Long-term inhibition of NO synthase induces cardiac hypertrophy with a decrease in adrenergic innervation.

Data concerning the effect of NO on the function and structure of the heart are controversial. We have studied two main questions: (i) Does the heart muscle reflect the hypertension induced by long-term inhibition of NO synthase? (ii) Since the arginine-NO pathway is also operative in the autonomic nervous system, the second goal was to ascertain the possible changes of the adrenergic nervous system in the heart after long-term NO synthase inhibition. Wistar rats were administered L-NAME in drinking water (50 mg/kg bw/day) for 8 weeks. Systolic blood pressure and heart rate were monitored weekly. The heart/body weight ratio were determined at the end of experiment. The adrenergic nerve terminals visualized by histochemistry were counted according to Haug's point counting method. Blood pressure increased significantly in L-NAME-treated rats. No changes were found in the heart rate. Heart/body weight ratio increased markedly. Surprisingly, the density of adrenergic nerve terminals did not alter accordingly. The density of adrenergic nerve terminals in the left ventricle and septum decreased but no significant changes were found in the left atrium and the right ventricle. Hypertension due to NO deficiency induced cardiac hypertrophy that was characterized by a decline in the density of adrenergic innervation of the overloaded left ventricle and septum.

Biomechanical signals in the coronary artery triggering the metabolic processes during cardiac overload.

Peculiarities in structure and deformability of epicardial conduit coronary arteries are described. The thin wall of animal coronary artery contrasts the human coronary artery in which the remarkable wall thickness is due namely by the intima thickness. Deformation in length and diameter of conduit coronary arteries, due to the left and right ventricle volume increase, has been defined in non-beating canine heart. Ramus interventricularis anterior being firmly tethered to the myocardium undergoes about 3 times larger deformation than ramus circumflexus. In anaesthetized dogs a 30% increase in blood pressure, elicited by aortic constriction, induces an increase in diameter of coronary artery, in segment length, in blood flow and consequently in shear stress which represents a load for circumferentially running smooth muscle bundles, longitudinally running smooth muscle bundles, as well as for the endothelium. The above load lasting 4 h is already reflected by an increase in total RNA content and [14C] leucin incorporation in the left ventricle myocardium in the wall of ramus interventricularis anterior, not in ramus circumflexus. The findings fit completely with the different range of deformation of both the above coronary branches and indicates an increase in proteosynthesis not only in myocardium, but in ramus interventricularis anterior as well. An increase in ornithindecarboxylase activity in coronary wall leading to an increase in biogenic polyamines, is present in the case only, when blood pressure increase is induced by infusion of noradrenaline.

Instrumentation of ramus circumflexus compromises the innervation of coronary artery and myocardium.

A fine non-compressing methacrylate ring was implanted on the ramus circumflexus (RC) in 13 dogs. After the animals survived 7-14 days, the distribution and the rate of the Wallerian degeneration was studied in coronary arteries and myocardium by means of the formaldehyde induced fluorescence (FIF) technique and the transmission electron microscopy. The density of vasomotor innervation in the main arterial trunks was expressed as the number of transsected terminals per 1 mm2 of the transversally sectioned media; the density of adrenergic terminals in the myocardium was determined by the point-counting method. A compact scar encompassing the ring was found. The scar compressed some perivascular nerves. As a consequence, by about 40% of terminals degenerated in the RC. The innervation of the branching of this artery was compromised as well. Moreover, the density of nerve terminals in myocardium decreased around the whole circumference of the ventricles and in the septum, equally by 29%. The value was lower than that found after the instrumentation of ramus interventricularis anterior (50%). The results indicate (i) a compromise of the innervation of coronary artery and myocardium after the instrumentation of RC; (ii) the different decrease in density of nerve terminals in the myocardium suggests that majority of adrenergic nerve supply for ventricle myocardium is running along ramus interventricularis anterior.

Expression of phosphorylated high molecular weight neurofilament protein (NF-H) and vimentin in human developing dorsal root ganglia and spinal cord.

