Yoshiko Yamauchi

Changes in the fluctuation of interbeat intervals in spontaneously beating cultured cardiac myocytes: experimental and modeling studies.

Abstract. Isolated and cultured neonatal cardiac myocytes contract spontaneously and cyclically, and have the properties of a non-linear oscillator. In this study, we have analyzed the relationship between the fluctuation of contraction rhythm of spontaneously beating cultured cardiac myocytes, and the coupling strength among them. The coefficient of variation of contraction intervals increased transiently in the early stages of incubation, and then decreased almost monotonically with time. The contraction rhythm of the myocytes became synchronized in the late stage of the culture. The day on which synchronization occurred almost coincided with the day when the coefficient of variation reached its lowest value. In addition, we have performed a mathematical analysis using interacting Bonhoeffer–van der Pol oscillators to clarify the mechanisms underlying the changes in the fluctuation of contraction rhythm with time. As the coupling strength among oscillators increased, the coefficient of variation of oscillation periods increased temporarily, but then decreased rapidly when the oscillators showed synchronization. These results suggest that the changes in the fluctuation of beating rhythm result from the increase in strength of electrical coupling among spontaneously beating cardiac myocytes.

A new method for continuous measurement of regional cerebral blood flow using laser Doppler flowmetry in a conscious rat.

We have developed a new system to continuously measure regional cerebral blood flow (rCBF) in the cortex of a conscious animal. For this purpose, we used rats and laser Doppler flowmetry. Under pentobarbital anesthesia, the animals skull was opened making a small square hole 3 mm x 3 mm in size. A transparent acrylic plate was placed over the hole in the skull. A polyethylene cannula (inner diameter 1.0 mm, length 5.0 mm) was fixed on the plate as a guide for the laser Doppler flowmeter (LDF) probe (outer diameter 1.0 mm, length 5.5 mm). Both the plate and guide cannula were fixed to the skull by dental cement. Every day for the following two weeks after surgery, the conscious animal was placed in a hammock for recording rCBF. A LDF probe was freely attachable to the plate above the cortex via the guide cannula during measurement of rCBF. The rats were kept in a hammock with their legs firmly touching the floor during measurement of rCBF. It was possible to measure rCBF every day for about two weeks, and rCBF responded consistently to inhalation of 7% CO2 when the responses were expressed as percentages of the prestimulus control rCBF values. This system is recommended for the continuous measurement of rCBF in a conscious animal.

Ruthenium red-induced transition from ventricular fibrillation to tachycardia in isolated rat hearts:: possible involvement of changes in mitochondrial calcium uptake

INTRODUCTION Ventricular tachycardia (VT) is considered to be the most common precursor of ventricular fibrillation (VF) and sudden cardiac death. However, the mechanisms underlying the transition from VT to VF remain unclear despite more than a century of study. Here, we investigated whether perfusion of the heart with blockers of mitochondrial Ca(2+) uniporter changed the macrodynamics of the heart between VT and VF. METHODS The experiments were performed using Langendorff perfused isolated rat hearts in which left ventricular pressure (LVP) and left ventricular cardiomyogram (LVCMG) were measured. Sustained VT or VF was induced by burst pacing of the left ventricular muscles. RESULTS During pacing-induced sustained VF, perfusion of the heart with ruthenium red (RR) or Ru 360, blockers of mitochondrial Ca(2+) uniporter, resulted in the reversible conversion of VF to VT. In contrast, during pacing-induced sustained VT, perfusion of the heart with spermine, an activator of mitochondrial Ca(2+) uptake, resulted in the reversible conversion of VT to VF, and the effect was antagonized by cotreatment with RR. In addition, RR-induced conversion of VF to VT was antagonized by cotreatment with S(-)-Bay K8644 (Bay K), an activator of L-type Ca(2+) channels, suggesting that the inactivation of L-type Ca(2+) channels was responsible for the RR-induced effect on the macrodynamics of hearts. In fact, perfusion with verapamil, an antagonist of L-type Ca(2+) channels, during pacing-induced sustained VF, resulted in the conversion of VF to VT. CONCLUSION This study demonstrated that perfusion of isolated rat hearts with blockers of Ca(2+) uptake by mitochondria resulted in the reversible conversion of pacing-induced sustained VF to VT, suggesting that changes in mitochondrial Ca(2+) uptake were possibly involved in the transition between VT and VF.

