Pig Myocardium Research Articles

Impact of 13(S)-HPODE, a lipoxygenase product of linoleic acid, on calcium transport and Na,K-ATPase activity in the myocardial sarcolemma

In thisin vitro study using a purified sarcolemmic fraction of guinea pig myocardium, the 13(S)-hydroperoxide of linoleic acid (13-HPODE) increased in a dose-dependent manner the permeability of myocardial sarcolemma to Ca ions in concentrations above 10 μmol/liter, stimulated Na/Ca exchange there in concentrations from 0.1 to 10 μmol/liter, and exerted a digitalis-like action on sarcolemmic Na,K-ATPase in concentrations between 0.1 and 100 μmol/liter (IC50=20 μmol/liter). The results indicate that the linoleic acid hydroperoxide may be an effective modulator of sarcolemmic Ca2+ transport and of membrane-bound enzymes.

Does interstitial adenosine mediate acute hibernation of guinea pig myocardium?

Does interstitial adenosine mediate acute hibernation of guinea pig myocardium?

Does interstitial adenosine mediate acute hibernation of guinea pig myocardium?

The aim was to test the role of interstitial adenosine in protective downregulation of myocardial energy demand during myocardial hibernation. Isolated working guinea pig hearts, perfused with glucose fortified Krebs-Henseleit, were subjected to 60 min global low flow ischaemia followed by 30 min reperfusion. Left ventricular performance was assessed from heart rate-developed pressure product and pressure-volume work. Cytosolic energy level was indexed by creatine phosphate and ATP phosphorylation potentials measured in snap frozen myocardium. Lactate and purine nucleosides (adenosine, inosine) were measured in venous effluent. When coronary flow was lowered by 80% for 60 min, heart rate-pressure product and pressure-volume work fell 87% and 75%, respectively, and stabilised at these low levels, but fully recovered when flow was restored. Myocardial ATP phosphorylation potential fell by 67% during the first 10 min of ischaemia, but subsequently recovered to preischaemic levels despite continuing ischaemia, indicating down-regulation of myocardial energy demand. Lactate release increased about 10-fold during ischaemia and remained increased until reperfusion. Purine nucleoside release varied reciprocally with phosphorylation potential, peaking at 10 min of ischaemia, then gradually returning to the preischaemic level during the subsequent 50 min of ischaemia. The ecto 5'-nucleotidase inhibitor alpha,beta-methylene adenosine 5'-diphosphonate (50 microM) decreased ischaemic purine nucleoside release by 41%, but did not attenuate postischaemic contractile recovery. The unspecific adenosine receptor antagonist 8-p-sulphophenyl theophylline (8-SPT, 20 microM) doubled ischaemic lactate release and lowered coronary venous purine nucleoside release by 21%. 8-SPT increased phosphorylation potential at 10 min ischaemia relative to untreated hearts, but blunted the subsequent rebound of phosphorylation potential. 8-SPT treatment during ischaemia resulted in a significantly higher cytosolic phosphorylation potential at 30 min of reperfusion, but did not affect postischaemic contractile function. We conclude that activation of adenosine receptors results in recovery of cytosolic energy level of moderately ischaemic working myocardium, but this energetic recovery is not solely responsible for postischaemic contractile recovery.

Synthesis of [1- 11C], [2- 11C], [1- 11C]( 2H 3) and [2- 11C]( 2H 3)acetate for in vivo studies of myocardium using PET

