Nuclei and Nuclear Core Region

Nuclei of myocardial cells are often, but not always, located toward the center of the cell. Nuclei are more or less fusiform and conform to the longitudinal arrangement of major organelles such as myofibrils and mitochondria. The elongate form of myocardial nuclei is maintained in part by numerous microtubules at their periphery (see Section II,E), as well as by the surrounding myofibrils. Extending from each pole of the nucleus is a roughly conical region of myofibril-free myoplasm containing a variety of organelles, including the Golgi system, centrioles, mitochondria, endoplasmic reticulum, and lysosomal bodies. In atrial working cells (Fig. 3), the Golgi system is extensive, appearing in selectively contrasted thick sections as an array of tubules and sacs that proliferate at both nuclear poles and connect through elements that sweep longitudinally along the length of the nucleus (Rambourg et al., 1984). The nuclear pole regions of these atrial cells are characterized by prominent spheroidal, electron-opaque specific atrial granules (Fig. 3), which react strongly when exposed to antibodies prepared against the vasoactive substance atrial natriuretic protein (ANP). The region housing nuclei and their associated myoplasmic zones can be viewed as a sort of cell core, and a similar central region free of contractile elements is a characteristic of vascular smooth muscle cells (Forbes, 1982, 1995).

E. Fibrillar Components 1. Contractile Apparatus

As pointed out earlier, the collections of myocardial cell contractile proteins, known as myofibrils (or some times ''myofilamentous masses''), form the major portion of the muscle cell. Archetypal "tonic" skeletal muscle, such as the frog sartorius, exhibits an extreme degree of regularity of its myofibrils, each of which is cylindrical, uniform in diameter, and closely aligned with its neighbors in terms of sarcomere register. The presence of uniform, small-diameter myofibrils is said to constitute a Fibrillenstruktur, whereas larger, amorphous masses of myofilaments are termed Felderstruktur. In the fast-beating hearts of such small mammals as shrew and mouse, the myofibrils of working cardiomyocytes tend to be of smaller diameters (i.e., more nearly a Fibrillenstruktur; e.g., see Fig. 3) than those of cells in slower-beating hearts (e.g., dog, monkey), whose myofibrils are thicker and more variable in cross-sectional profile (Felderstruktur) (Figs. 4 and 5). An obvious consequence (or, perhaps, advantage) of having a contractile system of Fibrillenstruktur conformation is the conferral of a larger expanse of myofibrillar surface area per unit cell volume; applied to these myofibrillar surfaces is an extensive system of anastomosing tubules and saccules of sarcoplasmic reticulum, which is responsible for movements of calcium into and out of the underlying myofibrils—the very basis of the contractile cycle of the cell.

The geometric interposition of various contractile proteins (notably actin, myosin, troponin, tropomyosin, and a-actinin) gives myofibrils a characteristic repeating pattern of stripes or striations whose basic unit is known as the sarcomere (Figs. 1 and 4). Sarcomeres in mammalian ventricular cells have a characteristic resting length of approximately 2.2 ^m. Their most evident components are Z bands (''Z lines'', ''Z discs''), A bands, and I bands; less noticeable segments include the M-band-L-line complex, or ''pseudo-H zone'' at the middle of the sarcomere (Figs. 4 and 5). Although a sarcomere, sensu stricto, encompasses two ''half'' I bands, one A band, and the transversely bisected halves of two Z bands (Fig. 4), popular convention considers a sarcomere to be each segment bracketed by Z bands. A (''anisotropic'') bands are zones in which actin and myosin filaments overlap, whereas I (for ''isotropic'') bands represent sarcomere zones in which the actin filaments stand alone. At the midlevel of the sarcomere, where actin filaments do not extend in the relaxed myofibrils, a pair of relatively clear L lines flanks a dark M band, whose opacity is derived from the presence of myosin-to-myosin crossbridges. The presence of M bands in heart is in fact a sign of myocardial cell maturity; in rat heart, M bands appear only postnatally and are missing altogether from the embryonic heart.

The intensely opaque Z bands are composed largely of a-actinin and likely act as linchpins for the stabilization of sarcomere structure via anchoring of the actin

FIGURE 2 A so-called ''type I" conducting cell (Purkinje fiber) in rhesus monkey ventricle. The nucleus of this cell occupies a substantial area of this transverse profile, and the myoplasm is lucent, containing scattered small mitochondria (Mi) and myofibrils composed of contractile filaments so dispersed as to be practically invisible at this magnification. Other types of conducting cells in the same heart have well-developed myofibrils and can be discerned from working cells only on the basis of their subendocardial location, small size, and lack of T tubules. Scale bar: 5 ^m.

FIGURE 3 Working myocardial cell in left atrium of mouse heart. Structures found in the nuclear pole myoplasm distinguish it from a ventricular myocyte, including a well-developed Golgi apparatus (GA) (appearing in section as collections of saccules, tubules, and vesicles), as well as the definitive atrial hallmark, specific atrial granules (SAG) that form within the Golgi and contain concentrated atrial natriuretic peptide. Different stages of SAG formation are seen, including nascent granules with small cores and distinct halos (*). Mitochondria (Mi) mass within the perinuclear region. Atrial myofibrils (Mf) are typically slender. Nu, nucleus;Ly, lysosome. Scale bar: 1 ^m.

