The localization of thiamine pyrophosphatase activity in Meckel's cartilage cells during endochondral ossification

The cytochemical distribution of thiamine pyrophosphatase (TPPase) activity in Meckel's cartilage cells of the mouse embryo has been studied during the endochondral ossification. All the cartilage cells contain reaction product within the Golgi apparatus. In immature chondrocytes, at the reserve cell zone, TPPase activity is restricted to several inner cisternae of independent Golgi apparatus. In mature cells at the proliferative cell zone, several Golgi complexes form a Golgi network connecting with each other by the TPPase positive tubular stalks. Golgi cisternae, condensing vacuoles and vesicles also contain reaction product. In the hypertrophic chondrocytes located in the calcifying zone, their disorganized Golgi apparatus still retain reaction product. Some chondrocytes, even those located within calcified or opened lacunae, exhibit intact structures and normal cytochemical enzyme distribution. These data indicate the possibility that some chondrocytes may survive and contribute the formation of mandible.     

Schistosoma mansoni: cytochemistry and morphology of the gastrodermal Golgi Apparatus

The gastrodermal Golgi apparatus of adult Schistosoma mansoni displays two distinct morphologies. In one type, there is an identifiable cis (forming) face where vesicles from the endoplasmic reticulum fuse to form the cisternae. A morphological change occurs in the cisternae as the trans (emitting) face is approached with the cisternae becoming progressively flattened. The cisternae at the emitting face produce a membrane-bound secretory granule with moderately electron-dense contents and a vacuolar structure that may be analogous to a condensing vacuole as reported in several vertebrate secretory cells. In a second type, vesicles possessing a thicker membrane than those of the transfer vesicles are observed at the emitting face. They are not observed when the secretory granules are present. Several cytochemical markers were used to aid in studying the polarity of the Golgi apparatus. Enzymes studied were thiamine pyrophosphatase (TPPase) (EC 3.6.1.1), nucleoside diphosphatase (NDPase) (EC 3.6.1.6) using uridine diphosphate as a substrate, and nicotinamide adenine dinucleotide phosphatase (NADPase) (EC 3.1.3.2). Reaction products from all enzyme markers were observed in the cisternae and, to some extent, in the transfer vesicles. At times, NADPase and TPPase reaction products were observed in all cisternae and in the transfer vesicles of the Golgi. When this distribution was evident, the latter vesicles were observed in clusters occasionally fusing with lipid-like globules dispersed throughout the gastrodermis. Heterogeneity in cisternae was observed when NDPase, TPPase, and osmium reduction techniques were used. NDPase activity was limited to the middle cisternae while reduced osmium was observed in the outer two cisternae and in some transfer vesicles. TPPase reaction product was also observed in the secretory granules and in the condensing vacuoles. It is hypothesized that a functional bipolarity may be demonstrated by the Golgi. Under certain stress conditions, the forming face of the Golgi may package lysosomal enzymes while the emitting region of the Golgi appears to be responsible for the packaging of the secretory granules. The fusion of transfer vesicles and, at times, secretory granules with lipid-like globules is postulated to represent a mechanism by which enzymes may be transported to the lumen of the cecum.     

Phosphatase activities and osmium reduction in cell organelles ofMicrasterias americana

Summary Organelles in resting and growing cells ofMicrasterias americana were examined using electron microscopy after cytochemical procedures for four kinds of phosphatases, acid phosphatase (ACPase), alkaline phosphatase (ALPase), thiamine pyrophosphatase (TPPase), and inosine diphosphatase (IDPase), and osmium tetroxide reduction. Special attention was paid to activities in the Golgi apparatus.In resting cells, positive reactions for ACPase and TPPase were observed in all cisternae of the dictyosome, especially in the peripheral parts. A positive IDPase reaction was seen in one central cisterna and was frequent in the distal-most cisterna. Reduction of osmium tetroxide was seen in the proximal cisternae.In early growing cells, the dictyosomes gave positive reactions for ACPase in the proximal cisternae and the distal cisterna, while in late growing cells only in proximal cisternae. Both in early and late growing cells, the dictyosomes were positive for TPPase and IDPase in the distal cisternae and vesicles derived from the distal cisternae, and for the reduction of osmium tetroxide in the proximal cisternae. ALPase activity was detected in the growing cell wall but not in the dictyosome.     

