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W J Larsen

Publications and source records attributed to W J Larsen.

At least 37 records · Page 2Linked to original sources
PubMed1990

Hyaluronic acid synthesis and gap junction endocytosis are necessary for normal expansion of the cumulus mass.

The application of a quantitative videographic technique has provided an opportunity to compare the quantitative volumetric expansion of cultured oocyte complexes (COCs) to quantitative changes in gap junction down-regulation and hyaluronic acid synthesis and to investigate the effects of physiological agents that influence these processes. Results of these experiments support the idea that the down-regulation of cumulus gap junctions is required for the initial phase of cumulus cell disaggregation and confirm earlier reports that hyaluronic acid synthesis plays a major role in additional expansion of the cumulus. These studies also provide evidence that the degree of expansion observed in culture lacking substrates of hyaluronic synthesis is significantly attentuated when compared with expansion occurring in vivo and that the failure of cultured complexes to expand maximally can be overcome by the addition of substrates of hyaluronic acid synthesis to the culture medium.

Animals↗
PubMed1990

Preendocytotic alterations in cumulus cell gap junctions precede meiotic resumption in the rat cumulus-oocyte complex.

Cumulus cells in the mammalian ovary are normally connected to each other and to their enclosed oocyte by an extensive network of gap junctions (GJs). We have shown that the loss of cumulus cell GJs is correlated temporally with meiotic resumption in the intact preovulatory rat follicle (Larsen et al., 1986). Here we describe morphological changes in GJ particle packing patterns (PPPs) that occur prior to GJ loss and meiotic resumption in hormonally stimulated rat cumulus-oocyte complexes (COCs). In the PMSG-primed rat, 89% of the cumulus cell GJ area detected by freeze-fracture electron microscopy consists of tightly packed junctional particles: 4% exhibit loose PPPs of randomly dispersed particles; and 7% contain a mixture of both tight and loose PPPs. One to 2 hr after stimulation with hCG, the area of GJs containing tight PPPs drops by 50%-60%, while junctions exhibiting loosely organized and mixed patterns increase concomitantly. These shifts in PPPs are accompanied by the appearance of unusual particle-free areas of puckered or ruffled nonjunctional membrane at the GJ periphery. Cumulus cell GJs from isolated COCs incubated in FSH-containing medium demonstrate a similar shift in PPPs prior to meiotic resumption. The appearance of fusing areas of particle-free nonjunctional membrane at the GJ periphery in vitro is correlated with GJ loss and is not seen in COCs treated with dihydrocytochalasin B to inhibit endocytotic removal of cumulus GJs. The structural and temporal nature of these morphological observations supports the hypothesis that interruption of junctional communication plays a role in meiotic maturation of the preovulatory oocyte.

Animals↗
PubMed1989

Meiotic resumption and gap junction modulation in the cultured rat cumulus-oocyte complex.

Intercellular communication within the ovarian follicle has been implicated in the control of meiotic arrest and maturation in the mammalian oocyte. We have shown that a rapid down-regulation of cumulus cell gap junctions is correlated temporally with meiotic resumption in the intact rat follicle [Larsen et al., Dev Biol, 113:517-521]. Here this relationship has been analyzed further by incubating isolated rat cumulus-oocyte complexes (COCs) with agents known to influence germinal vesicle breakdown (GVBD) or that have been shown to modulate gap junction turnover in vitro. Quantitative freeze-fracture analysis revealed that cumulus cell gap junction membrane decreased significantly prior to the initiation of GVBD in COCs incubated in medium lacking serum or other additives. The addition of serum and follicle-stimulating hormone, an experimental condition that delayed GVBD, accelerated and augmented gap junction loss at both the cumulus cell and oocyte surface. The continuous elevation of cyclic adenosine monophosphate levels, which stimulates gap junction formation in other systems, maintained meiotic arrest but did not interfere with gap junction loss. Conversely, the complete inhibition of junctional loss by a microfilament destabilizing agent, dihydrocytochalasin B, did not alter the course of GVBD normally seen in its absence. Subsequent freeze-fracture analysis and dye coupling experiments confirmed that cumulus and oocyte gap junctions in these preparations were intact and functional during the period of meiotic resumption. These findings suggest that factors other than cumulus and oocyte gap junction turnover are required for the control of meiotic arrest and maturation in the isolated COC. However, these results do support our earlier suggestion that gap junction loss within the cumulus oophorus is instrumental in isolating the oocyte from the regulatory influence of its underlying membrana granulosa cells during meiotic maturation in the intact preovulatory follicle [Larsen et al., Dev Biol, 122:61-71].

