Muneshige Kaibara

Relationship Between Atrial Conduction Defects and Fractionated Atrial Endocardial Electrograms in Patients with Sick Sinus Syndrome

The relationship between abnormal atrial electrograms (AAE) recorded during sinus rhythm by endocardial calheter mapping of the right atrium and the afrial conduction defects of sinus impulses or single atrial extrastimuli was investigated in 44 patients with sick sinus syndrome. The patients were divided into two groups on the basis of the presence (n = 29) or absence (n = 15) of AAE recorded during sinus rhythm. The P wave duration in the AAE (+) Group patients was 137 ± 14 msec, and 125 ± 15 msec in (he AAE (−) Group; P < 0.02. The intraatrial conduction time of sinus impulses in the AAE (+) Group was 54 ± 12 msec, and 39 ± 9 msec in the AAE (−) Group; P < 0.001. The interatrial conduction time in the AAE (+) Group was 101 + 14 msec, and 78 ± 16 msec in the AAE (−) Group; P < 0.001. In the AAE (+) Group, H (38%) patients ha d a sinus node recovery time > 4 seconds, whereas in the AAE (−) Group there was only one (6%) patient; P < 0.03. AAE showed a specificity of 93% and a positive predictive accuracy of 91% in predicting inducibility of atrial fibrillation. The sensitivity was 35% and the negative predictive accuracy was 42%. Sustained atrial fibrillation was induced in ten (35%) patients of the AAE (+) Group, and in one (7%) patient of the AAE (−) Group; P < 0.05. These data suggest that in patients with sick sinus syndrome who possess abnormal endocardial eJectrograms in sinus rhythm within the right atrium have: (1) a significantly longer P wave duration: (2) a significantly longer intraatrial and interafrial conduction time of sinus impulses; and (3) a significantly greater sinus node dysfunction and higher incidence of induction of sustained atriai fibrillation. It is concluded that there are significantly greater atrial conduction defects in patients with sick sinus syndrome who possess AAE within the right atrium during sinus rhythm.

Functions of peripheral 5-hydroxytryptamine receptors, especially 5-hydroxytryptamine4 receptor, in gastrointestinal motility.

Abstract: The multiple 5-hydroxytryptamine (5-HT, serotonin) receptor subtypes are distinguished. In this article, we described mainly the 5-HT4 receptor of four subtypes of functional 5-HT receptors, 5-HT1, 5-HT2, 5-HT3, and 5-HT4, recognized in the gastrointestinal tract. In-vivo microdialysis experiments determined that activation of the 5-HT4 receptor stimulated intestinal motor activity associated with a local increase in acetylcholine (ACh) release from the intestinal cholinergic neurons in the whole body of dogs. The 5-HT4 receptor-mediated response of ACh release in the antral, corporal, and fundic strips isolated from guinea pig stomach corresponds to the presence of 5-HT4 receptor in the myenteric plexus. In-vitro receptor autoradiograms of the stomach and colon indicate that the distribution of 5-HT4 receptors in human tissues is similar to that in the guinea pig, although density of 5-HT4 receptors in the myenteric plexus of human tissues is lower than that in guinea pig tissues. The 5-HT4 receptors located in the myenteric plexus may participate in gastrointestinal motility, and thus the 5-HT4 agonists and antagonists may be available for treatment of dysfunction of gastrointestinal motility.

Tissue extract recovers cardiac calcium channels from ‘run-down’

Effects of cardiac-tissue extract on the activity of L-type Ca2+ channels were investigated in guinea-pig ventricular myocytes with the patch-clamp method. In most patches, Ca2+-channel current recorded with a pipette solution containing 50 mM Ba2+ and 3 μM Bay K 8644 ran down within 5 min after excision of the patches into a solution containing EGTA. This run-down of Ca2+ channels was prevented when pathches were excised into a solution containing a supernatant fraction of homogenate of guinea-pig or bovine heart. Furthermore, this tissue extract was able to restore channel activity after run-down. This channel-activating effect of the extract was abolished by heat treatment or trypsin digestion. Fractionation of the extract by gel filtration suggested that the channel-activating factor(s) had an apparent molecular weight of 2–3×105. These results suggest that some cytoplasmic protein(s) maintains the activity of the cardiac L-type Ca2+ channel.

The inhibitory effects of tramadol on muscarinic receptor-induced responses in Xenopus oocytes expressing cloned M(3) receptors.