Thiamine pyrophosphatase activity in secretory cells of the lateral prostate and seminal vesicle of normal and castrated guinea pigs and castrates treated with oestradiol

Summary Ultrastructural localization of thiamine pyrophosphatase (TPPase) activity was studied in secretory cells of the lateral prostate and seminal vesicle of normal and castrated guinea pigs and castrates treated with 17-oestradiol benzoate. The present study has demonstrated that TPPase reaction product is consistently localized in the three to four trans cisternae of Golgi complexes in both the lateral prostate and the seminal vesicle. The reaction was intense and the reaction product often filled the cisternae completely.After castration there was a decrease in TPPase activity in both glands as revealed by the reduction in the amount of the reaction product which was found mainly in one to two trans cisternae of the regressed Golgi complex. The reaction product changed from a dense to a more particulate or granular pattern or to discrete deposits of high electron-density.Administration of 17-oestradiol benzoate to the castrates caused changes in the localization and patterns of distribution of TPPase. In the lateral prostate there was an apparent increase in TPPase activity. The reaction product was found in two to four trans cisternae and occasionally in the trans-most cisternae of the dilated Golgi complex. The reaction product appeared as discrete, dense coarse precipitates. In the seminal vesicle TPPase reaction product was consistently found in one to two trans cisternae in cells with larger Golgi complexes. However, almost all cisternae of the smaller Golgi complexes were TPPase-positive. The cytochemicl results of the present study suggest that TPPase activity and possibly the process of glycosylation in secretory cells of the lateral prostate and seminal vesicle may have been affected after castration and after oestradiol administration.     

Cytochemical localization of thiamine pyrophosphatase and nicotinamide adenine dinucleotide phosphatase activities in rat epiphyseal chondrocytes

Two phosphatase activities, which have been reported to be associated with the Golgi apparatus in several cellular types, have been cytochemically demonstrated in rat epiphyseal cartilage. This was the case for thiamine pyrophosphatase (TPPase) which was detected in Golgi trans face cisternae and also in nascent or immature secretory granules of chondrocytes. beta-Nicotinamide adenine dinucleotide phosphatase (beta-NADPase), on the other hand, was localized mainly in the endoplasmic reticulum region of both proliferative and hypertrophic chondrocytes. Most of the beta-NADPase reaction was shown to be associated with the cytoplasmic side of the rough endoplasmic reticulum membranes and also partially dispersed throughout the cytosol background. We suggest that beta NADPase in chondrocytes could be an enzyme with different properties from that described in other secretory cells.     

The localization of thiamine pyrophosphatase activity in the acinar cells of stimulated and non-stimulated sublingual glands of the rat

Summary The distribution of thiamine pyrophosphatase (TPPase) activity in the acinar cells of the rat sublingual gland has been studied at various stages of the secretory cycle following stimulated secretion. The rats were stimulated to secrete by an intraperitoneal injection of isoproterenol and pilocarpine. In non-stimulated glands, TPPase activity is detected mainly in 3–4 cisternae at the inner concave side of the Golgi complex and in some adjacent condensing vacuoles as in other cells. In the acinar cells 1 to 2 h after stimulation, however, reaction product for the same enzyme activity is detected in the cisternae at the outer aspect, as well as the inner aspect, of the Golgi complex and even in the cisternae of the endoplasmic reticulum (ER). About 4 h after stimulation, TPPase activity becomes concentrated in 3–4 disternae at the inner concave side of the Golgi complex as in the acinar cells under non-stimulated conditions. Morphological observations of the acinar cells 1 to 2 h after the stimulation have indicated that the reorganization of the Golgi complex and ER is a major event which occurs at this stage. It is possible that this cellular event is related to the occurrence of TPPase activity in those sites which normally show negative reaction in non-stimulated state.     

