Catecholamine-induced cardiac hypertrophy uncouples beta-adrenoceptors from slow calcium channels.

Changes in the parameters of Ca(2+)-dependent slow action potentials (APs) and in their sensitivity to noradrenaline, forskolin, dibutyryl-cAMP and extracellular Ca2+ concentration were studied and compared in left ventricular trabeculae from normal control rats and rats with cardiac hypertrophy. Cytochemical studies were also carried out to determine changes in the activity of membrane-bound adenylate cyclase. Hypertrophy was induced by administration of 5 mg/kg isoproterenol once daily for 7 days. In hypertrophied cardiac muscle, the overshoot of the slow APs was increased by 75%, the maximum rate of rise (Vmax) increased by 76% and the AP duration at 50% repolarization (APD50) prolonged by 56%. The Vmax, an indicator of the slow inward Ca2+ current, increased, in a dose-dependent manner, in response to the beta-adrenoceptor agonist noradrenaline, the adenylate cyclase activator forskolin, the protein kinase activator cAMP and elevated Ca2+ concentration in normal control preparations, whereas in hypertrophied myocardium, the beta-agonist noradrenaline and the adenylate cyclase activator forskolin had no effect. In cytochemical studies with ATP as substrate, adenylate cyclase activity was localized in the sarcolemma, and significantly fewer reaction products appeared on the outer side of the cell membrane in hypertrophied myocytes than in control myocytes. The results suggest that catecholamine-induced cardiac hypertrophy damages the catalytic subunit of membrane-bound adenylate cyclase, thus uncoupling beta-adrenoceptors from slow Ca2+ channels in the transmembrane signalling process.

Ultrastructural and electrophysiological alterations during the development of catecholamine-induced cardiac hypertrophy and failure.

Cardiac hypertrophy and failure were induced in male Wistar rats by daily administration of 5 mg/kg isoproterenol for three weeks. Age-matched animals were used as normal control. To estimate the degree of hypertrophy, the wet heart weight (HW) to body weight (BW) ratio (HW/BW) was used as an index of the myocardial enlargement. By the 7th day of the treatment, the HW/BW ratio was increased to 4.24, as compared with the control value of 3.11. In this early stage of cardiomyopathy, the structure was characterized with small necrotic foci, enlarged myofilaments and swollen mitochondria. The electrical activity showed broadened action potentials with an elevated plateau phase, and increased membrane resistance and time constant. The amplitude of the twitch contractions was elevated. Continuing the treatment of the animals with catecholamine caused a decompensated heart failure by the 21st day. In this late stage, many and large necrotic foci could be observed in the myocardium. The mitochondria were fragmented, and the resistance of the sarcolemma decreased, and the electrical and contractile activity suppressed. The results indicate that an electrically and structurally compensated cardiac hypertrophy model can be produced by a short-term treatment of the animals with isoproterenol, while a long-term treatment causes a decompensated heart failure.

Localization of antigenic components in proteoglycan aggregate of bovine nasal cartilage.

Proteoglycans and glyco(link)proteins are demonstrated in the cartilage matrix using immunohistochemical reactions, ruthenium red staining and concanavalin A-peroxidase procedure. Specific antibodies against proteoglycan monomers revealed a loose matrix structure in the interterritorial area of nasal cartilage. Thin filaments of 49-87 nm in length with a knob on one end corresponding to the protein core of proteoglycan monomers were found in irregular contacts with collagen fibres. Following hyaluronidase digestion the immunohistochemical reactions became more intense, and the matrix structure is suggestive of a network of single filaments which are presumably coupled together longitudinally at the sites of small matrix granules. These matrix granules proved to be glyco(link)proteins of proteoglycan aggregate. Immunohistochemical reactions combined with other methods can reveal an in situ structure of proteoglycan aggregate of hyaline cartilage, which contributes substantially to what has been known about the proteoglycan aggregates on the basis of physico-chemical data and has been verified in monomolecular electron microscopic specimens. The enzymatic treatments of cartilage slices suggest that some of the partially digested proteoglycan monomers are required to be present for the preservation of the structural integrity of cartilage tissue.

Primary afferent terminals in the nucleus of the solitary tract of the frog: an electron microscopic study.

In the frog solitarius nucleus, primary afferent terminals of the facial and glossopharyngeal-vagal nerves were identified with cobalt labelling and electron microscopy. The labelled terminals were grouped in two main categories, one with small (1-2 micron) and pale terminals, and another with large (3-5 micron) and dark terminals. The small terminals greatly outnumbered the large ones. In addition many terminals intermediate in size and staining reactions were found. All kinds of labelled boutons contained medium-size clear synaptic vesicles, among which dense-core vesicles of the smaller type frequently occurred. The labelled primary afferent terminals established axo-dendritic contacts of the asymmetric type. Close to these contact sites they were themselves very frequently contacted by a profile interpreted as presynaptic in relation to them. Such profiles contained spherical, pleomorphic (including dense-core) or flattened vesicles; a fourth kind was interpreted as presynaptic dendrites. It is concluded that viscerosensory fibres, as opposed to somatosensory fibres, predominantly generate small and lightly stained terminals. It is likely that the effect of synaptic transmission at the solitarius tract terminals is modulated in a very versatile manner by the various presynaptic profiles converging on these terminals.