Animals↗
PubMed1988

Roles of cell junctions in gametogenesis and in early embryonic development.

Recent reviews of the role of cell junctions in development have focused primarily upon functions related to the relatively subtle physiological modulation of their subunits in relation to fundamental developmental processes in a wide variety of organisms. There is, however, considerable support from numerous laboratories that the more radical modulation of the presence and number of junctional subunits in many diverse tissues may play a pivotal role in a wide spectrum of developmental phenomena ranging from gametogenesis to organogenesis. Since a great deal of recent interest in this latter subject has concentrated upon vertebrate systems including mammals, this review will examine the functional significance of the modulation of gap junctions, tight junctions and desmosomes in a developing idealized mammalian system from gamete formation to tissue and organ differentiation during embryo-genesis.

Animals↗
PubMed1987

Differential modulation of rat follicle cell gap junction populations at ovulation.

The hypothesis proposed in the late 1970s that meiotic resumption in mammalian oocytes might result from the disruption of gap junction communication between follicle cells and the oocyte has not been supported by metabolic cooperation experiments which demonstrate that exogenous tracer transfer from the cumulus oophorus to the oocyte does not decrease until several hours after germinal vesicle breakdown (GVBD). Since these studies utilized isolated cumulus-oocyte complexes for their measurements, however, they excluded from consideration the possible effect of separation of the cumulus oophorus from the membrana granulosa which was required for this assay. We considered the possibility that the disruption of cumulus junctions within the intact follicle could mimic this experimental manipulation and previously reported that cumulus gap junctions were dramatically down-regulated during the period of GVBD in vivo. In the present study, we have utilized quantitative morphometric techniques to analyze the responses of other gap junction populations in intact preovulatory rat follicles to an ovulatory stimulus and demonstrate now that membrana granulosa, cumulus, and cumulus-oocyte gap junctions are down-regulated at different times and rates during the preovulatory period. Although membrana gap junctions are down-regulated during the period of meiotic resumption, their loss is not as rapid or as complete as in the cumulus oophorus. Cumulus-oocyte gap junctions are down-regulated after meiosis resumes but during the same period other investigators have demonstrated a reduction in metabolite transfer between the cumulus oophorus and the oocyte. Our results are interpreted to suggest that the cumulus oophorus may regulate the conduction of meiosis inhibitory signals between the membrana granulosa and the oocyte.

Animals↗
PubMed1986

Interaction of lipoprotein lipase with phospholipid vesicles: effect on protein and lipid structure.

The interaction of lipoprotein lipase (LpL) and a nonhydrolyzable phosphatidylcholine, 1,2-ditetradecyl-rac-glycero-3-phosphocholine (C14-ether-PC), has been studied by several physical methods. Analysis of the circular dichroic spectrum of LpL gave the following fractional conformation: 35% alpha-helix, 30% beta-pleated sheet, and 45% remaining structure. No significant change in the circular dichroic spectrum of LpL was observed on addition of C14-ether-PC vesicles. The quenching of LpL fluorescence by acrylamide and iodide ion was decreased only slightly by addition of C14-ether-PC vesicles. Addition of LpL to sonicated C14-ether-PC vesicles containing entrapped carboxyfluorescein caused the release of less than 15% of the vesicle contents in 20 min, indicating that the enzyme did not disrupt the structure of the lipid. In contrast, greater than 80% of the vesicle contents were released with the addition of apolipoprotein A-I to an identical vesicle preparation. The temperature dependence of the fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene incorporated into C14-ether-PC vesicles was not significantly altered by the addition of LpL. When LpL is added to vesicles, the bilayer structure of the vesicles is not disrupted as observed by freeze-fracture electron microscopy. However, at low ionic strength (0.1-0.25 M NaCl) significant aggregation of intact vesicles is observed by light scattering and electron microscopy. Vesicle aggregation is prevented and reversed by 1 M NaCl and by heparin. These data demonstrate that LpL binds to the surface of a lipid interface, without dramatic changes in lipid bilayer or protein structure.

Animals↗
PubMed1986

A dramatic loss of cumulus cell gap junctions is correlated with germinal vesicle breakdown in rat oocytes.