Effects of secretory stimulation on the Golgi apparatus and GERL of rat parotid acinar cells

The structure and cytochemistry of the Golgi apparatus and GERL of rat parotid acinar cells was studied after in vivo secretory stimulation with isoproterenol. Discharge of mature secretory granules was complete within 1 hr after isoproterenol injection, but immature granules in the Golgi region or near the lumen were not released. At early times (1-5 hr) after isoproterenol, acid phosphatase (AcPase) activity was markedly increased in GERL and immature secretory granules compared to uninjected controls. GERL appeared increased in extent and numerous continuities with immature granules were observed. Reaccumulation of mature secretory granules was first evident at 5 hr, and was almost complete by 16 hr after isoproterenol. Thiamine pyrophosphatase (TPPase) activity, normally restricted to the trans Golgi saccules, was frequently present in immature granules during this time. Narrow cisternae resembling GERL, occasionally in continuity with immature granules, also contained TPPase reaction product. By 16-24 hr after stimulation, the activity and distribution of AcPase and TPPase were similar to control cells. These results demonstrate the dynamic nature of the Golgi apparatus and GERL in parotid acinar cells, and emphasize the close structural and functional relationship between these two structures.     

Golgi apparatus, GERL, and secretory granule formation within neurons of the hypothalamo-neurohypophysial system of control and hyperosmotically stressed mice

The vasopressin-producing neurons of the hypothalamo-neurohypophysial system are a particularly good model with which to consider the relationship between the Golgi apparatus nd GERL and their roles in secretory granule production because these neurons increase their synthesis and secretion of vasopressin in response to hyperosmotic stress. Enzyme cytochemical techniques for acid phosphatase (AcPase) and thiamine pyrophosphatase (TPPase) activities were used to distinguish GERL from the Golgi apparatus in cell bodies of the supraoptic nucleus from normal mice, mice hyperosmotically stressed by drinking 2% salt water, and mice allowed to recover for 5-10 d from hyperosmotic stress. In nonincubated preparations of control supraoptic perikarya, immature secretory granules at the trans face of the Golgi apparatus were frequently attached to a narrow, smooth membrane cisterna identified as GERL. Secretory granules were occasionally seen attached to Golgi saccules. TPPase activity was present in one or two of the trans Golgi saccules; AcPase activity appeared in GERL and attached immature secretory granules, rarely in the trans Golgi saccules, and in secondary lysosomes. As a result of hyperosmotic stress, the Golgi apparatus hypertrophied, and secretory granules formed from all Golgi saccules and GERL. Little or no AcPase activity could be demonstrated in GERL, whereas all Golgi saccules and GERL-like cisternae were TPPase positive. During recovery, AcPase activity in GERL returned to normal; however, the elevated TPPase activity and secretory granule formation seen in GERL-like cisternae and all Golgi saccules during hyperosmotic stress persisted. These results suggest that under normal conditions GERL is the predominant site for the secretory granule formation, but during hyperosmotic stress, the Golgi saccules assume increased importance in this function. The observed cytochemical modulations in Golgi saccules and GERL suggest that GERL is structurally and functionally related to the Golgi saccules.     

Phosphatase localization in the endomembrane system of the dinoflagellate Crypthecodinium cohnii

The distribution of four enzymes within the endomembrane system of the protist Crypthecodinium cohnii has been determined using cytochemical localizations with lead as a capture agent. Nucleoside diphosphatase (NDPase) activity, using inosine diphosphate (IDP) and thiamine pyrophosphate (TPP) as substrates, was observed in the Golgi apparatus, with a gradient of increasing reaction product noted in some cells from the cis to trans cisternae. Tubules and vesicles associated with the trans cisternae also contained reaction product. The endoplasmic reticulum exhibited a high activity of glucose-6-phosphatase [with glucose-6-phosphate (G-6-P) as substrate]. Traces of reaction product were also observed in the cis-most and trans-most cisternae of the dictyosomes. Activity of acid phosphatase (AcPase) was observed in Golgi cisternae as well as in associated cytoplasmic vesicles. Heaviest deposition was localized in medial and trans dictyosome cisternae. The cytoplasmic system of flattened vesicles subtending the surface membranes in these cells did not exhibit reactivity with any of the substrates used. The distribution of these enzymes in this algal cell appears similar to that observed in animal cells and suggests that these enzymes may represent markers for algal cell endomembrane compartments.     

