Changes in the random distribution of sialic acid at the surface of the myeloid sinusoidal endothelium resulting from the presence of diaphragmed fenestrae

Frog exocrine pancreatic tissue was studied in vitro under conditions which maintain the differences between tissues from fasted and fed animals. Sodium dodecyl sulfate (SDS) gel electrophoresis after labeling with [14C]amino acids showed that feeding stimulated the synthesis of secretory proteins to the same relative degree as the overall protein synthesis. The intracellular transport of secretory proteins was studied by electronmicroscopy autoradiography after pulse-labeling with [3H]leucine. It was found that the transport route is similar under both feeding conditions. After their synthesis in the rough endoplasmic reticulum (RER), the proteins move through the peripheral elements and cisternae of the Golgi system into the condensing vacuoles. The velocity of the transport increases considerably after feeding. When frogs are fasted, the release of labeled proteins from the RER takes greater than 90 min, whereas after feeding, this happens within 30 min. Comparable differences were observed for transport through the Golgi system. The apparent differences between the frog and mammalian pancreas in the regulation of synthesis, intracellular transport, and secretion of proteins are discussed.     

A morphometrical study of the exocrine pancreatic cell in fasted and fed frogs

The influence of feeding on the ultrastruct of the frog exocrine pancreatic cell was studied by morphometrical procedures. Volume and surface of various cell structures were measured and expressed per unit cell volume. The average cellular size was not influenced by feeding. Though protein synthesis changes 5-to 10-fold (van Venrooij, W. J., and C. Poort. 1971. Biochim. Biophys. Acta. 247:468-470), no significant differences were observed in the amount of membrane that constitutes the rough endoplasmic reticulum (RER) and that represented the major part of total cellular membranes. The appearance of the RER changed. When fasted, most of its membrane was arranged in stacks of tightly packed, narrow cisternae. Within 4 h after feeding, these cisternae were separated and irregularly dilated, and ribosomes became ordered in typical rosettes on their surface. The total volume of the Golgi system increased twofold after feeding. The vesicular and tubular elements at the Golgi periphery did not change, but the volumes of the Golgi cisternae and the condensing vacuoles increased 2.5- and 6-fold, respectively. The increased in the amount of membrane present in these structures was only 1.6- and 3.5-fold, which reflects the more distended appearance of the cisternae and the rounded shape of the condensing vacuoles after feeding. Feeding halved the number of secretory granules per cell, and signs of exocytosis were more common than in fasted animals. These findings suggest that, in the frog pancreatic cell, fluctuations in the production of secretory proteins are not accompanied by an important breakdown and renewal of cellular membranes. This may favor a rapid and strong response of the cell to feeding.     

Fine structural localization of thiamine pyrophosphatase and acid phosphatase activities in the mouse pancreatic acinar cell

The fine structural localization of thiamine pyrophosphatase (TPPase) and acid phosphatase (AcPase) was examined in pancreatic acinar cells of fasting and fed mice. The results were not affected by these conditions. TPPase activity was positive in two and sometimes three cisternae of the inner Golgi lamellae as well as in the condensing vacuoles of the trans area, but negative in the rigid lamellae and small vesicles of the trans area. AcPase activity was demonstrated in two and sometimes three cisternae of inner Golgi lamellae, condensing vacuoles, rigid lamellae, lysosomes and smooth or coated vesicles in the trans area. The inner Golgi lamellae and the condensing vacuoles were positive for both enzyme activities. From these facts, the lysosome is considered to be formed not only in the GERL system but also through the rough endoplasmic reticulum-Golgi apparatus route. It is reasonable to consider that Novikoff's GERL is not independent from the Golgi apparatus but represents a part of this organelle.     

