John S. Cameron

Delayed afterdepolarizations and triggered activity induced in feline Purkinje fibers by alpha-adrenergic stimulation in the presence of elevated calcium levels.

We studied the ability of alpha-adrenergic stimulation to induce delayed afterdepolarizations and triggered activity in Purkinje fibers from cat hearts in the presence of an elevated Ca++ concentration. Delayed afterdepolarizations could not be induced at drive cycle lengths of 200 to 500 msec in the presence of extracellular Ca++ concentrations of 2.7 to 8.1 mM. However, the addition of 10(-5)M phenylephrine in the presence of 5 X 10(-7)M propranolol elicited delayed afterdepolarizations in eight of 10 preparations at a Ca++ concentration of 8.1 mM; nondrive-triggered action potentials were recorded from three of the preparations. These afterpotentials were completely suppressed by 5 X 10(-7)M prazosin or 10(-6)M phentolamine. In the presence of 5 X 10(-7)M propranolol, 10(-5)M phenylephrine prolonged action potential duration and this effect was suppressed by 5 X 10(-7)M prazosin. Methoxamine, at a concentration of 5 X 10(-6)M, was also observed to potentiate delayed afterdepolarizations in all of three preparations studied. These results demonstrate that alpha-adrenergic stimulation can induce afterpotentials in the presence of elevated Ca++ levels in cat hearts. Stimulation of alpha-adrenoceptors may be responsible for arrhythmias under Ca++-loaded conditions such as ischemia and coronary reperfusion.

Electrophysiologic consequences of chronic experimentally induced left ventricular pressure overload

Cardiac electrophysiologic alterations were evaluated 1 to 8 months after partial supracoronary aortic constriction in cats. This procedure induced left ventricular systolic hypertension and hypertrophy with marked connective tissue infiltration. In situ, premature ventricular complexes were observed during or after vagal slowing of sinus rate in 8 (26%) of the 31 experimental animals, while an additional 3 of the 31 developed ventricular fibrillation. No arrhythmias were recorded in 31 normal or 7 sham-operated cats. In vitro, 29% of the left ventricular preparations from cats with pressure overload and 5% from control cats showed spontaneous ectopic activity. During stimulation at cycle lengths of 800 to 1,000 ms, multiple site impalements of subendocardial muscle cells within fibrotic regions revealed heterogeneous electrical abnormalities. These included short action potential duration, low amplitude action potentials generated from low resting potentials, split upstrokes and electrically silent areas. Impalements in nonfibrotic areas of the left ventricle showed prolongation of muscle action potential duration. Long-term disturbances in cellular electrophysiologic properties may favor the development of arrhythmias and thereby contribute to sudden cardiac death in left ventricular hypertension and hypertrophy.

Dissimilarities in the electrophysiological abnormalities of lateral border and central infarct zone cells after healing of myocardial infarction in cats

We studied the characteristics of an electrophysiological border zone detected after healing of experimental myocardial infarction in cats. Thirty-two isolated left ventricles were studied in tissue bath 2–7 months after distal left coronary artery ligation. Action potentials were recorded from endocardial ventricular muscle cells in normal, lateral border and central infarct zones. Action potential duration was prolonged in central infarct zone cells, while action potentials of lateral border zone cells had the shortest duration. Ventricular muscle cells in the border zone also had lower resting potential, action potential amplitude and Vmax. Slowly rising action potentials (Vmax ≪ 20 V/ sec) were noted in central infarct zone cells, but more consistently in border zone cells. Functional refractory period of cells in central infarct zone was significantly longer than that recorded from border and normal zone cells. Post-repolarization refractoriness occurred in the majority of border zone cells. Failure of a border zone cell to respond to a premature stimulus during repetitive activity was observed in ten of the 22 preparations in which repetitive activity could be induced. Furthermore, when the coupling interval between driving and premature stimuli was shortened, border zone cells were first to fail to be excited by the premature stimulus. These data indicate that conduction was impaired in the border zone, whereas normal conduction was still possible in central infarct and normal areas. The electrophysiological abnormalities in the endocardial lateral border zone cells of the healed myocardial infarction appear to be the most severe, and the border zone may play an important role in chronic electrophysiological instability observed both in situ and in vitro.

A role for nitric oxide in hypoxia-induced activation of cardiac KATP channels in goldfish (Carassius auratus)