Effects of brefeldin A on the Golgi apparatus, the nuclear envelope, and the endoplasmic reticulum in a green alga,Scenedesmus acutus

Summary The effects of brefeldin A (BFA) on the structure of the Golgi apparatus, the nuclear envelope, and the endoplasmic reticulum (ER), and on the thiamine pyrophosphatase (TPPase) activity in these organelles were examined in a green alga,Scenedesmus acutus, to obtain evidence for the existence of a retrograde transport from the Golgi apparatus to the ER via the nuclear envelope. InScenedesmus, Golgi bodies are situated close to the nuclear envelope throughout the cell cycle and receive the transition vesicles not directly from the ER, but from the nuclear envelope. BFA induced the disassembly of Golgi bodies and an increase in the ER cisternae at the trans-side of decomposed Golgi bodies in interphase cells and multinuclear cells before septum formation. The accumulated ER cisternae connected to the nuclear envelope at one part. TPPase activity was detected in all cisternae of Golgi bodies, but not in the nuclear envelope or the ER in nontreated cells. On the contrary, in BFA-treated cells, TPPase activity was detected in the nuclear envelope and the ER in addition to the decomposed Golgi bodies. When septum-forming cells were treated with BFA, the disassembly of Golgi bodies was less than that in interphase cells, and TPPase activity was detected in the Golgi cisternae but not in the nuclear envelope or the ER. These results suggest mat BFA blocks the anterograde transport from the nuclear envelope to the Golgi bodies but does not block the retrograde transport from the Golgi bodies to the nuclear envelope in interphase and multinuclear cells.Abbreviations BFA brefeldin A - ER endoplasmic reticulum - TPPase thiamine pyrophosphatase     

The occurrence of the Golgi apparatus in mouse oocytes. Demonstration of thiamine pyrophosphatase activity

Location of thiamine pyrophosphatase activity as a marker of the Golgi apparatus was studied ultracytochemically in mouse oocytes with germinal vesicle (OGV), oocytes at metaphase I (OMI) and oocytes at metaphase II (OMII), and further in cells of the respective cumulus oophorus serving as comparative objects. TPPase activity in cumulus oophorus cells and in OGV was found exclusively in the Golgi apparatus. In OMI the reaction product of TPPase activity was observed in isolated smooth vesicles, and in only one case in structures identifiable as the Golgi apparatus. In OMII the occurrence of TPPase activity was also recorded in isolated smooth vesicles in cortical cytoplasm and further, exceptionally, in smooth concentrically arranged vesicles or tubules. The TPPase activity was not present in vesicular complexes. The results have shown that after the resumption of meiosis the occurrence of the reaction product of TPPase activity drops abruptly due to the reduction of the Golgi apparatus. Changes affecting the Golgi apparatus after the resumption of meiosis are related to the loss of the nucleus after the germinal vesicle breakdown.     

Cellular compartments in mitotic cells: ultrahistochemical identification of Golgi elements in PtK-1 cells

This investigation focuses on the identification, distribution, and transport of intracellular membrane systems during mitosis. The membranes of the Golgi apparatus can be identified cytochemically by staining for acid phosphatase (acPase) and thiamine pyrophosphatase (TPPase) activity. Using this approach we are able to study the disintegration of the Golgi apparatus during mitosis and to follow the dislocation as well as the organized reappearance of Golgi elements after the completion of mitosis. We are able to demonstrate that during mitosis the activity of both enzymes is strong enough to react with the substrate applied during the staining procedure. Furthermore, we observe a characteristic pattern of membrane distribution in mitotic cells. During interphase the TPPase reaction is characteristically limited to one or two cisternae of a dictyosomal stack. The acPase reaction stains the membranes of the total stack, of the GERL, of some vesicles and cisternae near the dictyosomes and lysosomes. After the mitotic breakdown of the dictyosomal stacks the forming vesicles still stain positively and are distributed over the entire cytoplasm. At late anaphase and early telophase the enzyme activity occurs not only in the reconstituting dictyosomes but also in the nuclear envelope and in some ER cisternae. The extended spectrum of membrane structures indicating Golgi enzyme activity becomes obvious. This phenomenon favors the idea that at least some functions of the Golgi apparatus persist during mitosis.     