Tramadol is a widely used analgesic, but its mechanism of action is not completely understood. Muscarinic receptors are involved in neuronal function in the brain and autonomic nervous system, and much attention has been paid to these receptors as targets of analgesic drugs in the central nervous system. In this study, we investigated the effects of tramadol on type-3 muscarinic (M3) receptors using the Xenopus oocyte expression system. Tramadol (10 nM–100 &mgr;M) inhibited acetylcholine-induced currents in oocytes expressing M3 receptor. Although GF109203X, a protein kinase C inhibitor, increased the basal current, it had little effect on the inhibition of acetylcholine-induced currents by tramadol. Moreover, tramadol inhibited the specific binding sites of [3H]quinuclidinyl benzilate. These findings suggest that tramadol at clinically relevant concentrations inhibits M3 function via quinuclidinyl benzilate-binding sites. This may explain the modulation of neuronal function and the anticholinergic effects of tramadol.

Involvement of Na(+)-H+ antiporter in regulation of L-type Ca2+ channel current by angiotensin II in rabbit ventricular myocytes.

The present study investigated the possible involvement of a Na(+)-H+ antiporter in the regulation of L-type Ca2+ channels by angiotensin II (Ang II) in isolated rabbit ventricular cardiac myocytes by using both cell-attached and whole-cell patch-clamp current recording techniques. In cell-attached patch-clamp current recordings, an increase in the open-state probability of the Ca2+ channel (144.8 +/- 9.8% [mean +/- SEM], n = 11) was seen after exposure of the cells to Ang II (100 nmol/L). This effect was inhibited by pretreatment with losartan (10 mumol/L), a synthetic antagonist of the AT1 receptor. 5(N,N-Dimethyl)amiloride (100 mumol/L), an amiloride analogue, as well as Na(+)-deficient bath solution abolished Ang II-induced stimulation of the Ca2+ channel activities. In whole-cell patch-clamp current recordings, Ang II also increased the L-type Ca2+ current when a pipette solution of pH 7.1 containing 5 mmol/L HEPES (139 +/- 5%, n = 4) was used but did not significantly increase the current when a pipette solution of pH 7.5 containing 5 mmol/L HEPES or a pipette solution of pH 7.1 containing 30 mmol/L HEPES was used. These results suggest that Ang II-induced stimulation of the Ca2+ channels is mediated by a Na(+)-H+ antiporter and therefore provide a novel insight into signal transduction of Ang II receptor stimulation in cardiac myocytes.

Run-down of the cardiac Ca2+ channel: characterization and restoration of channel activity by cytoplasmic factors

Abstract Possible mechanisms for run-down in the Ca2+ channel, such as proteolysis or dephosphorylation of the channel, were examined in guinea-pig ventricular myocytes. The Ca2+ channel current, recorded in inside-out patches using a pipette solution containing 50 mM Ba2+ and 3 μM Bay K 8644, ran down with a mean survival time of 2.35 min. The survival time was not significantly affected by adenosine triphosphate (ATP) (3 mM), 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA) (2 mM), isoprenaline (l–5 μM), phosphate (l20 mM) and leupeptin (l0 μM). Stimulation of guanosine triphosphate (GTP)-binding proteins was also ineffective. The catalytic subunit of adenosine 3′,5′-cyclic monophosphate (cAMP)-dependent protein kinase (PKA, 0.5–2 μM) slightly and transiently increased channel activity, but had minimal effects on the channel when applied after complete run-down. On the other hand, cytoplasm from the heart, skeletal muscle, brain and liver, but not kidney, induced channel activity. There was a positive correlation between NPo (the product of the number of channels N and the open probability Po) value before run-down and that after the application of cytoplasm, suggesting that the activity of once-active channels was restored ba the exogenous cytoplasm. The potency of cytoplasm in tissues in inducing channel activity was not related to PKA activity nor to the number of dihydropyridine binding sites. These results suggest that the run-down of the cardiac Ca2+ channel is not mediated by dephosphorylation or proteolysis of the channel, but involves other factor(s), possibly interaction of the channel protein with a cytoplasmic regulatory protein.