FIGURE 2 A so-called ''type I" conducting cell (Purkinje fiber) in rhesus monkey ventricle. The nucleus of this cell occupies a substantial area of this transverse profile, and the myoplasm is lucent, containing scattered small mitochondria (Mi) and myofibrils composed of contractile filaments so dispersed as to be practically invisible at this magnification. Other types of conducting cells in the same heart have well-developed myofibrils and can be discerned from working cells only on the basis of their subendocardial location, small size, and lack of T tubules. Scale bar: 5 ^m.

FIGURE 3 Working myocardial cell in left atrium of mouse heart. Structures found in the nuclear pole myoplasm distinguish it from a ventricular myocyte, including a well-developed Golgi apparatus (GA) (appearing in section as collections of saccules, tubules, and vesicles), as well as the definitive atrial hallmark, specific atrial granules (SAG) that form within the Golgi and contain concentrated atrial natriuretic peptide. Different stages of SAG formation are seen, including nascent granules with small cores and distinct halos (*). Mitochondria (Mi) mass within the perinuclear region. Atrial myofibrils (Mf) are typically slender. Nu, nucleus;Ly, lysosome. Scale bar: 1 ^m.

Lysosomal Glycogen

FIGURE 4 Rhesus monkey right papillary. Relaxed myofibrils, sectioned longitudinally, display the distinct pattern of sarcomere units. Each sarcomere is delimited by opaque Z bands and also contains a dark central A band and two "half" I bands (entire I band denoted by I). The middlemost set of stripes in the sarcomere is known in heart as the "pseudo-H" zone (psH). Side-to-side register is not always exact between adjacent myofibrils (also see Fig. 1). Sarcomere patterning has its basis in organized arrays of proteinaceous filaments: actin stands alone in I bands, overlaps with myosin in all but the pseudo-H zone of the A band, and inserts into the dark substance of the Z band. Certain myocardial cell structures appear preferentially adjacent to the Z band, among them vesicles of corbular SR (C-SR), a form of "extended" junctional sarcoplasmic reticulum. Scale bar: 1 ^m.

FIGURE 5 Rhesus papillary in transverse section. Because of differences in myofibrillar register across the cell, all levels of the sarcomere are represented in this field. Z band material, at the right, the most opaque component of the myofibril, is often closely associated with tubules of sarcoplasmic reticulum (SR). The most lucent portion is the I band, where only thin actin filaments appear, whereas the A bands comprise both actin and myosin arranged in a geometric pattern. Actin does not extend into the midlevel of the sarcomere (the pseudo-H zone), and there thick myosin filaments appear either in isolation (*) or in a dinctinct pattern of hexagons formed by myosin-to-myosin crossbridges (example circled). Mitochondria (Mi) are massed in intermyofibrillar spaces, and in monkey heart are characterized by dense matrices and distinct scalloped internal membranous shelves (cristae). Scale bar: 1 ^m.

within Z band material, which in transverse view is revealed as structurally complex latticeworks (Fig. 5). Z band material may possess additional functions. The Z-line levels of cardiac myofibrils seem to have a definite organizing effect on the neighboring myoplasm, so that certain structures (Z tubules of sarcoplasmic reticulum, transverse tubules, bundles of intermediate filaments) are preferentially oriented at or across Z lines, whereas mitochondria are largely excluded from these regions of "Z-line myoplasm'' (Forbes and Sperelakis, 1980). Overproduction of Z-line substance is a hallmark of ventricular hypertrophy, and similar rod-like, paracrystalline accumulations, known collectively as ''nemaline bodies,'' occur normally in some conducting system cells (see, e.g., Figs. 1.44 and 1.45 of Forbes and Sperelakis, 1995).

2. Structural Fibrils

Filamentous structures that do not actively contribute to the contractile process, but instead support the shapes of the cell components, thence the cells themselves, constitute the so-called cytoskeleton. In heart, the cy-toskeleton is made up of microtubules and intermediate filaments. As mentioned earlier in the section on nuclei, microtubules frequently appear next to nuclear edges. This is best seen in strict transverse views of cells, where cross-sectioned microtubule profiles lie adjacent to the nuclear surface. This orientation is quite constant in some species such as mouse, suggesting that a framework of longitudinally disposed microtubules surrounds each myocardial nucleus. Microtubules form helical en-wrapments around individual myofibrils (Goldstein and Entman, 1979), and thin sections that graze myofibrillar surfaces often reveal microtubules coursing across I- or Z-line levels, at oblique or nearly right angles to the long axes of the myofibrils.

Intermediate filaments—so named because their average diameter of ca. 10 nm is roughly intermediate between the diameters of myosin and actin—have been demonstrated in a wide variety of cell types, and wherever found consistently serve a structural role. Although similar in general appearance in a variety of different cells, biochemically different categories of intermediate filaments have been identified, including desmin, vimen-tin, keratin, and neurofilamin. The desmin type of intermediate filament appears to predominate in cardiac myocytes, and in them is often encountered running transversely, either as individual filaments or in the form of meshworks surrounding myofibrils at Z-line levels. These fibrillar bundles, containing as many as 50 intermediate filaments, may also attach to the inner sarco-lemmal surface and the nuclear envelope, suggesting the existence of multiple nets that stabilize the cell transversely in a series of parallel, semirigid strata. Another prominent site within the cardiac muscle cell with which intermediate filaments are specifically associated is the intercalated disc, specifically the intracellular plaques of desmosomes (see Section II,H).

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