Four isotopically-labelled acetates ([1- 11C], [2- 11C], [1- 11C]( 2H 3) and [2- 11C]( 2H 3)acetate) were synthesized and used in positron emission tomography (PET) studies of pig myocardium. The [1- 11C]acetates were synthesized by carboxylation of the appropriate 1H or 2H methyl Grignard reagents immobilized on a C 2 solid phase extraction column (SPE). Purification by reverse-phase HPLC, resulted in 35–45% decay-corrected radiochemical yield with a total synthesis time of 25 min, and a radiochemical purity higher than 99%. The [2- 11C]acetates were synthesized by carboxylation of 11C-labelled 1H or 2H methyl lithium. Purification as above resulted in 35–55% decay-corrected radiochemical yield with a total synthesis time of 30 min, and a radiochemical purity higher than 99%. Position-specific labelling was assessed by 13C-labelling and NMR. Multiple isotopic labelling by the combination of position-specific 11C-labelling and 2H substitution, has the potential to highlight different aspects of a complex biochemical system using a selected set of tracers in comparative PET studies. An illustration of this principle is given using acetate, where citric acid cycle metabolism results in a position-specific kinetic for the 11C-label, and deuteration opens up the possibility for the proton-abstracting processes within the citric acid cycle to be assessed.

On the Reversal of Myocardial Stunning: A Role for Ca2+‐Sensitizers

The mechanism underlying myocardial stunning is still unknown, but current views hold that generation offree radicals and disturbances in the calcium homeostasis, mechanisms which are not mutually exclusive, are the two most likely causes of prolonged postischemic dysfunction. 1.2 Several groups of investigators have indeed shown that the capacity of cardiac sarcoplasrnic reticulum (SR) to sequester Ca2+ decreases during i~chernia.~.~ In a recent study we have shown that the phosphorylation rate of phospholamban was unchanged, and that Ca2+ uptake by the sarcoplasmic reticulum was even slightly increased in myocardium of intact open-chest pigs, while function was still reversibly depressed.5 These data and earlier work by Marban and coworkers6 in isolated stunned ferret hearts strongly suggest that a decreased response of the myofilaments to Ca2+ rather than a change in the active Ca2+ transport of the SR is involved in the contractile dysfunction of stunned myocardium. In vivo evidence of this hypothesis has been very difficult to obtain because available agents that increase the responsiveness of the myofilaments to Ca2+ usually also increase contractility via inhibition of phosphodiesterase. We now report on the effects of the thiadiazinone derivative EMD 60263, which in in vitro experiments has been shown to be a potent Ca2+ sensitizer devoid of any phosphodiesterase inhibiting properties (Ravens et al., unpublished data). The compound also affects the delayed rectifier current I K ~ in a way characteristic for a class I11 antiarrhythmic action. The latter property may lead to a reduction in heart rate.

Differential Effects of BDF 9148 and DPI 201–106 on Action Potential and Contractility in Rat and Guinea Pig Myocardium

The effects of BDF 9148 and its parent compound DPI 201-106 were compared in guinea pigs and rat cardiac tissue because these tissues possess long and short action potentials (AP), respectively, and have different excitation-contraction coupling mechanisms. Conventional electrophysiologic techniques were used to study drug effects on AP, sodium current, and force of contraction (Fc). In guinea pig and rat papillary muscle, BDF 9148 and DPI 201-106 increased Fc; BDF 9148 was more potent than DPI 201-106. In guinea pig, but not in rat muscle, DPI 201-106 significantly prolonged AP duration (APD) to a greater degree than did BDF 9148. In rat cardiac muscle, both agents were more potent but increased Fc to a lesser degree than in guinea pig cardiac muscle and led to Ca2+ overload, as evidenced by after-contractions and contracture. BDF 9148 failed to increase Fc at low frequencies in guinea pig but was effective at all frequencies in rat muscle. In isolated myocytes, substance effects on sodium current were similar in both species. In guinea pigs, but not in rats, DPI 201-106 prolongs APD to a greater degree than does BDF 9148. Both agents produce a greater positive inotropic effect in guinea pigs than in rats, which may be due to species differences in excitation-contraction coupling.

Ischemic cardioprotection by ATP-sensitive K+ channels involves high-energy phosphate preservation.