Concentration of amylase along its secretory pathway in the pancreatic acinar cell as revealed by high resolution immunocytochemistry

Summary The modified protein A-gold immunocytochemical technique was applied to the localization of amylase in rat pancreatic acinar cells. Due to the good ultrastructural preservation of the cellular organelles obtained on glutaraldehyde-fixed, osmium tetroxide-postfixed tissue, the labelling was detected with high resolution over the cisternae of the rough endoplasmic reticulum (RER), the Golgi apparatus, the condensing vacuoles, the immature pre-zymogen granules, and the mature zymogen granules. Over the Golgi area, the labelling was present over the transitional elements of the endoplasmic reticulum, some of the smooth vesicular structures at thecis- andtrans-faces and all the different Golgi cisternae. The acid phosphatase-positive rigidtrans-cisternae as well as the coated vesicles were either negative or weakly labelled. Quantitative evaluations of the degree of labelling demonstrated an increasing intensity which progresses from the RER, through the Golgi, to the zymogen granules and have identified the sites where protein concentration occurs. The results obtained have thus demonstrated that amylase is processed through the conventional RER-Golgi-granule secretory pathway in the pancreatic acinar cells. In addition a concomitance has been found between some sites where protein concentration occurs: thetrans-most Golgi cisternae, the condensing vacuoles, the pre- and the mature zymogen granules, and the presence of actin at the level of the limiting membranes of these same organelles as reported previously (Bendayan, 1983). This suggests that beside their possible role in transport and release of secretory products, contractile proteins may also be involved in the process of protein concentration.     

Effect of colchicine on the Golgi complex of rat pancreatic acinar cells

Colchicine administered to adult rats at a dosage of 0.5 mg/100 g of body weight effected a disorganization of the Golgi apparatus in pancreatic acinar cells. The results obtained after various periods of treatment (10 min to 6 h) showed (a) changes in all components of the Golgi complex, and (b) occurrence of large vacuoles that predominated in cytoplasmic areas outside the Golgi region. The alterations in Golgi stacks concerned elements of the proximal and distal side: (a) accumulation of transport vesicles, (b) formation of small, polymorphic secretion granules, and (c) alterations in the cytochemical localization of enzymes and reaction product after osmification. Transport vesicles accumulated and accompanied short, dilated cisternae, which lack mostly the reaction products of thiamine pyrophosphatase, inosine diphosphatase, and acid phosphatase, and osmium deposits after prolonged osmification. After 4 to 6 h of treatment, accumulated transport vesicles occupied extensive cellular areas; stacked cisternae were not demonstrable in these regions. The changes on the distal Golgi side included GERL elements: condensing vacuoles were diminished; they were substituted by small, polymorphic zymogen granules, which appeared to be formed by distal Golgi cisternae and by rigid lamellae. Unusually extended coated regions covered condensing vacuoles, rigid lamellae, and polymorphic secretion granules. A cytochemical distinction between Golgi components and GERL was possible neither in controls nor after colchicine treatment. The cytochemical alterations in Golgi components were demonstrable 20-30 min following administration of colchicine; at 45 min, initial morphological changes--augmentation of transport vesicles and formation of polymorphic zymogen granules--became apparent. 20 min after administration of colchicine, conspicuous groups of large vacuoles occurred. They were located mostly in distinct fields between cisternae of the endoplasmic reticulum, and were accompanied by small osmium--reactive vesicles. Stacked cisternae were not demonstrable in these fields. Vacuoles and vesicles were devoid of reaction products of thiamine pyrophosphatase, inosine diphosphatase, and acid phosphatase. The results provide evidence that formation of stacked Golgi cisternae is impaired after colchicine treatment. The colchicine--induced disintegration of the Golgi complex suggests a regulatory function of microtubules in the organization of the Golgi apparatus.     

Immunolocalization of the oligosaccharide trimming enzyme glucosidase II

We used immunoelectron microscopy to localize glucosidase II in pig hepatocytes. The enzyme trims the two inner alpha 1,3-linked glucoses from N-linked oligosaccharide precursor chains of glycoproteins. Immunoreactive enzyme was concentrated in rough (RER) and smooth (SER) endoplasmic reticulum but not detectable in Golgi apparatus cisternae. Transitional elements of RER and smooth membraned structures close to Golgi apparatus cisternae contained labeling for glucosidase II. Specific labeling was also found in autophagosomes. These results indicate strongly that glucosidase II acts on glycoproteins before their transport to, and processing in Golgi apparatus cisternae, and suggest that an important transitional region for glucosidase II exists between RER and Golgi apparatus cisternae. Degradation in autophagolysosomes could form a normal catabolic pathway for glucosidase II.     

