ブタの卵母細胞と初期胚におけるグルコース-6-リン酸脱水素酵素の組織化学的研究

体外で成熟過程にあるブタの卵母細胞、1細胞期から拡張胚盤胞期までの体外受精に由来するブタの卵子と初期胚について、グルコース-6-リン酸脱水素酵素(G-6-PDH)の活性を組織化学的に検出した。G-6-PDH活性は、胞状卵胞から採取直後の卵母細胞では強く、この強い活性は培養後44時間の卵母細胞まで維持された。また、この酵素の活性は、媒精後の受精卵子でも強かったが、2細胞期から16細胞期の胚ではやや弱まり、弱度ないし強度となった。桑実胚期以降、酵素活性はさらに弱まり、活性を示さない胚も出現するとともに、拡張胚盤胞では活性はまったくみられなかった。以上の結果と従来のステロイド代謝能の結果とを考え合わせると、成熟過程にあるブタの卵母細胞と発生過程にあるブタの初期胚は、G-6-PDHの作用によって産生したNADPHをステロイドの生合成のために利用していることが推察された。The activity of gucose-6-phosphate dehydrogenase (G-6-PDH) in porcine oocytes and embryos was histochemically examined by the Rudolph and Klein method. Strong activity of G-6-PDH was observed in oocytes cultured for maturation and in fertilized oocytes, and blue diformazan granules produced by the enzyme reaction were spread evenly throughout the cytoplasm. In embryos, the activity somewhat decreased at the 2-cell stage, and the such activity was maintained up to the 16-cell stage. The activity weakened in embryos at the morula and early blastocyst stages, and some of them showed no enzyme activity. The enzyme activity completely disappeared from expanded blastocysts. In cleaved embryos, diformazan granules were distributed throughout the cytoplasm of blastomeres, while the amount of the granules differed among blastomeres. In early blastocysts, the granules were distributed in the cytoplasm of inner-cell-mass cells, but not in the cytoplasm of trophoblasts. The results obtained from this investigation and former studies concerning steroid metabolism seem to suggest that porcine oocytes and preimplantation embryos utilize NADPH produced by G-6-PDH for biosynthesis of steroids.departmental bulletin pape

Conduction velocities and changes in regional <i>I</i>_Na in cardiomyocytes in each myofiber model.

<p>(A) and (B), Relative ratios of CV (%CV) normalized by CV in the myocardial fiber with 50%<i>g</i>_Na,JM and 50%<i>g</i>_Na,LM as a function of %<i>g</i>_Na,JM or %<i>g</i>_Na,LM in the cleft (A) and non-cleft (B) models. (C), Peak values of post-junctional <i>I</i>_Na (post-<i>I</i>_Na,JM), pre-junctional <i>I</i>_Na (pre-<i>I</i>_Na,JM), and lateral <i>I</i>_Na (<i>I</i>_Na,LM) in a myocyte in the cleft model in which the Na⁺ channels are fixed at 50%<i>g</i>_Na,JM with 50%<i>g</i>_Na,LM, 30%<i>g</i>_Na,LM, or 10%<i>g</i>_Na,LM (<i>left</i>) and at 50%<i>g</i>_Na,LM with 50%<i>g</i>_Na,JM, 30%<i>g</i>_Na,JM, or 10%<i>g</i>_Na,JM (<i>right</i>). (D), Peak values of post-<i>I</i>_Na,JM, pre-<i>I</i>_Na,JM, and <i>I</i>_Na,LM in a myocyte in the non-cleft model with 50%<i>g</i>_Na,JM and 50%<i>g</i>_Na,LM, 30%<i>g</i>_Na,LM, or 10%<i>g</i>_Na,LM (<i>left</i>), and 50%<i>g</i>_Na,LM with 50%<i>g</i>_Na,JM, 30%<i>g</i>_Na,JM, or 10%<i>g</i>_Na,JM (<i>right</i>). The peak values of <i>I</i>_Na in each membrane segment were measured in the middle of the myofiber.</p

Reentry induction by Na⁺ channel blockade in myocardial ring models.

<p>(A), Phase diagram of %<i>G</i>_Na block vs. S1–S2 interval showing proarrhythmic events under Na⁺ channel blockade in the myocardial ring model. Right bar (control): responses to S2 stimulus in the myocardial ring constructed of only NZ myocytes under 50%<i>G</i>_Na block. (B), Examples of AP propagation in response to S1–S2 interval. Arrows and black short bars indicate the directions of AP propagation and entrance block, respectively.</p

Destabilization of action potential propagation by Na⁺ channel blockade.

<p>(A), Conduction velocity (CV) as a function of %<i>G</i>_Na block. Open circles with the dashed lines denote CV under AP alternans. The relationship between the excitation conduction mode and %<i>G</i>_Na block is represented by the bottom bars. SC, stable conduction; UC, unstable conduction (e.g., AP alternans, or 2∶1 or 3∶1 conduction); CB, complete conduction block. (B), Examples of AP propagation observed in NZ (i) and IBZ (ii) myofibers. (C)–(E), Examples of 2∶1 conduction by 55%<i>G</i>_Na block (C), 3∶1 conduction by 56% <i>G</i>_Na block (D), and complete conduction block by 60% <i>G</i>_Na block (E) in the IBZ.</p

Cleft and non-cleft models of myocardial fibers and rings.