SUMMARY Hypoxia-induced shortening of cardiac action potential duration (APD) has been attributed in mammalian hearts to the activation of ATP-sensitive potassium (KATP) channels. Since KATP channels are also present at high densities in the hearts of vertebrate ectotherms, speculation arises as to their function during periods of reduced environmental oxygen. The purpose of the present study was to determine whether nitric oxide (NO) plays a role in cardiac sarcolemmal KATP channel activation during hypoxia in a species with a high degree of tolerance to low oxygen environments: the goldfish (Carassius auratus). Conventional intracellular and patch-clamp recording techniques were used to record responses from excised ventricles or isolated ventricular myocytes and inside-out patches, respectively, from fish acclimated at 21°C. During moderate, substrate-free hypoxia (6.1±0.2 kPa), ventricular APD was significantly shortened at 50% and 90% of full repolarization, a response that was reversible upon reoxygenation and blocked by the KATP channel antagonist BDM. Under normoxic conditions, APD was also reduced in the presence of the NO-donor SNAP (100 μmol l-1). In cell-attached membrane patches, sarcolemmal KATP channel activity was enhanced after 10 min hypoxia, an effect that was reduced or eliminated by simultaneous exposure to BDM, to the guanylate cyclase inhibitor ODQ or to the NO synthase inhibitor l-NAME. In cell-free patches, KATP channel activity was abolished by 2 mmol l-1 ATP but increased by SNAP; the cGMP analog 8-Br-cGMP (200 μmol l-1) also enhanced activity, an effect that was eliminated by BDM. Our data indicate that NO synthesized in cardiac myocytes could enhance sarcolemmal KATP channel activation during moderate hypoxia in goldfish. This response may serve a cardioprotective role by helping to conserve ATP or by reducing intracellular Ca2+ accumulation.

Cardioprotective effects of KATP channel activation during hypoxia in goldfish Carassius auratus

SUMMARY The activation of ATP-sensitive potassium (KATP) ion channels in the heart is thought to exert a cardioprotective effect under low oxygen conditions, possibly enhancing tolerance of environmental hypoxia in aquatic vertebrates. The purpose of this study was to examine the possibility that hypoxia-induced activation of cardiac KATP channels, whether in the sarcolemma (sarcKATP) or mitochondria (mitoKATP), enhances viability in cardiac muscle cells from a species highly tolerant of low oxygen environments, the goldfish Carassius auratus. During moderate hypoxia (6–7 kPa), the activation of sarcKATP channels was indicated by a reduction in transmembrane action potential duration (APD). This response to hypoxia was mimicked by the NO-donor SNAP (100 μmol l–1) and the stable cGMP analog 8-Br-cGMP, but abolished by glibenclamide or l-NAME, an inhibitor of NO synthesis. The mitoKATP channel opener diazoxide did not affect APD. Isolated ventricular muscle cells were then incubated under normoxic and hypoxic conditions. Cell viability was decreased in hypoxia; however, the negative effects of low oxygen were reduced during simultaneous exposure to SNAP, 8-Br-cGMP, and diazoxide. The cardioprotective effect of diazoxide, but not 8-Br-cGMP, was reduced by the mitoKATP channel blocker 5-HD. These data suggest that hypoxia-induced activation of sarcKATP or mitoKATP channels could enhance tolerance of low-oxygen environments in this species, and that sarcKATP activity is increased through a NO and cGMP-dependent pathway.

ATP-sensitive K+ Channels in Cardiac Muscle from Cold-Acclimated Goldfish: Characterization and Altered Response to ATP

ATP-sensitive potassium channels (K(ATP)) play an important, if incompletely defined, role in myocardial function in mammals. With the discovery that K(ATP) channels are also present at high densities in the hearts of vertebrate ectotherms, speculation arises as to their function during periods of cold-acclimation and depressed ATP synthesis. We used single-channel and intracellular recording techniques to examine the possibility that channel activity would be altered in cardiac muscle from goldfish (Carassius auratus) acclimated at 7+/-1 degrees C relative to control (21+/-1 degrees C). As previously observed in mammals, K(ATP) channels in isolated ventricular myocytes were inwardly rectified with slope conductances of 63 pS. However, channel mean open-time and overall open-state probability (Po) were significantly increased in cells from the cold-acclimated animals. In addition, K(ATP) channels in cells from fish acclimated at 7 degrees were nearly insensitive to the inhibitory effects of 2 mM ATP, whether studied at 7 or at 21 degrees C. Transmembrane action potential duration (APD) in hearts of cold-acclimated fish studied at 21 degrees was significantly shorter than that observed in hearts of warm-acclimated fish at the same temperature; this difference was eliminated by the K(ATP) channel antagonist glibenclamide (5 microM). These data suggest that K(ATP) channels in the hearts of cold-acclimated animals are more active and less sensitive to ATP-inhibition than those in warm-acclimated fish, possibly reflecting a functional adaptation to promote tolerance of low temperatures in this species.

Cellular electrophysiologic changes and "arrhythmias" during experimental ischemia and reperfusion in isolated cat ventricular myocardium.