Ultrastructural immunocytochemical localization of cyclic AMP-dependent protein kinase regulatory subunits in rat parotid acinar cells

Polyclonal antibodies to types I and II regulatory (R) subunits of cyclic AMP-dependent protein kinase (cA-PK) were utilized in a post-embedding immunogold-labeling procedure to localize these proteins in rat parotid acinar cells. Both RI and RII were present in the nuclei, cytoplasm, rough endoplasmic reticulum (RER), Golgi apparatus, and secretory granules. In the nuclei, gold particles were mainly associated with the heterochromatin. In the cytoplasm, the label was principally found in areas of RER. Most gold particles were located between adjacent RER cisternae or over their membranes and attached ribosomes; occasional particles were also present over the cisternal spaces. Labeling of the Golgi apparatus was significantly greater than background, although it was slightly lower than that over the RER cisternae. In secretory granules, gold particles were present over the granule content; no preferential localization to the granule membrane was observed. Morphometric analysis revealed equivalent labeling intensities for RI and RII in the cytoplasm-RER compartment. Labeling intensities for RII in the nuclei and secretory granules were about 50% greater than in the cytoplasm-RER, and 3 to 4-fold greater than values for RI in these two compartments. Electrophoresis and autoradiography of the postnuclear parotid-tissue fraction, the contents of purified secretory granules and saliva collected from the main excretory duct, after photoaffinity labeling with [32P]-8-azido-cyclic AMP, revealed the presence of R subunits. Predominantly RII was present in the granule contents and saliva, while both RII and RI were present in the cell extracts. Additionally, R subunits were purified from saliva by affinity chromatography on agarose-hexane-cyclic AMP. These findings confirm the localization of cA-PK in parotid cell nuclei and establish the acinar secretory granules as the source of the cyclic AMP-binding proteins in saliva.     

Fine structural localization of thiamine pyrophosphatase and acid phosphatase activities in the mouse pancreatic acinar cell

Summary The fine structural localization of thiamine pyrophosphatase (TPPase) and acid phosphatase (AcPase) was examined in pancreatic acinar cells of fasting and fed mice. The results were not affected by these conditions. TPPase activity was positive in two and sometimes three cisternae of the inner Golgi lamellae as well as in the condensing vacuoles of the trans area, but negative in the rigid lamellae and small vesicles of the trans area. AcPase activity was demonstrated in two and sometimes three cisternae of inner Golgi lamellae, condensing vacuoles, rigid lamellae, lysosomes and smooth or coated vesicles in the trans area. The inner Golgi lamellae and the condensing vacuoles were positive for both enzyme activities. From these facts, the lysosome is considered to be formed not only in the GERL system but also through the rough endoplasmic reticulum-Golgi apparatus route. It is reasonable to consider that Novikoff's GERL is not independent from the Golgi apparatus but represents a part of this organelle.This study was supported by a grant from the Japan Educational Ministry     

Synthesis and intracellular transport of proteins in the exocrine pancreas of the frog (Rana esculenta)

Summary Frog pancreatic tissue was pulse-labelled in vitro with ³H-leucine and protein transport was studied in exocrine cells by electron microscope autoradiography. The proteins appeared to be synthesized in the RER and transported to the secretory granules along a similar route and with the same velocity as previously described under in vitro conditions.Evidence was obtained for the involvement of the vesicular and tubular elements at the periphery of the Golgi system in transferring protein from the RER to the Golgi cisternae.Kinetics of the release of newly synthesized proteins from the RER and their appearance in the condensing vacuoles are discussed and related to results reported from other tissues.The transport velocity in this poikilothermic system was studied in relation to the incubation temperature and compared with results reported from its mammalian counterpart. At temperatures between 20 and 30° C intracellular protein transport occurs faster in the frog than in the Guinea pig pancreas. At higher temperature the transport process was severely disturbed in the frog.     