We previously demonstrated that ATP-sensitive K+ channels (KATP) protect the guinea pig myocardium against ischemia-reperfusion injury (Cole et al., Circ. Res. 69: 571-581, 1991), but the cellular alterations leading to ischemic injury affected by KATP remain to be defined. This study investigates the relationship between activation of KATP and preservation of high-energy phosphates during global no-flow ischemia in arterially perfused guinea pig right ventricular walls. Electrical and mechanical activity were recorded via intracellular microelectrodes and a force transducer. Glibenclamide (10 and 50 microM) and pinacidil (10 microM) were used to modulate KATP. ATP and creatine phosphate (CP) levels were determined at the end of no-flow ischemia by enzymatic analysis. Preparations were subjected to 1) 20 min no-flow +/- glibenclamide (10 or 50 microM), 2) 30 min no-flow +/- pinacidil (10 microM) or pinacidil (10 microM) and glibenclamide (50 microM), or 3) 40 or 50 min of control perfusion before rapid freezing in liquid nitrogen. Pinacidil (10 microM) enhanced ischemic shortening of action potential duration (APD) and early contractile failure, prevented ischemic contracture, and inhibited high-energy phosphate depletion during ischemia. Glibenclamide (50 microM) inhibited the effects of pinacidil (10 microM) on electromechanical function and preservation of ATP and CP. Glibenclamide (10 microM) alone inhibited the early decline in APD and produced earlier ischemic contracture but did not enhance ATP or CP depletion compared with untreated tissues during 20 min of no-flow. Glibenclamide (50 microM) produced a greater inhibition of APD shortening in early ischemia, further decreased the latency to ischemic contracture, and caused enhanced ischemic depletion of ATP. The data indicate the changes in electrical activity induced by KATP indirectly preserve high-energy phosphates and reduce injury associated with ischemia. However, the data also suggest the possible presence of additional mechanisms for cardioprotection by KATP.

Short-term biventricular circulatory support in the regionally stunned pig myocardium.

Mechanical circulatory support is only meaningful when reversible myocardial damage exists. Prompt biventricular assistance during reperfusion for six hours was tried in the regionally sublethally injured pig myocardium which has perspective for recovery. Postmortem tissue was examined with a quickly performed histochemical technique for tissue viability. Animals underwent five coronary occlusions (5 min occlusion/10 min reperfusion). Postischemic systolic wall thickening, observed by direct echocardiography, fell about 50% (p < 0.05 vs base line) before treatment which paralleled cardiac inosine and lactate efflux. Biventricular unloading (n = 5) resulted in contractile recovery to 94 +/- 19% (p < 0.05 vs stunning), untreated pigs remained depressed (33 +/- 12%, n = 5). During circulatory support the arterial lactate levels rose three-fold, concomitant with a drop in pH of 0.2 units (p < 0.05). Post-mortem histochemistry showed scattered areas with decreased activities of cytoplasmic lactate dehydrogenase and mitochondrial beta-hydroxybutyrate dehydrogenase, irrespective of treatment. Transmission electron microscopy revealed irreversible damage to mitochondria in depleted areas in contrast to areas where enzyme activity was present. We conclude that prompt volume unloading during reperfusion accelerated contractile recovery of sublethally postischemic myocardium.

β-Adrenergic receptors, voltage-operated Ca 2+-channels, nuclear triiodothyronine receptors and triiodothyronine concentration in pig myocardium after long-term low-dose amiodarone treatment