The expression of vimentin and the phosphorylated variant of high molecular weight neurofilament protein (NF-H) was studied in developing human fetal dorsal root ganglia and spinal cord. The technique used for examination of cryosections was double-label fluorescence with monoclonal antibodies. Both proteins were present in the nerve fibres inside the ganglia of 6- and 8-week-old embryos. During further development the expression of vimentin continued to increase in the satellite cells, but was found to be decreasing in the ganglion cells. Phosphorylated NF-H was found in the processes of ganglion cells, as well as in the perikarya at all developmental stages. In the spinal cord of 6- and 8-week-old embryos, phosphorylated NF-H protein was found in the longitudinal fibres of the marginal layer and in processes of the mantle zone; some of the fibres also contained vimentin. Later the co-expression of the two proteins ceased and vimentin was found only in glial and mesenchymal derivatives. Phosphorylated NF-H was located, at all developmental stages, in the axons of both white and grey matter, but not in the neuronal perikarya. The results indicate that phosphorylation of the NF-H in human dorsal root ganglia starts in the perikarya of the ganglion cells while in the ganglion cells of the spinal cord it takes place in the axons.

Contractility of the rabbit abdominal aorta 4 days after endothelium denudation.

The purpose of this study was to evaluate contractility of the rabbit abdominal aorta 4 days after de-endothelialization by balloon catheter. The isometric tension of ring-segments in response to vasoactive agents was monitored. A significant enhancement of contraction to noradrenaline and serotonin was found in aortas 4 days after endothelium denudation as compared with controls with endothelium. The enhancement, however, did not differ from that found already in acutely denuded vessels (immediate denudation). No significant difference in contractility to potassium chloride was found in either group of denuded preparations as compared with controls. The sensitivity to all three vasoactive agents (EC50) was not influenced by denudation. These results indicate that changes in the contractility of denuded vessels are predominantly a consequence of lacking the endothelium as a producer of endothelium-derived relaxing factor. The access of mitogens to the media does not seem to interfere with the magnitude of contraction 4 days after denudation.

Diminished release of atrial natriuretic factor in calves with a total artificial heart.

Venous hypertension after total artificial heart (TAH) replacement, a common problem in humans as well as experimental animals, is thought to be related to reduced release of atrial natriuretic factor (ANF) due to atrial damage. To verify this hypothesis, we stimulated the release of ANF in 6 calves before (-7 days) and after TAH replacement (+25, +50, +80, and +100 days) by the rapid (+ = 10 min) infusion of 2 L of Ringer's solution. In normal calves (-7 days) this procedure induced a rise in plasma concentration of ANF from 18.5 +/- 12 to 31.6 +/- 12 pmol/L (p < 0.05). After TAH the ANF release continuously decreased (analysis of variance, p < 0.02). Thus, ANF increased by 9.8 +/- 9.3, 6.8 +/- 17.5, 0.4 +/- 5.2, and 0.5 +/- 3.2 pmol/L at days 25, 50, 80, and 100, respectively. The central venous pressure increased by 3.4 +/- 1.5 mm Hg (before TAH) and by 6 +/- 2.9 mm Hg (after TAH) during this procedure. A decrease in plasma concentrations of aldosterone, most likely due to extracellular volume expansion and a decrease in total protein plasma concentrations and in hematocrit due to the dilution of blood, was seen in each experiment. Necropsy demonstrated massive atrial dilatation and myocardial dystrophy with atrial fibrosis. ANF granules were present to a small extent in every calf. We conclude that rapid intravenous infusion is an adequate stimulus to release ANF in normal calves. After TAH implantation the release of ANF by high venous pressure declines although ANF granules are still present in the damaged atrial stumps. Therefore, the loss of stimulated ANF could contribute to the observed increase in venous pressure in TAH calves although other pathophysiologic mechanisms cannot be excluded.

[Hemostatsis during liver resection using the Leister Hot-Jet coagulator]. / Stavení krvácení pri resekci jater pomocí Leister Hot-Jet koagulátoru.

The authors present their own experience with the use of a hot-jet coagulator in resections of the liver. The principle is based on contact-free transmission of thermal energy by means of a current of hot air to achieve haemostasis and tissue coagulation. They mention the use of the apparatus in animal experiments and in resections of the liver in nine patients. They recommend the procedure as one possibility of haemostasis where conventional methods do not affect haemostasis.