Antibodies to the Golgi complex and the rough endoplasmic reticulum

Rabbits were immunized with membrane fractions from either the Golgi complex or the rough endoplasmic reticulum (RER) by injection into the popliteal lymph nodes. The antisera were then tested by indirect immunofluorescence on tissue culture cells or frozen, thin sections of tissue. There were may unwanted antibodies to cell components other than the RER or the Golgi complex, and these were removed by suitable absorption steps. These steps were carried out until the pattern of fluorescent labeling was that expected for the Golgi complex or RER. Electron microscopic studies, using immunoperoxidase labeling of normal rat kidney (NRK) cells, showed that the anti-Golgi antibodies labeled the stacks of flattened cisternae that comprise the central feature of the Golgi complex, many of the smooth vesicles around the stacks, and a few coated vesicles. These antibodies were directed, almost entirely, against a single polypeptide with an apparent molecular weight of 135,000. The endoplasmic reticulum (ER) in NRK cells is an extensive, reticular network that pervades the entire cell cytoplasm and includes the nuclear membrane. The anit-RER antibodies labeled this structure alone at the light and electron microscopic levels. They were largely directed against four polypeptides with apparent molecular weights of 29,000, 58,000, 66,000, and 91,000. Some examples are presented, using immunofluorescence microscopy, where these antibodies have been used to study the Golgi complex and RER under a variety of physiological and experimental condition . For biochemical studies, these antibodies should prove useful in identifying the origin of isolated membranes, particularly those from organelles such as the Golgi complex, which tend to lose their characteristic morphology during isolation.     

Herpes simplex virus 1 envelopment follows two diverse pathways

Herpesvirus envelopment is assumed to follow an uneconomical pathway including primary envelopment at the inner nuclear membrane, de-envelopment at the outer nuclear membrane, and reenvelopment at the trans-Golgi network. In contrast to the hypothesis of de-envelopment by fusion of the primary envelope with the outer nuclear membrane, virions were demonstrated to be transported from the perinuclear space to rough endoplasmic reticulum (RER) cisternae. Here we show by high-resolution microscopy that herpes simplex virus 1 envelopment follows two diverse pathways. First, nuclear envelopment includes budding of capsids at the inner nuclear membrane into the perinuclear space whereby tegument and a thick electron dense envelope are acquired. The substance responsible for the dense envelope is speculated to enable intraluminal transportation of virions via RER into Golgi cisternae. Within Golgi cisternae, virions are packaged into transport vacuoles containing one or several virions. Second, for cytoplasmic envelopment, capsids gain direct access from the nucleus to the cytoplasm via impaired nuclear pores. Cytoplasmic capsids could bud at the outer nuclear membrane, at membranes of RER, Golgi cisternae, and large vacuoles, and at banana-shaped membranous entities that were found to continue into Golgi membranes. Envelopes originating by budding at the outer nuclear membrane and RER membrane also acquire a dense substance. Budding at Golgi stacks, designated wrapping, results in single virions within small vacuoles that contain electron-dense substances between envelope and vacuolar membranes.     

Disproportional immunostaining patterns of two secretory proteins in guinea pig and rat exocrine pancreatic cells. An immunoferritin and fluorescence study

Amylase (Am) and chymotrypsinogen (Chtg) were demonstrated in rat and guinea pig exocrine pancreatic cells by immunofluorescence and immunoferritin cytochemistry on thin and ultrathin frozen sections. We describe two observations indicating that Am and Chtg may behave differently in the pre-Golgi phase of their intracellular transport. Firstly, aggregates of material within the RER cisternae of the guinea pig (so-called intracisternal granules) reacted strongly with anti-Chtg, but showed no affinity for anti-Am. Secondly, in both rat and guinea pig, the increase in labeling intensity from cytoplasm (RER) to secretory granules was larger for Chtg than for Am. We hypothesize that the two proteins do not travel in-parallel towards the Golgi complex. Compared with Chtg, Am would lag behind in the RER cisternae.     

The mystery of the unstained Golgi complex cisternae

The Champy-Maillet OsKI reaction has been used upon Golgi complexes to show two kinds of staining. It stains material being processed as it passes along the secretory pathway of the rough endoplasmic reticulum (RER) and Golgi cisternae (GC) up to crystallization in secretory vesicles. It also stains separately the environment within parts of the GC. This GC staining may occur in all compartments (transition vesicles, saccules, condensing vacuoles), but it is characteristically missing from any one of them. The unstained cisternae may be explained if outer saccules are made from either stained or unstained transition vesicles, both of which occur. The presence of empty, unstained transition vesicles is dictated by the surface to volume ratios of microvesicles in relation to saccules. Most transition vesicles must return their membrane to the endoplasmic reticulum, but from time to time it is presumed that they fuse to make a saccule. Saccules, stained and unstained, then mature through the stack. OsKI reactions with tissues and test molecules suggest that in the RER and GC the stain detects labile--S . S--bridges before they lock the tertiary configuration of proteins.     