Similar features during chronic amiodarone treatment and hypothyroidism suggest that amiodarone induces a state of "triiodothyronine (T3)-resistance" or "cardiac hypothyroidism", which may predispose the heart to pump failure under conditions with severe strain, such as recovery after cardiac surgery. Disagreements exists as to how amiodarone, and possibly its main metabolite desethylamiodarone, act upon the various receptor systems in the heart. The aim of the present study was to elucidate whether chronic amiodarone treatment leads to a functional reduction in the number of myocardial nuclear T3 receptors, the myocardial tissue T3 concentration and the number of beta-receptors and voltage-operated Ca(2+)-channels. Finally, special attention was drawn to any changes that could contribute to explain previous reports on reduced haemodynamic reserve in animals exposed to severe cardiac strain, such as cardiac surgery. Pigs (72 +/- 2 kg) were assigned randomly to amiodarone treatment (20 mg.kg-1.day-1 for 30 +/- 1 days, N = 8); controls received no medical treatment (N = 6). The left ventricle was evaluated for beta-adrenergic receptors, voltage-operated Ca(2+)-channels, T3 nuclear receptors and tissue T3 concentration. Maximum binding capacity for beta-receptors and Ca(2+)-channels was reduced in amiodarone-treated pigs (by 38%, p < 0.05, and by 52%, p < 0.01) and correlated with tissue drug concentrations for both receptor types (p < 0.05). No changes were observed concerning nuclear T3 receptors. In vitro competition studies revealed that amiodarone, but not desethylamiodarone, possessed binding properties to Ca(2+)-channels, whereas neither of the compounds bound to beta-receptors. Desethylamiodarone, but not amiodarone, competitively inhibited T3 binding to its nuclear receptors. Myocardial T3 was undetectable (< 0.05 nmol/kg wet wt) in amiodarone-treated pigs. From our observations we suggest that the active metabolite desethylamiodarone, rather than the parent drug, is mainly responsible for the observed local hypothyroid-like effects during amiodarone treatment. The observed changes after treatment with low-dose amiodarone in pigs are likely to have biological implications. Functionally, the changes may imply reduced cardiac reserve during conditions of extraordinary strain.

RWJ-24517, a Positive Inotropic Agent, Has Novel Effects on Action Potentials in Guinea Pig Myocardium

RWJ-24517 is a positive inotropic agent whose mechanism of action is under investigation. We examined the effects of RWJ-24517 on guinea pig papillary muscle action potentials and myofilament response to Ca2+. RWJ-24517 increased the fast action potential duration (APD) in a dose-dependent fashion but had no effect on the myofilament response. Tetraethylammonium (TEA 10 mM), which in itself slightly prolonged the control AP, completely blocked the increase in APD75 (APD at 75% of repolarization) and suppressed the increase in APD25 induced by RWJ-24517 (10 microM). Verapamil (5 microM) had little effect on control APs but did decrease the prolongation of the AP induced by RWJ-24517. The increase in APD25 induced by RWJ-24517 was not completely blocked by TEA even with addition of verapamil or 4-aminopyridine (2 mM). RWJ-24517 enhanced Ca(2+)-dependent slow APs elicited by 0.1 microM isoproterenol in preparations depolarized by high extracellular K+ (25 mM), but had no effects on slow APs elicited by 10 mM TEA. These results suggest that the primary electrophysiologic effect of RWJ-24517 is a substantial AP prolongation, which appears to occur largely through a mechanism which is likely to involve inhibition of Ca(2+)-dependent K+ channels. It also appears to have additional effects on some other channels (possibly Na+ channels), which may contribute to the positive inotropic action of RWJ-24517.

Species comparison of adenosine and β-adrenoceptors in mammalian atrial and ventricular myocardium

The antagonist radioligand 1,3-[ 3H]dipropyl-8-cyclopentylxanthine ([ 3H]DPCPX) was used to characterize adenosine A 1 receptors in membrane preparations from atrial and ventricular myocardium of rat, rabbit, guinea pig and pig. K d values in crude membranes from guinea pig atria and ventricles (3.3 and 3.0 nM) were higher than those in the other species (ranges, 1.5–1.8 and 1.5–1.9 nM). B max values were greater in atria than in ventricles in all four species, and in atria and ventricles of guinea pig (76 and 34 fmol/mg), than in the other species (ranges, 15–17 and undetectable to 12 fmol/mg). In contrast, guinea pig K d and B max values for β-adrenoceptors, which were labelled with (−)3-[ 125I]iodocyanopindolol, fell within the range of values for the other three mammalian species. In semipurified membrane preparations from pig, [ 3H]DPCPX and the agonist radioligand [ 125I]-N 6-4-aminobenzyladenosine appeared to label a similar population of receptors and gave comparable K d values in atria (0.73 and 0.66 nM) and in ventricles (0.57 and 0.70 nM). In semipurified preparations from pig, the agonist R-(−)-N 6-2(-phenylisopropyl)adenosine (R-PIA) displaced [ 3H]DPCPX in a manner consistent with the presence of both high- and low-affinity adenosine A 1 receptors. The data from this study indicate that the density of adenosine A 1 receptors in atria is greater than in ventricles, but similar K d values suggest that the A 1 receptor population is the same in the two cardiac tissues. Also, the data demonstrate that the [ 3H]DPCPX antagonist binding characteristics of guinea pig myocardium differ from those in rat, rabbit and pig.