Expression of high-molecular-weight neurofilament protein and vimentin in human developing spinal cord and dorsal root ganglia.

Expression of intermediate filament proteins in developing human dorsal root ganglia and spinal cord was studied using double-label immunofluorescence on frozen sections with monoclonal antibodies to vimentin and high-molecular-weight neurofilament protein (NF-H). Both proteins were found in the nerve fibres inside the ganglia of 6- and 8-week embryos. In 9-14-week embryos the NF-H protein was found in the perikarya of ganglion cells. Vimentin was only sporadically present in these cells, but was accumulated in connective tissue cells. In the following development the NF-H protein expression was limited to the ganglion cells and processes, and vimentin was confined to the satellite cells and mesenchymal derivatives. In the spinal cord of the 6-week and 8-week human embryos NF-H protein was found in the longitudinal fibres of the marginal layer and in a loose network of nerve fibres in the mantle zone. Some of the nerve fibres were also vimentin positive. The coexpression of both proteins stopped during weeks 9-10. In 30-week foetuses vimentin was only found in glial cells and mesenchymal derivatives, while NF-H protein was in the axons of both white and grey matter but not in neuronal perikarya.

Evaluation study of calves with total artificial heart (TAH) surviving for 218-293 days of pumping.

Evaluation of five longest surviving calves among a total of 54 longterm survivors, at the Brno Research Centre was performed. These calves were treated by administration of antihypertensives and partially by electrical stimulation of the right atrium to reduce central venous pressure (CVP). Treatment for CVP was effective, as compared with two groups of 5 calves each not treated in this way, and it is assumed to be an important factor for prolongation of survival. Mineralization of driving diaphragms was present in all these calves. No preventive measures of this pathologic change were performed in this group. Another complication is hemolysis, which is partly caused by immune mechanisms initiated by administration of alpha-methyl dopa and partly by mechanical destruction due to a heavily mineralized driving diaphragm. The prevention of mineralization is another important problem for future research. Other problems, concerning the survival of TAH recipients are discussed.

Mineralization of polyurethane membranes in the total artificial heart (TAH): a retrospective study from long-term animal experiments.

The degree of mineralization of TAH membranes in animals is directly related to the survival time of animals with a TAH. Large differences in mineralization of the membranes among individual animals have been observed. These differences can be explained by the simultaneous presence of infection on biomaterials (polyurethane) as well as by different levels of natural inhibitors of mineralization. No positive influence of applying sodium warfarin for the inhibition of the mineralization was observed.

Development of the adrenergic innervation in the myocardium and coronary arteries of the dog.

The development of the adrenergic cardiac innervation was studied in premature dog fetuses, puppies and adult dogs by means of the formalin-induced fluorescence technique. A point-counting technique was used to evaluate the density of innervation. Two types of fluorescent profiles can be observed in the heart during development: (1) sprouting axons, and (2) beaded terminals. The axonal fluorescence disappears in adult neurons. A different morphology and a different time course of development enable to study separately the innervation of the myocardium (cardiomotor innervation) and that of the vessels (vasomotor innervation). The late prenatal innervation is very poor (0.1 hit). The first but very scant cardiomotor terminals appear in this period. A mature cardiomotor innervation is found in 4-month-old puppies [1.5 +/- 0.3 (SD) hits]. The vasomotor innervation is shifted to the right. The development of beaded vascular terminals begins and matures 1-2 weeks later. The growing fluorescent axons reveal that the myocardium is supplied by axons of the cardiac plexus and of the perivascular nerves; the vascular wall, on the other hand, is supplied by the perivascular nerves only. The developmental, spatial and morphological differences in innervation suggest that two different types of neurons exist in the sympathetic ganglia: (1) neurons innervating the vessels (coronaromotor neurons), and (2) neurons innervating the myocardium (cardiomotor neurons).

Recent efforts in artificial heart research in Czechoslovakia.