Coupling between respiratory and locomotor rhythms during fictive locomotion in decerebrate cats

Fictive locomotion of the hindlimb was evoked by stimulation of the mesencephalic locomotor region (MLR) in immobilized, decerebrate cats. Fictive respiration can also be obtained in such a preparation after bilateral vagotomy. A cross-correlation technique was used to evaluate the strength of the coupling between the locomotor and respiratory rhythms. This study demonstrated that there was a locomotor-respiratory coupling of central origin and the strength of the coupling varied depending on the level of end-tidal pCO2, reflecting the arterial CO2 tension.

Fluctuations of Contraction Rhythm During Simulated Ischemia/Reperfusion in Cultured Cardiac Myocytes from Neonatal Rats

Cardiac ischemia results in a rapid decrease of intracellular pH and in the rise of intracellular Ca 2+, changes that have been shown to reduce intercellular communication via gap junctions (GJ) between cardiac myocytes. Ischemia also results in electrical instability probably caused by the reduced GJ permeability contributing to an increased vulnerability to arrhythmias. This study aims at elucidating whether the fluctuations of contraction rhythm of spontaneously beating cardiac myocytes in culture changes during simulated ischemia/reperfusion. The coefficient of variation (CV) of contraction intervals, reflecting the fluctuation of contraction rhythm, increased significantly during simulated ischemia/reperfusion. However, the contraction rhythm of the cardiac myocytes in an aggregate remained synchronized during simulated ischemia/reperfusion. In contrast, pharmacological blockade of GJ with 12-doxyl stearic acid, a blocker of GJ permeability, resulted in the de-synchronization of contraction rhythm and in an increase in the CV of contraction intervals in normoxic conditions. The present findings lead to the suggestion that GJ remained open during simulated ischemia/reperfusion, and that a mechanism other than electrical uncoupling between myocytes contributed to the observed increase in the fluctuation of beating rhythm during ischemia.

Parallel suppression of extensor muscle tone and respiration by stimulation of pontine dorsal tegmentum in decerebrate cat

This paper describes the pontine brainstem area responsible for the suppression of postural muscle tone as well as of respiration in acute precollicular-postmammillary decerebrate (mesencephalic) cats. Stimulation of the dorsal part of the pontine tegmentum (DTF) along the midline (P4-P7, H-5 to H-6) decreased the bilateral tone of the hindlimb extensor muscles and the diaphragmatic activity. Tonic discharges of the extensor muscles were suppressed by DTF stimulation and the suppression of muscle activity continued for more than 5 min after termination of the stimulation. In contrast, the suppression of the diaphragmatic activity, which resulted in apnea in some of the animals tested, resumed in spite of the continuation of the stimulation. However, the rebound augmentation of the diaphragmatic activity appeared immediately after the termination of the stimulation. The existence of such a rebound phenomenon suggested that the suppressive effects on the diaphragmatic activity persisted during the entire period of the stimulation. The recovery of respiratory movements during the stimulation led us to suggest that the strong respiratory drives emerge to overcome the exerted DTF-elicited suppressive effects on respiration. In the paralyzed and vagotomized animal, the DTF-elicited suppressive effects on phrenic neural discharges were minimal when the end-tidal pCO2 was set at a higher level than during spontaneous breathing.

Increased vulnerability to ischemia/reperfusion-induced ventricular tachyarrhythmias by pre-ischemic inhibition of nitric oxide synthase in isolated rat hearts

