The arthropod gap junction and pseudo-gap junction: Isolation and preliminary biochemical analysis

Summary The hepatopancreas of the crayfish, Procambarus clarkii, contains an unusual abundance of gap junctions, suggesting that this tissue might provide an ideal source from which to isolate the arthropod-type of gap junction. A membrane fraction obtained by subcellular fractionation of this organ contained smooth septate junctions, zonulae adhaerentes, gap junctions and pentalaminar membrane structures (pseudo-gap junctions) as determined by electron microscopy. A further enrichment of plasma membranes and gap junctions was achieved by the use of linear sucrose gradients and extraction with 5 mM NaOH. The enrichment of gap junctions correlated with the enrichment of a 31 Kd protein band on polyacrylamide gels. Extraction with 20 mM NaOH or 0.5% (w/v) Sarkosyl NL97 resulted in the disruption and/or solubilization of gap junctions. Negative staining revealed a uniform population of 9.6 nm diameter subunits within the gap junctions with an apparent sixfold symmetry. Using antisera to the major gap junctional protein of rat liver (32 Kd) and to the lens membrane protein (MP 26), we failed to detect any homologous antigenic components in the arthropod material by immunoblotting-enriched gap junction fractions or by immunofluorescence on tissue sections. The enrichment of another membrane structure (pseudo-gap junctions), closely resembling a gap junction, correlated with the enrichment of two protein bands, 17 and 16Kd, on polyacrylamide gels. These structures appeared to have originated from intracellular myelin-like figures in phagolysosomal structures. They could be distinguished from gap junctions on the basis of their thickness, detergent-alkali insolubility, and lack of association with other plasma membrane structures, such as the septate junction. Pseudo-gap junctions may be related to a class of pentalaminar contacts among membranes involved in intracellular fusion in many eukaryotic cell types. We conclude that pseudo-gap junctions and gap junctions are different cellular structures, and that gap junctions from this arthropod tissue are uniquely different from mammalian gap junctions of rat liver in their detergentalkali solubility, equilibrium density on sucrose gradients, and protein content (antigenic properties).     

Morphometrical analysis of the gap-junctional area in parenchymal cells of the rat liver after administration of dibutyryl cAMP and aminophylline

Summary In view of the presumed involvement of gap junctions in the coordination of metabolic activities, the influence of cAMP as a regulatory signal of cell metabolism on gap junctions of hepatocytes has been examined. Male rats received two intraperitoneal doses of 10 mg dibutyryl cAMP/100 g body weight with a time interval of 2.5 h and were decapitated 2.5 h later. After this 5-h interval, analysis of freeze-fracture replicas of fixed liver tissue revealed an increase in the mean (± SEM) gap-junctional membrane portion on the lateral hepatocyte membranes from 0.049 + 0.003 (n = 66) in controls to 0.061 ± 0.003 (n = 70) in treated rats, while the configuration of the connexons appeared unaltered. This effect could not be reinforced by prior administration of aminophylline: the relative gapjunctional area is similarly extended from 0.054 ± 0.003 (n = 126) in the control group to 0.065 ± 0.004 (n = 105) in the experimental animals. Probing for the time course of the junctional response, a group of rats was sacrificed 3 h after the onset of treatment. Already within this time, the gapjunctional area is augmented from 0.042 ± 0.004 (n = 63) in the concurrent controls to 0.069 ± 0.006 (n = 42) in the treated rats. These statistically significant increases in area may suggest a stimulating effect of cAMP on gap junctions of hepatocytes in vivo.This investigation was supported by grant No. 3.0059.81 (to D.W.S.) from the Fund for Medical Scientific Research (Belgium)     

Seedling recruitment ofChamaecyparis obtusa andSciadopitys verticillata in different microenvironments in an old-growthSciadopitys verticillata forest

Seedling recruitment ofChamecyparis obtusa andSciadopitys verticillata in different microenvironments in an old-growthS. verticillata forest was studied for five years. Exposed mineral soil beneath a gap, which was newly created by a typhoon, supported the best seedling emergence forC. obtusa. S. verticillata seemed to require no specific microenvironment for its seedling emergence. Mortality of seedlings was higher for both species beneath a closed canopy, where all new seedlings ofC. obtusa died within a year after their emergence. Beneath the gap, mortality of seedlings was generally lower for both species, regardless of the presence of litter on the ground. The occurrence of saplings was more frequent forC. obtusa beneath the gaps and forS. verticillata beneath a closed canopy. Difference of seedling recruitment behaviour could maintain the coexistence ofC. obtusa andS. verticillata in the canopy layer of this forest in a non-equilibrium state caused by occasional disturbance.     