Cytochemical studies on the internal polarity of the golgi apparatus and the relationship between this organelle and GERL

Summary Cytochemical studies were performed to clarify the occurrence of an internal polarity of the Golgi apparatus and the relationship between this organelle and GERL in many kinds of cells having different morphologies and functions. The fine structural localizations of thiamine pyrophosphatase (TPPase) and acid phosphatase (AcPase) were examined in anterior pituitary cells, thyroid epithelial cells, gastric chief and parietal cells, duodenal absorptive epithelial cells, hepatocytes, adrenal cortical and medullary cells of mice, and thyroid epithelial cells of domestic fowls. TPPase activity is usually localized in the cisternae of 1–3 stacks and vesicles on the trans-side of the Golgi apparatus of all the cells examined, and in some immature secretory granules of anterior pituitary cells and of gastric chief cells. Rigid lamellae and multivesicular bodies are rarely positive to this reaction, in several kinds of cells. AcPase activity was usually demonstrable in the cisternae of 1–3 stacks and vesicles on the trans-side of the Golgi apparatus, and also in rigid lamellae, coated vesicles, multivesicular bodies and lysosomes in all varieties of cells studied. Some immature secretory granules are positive to the AcPase reaction in anterior pituitary cells and gastric chief cells. The areas positive for both enzyme activities were partially or almost completely overlapping in all the cells examined, though there were minor variations among them. The grades of overlap are classified into three types. Prolonged osmication was performed on thyroid epithelial cells, duodenal absorptive epithelial cells, hepatocytes, adrenal cortical cells, Leydig cells, the epithelial cells of the vas deferens and the theca cells of mice. Cisternae of 1–3 stacks on the cis-side of the Golgi apparatus of all the cells examined were stained with osmium tetroxide. In all these cells we observed that the Golgi apparatus has an internal polarity and that GERL is a part of this organelle in cytochemical respects.This study was supported by grants from the Japan Ministry of Education     

Morphological and cytochemical alterations of the Golgi apparatus and GERL in rat parotid acinar cells during ethionine intoxication and recovery

The present electron microscopic cytochemical investigation was undertaken to characterize the alterations in the golgi apparatus and GERL of rat parotid acinar cells during ethionine intoxication and recovery. Although the Golgi apparatus and GERL were reduced in size, and some broadening of the Golgi saccules occurred as the result of ethionine treatment, the relative localization of thiamine pyrophosphatase (TPPase) activity in the Golgi saccules, and acid phosphatase activity (AcPase) in GERL, remained unchanged. Shortly after ethionine treatment was stopped, a dramatic redistribution of enzyme activities was noted. Within the first 24 hours of recovery, the Golgi apparatus began to enlarge, and the content of secretory granules increased. By day 3 of recovery, cisternae morphologically identifiable as GERL and forming secretory granules possessed TPPase activity, while AcPase activity was virtually undetectable. After seven days of recovery, the Golgi apparatus and GERL appeared both morphologically and cytochemically normal. The enzyme modulation observed during recovery may be correlated with increased secretory granule production. Furthermore, the presence of TPPase activity in GERL and forming secretory granules lends support to the suggestion that GERL may be derived from the trans Golgi saccule.     