Interactions between the cumulus-oophorus and the oocyte have been implicated in the regulation of meiotic maturation. Quantitative analysis of freeze-fractured rat cumulus-oocyte complexes reveals that the net area of cumulus cell gap junction membrane decreases about 15-fold, 2-3 hr following an ovulatory stimulus. This dramatic loss of gap junctions is temporally correlated with germinal vesicle breakdown and cumulus expansion, and is discussed with respect to meiotic maturation and ovulation of the mammalian oocyte.

Animals↗
PubMed1984

An in vitro model for studies of intercellular communication in cultured rat anterior pituitary cells.

The formation of intimate associations among different hormone-secreting cells within the rat adenohypophysis may serve as a possible site for physiologic regulation. In this report we describe a high density plating method which enables us to study cell-to-cell interactions within anterior pituitary cell cultures. Trypsin-dispersed pituitary cell suspensions attach rapidly (within 6 hr) and quantitatively (95-97%) to glass or plastic surfaces when plated in medium containing microM calcium concentrations (pH 7.6-7.8). Freshly plated cell suspensions obtained from female pituitary glands contained subpopulations of mammotrophs 49.3%, somatotrophs 30.3%, gonadotrophs 12.6%, corticotrophs 3.4% and thyrotrophs 1.5%. Epithelial cell colonies were formed during a 3-day culture period as the cells flattened and re-established contacts with neighboring cells. Freeze-fracture electron microscopic analysis of these colonies produced morphological evidence for direct intercellular contacts among the hormone-secreting cells. Large areas of tight junctions and small gap junctions were identified on the membranes of the epithelial cells within these colonies. Cells which contained tight junctions usually contained microvilli and morphological signs of active hormone secretion. Small junctional plaques containing tightly packed intramembrane particles were also occasionally found on the membranes of cells which were actively secreting pituitary hormones. The high density plating procedure which is described in this report provides greater opportunity for cell-cell interaction and thus may prove to be a useful model for evaluating the role of intercellular communication within this tissue.

Animals↗
PubMed1983

Interaction of filipin with junctional membrane at different stages of the junction's life history.

The relationship of filipin-sterol complexes to tight and gap junctions during their formation, maturation, internalization, and degradation was studied in separate cell lines. Filipin-sterol complexes tended to be excluded from mature junctions in tight junction forming COLO 316 cells and gap junction forming SW-13 cells. Once internalized, unlabeled junctional membrane appeared to fuse with heavily labeled vesicles, presumably lysosomes. Although the absence of filipin-sterol complexes from junctional membrane does not necessarily reflect the absolute sterol content of this membrane, the fact that filipin-sterol complexes are largely excluded from these areas indicates that this membrane is different from surrounding membrane. The absence of filipin-sterol complexes also permits the visualization of 'mixing' of this specialized unlabeled membrane domain with other filipin labeled membrane systems.

Adrenal Gland Neoplasms↗
PubMed1983

Biological implications of gap junction structure, distribution and composition: a review.

Traditionally, all gap junctions have been considered to be identical in structure and function throughout the animal kingdom. Functions ascribed to these membrane specializations have been fundamental and have not been thought to differ significantly with respect to their mechanism of action. More recent studies support the view, however, that structural and compositional diversity may reflect significant functional differences between gap junctions in different classes of tissue but no clear and definitive patterns have yet emerged. This review does not attempt to comprehensively analyze the totality of the vast gap junction and coupling literature but focuses instead upon those recent observations which raise new questions related to the biological activities of gap junctions in different tissues.

Freeze Fracturing↗
PubMed1981

Endocytosis of cell-cell junctions and spontaneous cell disaggregation in a cultured human ovarian adenocarcinoma. (COLO 316).

Although cultured COLO 316 human ovarian adenocarcinoma cells are joined by extensive tight junctions and numerous demosomes in confluent monolayers, viable cells may be spontaneously released into the nutrient medium. Intracytoplasmic vesicles containing tight junctional and desmosomal elements were identified in freeze-fracture and thin section preparations of the released cells and some vesicles exhibited structural signs of degradation. Possible mechanisms for tight junctional and desmosomal interiorization and the possible relationship between junctional interiorization and certain malignant behaviors are discussed.

Adenocarcinoma↗
PubMed1981

Gap junction assembly and endocytosis correlated with patterns of growth in a cultured adrenocortical tumor cell (SW-13).