High Selectivity for Inhibition of Phosphodiesterase III and Positive Inotropic Effects of MCI-154 in Guinea Pig Myocardium

MCI-154 (0.3-100 microM) exerted a concentration-dependent positive inotropic effect in isolated guinea pig papillary muscles (EC50 0.8 microM). The efficacy of MCI-154 (253% of predrug value) was 1.7-fold higher than that of saterinone but comparable to that of milrinone. Carbachol markedly reduced the increase in force of contraction (FOC) of MCI-154. In intact contracting papillary muscles, the positive inotropic effect was accompanied by an increase in cyclic AMP content to 0.78 +/- 0.09 pmol/mg wet weight (n = 10), corresponding to 150% of the basal value (0.51 +/- 0.05 pmol/mg wet weight, n = 21) in the presence of submaximal cyclic AMP phosphodiesterase (PDE) isoenzyme III inhibiting concentrations of MCI-154 (30 microM). MCI-154 (1-1,000 microM) concentration-dependently inhibited the activity of PDE III from homogenates of guinea pig myocardium. The IC50 was 3.8 microM. PDE I, II, and IV were not significantly affected up to 100 microM (PDE I and IV) and up to 1,000 microM (PDE II). In comparison, milrinone and saterinone were PDE III/IV-selective PDE inhibitors. Rolipram inhibited PDE IV only. IBMX and theophylline were nonselective PDE inhibitors. MCI-154 had only a marginal positive chronotropic effect. The frequency of spontaneously beating right auricles from guinea pig heart was increased by 8.7% at most (n = 5). MCI-154 increased Ca2+ sensitivity in chemically skinned porcine ventricular muscle fibers.(ABSTRACT TRUNCATED AT 250 WORDS)

Identification of a specific radioligand for the cardiac rapidly activating delayed rectifier K+ channel.

Class III antiarrhythmic drugs show promise as effective treatments for the suppression of potentially lethal cardiac arrhythmias. Dofetilide (UK-68,798), is a potent class III antiarrhythmic agent that is presently under clinical investigation. The objective of this study was to determine whether [3H]dofetilide could be used as a specific radioligand for the rapidly activating delayed rectifier K+ channel of the heart. We find that [3H]dofetilide binds to high-affinity sites on guinea pig cardiac myocytes. Competition studies using unlabeled dofetilide indicate that binding is characterized by an IC50 of 100 +/- 30 nM (mean +/- SD, n = 13). Scatchard analyses of binding indicate a Kd of 70 +/- 6 nM and a maximal binding capacity of 0.30 +/- 0.02 pmol/mg protein. [3H]Dofetilide is displaced from guinea pig myocytes by dofetilide, clofilium, quinidine, sotalol, and sematilide with a rank order of potency that correlates with functional blockade of the rapidly activating delayed rectifier K+ current (correlation coefficient, 0.951; slope, 0.99 +/- 0.19; p = 0.014). High-affinity [3H]dofetilide binding is not detected in rat myocytes, which are devoid of delayed rectifier K+ current. We conclude that [3H]dofetilide specifically binds to sites associated with the rapidly activating delayed rectifier K+ channel of guinea pig myocardium.

