82 SUBCELLULAR PARTICLES 



Dr. Palade: /) The periphery of myocardial infarcts may be a good test material for 

 the old assumption of direct transformation of mitochondria into lipid inclusions but, 

 as far as I know, such material has not yet been studied by electron microscopy. In 

 the ischemic myocardium, however, one would deal with a degenerative process, rather 

 than with a normal physiological event. Even in this case, the relationship between the 

 rate of lipid transport and the rate of lipid oxidation may have some bearing on the 

 accumulation of fat droplets. 2) I believe that there is at present good evidence for 

 the existence of two distinct mitochondrial membranes. The inner one is frequently 

 continuous with the cristae mitochondriulcs (which appear as its infoldings) and cor- 

 responds to what Dr. Novikoff calls "the outer limit of the mitochondrial body." The 

 outer membrane — Dr. Novikoff and I agree on its existence — separates the mitochon- 

 drion from the cytoplasmic matrix. The hypothesis that the oxidative enzymes are 

 located in the internal membrane and in its cristae is supported by a number of 

 observations (cf. 25). A final answer may be obtained by comparing the concentration 

 of oxidative enzymes with the amount of internal and external membrane per unit 

 volume in a representative series of mitochondria that should include specimens with 

 few and with numerous cristae. Evidence obtained by dismantling isolated mitochondria 

 (cf. 43, 45) cannot solve this problem, because the typical mitochondrial organization 

 is lost in the process, and the derivation of the small, membrane-bound vesicles finally 

 obtained cannot be decidedly ascertained. These 'submitochondrial' vesicles may be 

 derived from both the outer and the inner membrane, but most of them are probably 

 formed at the expense of the latter, since there is more of inner than of outer mem- 

 brane in all mitochondria. 



Dr. Littlefield: You certainly have beautifully correlated evidence on the direction 

 of flow of protein synthesis in the microsomal fractions from your morphologic and 

 in vivo labeling experiments. I wonder if you and Dr. Siekevitz have been able to show 

 any movement of labeled protein in homogenates? 



Dr. Palade: With our material, i.e., the pancreas of the adult guinea pig, we were 

 not able to obtain a system capable of incorporating amino acids into proteins in vitro. 

 Moreover, we have not carried out experiments designed to explore the transfer in vitro 

 of proteins labeled in vivo. 



Dr. M.\rshak: Since the intracisternal granules are actually located in the cisternal 

 space, i.e., with their surfaces rather far removed from the RNP granules which pre- 

 sumably synthesize the protein making up the intracisternal granules, is it not necessary 

 also to postulate an additional mechanism aggregating the protein found and removing 

 it from the site of synthesis? 



Dr. Palade: It is indeed necessary to postulate a mechanism by which the product is 

 removed from the site of synthesis (presumably the attached RNP particles) and, in 

 addition, transported across the membrane which separates the cytoplasmic matrix from 

 the intracisternal spaces. After being segregated within these spaces, the zymogen 

 molecules probably aggregate into intracisternal granules when a suitable concentra- 

 tion is reached. The biochemical and physiological advantages brought about by the 

 segregation of the newly synthesized zymogens are easily understood, but the mech- 

 anisms involved remain, for the moment, entirely unknown. 



Dr. Prosser: /) Have you evidence for localization of amylase and lipase similar to 