Immunocytochemical localization of Mullerian inhibiting substance in the rough endoplasmic reticulum and Golgi apparatus in Sertoli cells of the neonatal calf testis using a monoclonal antibody

Mullerian Inhibiting Substance (MIS) has been localized in the Sertoli cells of the neonatal calf testis using preembedding immunoperoxidase techniques and a monoclonal antibody which almost completely blocks the biological activity of MIS. Both the peroxidase-labeled antibody method using a peroxidase-conjugated F(ab')2 fragment of IgG as a second antibody and the unlabeled antibody peroxidase-antiperoxidase (PAP) method using Fab fragments of the PAP complex were employed. With both methods, MIS was demonstrated within the cisternae of the rough endoplasmic reticulum (RER) and the Golgi apparatus. In the Golgi, MIS was concentrated in the transmost cisternae especially at their peripheral expansions. This study indicates that MIS is synthesized in the RER and transported to the Golgi apparatus, presumably for glycosidation, before secretion from Golgi derived vacuoles.     

An electron microscopic study of hemoglobin synthesis in the marine annelid,Amphitrite ornata (Polychaeta: Terebellidae)

The heart-body of the marine worm Amphitrite, located within the supraesophageal dorsal vessel, is in the form of a cylinder the thin wall of which is deeply corrugated by luminal projections and folds along its entire length. It is anchored in places to the luminal surface of the dorsal vessel by an extracellular matrix containing collagen fibers. The luminal surfaces of both the heart-body and the dorsal vessel are covered by a basement membrane-like vascular lamina which in turn supports a discontinuous pseudoendothelium of littoral hemocytes. The cells of the heart-body constitute a pseudostratified, high columnar epithelium. They possess extensive rough endoplasmic reticulum (RER), a well developed Golgi zone, ferritin particles and granules, and several types of membrane-bound inclusions. Hemoglobin molecules identical to those in the circulation lie within cytoplasmic, membrane-bound vesicles. Analysis of our electron micrographs suggests the following sequence of hemoglobin production and secretion: Large quantities of a moderately dense flocculent material, probably globin, are synthesized in RER and move to the Golgi zone within partly rough- and partly smooth-surfaced transitional cisternae; small transport vesicles, formed from Golgi cisternae that have fused with transitional cisternae, convey the flocculent material from the convex to the concave face of the Golgi complex; a similar flocculent material and an amorphous, highly dense material are processed in the Golgi complex and are transferred to condensing vacuoles in which clearly identifiable hemoglobin molecules are first observed. Mature secretory vesicles containing only hemoglobin migrate to the cell periphery and discharge their contents by exocytosis. Hemoglobin molecules then cross the vascular lamina to reach the circulation.     

Temperature and energy dependence of secretory protein transport in the exocrine pancreas.

Pancreatic lobules pulse-labeled with [3H]leucine have been incubated at temperatures between 0 and 37 degrees C in the presence or absence of ongoing oxidative phosphorylation. Subcellular fractionation methods and electron microscopic autoradiography have been used to monitor the progress of intracellular transport of newly synthesized secretory proteins. Over the period studied, exit from the rough endoplasmic reticulum (RER) occurs only at greater than 10 degrees C while traversal of the Golgi complex and entry into condensing vacuoles requires greater than 22 degrees C. Both steps of transport require ongoing ATP production. Incubation at 10 or 20 degrees C does not diminish ATP levels, relative to 37 degrees C controls. Remarkable and unprecedented alterations of the ultrastructure of transitional elements of the RER accompany the arrest of exit from the RER: at 10 degrees C transitional elements are much more numerous and longer than in controls; in the absence of ATP production they are essentially absent. These observations are interpreted in terms of a cyclic model of RER-to-Golgi vesicular traffic. Inhibition of ATP production also causes an increase in the rigid cisternae and coated elements in the distal Golgi area.     