Characterization of coexistent histamine H1- and H2-receptor binding sites in the purified guinea pig myocardial membranes from ventricles

In the present work, we identified and characterised histamine H1-and H2-receptors in highly purified myocardial membranes isolated from female guinea pig ventricles. We determined the binding parameters for the interactions of3H-mepyramine with the histamine H1-receptor binding site and3H-tiotidine with the histamine H2-receptor binding site. Binding of both ligands in our study was saturable, reversible and of high affinity. Scatchard's analysis of the specific3H-mepyramine binding revealed the existence of high and low affinity binding sites with apparent KD values of 0.4 nM and 4.5 nM, respectively. The density of binding sites (Bmax) was 100 fmol/mg protein for the high and 466 fmol/mg protein for the low affinity binding site.3H-tiotidine binds to a single population of binding sites with a KD of 1.0 nM and a Bmax of 27 fmol/mg protein. These data suggest that both histamine H1-and H2-receptors coexist in the guinea pig myocardium with a significantly higher prevalence of the histamine H1-receptor population.

Myocardial Phosphoinositides Do Not Share the Same Fatty Acid Profile

It is generally assumed that the fatty acid compositions of the phosphoinositides are identical. To investigate this in myocardium, inositol lipids extracted from rat and pig ventricular homogenates were absorbed to neomycin-coated glass heads, eluted and quantitated by fatty acid analysis after thin-layer chromatography. The percentages of stearic, oleic, linoleic and arachidonic acid (20:4 n-6) in the rat were, respectively, 49, 4, 7 and 26 for phosphatidylinositol, 62, 1, 4 and 18 for phosphatidylinositol-4-monophosphate and 63,2,4, 18 for phosphatidylinositol-4,5-bisphosphate. Equal distribution patterns of fatty acids were found in homogenate and sarcoplasmic reticulum of pig myocardium. Cultured rat ventricular myocytes were utilized to study the incorporation (25 h) of [ 14C]20:4 n-6 relative to that of myo-[ 3H]inositol into phosphatidylinositol and phosphatidylinositol-4,5-bisphosphate which were, respectively, 1.61 and 1.22. The data indicate that in myocardium phosphatidylinositol-4,5-bisphosphate represents a relatively modest source of 20:4 n-6.

Биосинтез белка в бесклеточных системах из миокарда свиньи при аноксии

Биосинтез белка в бесклеточных системах из миокарда свиньи при аноксии

Ultrasound myocardial integrated backscatter signal processing: Frequency domain versus time domain

In the literature, different forms of measuring the ultrasound power returned by myocardial tissue are reported. Frequency domain methods will give the maximum frequency information, whereas time domain methods are limited in bandwidth, but more practical to realize. It was the purpose of this study to compare the various methods of signal processing. High frequency ultrasound signals from a pig's myocardium, digitally recorded during normal contractile performance, were analyzed by six different methods of signal processing to obtain estimates of backscatter power. The myocardial tissue characterization parameters studied were the integrated power as well as its cyclic variation during the cardiac cycle. A total number of 8109 ultrasound traces obtained in 16 pigs were processed. The study included three signal processing methods in the frequency domain: frequency compensated integrated backscatter calculated over both a large (4 MHz, method 1) as well as a small frequency bandwidth (2 MHz, method 2) and uncompensated integrated backscatter (method 3), and three methods in the time domain: high frequency signal squared and integrated (method 4), mean rectified signal level (method 5) and mean signal level after logarithmic compression and envelope detection (method 6). The random measurement variation (including beat-to-beat variation) was analyzed as well as the paired differences of the backscatter parameters obtained by the respective methods as compared with the only theoretically correct method in the time domain (method 4). The magnitudes of the random measurement variation expressed as a standard deviation (SD) were comparable (range 0.93-1.2 dB) except for method 6 (0.61 dB), where the measurement variation is decreased by the logarithmic compression.(ABSTRACT TRUNCATED AT 250 WORDS)