Acetazolamide acts directly on the human skeletal muscle chloride channel

Acetazolamide, a carbonic anhydrase inhibitor, is used empirically in neuromuscular diseases with episodic ataxia, weakness, and myotonia, although not all of the mechanisms responsible for its therapeutic effects are understood. To elucidate whether acetazolamide acts directly on the human skeletal muscle voltage‐gated chloride channel (ClC‐1), which is associated with myotonia, we evaluated the effects of acetazolamide on ClC‐1 expressed in cultured mammalian cells, using whole‐cell recording. Acetazolamide significantly shifted the voltage dependency of the open probability (Po) toward negative potentials in a dose‐dependent manner, resulting in an increase of chloride conductance at voltages near the resting membrane potential. This effect was attenuated when using a pipette solution containing 30 mmol/L Hepes. These results suggest that acetazolamide can influence the voltage‐dependent opening gate of ClC‐1 through a mechanism related to intracellular acidification by inhibiting carbonic anhydrase, and that the therapeutic effects of acetazolamide in neuromuscular diseases may be mediated by activation of ClC‐1. Muscle Nerve, 2006

Involvement of cholinergic neurons in orexin-induced contraction of guinea pig ileum

The mechanism underlying orexin-induced contraction was examined in isolated preparations of guinea pig ileum, in relation to cholinergic transmission. Orexin-A caused contraction of ileal strips in a concentration-dependent manner. 1-(2-Methylbenzoxazol-6-yl)-3-[1,5]napthyridin-4-yl-urea hydrochloride (SB-334867-A) antagonized the orexin-A-induced contraction, with no effects on the acetylcholine-induced contraction and twitch contractions. The orexin-A-induced contraction was inhibited by tetrodotoxin and atropine, but not by hexamethonium, an antagonist of vasoactive intestinal peptide and a mixture of 5-hydroxytryptamine receptor antagonists. Orexin-A evoked an outflow of [3H]acetylcholine from the ileal strips preincubated with [3H]choline, in a concentration-dependent manner, and the orexin-A-evoked outflow was inhibited by tetrodotoxin, indicating that the outflow of [3H]acetylcholine originates from the nerve terminals. The orexin-A-evoked outflow of [3H]acetylcholine was antagonized by SB-334867-A. Thus, orexin-A evokes the release of acetylcholine from the enteric cholinergic neurons due to stimulation of the orexin-1 receptors and then causes contractions of guinea pig ileum.

Activation of inwardly rectifying K+ channels by GABA-B receptors expressed in Xenopus oocytes.

IN Xenopu s oocytes coinjected with poly(A)+ RNA derived from the rat cerebellum and cRNAs for the cloned G protein-gated inwardly rectifying K+ channel (GIRK), GIRK1 and GIRK2, the GABA-B agonist baclofen elicited inwardly rectifying K+ currents. The inward K+ currents elicited by baclofen were inhibited by the selective GABA-B antagonists 2–OH saclofen and CGP 35348, and by the GIRK inhibitor Ba2+. In contrast, baclofen caused no currents in oocytes injected with the cerebellar poly(A)+ RNA alone, the poly(A)+ RNA and cRNA for GIRK1 or GIRK2, or only cRNAs for GIRK1 and GIRK2. These findings indicate that GABA-B receptors in the rat cerebellum were functionally expressed in Xenopu s oocytes and activated the cloned GIRKs composed of GIRK1 and GIRK2 as heteromultimers.

Regional difference in correlation of 5-HT4 receptor distribution with cholinergic transmission in the guinea pig stomach.

Localization and function of 5-HT4 receptors in the stomach were examined in mucosa-free preparations of antrum, corpus and fundus from guinea pig stomach by determination of acetylcholine release and in vitro receptor autoradiography. Specific [125I]SB207710, (1-n-butyl-4-piperidinyl) methyl-8-amino-7-iodo-1,4-benzodioxane-5-carboxylate, binding sites were detected in 3 regions of the stomach. High densities of binding were observed in the myenteric plexus of antrum and corpus, but not fundus. In mucosa-free preparations treated with 5-HT1, 5-HT2 and 5-HT3 receptor antagonists, 5-HT (10(-8)-10(-6) M) potentiated the electrically stimulated (0.5 Hz, 1 ms) outflow of [3H]acetylcholine from antrum and corpus strips preloaded with [3H]choline, but not from fundus strips, and the potentiation was antagonized by SB204070, (1-n-butyl-4-piperidinyl) methyl-8-amino-7-chloro-1,4-benzodioxane-5-carboxylate. Thus, 5-HT4 receptors are located on myenteric cholinergic neurons in the antrum and corpus of guinea pig stomach and their activation evokes the release of acetylcholine.