Kaichiro Kamiya

The Effect of β-adrenargic Agonists on Ca^2+ Sensitivity in Tracheal Smooth Muscle

To determine whether a reduction in the sensitivity to Ca 2 + is involved in relaxation by β-adrenergic receptor agonists, we examined correlation between force generation and intracellular Ca 2 + concentration in the inhibitory effects of isoproterenol (ISO) on methacholine (MCh)-induced contraction. Fura-2 was loaded in tracheal smooth muscle of guinea pigs and tension and fluorescence ratio F 3 4 0 /F 3 8 0 as an index of intracellular Ca 2 + concentration were recorded simultaneously. Addition of 0.3 μM ISO caused an inhibition in 1 μM MCh-induced contraction and a reduction in intracellular Ca 2 + concentration. The reduction in intracellular Ca 2 + concentration was, however, much smaller than that of tension. In conclusion, Ca 2 + sensitivity in addition to Ca 2 + influx plays an important role in β-adrenergic action in airway smooth muscle.

Limited Association of Connexin43 and ZO-1 in the Intercalated Disks of Adult Rat Ventricular Myocardium

Recent biochemical studies have suggested that zonula occludens-1 (ZO-1), a PDZ domain-containing protein may play an important role in targeting the major gap junction protein, Cx43 to the intercalated disk region of cardiac muscle. Information for in vivo is, however, still limited. We investigated the distribution of Cx43 and ZO-1 proteins in the ventricular myocardium of adult rats using high-resolution analysis with a confocal microscopy. Immunolabeling spots for Cx43 and ZO-1 were largely confined to the intercalated disk regions of ventricular muscles. ZO-1 was also localized mainly in the intercalated disk region. These spots were a marked difference in the positions, indicating low levels of colocalization of Cx43 and ZO-1 immunolabled spots in the intercalated disks. These results suggest that ZO-1 does not play an important role in intercalated disk-targeting of Cx43 molecules in the normal ventricular myocardium.

Virtual electrode polarization-induced reentrant activity

It is known that electric shocks create a characteristic pattern of polarization (virtual electrode polarization) in the myocardium but its roles on post-shock excitations remain to be clarified. We have optically mapped electrical activity of the left ventricular subepicardial layer of Langendorff-perfused rabbit hearts endocardialy frozen. Monophasic anodal and cathodal shocks were applied during the plateau or repolarization phase of the action potential. Electric shocks (20 V, 10 ms) resulted in a dog-bone pattern of polarization around the stimulation electrode and the withdrawal of the shocks were followed by break excitations. Spread of the break excitations were partially blocked by refractory of the preceding excitation, and slow conduction around the functional line of block initiated reentrant excitations. These results suggest that break excitations arising from the virtual electrode polarization combined with spatial heterogeneity in repolarization of the preceding excitation play important roles in the genesis of post-shock reentrant activities.

Role of Ca^ Release from Sarcoplasmic Reticulum in Pacemaker Activity of the Sinoatrial Node

Recent studies using confocal microscopy combined with patch clamping on single sinoatrial (SA) node pacemaker cells suggest that Ca 2 + release from the sarcoplasmic reticulum (SR) during diastole may play a prominent role in the late phase of pacemaker depolarization. The present study was designed to test this hypothesis in the intact SA node. We investigated the effects of a high concentration of ryanodine, which is known to disable SR Ca 2 + release, on spontaneous activity of isolated rabbit right atrium including the whole SA node by using an extracellular potential mapping technique. Inhibition of SR Ca 2 + release by 30 μM ryanodine caused only a moderate reduction of the spontaneous firing rate (by 20.0′2.8 %, n=4) of the intact SA node. This observation is inconsistent with previous data obtained from single pacemaker cells. Physiological significance of SR Ca 2 + release in the regulation of SA node pacemaker activity is still unsettled.

mRNA levels of ERG, KVLQT1 and minK in rabbit right and left ventricles

Our previous study showed that two components of the delayed rectifier potassium current, I K r and I K s , were heterogeneously distributed in right (RV) and left (LV) ventricles in rabbit hearts. However, the mechanisms of heterogeneous distribution of these channels were not known. In the present study, we investigated the encoding mRNA levels of I K r (ERG) and I K s (KVLQT1 and minK) in RV and LV. By using a quantitative real-time PCR method, we found mRNA levels were not significantly different between two ventricles in ERG (RV: 1103 ′ 218 molecules/10 5 GAPDH molecules, LV: 886 ′ 155, n=4), KVLQT1 (RV: 645 ′ 113, LV: 509 ′ 170, n=4) and minK (RV: 209 ′ 33, LV: 185 ′ 47, n=4). These results suggest that heterogeneous distribution of I K r and I K s in RV and LV could not be explained by the mRNA levels in ventricles. Other unknown factor may underlie this phenomenon.

Dephosphorylation of Connexin43 Associated with Ventricular Hypertrophy

Altered expression and distribution of gap junctions in hypertrophied hearts may provide a potential substrate for abnormal conduction. We investigated changes of phosphorylation state of connexin43 (Cx43) in hypertrophied rat right ventricles secondary to pulmonary hypertension induced by monocrotaline (MCT) using western blot analysis. In normal right ventricular myocardium, most of Cx43 was phosphorylated. In hypertrophied ventricles of MCT-treated rats, the non-phosphorylated isoform of Cx43 increased and the phosphorylated isoform of Cx43 decreased with no significant a change in the total amount of Cx43 protein. These results suggest that dephosphorylation of Cx43 in MCT-induced right ventricular hypertrophy may be involved in gap junction disorganization.