Clathrin-coated vesicular transport of secretory proteins during the formation of ACTH-containing secretory granules in AtT20 cells

We have studied by electron microscopy and immunocytochemistry the formation of secretory granules containing adrenocorticotropic hormone (ACTH) in murine pituitary cells of the AtT20 line. The first compartment in which condensed secretory protein appears is a complex reticular network at the extreme trans side of the Golgi stacks beyond the TPPase-positive cisternae. Condensed secretory protein accumulates in dilated regions of this trans Golgi network. Examination of en face and serial sections revealed that "condensing vacuoles" are in fact dilations of the trans Golgi network and not detached vacuoles. Only after presumptive secretory granules have reached an advanced stage of morphological maturation do they detach from the trans Golgi network. Frequently both the dilations of the trans Golgi network containing condensing secretory protein and the detached immature granules in the peri-Golgi region have surface coats which were identified as clathrin by immunocytochemistry. Moreover both are the site of budding (or fusion) of coated vesicles, some of which contain condensed secretory protein. The mature granules below the plasma membrane do not, however, have surface coats. Immunoperoxidase labeling with an antiserum specific for ACTH and its precursor polypeptide confirmed that many of the coated vesicles associated with the trans Golgi network contain ACTH. The involvement of the trans Golgi network and coated vesicles in the formation of secretory granules is discussed.     

The origin of the zymogen granule membrane of the pancreatic acinar cell as examined by ultrastructural cytochemistry of acid phosphatase, thiamine pyrophosphatase, and ATP-diphosphohydrolase activities

Cytochemical distributions of acid phosphatase, thiamine pyrophosphatase, and ATP-diphosphohydrolase activities have been examined on thin sections of rat pancreas and on isolated zymogen-granule membranes. Acid phosphatase was found in the rigid lamellae separated from the Golgi stacked cisternae, in condensing vacuoles, and in the trans-saccules of Golgi apparatus; it was not detected in purified zymogen-granule membranes. Thiamine pyrophosphatase was detected in trans-saccules of the Golgi apparatus, in purified zymogen-granule membranes, and in the plasmalemma of the acinar cell. It was absent in condensing vacuoles. The ATP-diphosphohydrolase activity has a distribution similar to thiamine pyrophosphatase. These observations illustrate the similarity between the trans-saccules of the Golgi apparatus and the membrane of mature zymogen granules and the disparity between the latter membrane and the membrane of the condensing vacuole. They suggest that the condensing vacuole might not be the immediate precursor of the zymogen granule as commonly assumed. An alternative possibility would be that condensing vacuoles would fuse with the trans-saccule (transition) of the Golgi apparatus which in turn would form mature zymogen granules.     

The Golgi apparatus and lysosomes of rat pancreatic acinar cells following refeeding

Summary The short term effects of refeeding on the Golgi apparatus and lysosomes of the rat exocrine pancreas were evaluated by ultrastructural, morphometric and cytochemical methods. Ten minutes after refeeding, there was a significant enlargement of Golgi cisternae and a significant increase, compared with the controls, in the number of condensing vacuoles and lysosomes. These modifications were accompanied by the appearance of acid phosphatase activity in stacked Golgi cisternae (as well as GERL) of some cells. One hour after refeeding, there were about the same numbers of condensing vacuoles and lysosomes as in the control; Golgi cisternae were still significantly enlarged, compared with the controls, but they were no longer reactive for acid phosphatase. In both fasting and refed animals, acid phosphatase activity was demonstrable in tubular lysosomes.The data are interpreted in terms both of membrane disposal and recycling, leading to enhanced formation of zymogen granules, during physiologically stimulated secretion.     

Intracellular transport of proteins in active and resting secretory cells of the venom gland of Vipera palaestinae

The intracellular transport of venom proteins has been studied in active and resting venom glands of the snake Vipera palaestinae by electron microscope radioautography after an intra-arterial injection of [3H]leucine. In the active gland, most of the label is initially (10 min) found over the RER. By 30 min, the relative grain density of the Golgi complex reaches its maximum, with concomitant increase in the labeling of the condensing vacuoles. Later on, a steep increase in radioactivity of the secretory granules is observed. At 3 h, these granules, which comprise about 2% of the cell volume, contain 22% of the total grains. At the following hour, their labeling declines and at the same time the radioactivity of the secreted venom is increased. It is concluded that, in the active cell, venom proteins are transported via the Golgi apparatus into membrane-bounded granules which are the immediate source of the secreted venom. An alternative pathway, which involves the RER cisternae as a storage compartment, seems unlikely, since incorporated label does not accumulate in this compartment after prolonged postpulse intervals. The route of intracellular transport of proteins in the resting glands is similar to that of the active ones, but the rate of synthesis and transport is much slower. The present results and earlier data, thus, show that the increase in the rate of secretion after initiation of a new venom regeneration cycle is the result of accelerated rates of both synthesis and transport.