Origin and fate of autophagosomes in Leydig cells of normal adult rats.

Abstract

Autophagosomes were observed frequently in electron microscope photographs of Leydig cells from normal adult rat testis. Their formation, evolution and fate were analyzed morphologically in preparations treated to show cytidine monophosphatase (CMPase) and glucose-6-phosphatase (G-6-Pase) activities and in animals sacrificed at various time intervals ranging from 5 min to 6 hrs after a single intratesticular injection of cationic ferritin. Analysis of the morphologic data led to the following interpretation and model. Preautophagosomal structures appeared as flattened, elongated membranous profiles. These expanded, took on a C-shape and fused at their edges to demarcate a small cytoplasmic territory containing normal-looking smooth endoplasmic reticulum (ER) and mitochondria. Such early autophagosomes were thus delimited by two membranes separated by a narrow lumen. Following fusion of these elements with secondary lysosomes, the space between the two membranes increased in size, the inner membrane disintegrated and the enclosed organelles no longer could be identified. The late autophagosomes then reached the cell surface and appeared to exocytose their residual content. In contrast to secondary lysosomes and trans-Golgi elements, which were CMPase-positive, the preautophagosomal flattened membranous elements and early autophagosomes were CMPase-negative. The late autophagosomes on the contrary were CMPase-positive. While ER cisternae were G-6-Pase-positive, the pre-, early and late autophagosomal structures were unreactive for this enzyme. Cationic ferritin tracer experiments showed that only late autophagosomes became labeled with cationic ferritin following their fusion with secondary lysosomes into which the tracer had accumulated following its endocytosis from the cell surface.(ABSTRACT TRUNCATED AT 250 WORDS)