Biophysical parameters of radiofrequency catheter ablation

Radiofrequency catheter ablation has been shown to be an effective treatment for patients with accessory pathways in Wolff-Parkinson-White syndrome and other supraventricular tachycardias. However, the biophysical parameters used so far in vivo did not correlate to the size of myocardial lesions and provided no information about the myocardial wall contact of the electrode. In this study 104 radiofrequency applications were performed on excised pig myocardium in circulating heparinized pig blood as well as in blood alone, and root mean square (rms) voltage, root mean square current and phase angle were measured using a specially developed device. The calculated effective power and output power differed by only 2–7% when measured at the point of maximum current during coagulation. A drop of current following a rise in impedance led to a phase displacement of more than 80° and thereby to a drop of effective power to 17% of the output power. Hence, apparent output power consists mainly of ineffective power. The time dependent variations of phase angle, impedance and current were found to be useful for distinguishing between the media blood and myocardium. These results show that physical parameters measured during radiofrequency catheter ablation may help to control electrode position in the clinical situation and reduce the number and duration of ineffective energy applications.

Characteristics of depressing effect of cycloprotobuxine-A on the maximal velocity of depolarization in myocardium

The characteristics of the depressing effect of cycloprotobuxine-A on V max in isolated guinea pig myocardium were investigated with a standard microclcctrode technique. Cycloprotobuxinc-A 3–30 μmol/l produced a concentration-dependent decrease in V max, associated with a prolongation of the action potential duration. At 3 μmol/l, a small resting block of V max. but a pronounced use-dependent block was found. This use-dependent block increased progressively as stimulation frequency increased from 0.5 to 4.0 Hz. The time constant and rate for onset of use-dependent block were 2.0–7.3 s and 0.11–0.25 AP 1 (reciprocal action potential number), respectively. The time constant and half-life for recovery from use-dependent block were 11.8 and 8.1 s. Thus, the kinetics of the depressing effect of cycloprotobuxine-A on V max were intermediate. In muscles depolarized by high [K +] 0, the resting block was enhanced slightly, while use-dependent block was greatly augmented. These results suggested that cycloprotobuxine-A could have a low affinity for resting Na + channels but a much higher affinity for activated and/or inactivated Na + channels, and that the use-, frequency- and voltage-dependent effects of cycloprotobuxine-A on V̇ max may play an important role in preventing the development of cardiac arrhythmias.

Absence of mitochondrial β-adrenoceptors in guinea pig myocardium: Evidence for tissue disparity

1. 1. Binding sites in guinea pig myocardial tissue labelled by (−)-[ 125I]cyanopindolol (CYP) were investigated using differential centrifugation and autoradiographic techniques. Autoradiographs of myocardial sections (0.1 μm) indicated (−)-[ 125I]CYP binding to sarcolemmal membrane. A low density of binding sites was observed to mitochondria. 2. 2. Binding studies were performed in subcellular fractions. The density of binding sites in the mitochondrial fraction (36.1 ± 9.4 fmol/mg protein) was <10% that in the sarcolemmal membrane (371.7 ± 38.2 fmol/mg protein). The β 1/ β 2-adrenoceptor subtype ratio in the mitochondrial fraction (83.3/16.7) was similar to that in the sarcolemmal fraction (87.1/12.9). 3. 3. Ouabain (100 μM), in the presence of sodium azide (0.4 mM), inhibited a Na +K + stimulated ATPase activity (1.0 ± 0.2 μmol Pi/mg protein/hr reduction), indicating a low but significant level of sarcolemmal contamination of the mitochondrial fraction. 4. 4. The study showed β-adrenoceptors in guinea pig heart are located primarily on the sarcolemmal membrane of myocardium. No evidence was obtained for β-adrenoceptors over mitochondria, as has been suggested in other tissues and species, but that this binding was to sarcolemmal inclusions in the mitochondrial fraction.