The Ultrastructure of Megakaryocytes

The cytoplasm of the megakaryocyte in the rat spleen possesses three zones, the perinuclear, intermediate and marginal. The perinuclear zone is characterized by the presence of Golgi apparatus, ribosomes, endoplasmic reticulum, and mitochondria. These organelles are found also in the more voluminous intermediate zone which in addition exhibits platelet granules and an extensive development of vesicles from smooth-surfaced endoplasmic reticulum to form demarcation membranes by coalescence. The marginal zone is almost devoid of the organelles and inclusions present elsewhere. Shedding of platelets appears to occur by extension of a paired demarcation membrane from the intermediate zone to the cell membrane and subsequent separation of its lamellae so that all of the essential organelles and inclusions of the intermediate zone may be included within the platelet. In addition, platelets contain vesicles which are probably pinocytotic in nature. Platelets are sometimes engulfed by the cytoplasm o f phagocytic cells. Numerous incidental observations on the ultrastructure of megakaryocytes and blood platelets have been reported (De Marsh et al., ’55; Pease, ’55; Rinehart, ’55; Goodman et al., ’57; Kisch, ’57; Mizuno et al., ’59; Jones, ’60). A detailed description of the ultrastructure of megakaryocytes by Yamada (’57) confirmed the earlier description of Heidenhain ( 1894) based on observation with the light microscope. Yamada subdivided the cytoplasm of the megakaryocyte into three zones. The innermost or perinuclear zone consisted of a narrow band of cytoplasm immediately surrounding the nucleus; the middle or intermediate zone was wide and distinguished by the presence of the platelet demarcation membrane system; and the outer or marginal zone was narrow and made up of a finely granular ground cytoplasm. Mitochondria, small vesicles and endoplasmic reticulum were observed throughout the perinuclear and intermediate zones. The Golgi apparatus and centriole were found chiefly in the perinuclear zone and in the area of cytoplasm encompassed by the lobated nucleus. Yamada described four stages in formation of the platelet demarcation membranes, namely the prevesicular, vesicular, membranous and residual. During the vesicular stage, vesicles about 400 A in diameter appeared in the intermediate zone ANAT. REC., 149: 251-268. and became aligned in rows. Fusion of these vesicles to form a system of paired membranes distingiushed the membranous stage. Liberation of platelets was believed to occur by extension of the demarcation membranes through the marginal zone to insertion into the plasma membrane with subsequent separation of the paired demarcation membranes. Following shedding of platelets the cell reverted to the residual stage. This report is concerned with the numerous aspects of the ultrastructure of megakaryoctyes and platelets which merit additional study including the distribution of ribosomes, the origin and structure of platelet granules,’ the structure of mitochondria, and the structure and distribution of the Golgi apparatus. MATERIALS AND METHODS Six female Sprague-Dawley rats of 200 gm body weight were used. While the animals were under ether anesthesia, small pieces of spleen were excised and fixed for four hours in chilled 1 or 2% OsOl buffered with 0.14 M veronal acetate to pH 7.4. Sucrose was added to the fixative to make a 4.5% solution. The tissues were dehyI Supported in part by research grants from the National Institutes of Health Al31(C9) and from the Upjohn Company. 2 The term “platelet granule” refers to the dense membrane-enclosed granules the diameter of which was approximately that of mitochondria.