The corpus luteum of the guinea pig. II. Cytochemical studies on the Golgi complex, GERL, and lysosomes in luteal cells during maximal progesterone secretion

This study characterizes the cytochemical properties of the Golgi complex, the structure which corresponds to Golgi complex-endoplasmic reticulum-lysosomes (GERL), and the granule population in luteal cells of guinea pigs at the time of maximum progesterone secretion, in material fixed by vascular perfusion, a method particularly suited for preserving both fine structure and enzyme activity. The distribution of several marker enzymes was determined by electron microscope cytochemistry. Acid phosphatase (ACPase) and arylsulfatase were used to identify structures containing lysosomal proteins. To resolve specific problems, additional cytochemical markers were employed: localization of thiamine pyrophosphatase (TPPase) (in the Golgi complex) and alkaline phosphatase (ALPase) (a plasma membrane marker), and prolonged osmication (a generally accepted method of marking the outer cisterna of the Golgi complex). The results demonstrate that at the time of peak steroid secretion the Golgi complex in luteal cells, in marked contrast to that of most other cell types, typically displays intense ACPase activity in all of its cisternae. Similarly, all Golgi cisternae stain after prolonged osmication and may show TPPase activity. On the other hand, GERL in luteal cells of this age, unlike that in most cells, commonly shows low levels of, or lacks, ACPase activity. However, GERL resembles that of other cell types in being TPPase-negative and in being unstained by treatment with aqueous OsO4. GERL and some Golgi cisternae are reactive for ALPase. The granule population in luteal cells of this stage consists of lysosomes, multivesicular bodies, electrontransparent vacuoles, and microperoxisome-like bodies. These results form a base line with which luteolytic changes described in the companion study (Paavola, L.G. 1978. The corpus luteum of the guinea pig. III. Cytochemical studies on the Golgi complex and GERL during normal postpartum regression of luteal cells, emphasizing the origin of lysosomes and autophagic vacuoles. J. Cell. Biol. 79:59--73.) can be compared.     

Membrane modification during secretory granule formation in rat somatotrophs

Somatotrophs from male rat anterior pituitary were used to investigate the formation of secretory granules. When enzymatically dispersed cells were incubated with cationized ferritin (CF) for 15 min, CF labeled immature secretory granules, but not mature granules of somatotrophs. Most immature granules labeled by CF transformed to the mature types within 120 min. This indicates that the fusion of endocytic vesicles with the immature granules occurs during the maturation process of secretory granules. The internalized CF was distributed not only in the immature secretory granules, but also in the peripheral region of trans Golgi cisternae or GERL. Enzyme cytochemistry revealed that acid phosphatase-positive cisternae (GERL) were the main site for secretory granule formation, and was devoid of thiamine pyrophosphatase (TPPase) activity. A small number of secretory granules were also present in the peripheral regions of TPPase-positive Golgi cisternae. The granule-forming sites, however, lacked TPPase activity, while the remaining region of the same cisterna showed the positive enzyme activity. This indicates that the granule-forming region at the periphery of Golgi cisterna is different from the remaining part of the same cisterna in terms of cytochemical properties. This probably results from the insertion of endocytic vesicle membrane, since the same granule-forming sites preferentially fused with CF-labeled small vesicles which lacked cytochemical TPPase activity. Taken together. Our results suggest that the membrane of secretory granules is modified during the granule formation, at least partly by the fusion of endocytic small vesicles with Golgi cisternae (or GERL), and with immature secretory granules.     

Ultrastructural demonstration of lectin binding sites in the Golgi apparatus of rat epiphyseal chondrocytes

Binding sites for wheat germ agglutinin (WGA), Dolichos biflorus agglutinin (DBA), Ricinus communis I agglutinin (RCA I) and Limax flavus agglutinin (LFA) have been ultrastructurally detected in rat epiphyseal chondrocytes by a post-embedding cytochemical technique using colloidal gold as marker. The four lectins labelled exclusively the Golgi apparatus of chondrocytes embedded in Lowicryl K4M resin by two different methods. WGA binding sites were localized in medial and trans cisternae as well as in immature secretory vesicles, whereas those for DBA were seen concentrated in cis and medial cisternae. Labelling with both RCA I and LFA lectins was distributed throughout all the cisternae of the Golgi stack, and the latter also in vesicles and tubules at the trans face. Neuraminidase pretreatment of the sections abolished LFA staining, decreased reaction with WGA and increased that with RCA I, while it did not affect DBA staining. After chondroitinase ABC treatment only the RCA I reaction was modified, revealing new binding sites in the trans Golgi face, secretory granules and extracellular matrix. These results indicate that the distribution of subcompartments in the Golgi apparatus of chondrocytes is different from that in cells secreting glycoproteins as major products.     