<p>(A), Schematic representation of a myocardial fiber comprising cylindrical 300 cells. (B) and (C), Schematic representations of the intercellular junction in the cleft (B) and non-cleft (C) models. (D), The AP of each membrane segment is represented by the modified Luo–Rudy dynamic (mLRd) ventricular myocyte model <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109271#pone.0109271-Faber1" target="_blank">[24]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109271#pone.0109271-Suzuki1" target="_blank">[25]</a>. (E), Schematic representation of the myocardial ring comprising 900 cells (a), and the pacing protocol (b). <i>G</i>_g, gap junctional conductance; <i>G</i>_j, radial cleft conductance; <i>G</i>_d, axial cleft conductance.</p

Ischemia-Related Subcellular Redistribution of Sodium Channels Enhances the Proarrhythmic Effect of Class I Antiarrhythmic Drugs: A Simulation Study

<div><p>Background</p><p>Cardiomyocytes located at the ischemic border zone of infarcted ventricle are accompanied by redistribution of gap junctions, which mediate electrical transmission between cardiomyocytes. This ischemic border zone provides an arrhythmogenic substrate. It was also shown that sodium (Na⁺) channels are redistributed within myocytes located in the ischemic border zone. However, the roles of the subcellular redistribution of Na⁺ channels in the arrhythmogenicity under ischemia remain unclear.</p><p>Methods</p><p>Computer simulations of excitation conduction were performed in a myofiber model incorporating both subcellular Na⁺ channel redistribution and the electric field mechanism, taking into account the intercellular cleft potentials.</p><p>Results</p><p>We found in the myofiber model that the subcellular redistribution of the Na⁺ channels under myocardial ischemia, decreasing in Na⁺ channel expression of the lateral cell membrane of each myocyte, decreased the tissue excitability, resulting in conduction slowing even without any ischemia-related electrophysiological change. The conventional model (i.e., without the electric field mechanism) did not reproduce the conduction slowing caused by the subcellular Na⁺ channel redistribution. Furthermore, Na⁺ channel blockade with the coexistence of a non-ischemic zone with an ischemic border zone expanded the vulnerable period for reentrant tachyarrhythmias compared to the model without the ischemic border zone. Na⁺ channel blockade tended to cause unidirectional conduction block at sites near the ischemic border zone. Thus, such a unidirectional conduction block induced by a premature stimulus at sites near the ischemic border zone is associated with the initiation of reentrant tachyarrhythmias.</p><p>Conclusions</p><p>Proarrhythmia of Na⁺ channel blockade in patients with old myocardial infarction might be partly attributable to the ischemia-related subcellular Na⁺ channel redistribution.</p></div

Conduction velocity restitution properties.

<p>Conduction velocity (CV) restitution curves in the NZ (A) and IBZ (B) myofiber models as a function of S1–S2 interval. Ten S1 stimuli of basic cycle length (400 ms) were applied transmembranously followed by an S2 stimulus with various coupling intervals.</p

Beating motion of the whole heart model.

<p>(A) The temporal variation of wall thickness of an actual healthy heart observed by Traill et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036706#pone.0036706-Traill1" target="_blank">[26]</a>. The four dashed red lines in (B) indicate the temporal variations of wall thickness of our simulation results with different MC rates (<i>A^mc</i> = 0.4, 0.5, 0.6., and 0.7). (C) Specified time-contraction curve; the blue chart is for atrial regions and the green chart is for ventricular regions. (D, E) Representative frames of the whole-heart simulation with different visualization. (F–H) Three interaction tools. (F, G) Cross sections and myocardial layers of the heart at time <i>t0</i> and <i>t3</i>. (H) Direct dragging tool. White arrows indicate the grabbed point and black arrows are dragged direction.</p

The effect of fiber-direction-dependent weights.

<p>We specified horizontal (B), vertical (C), and 45°-slanted (D) fiber orientation in a thick -sheet model (100 mm×100 mm×20 mm) (A). We then fixed the top regions and observed the resting shapes in a gravity field without activation. The resting shapes (B–D) show that the model is stiffer in the fiber direction than in the perpendicular direction. We specified gravity acceleration as 9.8 m/s², time step <i>h</i> = 0.005 s, and stiffness iteration <i>M</i> = 3. Note that we used small stiffness value so as to observe large deformations.</p

An overview of our kinematic approach.

<p>(A) Heart model. LA, RA, LV, and RV stand for left/right atrium, and left/right ventricle, respectively. (B) Target mesh model in 2D. Constructed local regions (D) are constructed along the fiber orientation so as to maintain their original volumes (E). (F, C)We deform global shape so as to satisfy the shapes of the constructed local regions as much as possible.</p