After seeding at subconfluent densities, human adrenocortical adenocarcinoma cells (SW-13) proliferate to form a typical epithelial monolayer upon the culture vessel substratum. Single cells of the confluent monolayer are spontaneously released into the medium, however, where they may remain isolated or reaggregate with other floating cells. In this study, we examined the growth patterns of these cells in culture and analyzed alterations of their most prominent intercellular contact specialization, the gap junction, as cellular relationship changed during growth and development of the cell culture. We report here details of gap junction assembly during cellular aggregation and correlate the apparent endocytosis of gap junctions with the spontaneous release of single cells from the monolayer into the medium.

Adenocarcinoma↗
PubMed1980

The bicellular and reflexive membrane junctions of renomedullary interstitial cells: functional implications of reflexive gap junctions.

Renomedullary interstitial cells are loosely organized within the interstitial space surrounding collecting ducts, limbs of Henle, and capillaries of the rat renal medulla. These cells possess long processes, which interact with each other and with cell bodies to form bicellular tight, intermediate, and gap junctions. In addition, both cell bodies and cell processes possess "reflexive" gap and intermediate junctions. Possible functions of renomedullary interstitial cell membrane junctions are discussed. Particular attention is given to a consideration of the functional significance of "reflexive" gap junctions.

Animals↗
PubMed1979

Evidence for the participation of actin microfilaments and bristle coats in the internalization of gap junction membrane.

Thin sections of rabbit granulosa, human SW-13 adrenal cortical adenocarcinoma, and mouse B-16 melanoma cells revealed an apparent single-layered basket of 4- to 7-nm filaments surrounding cytoplasmic gap junction vesicles. This interpretation was based upon apparent longitudinal, cross, and en face sections of structures surrounding the vesicle profiles in tissue treated with tannic acid-glutaraldehyde. In granulosa cells incubated with the S-1 fragment of heavy meromyosin, arrowhead-decorated filaments were observed at the periphery of the gap junction vesicles, suggesting that these filaments contained actin. In addition, we found that small gap junctional blebs and vesicles at the cell surface were coated with short electron-dense bristles similar in appearance to the cloathrin-containing coat of coated vesicles of nonjunctional membrane. The role of these and other cytoskeletal elements in the possible endocytosis of gap junction membrane is discussed.

Actins↗
PubMed1978

Gap junctions and ACTH sensitivity in Y-1 adrenal tumor cells.

Initial studies of adrenocorticotropin-sensitivity (ACTH-sensitive) and ACTH-insensitive Y-1 adrenal cortical tumor cell lines suggest a relationship between responsiveness to ACTH and the presence of gap junctions. An ACTH-sensitive clone of Y-1 cells possesses gap junctions and these junctions appear to enlarge with ACTH treatment. GAP junctions have not been observed, however, in an ACTH-insensitive clone of Y-1 tumor cells even when stimulated to produce cyclic adenosine monophosphate and steroids with cholera toxin.

Adrenal Gland Neoplasms↗
PubMed1978

Origin and fate of cytoplasmic gap junctional vesicles in rabbit granulosa cells.

Vesicles within the cytoplasm of the rabbit granulosa cell, in both thin section and freeze-fracture preparations, appear to consist of typical gap junctional membrane. In most cases these vesicles, like gap junctions at the cell surface, are composed of two membranes, but in rare cases may consist in part of single membrane, identified as gap junctional because of the presence of typical 8.5 nm P face particles and E face pits. Both thin section and freeze-fracture studies support the view that these vesicles may be produced by the endocytosis of junctional membrane from the cell surface as one cell invaginates into its neighbor at the junction. The presence of apparent acid phosphatase activity within the matrix of these gap junctional vesicles suggests that they may represent a stage in the specific degradation of gap junctional membrane.

Acid Phosphatase↗
PubMed1977

Intercellular communication and tissue growth: IX. Junctional membrane structure of hybrids between communication-competent and communication-incompetent cells.

The structure of the membrane junctions of the hybrid cell system, examined in the companion paper in respect to competence for communication through cell-to-cell membrane channels, is here examined by freeze-fracture electron microscopy. The junctions of the channel-competent parent cell and of the channel-competent hybrid cells present aggregates of intramembranous particles typical of "gap junction"; those of the channel-incompetent parent cell and channel-incompetent segregant hybrid cells do not. Competence for junctional communication and for gap junction formation are genetically related. The junctions of the intermediate hybrid cells with incomplete channel-competence (characterized by cell-to-cell transfer of small inorganic ions but not of fluorescein), present special intramembranous fibrillar structures instead of discrete gap-junctional particles. The possibility that these structures may constitute coupling elements with subnormal permeability is discussed in terms of incomplete dominance of the genetic determinants of gap junction.

Animals↗