The cellular electrophysiologic consequences of both regional and global experimental ischemia and reperfusion were studied in the isolated cat myocardium, using conventional microelectrode techniques. Oxygenated Tyrodes solution was perfused through the left anterior descending and circumflex coronary arteries, while the preparation was superfused with Tyrodes solution gassed with 95% nitrogen and 5% carbon dioxide. Electrophysiologic characteristics of endocardial muscle cells were normal during coronary perfusion. When perfusion was discontinued for 30 minutes, resting membrane potential was decreased by 21.6 +/- 4.1%, action potential amplitude was decreased by 29.1 +/- 8.6% and action potential duration was decreased by 54.1 +/- 12.5% (p less than 0.001). Ectopic activity occurred after 5 to 10 minutes of ischemia and was more frequent in regional than in global ischemia (p less than 0.05). Rapid ventricular activity was observed in only 5 (17%) of 29 preparations during ischemia, whereas it occurred in 24 (83%) of 29 preparations during reperfusion. Rapid ventricular activity began 5 to 40 seconds (mean 19) after the start of reperfusion, stopped spontaneously after a mean of 113 +/- 211 seconds and occurred after both regional and global ischemia. The cellular electrophysiologic changes induced by ischemia returned to baseline values within the next 5 minutes. Repeated ischemia and reperfusion runs reproduced the same electrophysiologic changes and rapid ventricular activity. Coronary perfusion with procainamide (20 mg/liter) aggravated the ischemic depressions of action potential amplitude and action potential duration and increased conduction delay during ischemia, but it did not prevent rapid ventricular activity induced by reperfusion. In contrast, verapamil (1 mg/liter) perfusion did not affect the changes in action potential variables during ischemia but prevented reperfusion-induced rapid ventricular activity. Perfusion with calcium ion (Ca2+)-free Tyrodes solution just before ischemia and during reperfusion slowed or prevented reperfusion-induced rapid ventricular activity, without affecting the action potential changes during ischemia. It is concluded that, in these isolated perfused ventricular muscle preparations, different mechanisms may be operative in ischemic and reperfusion arrhythmias and Ca2+ may play an important role in the development of arrhythmias during the reperfusion phase of ischemia/reperfusion sequences.

A clinically relevant liver injury grading system by CT, preliminary report

Current computed tomography (CT) grading scales are anatomic and do not reliably identify those liver injuries requiring intervention (surgery or angioembolization). We propose a clinically relevant CT grading system that could predict need for intervention. CT scans of 11 patients with hepatic injury were reviewed to establish criteria that correspond with intervention. Five features were identified that were associated with intervention: laceration in greater than or equal to three segments, laceration extending into the hilum, hemoperitoneum, active extravasation, and sentinel clot. Radiologists then evaluated the predictability of these criteria by analyzing 24 CT scans. Inter-observer agreement of the American Association for the Surgery of Trauma (AAST) grading system was compared to this new system. In the analysis of 24 CT scans, active extravasation and sentinel clot demonstrated the highest specificity for intervention. This new grading system had superior inter-observer agreement (k=0.56) as compared to the AAST grading system (k=0.47). Active extravasation and the presence of sentinel clot should form the foundation of a new liver grading system.

Regional variations in myosin heavy chain concentration after healing of experimental myocardial infarction in cats

Densitometric scanning of SDS-polyacrylamide gels was used to measure myosin heavy chain concentration in left ventricular specimens obtained from cat hearts 3 to 12 months after healing of small experimental myocardial infarctions. The study was designed to test the hypothesis that myosin concentration varies as a function of anatomic proximity to the infarct scar. Myosin heavy chain concentration was elevated in non-scarred areas adjacent to a healed infarct and normal in areas remote from the scar. The scar itself had reduced concentrations, reflecting the loss of muscle mass in this area. The increased myosin heavy chain concentration in regions adjacent to the scar may be an attempt to regulate or compensate for the decrease in mechanical function of the scarred area.

Regional reduction in ventricular norepinephrine after healing of experimental myocardial infarction in cats

Regional ventricular norepinephrine and myosin heavy chain concentrations were measured in two models of healed left ventricular myocardial infarction in cats. One model was characterized by a well-defined dense transmural scar (discrete myocardial infarction), while the other demonstrated a pattern of nontransmural diffuse patchy fibrosis in the infarct area (diffuse myocardial infarction). Norepinephrine and myosin heavy chain concentrations were measured in the scarred area, the non-infarcted zone surrounding the scar(s), and in sites remote from the scar. Corresponding tissue sites from unoperated animals and sham operated animals served as controls. Myosin heavy chain concentration was used as an index of surviving muscle mass to express norepinephrine concentration. Norepinephrine concentration, as a function of crude tissue mass, was significantly reduced in both the scarred tissues and the non-scarred tissues surrounding the scar in the discrete infarction model but was significantly reduced only in non-scarred tissues adjacent to the dense scar when expressed as a function of myosin heavy chain. The heavily scarred area of the discrete preparation approached normal values when corrected for myosin heavy chain content. The diffuse infarct preparation demonstrated normal norepinephrine concentration at all three sites studied, whether expressed as a function of tissue mass or myosin heavy chain. These data indicate a long-term regional reduction in norepinephrine concentration specific to non-infarcted tissues adjacent to a dense transmural myocardial infarction scar. This regional reduction in norepinephrine concentration corresponds to reported regions of increased sensitivity to sympathetic nerve stimulation in the discrete myocardial infarction model.