The authors present basic information about the technical development of the total artificial heart (TAH) and TAH drivers. Long-term experiments, in which the TAH TNS-Brno-II and VII and the Rostock hearts were implanted, are described. Long-term experiments were dedicated to analysis of specific pathophysiologic problems (venous hypertension, thrombogenesis, and calcification) in order to increase survival rates. Thrombogenesis has been partially solved in the TNS-BRNO-VII device by optimal TAH construction using an asymmetric driving diaphragm, the undulating motion of which produced excellent results. Two methods for prevention of increase in central venous pressure (CVP) are verified: administration of antihypertensive drugs and atrial electrical stimulation. Of 155 calves studied, 50 long-term experiments in calves and one in a goat were done. The survival of these animals extended from 31 to 293 days of pumping (average survival, 116.9 days). To prevent driving diaphragm calcification, we have incorporated exogenic inhibitors of calcification into the diaphragm biomaterial (polyurethane). A unique long-term experiment with intrathoracic TAH implantation in the goat is also described. In addition clinical use of the TAH as a bridge to transplantation is discussed. Parallel research with the left ventricular assist device (LVAD) system was studied experimentally, and twice used in patients. The TAH TNS-BRNO-VII/80/clin/ was used in four patients. It worked faultlessly for 22 hours to 10 days. More precise and specific indications for TAH implantation when used as a bridge to transplantation are needed.

Therapy of venous hypertension in calves with total artificial heart (TAH).

Vasomotor disregulation, preponderantly expressed by a pathological increase of central venous pressure (CVP) in calves with total artificial heart (TAH), starts to be evident from about the 50th day of pumping. The main cause of this state is an imbalance in cardiac receptor areas. Ventricular vasodepressor mechanisms are eliminated with the ventricular tissue, which is replaced by the artificial blood pump. In the stumps of both atria, which remain in situ, all neural elements disappear immediately after TAH implantation, but within two months they are fully regenerated. Regenerated atrial receptors are the starting points of afferent neural stimuli, which in the vasomotor center of the brainstem, increase the activity of the vasoconstricting functional component. A general tendency to vasoconstriction, now not well counterbalanced, increases, and the progressive venous hypertension causes loss of liver function and morphology. Two therapeutic approaches were tried: afferent therapy by atrial electrical stimulation, and efferent therapy by the administration of antihypertensives. Both kinds of this therapy were sufficiently effective in reducing CVP, protecting the liver, and prolonging average survival.

Sympathetic nervous system and age-dependent salt hypertension in Brattleboro rats.

The effects of early sympathectomy on the development of salt hypertension were studied in prepubertal and adult rats with hereditary diabetes insipidus (DI). Early guanethidine administration caused a pronounced and long-term destruction of sympathetic nervous system (SNS) in Brattleboro rats in which blood pressure (BP) was significantly decreased until the age of 22 weeks. This SNS impairment did not abolish the age-dependent BP response of salt-loaded rats that was still greater in young than in adult sympathectomized DI rats. BP of young uninephrectomized DI rats was higher in the late than in the early phase of salt hypertension development. The early sympathectomy lowered BP and increased mortality in all groups of saline drinking DI rats except young uninephrectomized animals in which hypertensive response was attenuated but not prevented. It could be suggested that 1) increased BP response of young rats to high salt intake occurs even in animals with attenuated principal pressor systems, 2) the effects of early sympathectomy on the development of salt hypertension depend on the actual hemodynamic pattern, and 3) moderate BP increase might be a part of homeostatic mechanisms defending the organism threatened by chronic salt overload.

Innervation of atrial stumps in calves living with the TNS-Brno II artificial heart.

The remnant stumps of the atria in six calves surviving with the artificial TNS-Brno II methacrylate heart for up to 5 months were examined by means of the Bodian silver-impregnating technique and by the formalin-induced fluorescence technique visualizing noradrenaline in monoaminergic nerve terminals. The degeneration of the nerve terminal system detected in animals surviving for 2 days and 1 week was followed by regeneration of the vegetative nerve system in four animals surviving for greater than 1 month. Moreover, thick myelinated nerves afferent in character were detected within the atrium of one animal surviving for 5 months.