Ductal Excretion of Neutral Red Lysosomes in the Mouse Pancreas

In earlier studies of the mouse pancreas (1), it was found microscopically in saline preparations of living tissue that, within 30 min after the injection of an optimum dose of neutral red, small dye granules appeared in the cytoplasm of the acinar cells. With time, the granules became larger, reached a peak development at about 7 hr, then gradually diminished in size and number until, at about 14 hr, the cell was clear of dye and again normal in appearance. Throughout this "neutral red granule cycle," the dye granules appeared to be confined to supraand paranuclear positions in the cell. In an effort to shed some light on the possible role of the Golgi apparatus in this process, the Aoyama silver impregnation method was applied to pieces of the same pancreas examined in the fresh at each of the intervals throughout the cycle (2). With this procedure, the acinar cell showed a gradual loss of the typical Golgi network after dye injection and the development of argentophil granules having many of the properties of the neutral red granules seen in the fresh tissue. The argentophil bodies were not observed in the acinar cell unless the animal had been injected with neutral red. The position, number, general configuration, and relative size of the argentophil bodies were those of the neutral red granules. Finally, the time-relationships in terms of the appearance, development, and disappearance of the argentophil bodies were identical with those of the neutral red granules (2, 3). The neutral red granules as seen in a cryostat-frozen section of pancreas 5 hr after dye injection (Fig. 1) may be compared with the argentophil bodies shown by the Aoyama technique applied to the same pancreas (Fig. 2). It was concluded from this study that the argentophil bodies demonstrated by the Aoyama method were the fixed-tissue counterparts of the neutral red granules seen in living cells. Recent electron microscope studies of similarly timed sequences after injection of neutral red have indicated that the dye granules are lysosomal in nature (4, 5), and are positive for acid phosphatase (6). The fate of the neutral red granules formed in the cytoplasm of pancreatic acinar cells has never been resolved. The possibilities have been considered that the dye is broken down within the granule, or is merely held temporarily in these saclike structures and then passes back into the serum for final excretion by another organ, e.g. the liver. In relation to this question, Covell in an early cytological study (7) reported that after pilocarpine stimulation the neutral red granules pass into the lumen of the pancreatic acinus and then into the ductules. In our studies of bits of fresh pancreas examined microscopically in saline, the dye granules always appeared confined to supraand paranuclear positions in the acinar cell. They were never seen mixed amongst the zymogen granules or near the apex of the cell. It was our conclusion, therefore, that the dye granules did not pass into the ducts. While our preparations of bits of fresh pancreas