Fine structure of dorsal root terminals in the dorsal horn of the frog spinal cord.

Dorsal root fibers were labelled with cobalt and their terminals were identified in an oval-shaped area and in its lateral extension of the dorsal horn. These areas receive mainly cutaneous afferents. A number of labelled terminals exhibited either an electron lucent and swollen, or an electron dense and shrunken form of degeneration. The majority of the labelled terminals could be classified into three groups. Type A terminals were small, less than 2 microns in diameter, and lightly stained. Type B terminals were larger and neurofilaments and an electron dense material resulted in a dark staining of the bouton. Type C terminals contained medium size granulated vesicles. The type A and type B terminals were frequently found in postsynaptic positions. The presynaptic profiles contained spheric or elliptic vesicles, or they exhibited the structural characters of presynaptic dendrites. Preterminal dorsal root fibers, identified on account of their cobalt label, were also found in postsynaptic positions to the same kinds of presynaptic profiles. It is concluded that cobalt labelling is sufficiently selective for a reliable identification of dorsal root terminals. No correlation was found between the different forms of degeneration and the different types of terminal boutons. The possible functional significance of the postsynaptic relation of dorsal root fibers and terminals to a variety of presynaptic profiles is discussed.

Histological and histochemical investigations of the epiphyseal cartilage in rats after administration of heparin, coumarin as well as coumarin and diphosphonate (EHDP).

Heparin and coumarin (Marcumar) as well as coumarin combined with a diphosphonate (EHDP) were examined histologically and histochemically with regard to their effect on the structure of the epiphyseal cartilage in rats. Topo-optical reactions were used to assess the submicroscopic structural changes in the developing trabeculae of metaphysis. The zone of resting cartilage and the zone of young proliferation cartilage in epiphyseal plates revealed a reduced cell count due to the influence of anticoagulants. With regard to the matrix of the epiphyseal plate, it may be stated that its ultrastructure appeared partly regular and that, on the other hand, large areas of degeneration were detected in the connective-tissue. The negative effect of anticoagulants on the ground substance could not be reduced definitely by means of a diphosphonate (EHDP) in conjunction with Marcumar which was administered simultaneously.

The localization of proteoglycans and glycoproteins in the hyaline cartilage.

Antibodies to proteoglycan (PG) and glycoprotein of bovine nasal cartilage were conjugated with fluorescein isothiocyanate and with horseradish peroxidase. Hyaluronidase digestion of cartilage tissue-specimens increased the intensity of immune reactions; pronase digestion or extraction with 4 M guanidinium chloride abolished the staining. In the intercellular matrix fine filaments beaded with small granules were seen forming an irregular network. The interstices of the network are filled with collagen fibers linked together by the filaments and granules. In view of the linear conformation of core proteins of PGs and the globular conformation of glycoproteins (link proteins), it may be supposed that the granules and filaments represent these two protein components of PG-aggregates. In chondrocytes a homogeneous staining was recorded in the endoplasmic reticulum, in the juxtanuclear areas and in several smooth-walled vesicles and elongated areas situating subjacent to the cell membrane. In contrast to the extracellular immune reactions, this homogeneous intracellular staining was never enhanced by hyaluronidase digestion. This is interpreted in the sense that conformation changes of molecules secreted, and the aggregation of PGs, occur extracellularly.

Influence of hypothermia on the generalized Shwartzman reaction.

In normothermic rabbits the generalized Shwartzman reaction can usually be elicited by two intravenous injections of endotoxin spaced 24 hr apart. The reaction could be prevented by cooling the rabbits. Hypothermia is effective only at the time of the second injection. The protective effect can clearly be demonstrated by gross and histologic examination. The possible mechanism of the protective action of hypothermia is discussed.

Effect of triiodothyronine (T3) thyroparathyroidectomy and mercaptoimidazole treatment on enchondral bone growth. II. Growth dynamics in the proximal growth organ of the tibia in young rats.

The 24 hour growth rate of the tibial proximal growth organ was investigated in triiodothyronine or mercaptoimidazole-treated, and in thyroidectomized male rats. Longitudinal growth of the tibia of the same period was also studied. After one week treatment, the 24 hour rate of growth in the triiodothyronine-treated rats was retarded as compared with the controls, whereas at the end of the 3rd and 5th weeks the growth rate of the treated animals exceeded that of the controls. The tibia was somewhat longer in the treated rats than in the untreated controls, though the difference was not significant. The growth rate in the growth organ of mercaptoimidazole-treated and thyroidectomized animals was generally higher than that measured in the controls. The length of the tibia was less than in the controls, but the difference was not significant. The phenomenon is attributed to the altered hormonal conditions.

Effect of triiodothyronine treatment, thyroparathyroidectomy and mercaptoiminazole treatment on enchondral bone growth. Changes in the histological structure of the growth organ.

Histological changes caused by triiodothyronine (T3) and mercaptoiminazole treatment as well as by thyroidectomy have been studied in the proximal growth organ of the tibia of growing rats. On triiodothyronine treatment morphometric examinations revealed an increased proliferation and resorption of cartilage associated with a transitory acceleration of linear bone growth. Administration of mercaptoiminazole and thyroidectomy inhibited cartilage proliferation and resorption resulting in a slowing down of bone growth.