Electron microscopy of insulin precipitates.

Morphologic characteristics of insulin precipitates were examined by both scanning and transmission electron microscopy. Insulin precipitate obtained after 1 wk of in vivo storage in an implanted dog reservoir was compared with insulin precipitate produced in vitro by isoelectric precipitation, acid-freeze-heat precipitation, and motion-induced precipitation. Insulin precipitate produced in vivo had several morphologic forms, with spherical-lamellar structures predominating. In vitro isoelectric precipitated insulin produced microcrystalline material, whereas acid-freeze-heat and motion-induced precipitated insulin were associated with elongated fibrils. The morphologic appearance of the in vivo precipitated insulin was not entirely reproduced by any of the three in vitro methods of insulin precipitation. We conclude that insulin precipitation in vivo is a process that may involve more than one of the known mechanisms by which insulin precipitates in vitro.

A method for culturing canine tracheal smooth muscle cells in vitro: morphologic and pharmacologic observations.

A method of culturing canine tracheal smooth muscle cells in vitro is described. The morphology of these cells is monitored up to 60 days in culture and selected stages are illustrated. The characteristics of these cells are numerous mechanical attachments, the presence of thick filaments in suitably processed cells, and their contractile response to in vitro administration of carbachol, a cholinomimetic drug. They also possess nexus formations and both thin (actin) filaments and 10-nm filaments. Mitosis is found in the nonconfluent preparations up to 16 days after culturing. Cultures of 2 to 8 days appear to be most useful as pharmacological test vehicles. This system will be used to explore the phenomenon of adrenergic beta-2 receptor desensitization in airway smooth muscle, to attempt to localize these receptor sites and to determine how receptor affinity and/or number may be regulating cell response to pharmacologic agents.

Ultrastructure of secondary neurulation in the chick embryo.

Formation of the future lumbosacral level of the spinal cord was studied in two-day-old chick embryos by light and electron (transmission and scanning) microscopy. A neurulation overlap zone occupied this level. The dorsal portion of the neural tube formed by bending of the neural plate and approximation and fusion of neural folds (i.e., by primary neurulation), and the ventral part formed during secondary neurulation by cavitation of an initially solid, compact mass of cells, the medullary cord, derived from the tail bud. Secondary neurulation involved four morphogenetic processes: (1) segregation of the cells of the prospective medullary cord from cells of adjacent regions, (2) formation of a precisely delimited medullary cord, (3) cavitation of the central portion of this cord, and (4) coalescence of all lumina into a single, central cavity. Cell segregation was associated with the formation of a layer of primarily extracellular materials between adjacent organ rudiments. The source and composition of these materials are unknown. Formation of the medullary cord entailed considerable elongation of the peripheral cells of this developing structure and the fabrication of small intercellular juctions, first at the basal (outer) ends of the elongating peripheral cells, and then at their apical (inner) ends. These arranged, columnar cells, having characteristics similar to those of the neural plate, and an inner cluster of irregularly shaped and arranged cells. Cavitation always occurred first at the junction between these two cellular populations. The central cells of the medullary cord also eventually elongated, like the peripheral cells, and may have been intercalated into the lateral walls of the developing neural tube as lumina coalesced.