting that cAMP degradation by phosphodiesterases (PDEs) may be low. Surprisingly, total suppression of PDE activity with a broad-spectrum PDE inhibitor, 3Ј-isobutylmethylxanthine (IBMX), produced a 9-fold increase in the cAMP level, doubled cAMP-mediated, protein kinase A-dependent phospholamban phosphorylation, and increased SANC firing rate by Ϸ55%, indicating a high basal activity of PDEs in SANCs. A comparison of specific PDE1 to -5 inhibitors revealed that the specific PDE3 inhibitor, milrinone, accelerated spontaneous firing by Ϸ47% (effects of others were minor) and increased amplitude of L-type Ca 2ϩ current (I Ca,L ) by Ϸ46%, indicating that PDE3 was the major constitutively active PDE in the basal state. PDE-dependent control of the spontaneous SANC firing was critically dependent on subsarcolemmal LCRs, ie, PDE inhibition increased LCR amplitude and size and decreased LCR period, leading to earlier and augmented LCR Ca 2ϩ release, Na ϩ /Ca 2ϩ exchange current, and an increase in the firing rate. When ryanodine receptors were disabled by ryanodine, neither IBMX nor milrinone was able to amplify LCRs, accelerate diastolic depolarization rate, or increase the SANC firing rate, despite preserved PDE inhibition-induced augmentation of I Ca,L amplitude. Thus, basal constitutive PDE activation provides a novel and powerful mechanism to decrease cAMP, limit cAMP-mediated, protein kinase A-dependent increase of diastolic ryanodine receptor Ca 2ϩ release, and restrict the spontaneous SANC beating rate. Key Words: sinoatrial node Ⅲ phosphodiesterase Ⅲ ryanodine receptors Ⅲ local Ca 2ϩ release T he sinoatrial (SA) node is the primary physiological pacemaker of the heart. The pacemaker action potential (AP) is initiated within the SA node center and then propagates to the atria and ventricle to initiate contraction. 1,2 Our recent studies have demonstrated that spontaneous firing of SA node pacemaker cells (SANCs) is controlled by local subsarcolemmal Ca 2ϩ releases (LCRs) from ryanodine receptors (RyR) that occur during the second half of spontaneous diastolic depolarization (DD), just prior to the AP upstroke. 3 LCRs activate inward Na ϩ /Ca 2ϩ exchange (NCX) current that accelerates the rate of DD, leading to an earlier occurrence of the subsequent spontaneous AP, ie, to an increase in the beating rate. 3 Although LCRs do not require membrane depolarization, 4 they are critically dependent on levels of cAMP and cAMP-mediated, protein kinase (PKA)-dependent phosphorylation, both of which are markedly higher in SANCs than in atrial or ventricular myocytes, because of constitutive activation of adenylyl cyclases (ACs). 5 T...

The kidney was recognized as a dominant organ for uric acid excretion. The main aim of the study demonstrated intestinal tract was an even more important organ for serum uric acid (SUA) lowering. Sprague-Dawley rats were treated normally or with antibiotics, uric acid, adenine, or inosine of the same molar dose orally or intraperitoneally for 5 days. Rat’s intestinal tract was equally divided into 20 segments except the cecum. Uric acid in serum and intestinal segment juice was assayed. Total RNA in the initial intestinal tract and at the end ileum was extracted and sequenced. Protein expression of xanthine dehydrogenase (XDH) and urate oxidase (UOX) was tested by Western blot analysis. The effect of oral UOX in lowering SUA was investigated in model rats treated with adenine and an inhibitor of uric oxidase for 5 days. SUA in the normal rats was 20.93±6.98 μg/ml, and total uric acid in the intestinal juice was 308.27±16.37 μg, which is two times more than the total SUA. The uric acid was very low in stomach juice, and attained maximum in the juice of the first segment (duodenum) and then declined all the way till the intestinal end. The level of uric acid in the initial intestinal tissue was very high, where XDH and most of the proteins associated with bicarbonate secretion were up-regulated. In addition, SUA was decreased by oral UOX in model rats. The results suggested that intestinal juice was an important pool for uric acid, and intestinal tract was an important organ for SUA lowering. The uric acid distribution was associated with uric acid synthesis and secretion in the upper intestinal tract, and reclamation in the lower.

Background Reversible N 6 -methyladenosine (m 6 A) modifications in messenger RNAs can be categorized under the field of “RNA epigenetics.” However, the potential role of m 6 A-related genes in gastric cancer (GC) prognosis has not been systematically researched. Aims This study was aimed at providing insights into the prognostic role of m 6 A-related gene expression, at both mRNA and protein levels. Methods Kaplan–Meier (KM) plotter database and The Cancer Genome Atlas (TCGA) database were used to explore the prognostic significance of individual m 6 A-related genes in overall survival (OS) and progression-free survival at the mRNA level. For independent validation, the protein level of genes significantly associated with prognosis in both databases was further detected in 450 paired GC and corresponding adjacent non-tumor tissues using tissue microarray (TMA)-based immunohistochemistry (IHC). The relationship between the FTO and ALKBH1 expression and the clinicopathological characteristics was explored. Results Among nine m 6 A-related genes, aberrantly high mRNA expression of FTO and ALKBH1 was associated with poor OS in the KM and TCGA cohorts. However, the TMA-IHC indicated that protein expression of FTO and ALKBH1 was markedly downregulated in GC tissues. A lower protein level of ALKBH1 was closely correlated with larger tumor sizes (≥ 5 cm) and more advanced TNM stages, while lower FTO protein expression was associated with shorter OS in GC patients. Conclusions Aberrant expression of demethylase genes, FTO and ALKBH1 , has a distinct prognostic value in GC patients, indicating that FTO and ALKBH1 may play vital roles in GC progression and metastasis. Electronic supplementary material The online version of this article (10.1007/s10620-018-5452-2) contains supplementary material, which is available to authorized users.

Basal cardiac pacemaker function is regulated by concurrent PDE3+PDE4 activation which operates in a synergistic manner via decrease in cAMP/PKA phosphorylation, suppression of LCR parameters, and prolongation of the LCR period and spontaneous SANC cycle length.