Cytochemistry of the Golgi apparatus in developing ovarian germ cells of the Syrian hamster

Summary A cytochemical study of the Golgi apparatus in the developing oocyte of the golden hamster was carried out using the TPPase, AcPase and zinc iodide-osmium tetroxide (ZnOs) techniques. Tissue from both immature and sexually mature animals was investigated.Peak TPPase activity was found in pre-growth oocytes in ovaries from sexually mature adults. Some activity was also present in SER in the peripheral cytoplasm of growing oocytes. AcPase activity was found only after the onset of oocyte growth. It was present in Golgi cisternae and associated vesicles and in some profiles of peripheral SER. No structures corresponding to GERL were identified. Strong staining with ZnOs was seen, at all stages studied, in certain Golgi vesicles and short tubules but not in the cisternae unless the oocyte was atretic. Weaker ZnOs staining was characteristic of ER throughout the oocyte.With all techniques there was a falling off of reactivity as oocyte size increased. Within a single oocyte some Golgi bodies were negative while others were positive, with both TPPase and AcPase techniques. This suggests that two or more functional types of this organelle are present within the developing oocytes.We would like to thank Dr. K.N. Christie for his interest and helpful suggestions regarding the enzyme techniques     

Prenatal exposure to alcohol alters the Golgi apparatus of newborn rat hepatocytes: a cytochemical study

The effect of prenatal exposure to ethanol on the Golgi apparatus of newborn rat hepatocytes has been studied cytochemically using several trans-Golgi markers (thiamine pyrophosphatase, uridine diphosphatase, inosine diphosphatase, acid phosphatase, and 5'-nucleotidase) as well as a cis-side marker (osmium impregnation). The amount of cerium phosphate formed in the cytochemical reactions was roughly quantitated by stereologic methods. The Golgi apparatus of about 40% of the hepatocytes appeared disorganized after alcohol treatment, and in the other 60%, the electron density of reaction product deposits for all phosphatases investigated was decreased. 5'-Nucleotidase was completely absent in cisternae of Golgi apparatus of treated cells. In control cells impregnated with osmium tetroxide, reduced osmium compounds were observed in most Golgi cisternae and in nearby vesicles. In contrast, only small vesicles appeared positive in treated hepatocytes. These results suggest that prenatal alcohol exposure alters some Golgi functions. Thus, the decrease in nucleoside diphosphatase and 5'-nucleotidase cytochemical activities after ethanol exposure strongly suggests that this treatment could affect glycosylation in the Golgi apparatus of newborn rat hepatocytes.     

Effect of monensin on the Golgi apparatus of absorptive cells in the small intestine of the rat

Summary The effect of short-time treatment with the ionophore monensin, administered intraluminally at concentrations of 5 and 10 M, was studied on the Golgi apparatus of absorptive cells in the small intestine of the rat. At 2–3 min after treatment most of the Golgi stacks exhibited dilated cisternae. At 4–5 min stacked cisternae were absent; they were replaced by groups of smooth-surfaced vacuoles. Dilatation and vacuolization occurred in the entire stacks without preferential effect on any particular Golgi subcompartment.Monensin did not influence the cytochemical Golgi reaction of thiamine pyrophosphatase and acid phosphatase. The characteristic staining pattern of these two enzymes in all Golgi cisternae of absorptive cells in the proximal small intestine, and the reactivity restricted to trans cisternae in distal segments of the small intestine, were unchanged after treatment with monensin. In the distal small intestine, the cytochemical pattern allowed the monensin-induced vacuoles to be attributed to the former cisor trans-Golgi face. Further, the cytochemical results demonstrate that vacuolization is not restricted to the stacked cisternae, but includes the trans-most cisterna. The latter, usually located at some distance from the Golgi stacks, has been defined as belonging to the GERL system in several types of cells. The clear response to monensin, an agent that selectively affects the Golgi apparatus, indicates common properties between trans-most and stacked Golgi cisternae.