INTRODUCTION The relationship between vulnerability to reperfusion-induced ventricular tachyarrhythmias, such as ventricular tachycardia (VT) and fibrillation (VF), and the endogenous activity of nitric oxide synthase (NOS) has not been well documented. The objective of the present study was to clarify whether the vulnerability to reperfusion-induced VT/VF changes with preishemic, sustained inhibition of NOS. METHODS The experiments were performed using Langendorff-perfused isolated rat hearts, in which left ventricular pressure (LVP) and left ventricular cardiomyograms (LVCMGs) were measured. RESULTS A pre-ischemic, sustained inhibition of NOS resulted in an increased vulnerability to reperfusion-induced VT/VF, and the increase was markedly attenuated by co-treatment with L-arginine or by post-ischemic treatment with 2,4-diamino-6-hydroxypyrimidine (DAHP), an inhibitor of tetrahydrobiopterin (BH(4)) synthesis. We then tried to elucidate whether nitric oxide (NO) and superoxide were produced during reperfusion, and ATP-sensitive potassium channels (K(ATP)), especially mitochondrial ATP-sensitive potassium channels (mitoK(ATP)), are involved in the increased vulnerability. Post-ischemic inhibition of NOS and treatment with a NO scavenger attenuated the increased vulnerability to reperfusion-induced VT/VF, but post-ischemic treatment with a superoxide scavenger did not. In addition, post-ischemic treatment with S-nitroso-N-acetyl-DL-penicillamine (SNAP), a NO donor, or with diazoxide, a selective opener of mitoK(ATP), increased the VT/VF duration during reperfusion. The increased vulnerability to VT/VF was attenuated by the treatment with a selective mitoK(ATP) blocker. CONCLUSION The results suggest that a pre-ischemic, sustained inhibition of NOS increases the vulnerability to reperfusion-induced VT/VF, and the NO-mitoK(ATP) pathway is one of the possible factors contributing to the increased vulnerability to VT/VF.

Spectral analysis on fluctuation of heat period in paralyzed, vagotomized, and unanesthetized decerebrate cats

Spectral analysis was performed on the heart period fluctuation in vagotomized, paralyzed, and unanesthetized decerebrate cats. The heart period was measured as the time interval between successive R waves of the electrocardiograms. When end-tidal PCO2 was set at the same level as that before immobilization, the power spectral density plot of the heart period fluctuation showed several distinct peaks: one peak corresponded to the frequency of the artificial ventilator and the others to its harmonics. In addition, the spectral density plot had another peak centered at the intrinsic respiratory frequency evaluated by recording efferent phrenic neural discharges. The amplitude of these spectral peaks tended to become greater when the end-tidal PCO2was increased by adding CO2 to the input gas. Our results, therefore, provide evidence that the heart period is modulated not only by the artificial ventilation rhythm but also by the centrally generated respiratory rhythm, and suggested that the strength of such central interactions between cardiac and respiratory rhythms varies depending on the end-tidal PCO2 level.

Inhibitory influences on hypoglossal neural activity by stimulation of midpontine dorsal tegmentum in decerebrate cat

In the spontaneously breathing decerebrate cat, the properties of the suppressive effects on hypoglossal nerve activity and on diaphragmatic activity elicited by stimulation of the midpontine dorsal tegmentum (DTF area) were analyzed. Stimulation simultaneously decreased the activities of the hypoglossal nerve as well as that of the diaphragm. However, the inhibitory influences on the above two kinds of activities were different in nature. Diaphragmatic activity, once suppressed by stimulation, recovered and gradually became greater in amplitude in spite of the continuation of stimulation. In contrast, DTF stimulation depressed tonic discharges of the hypoglossal nerve, and the decreased tonic nerve activity persisted after stimulation ended. Rhythmic hypoglossal activity, once suppressed by stimulation, reappeared during DTF stimulation. Such a rhythmic activity, however, vanished after the termination of stimulation, although the rhythmic diaphragmatic activity did not.

Neuronal origin of parallel suppression of postural tone and respiration elicited by stimulation of midpontine dorsal tegmentum in the decerebrate cat

This paper describes the possibility that rostal pontine neuronal structures cause the parallel suppression of postural tone and respiration evoked by stimulation of the dorsal tegmental field (DTF) of the pons in decerebrate cats. Stimulation of the DTF along the midline decreased both diaphragmatic activity and the bilateral tone of the hind-limb extensor muscles. Pontine neuronal structures located rostrally to the DTF, from which antidromically activated units could be recorded on stimulation of the DTF, were studied. Antidromic spikes were recorded in and near the nucleus reticularis pontis oralis. Tonic electrical stimulation of these sites caused parallel suppression of postural tone and respiration. These suppressive effects were almost similar to those elicited by the DTF stimulation.