The gap theory in forest dynamics

Since the late 1970s, ecologists interested in forest dynamics have focused their attention to the responses of individuals, populations and communities to “gaps” which are openings created in the forest canopy. This review intended to introduce some collective knowledge on major subjects of the gap theory in forest dynamics, in relation to gap-disturbance regimes, tree regeneration behaviour and community structure.     

外胚层细胞诱导后的间隙连接

用电镜观察了经受诱导作用之后胚胎细胞的冷冻蚀刻复型膜。和未经诱导作用的对照组比较,早期和中期神经胚的神经上皮细胞以及经过豚鼠骨髓粗提液(BME)——一种有效的异源中胚层诱导物——处理过的早期原肠胚外胚层,它们的间隙连接都处于活跃的动态状态。用图像分析仪测得的间隙连接连接子的排列密度,指出经受过诱导作用的三组分别和未经受诱导作用的对照组比较,计算求出P值,神经上皮两组和对照组的差别为非常显著,BME处理过的细胞和对照组的差别为显著。结合对照组与诱导后胚胎细胞间隙连接连接子的变化讨论了它们在信息传递上可能起的作用。     

Interneuronal and glial-neuronal gap junctions in the lamina ganglionaris of the compound eye of the housefly,Musca domestics

Summary The cell-body layer of the lamina ganglionaris of the housefly, Musca domestica, contains the perikarya of five types of monopolar interneuron (L1–L5) along with their enveloping neuroglia (Strausfeld 1971). We confirm previous reports (Trujillo-Cenóz 1965; Boschek 1971) that monopolar cell bodies in the lamina form three structural classes: Class I, Class II, and midget monopolar cells. Class-I cells (L1 and L2) have large (8–15 m) often crescentshaped cell bodies, much perinuclear cytoplasm and deep glial invaginations. Class-II cells (L3 and L4) have smaller perikarya (4–8 m) with little perinuclear cytoplasm and no glial invaginations. The midget monopolar cell (L5) resides at the base of the cell-body layer and has a cubshaped cell body. Though embedded within a reticulum of satellite glia, the L1–L4 monopolar perikarya and their immediately proximal neurites frequently appose each other directly. Typical arthropod (-type) gap junctions are routinely observed at these interfaces. These junctions can span up to 0.8 m with an intercellular space of 2–4 nm. The surrounding nonspecialized interspace is 12–20 nm. Freezefracture replicas of monopolar appositions confirm the presence of -type gap junctions, i.e., circular plaques (0.15–0.7 m diam.) of large (10–15 nm) E-face particles. Gap junctions are present between Class I somata and their proximal neurites, between Class I and Class II somata and proximal neurites, and between Class II somata. Intercartridge coupling may exist between such monopolar somata. The cell body and proximal neurite of L5 were not examined. We also find that Class I and Class II somata are extensively linked to their satellite glia via gap junctions. The gap width and nonjunctional interspace between neuron and glia are the same as those found between neurons. The particular arrangement and morphology of lamina monopolar neurons suggest that coupling or low resistance pathways between functionally distinct neurons and between neuron and glia are probably related to the metabolic requirements of the nuclear layer and may play a role in wide field signal averaging and light adaptation.     

Forest succession in the subalpine region of Mt. Fuji,Japan

Subalpine forest succession was studied on Mt. Fuji, Japan, where various types of forests in different successional phases occur owing to volcanic action. Ninety stands were subjected to ordination using an index (SI) defined by the relative basal area and the life span of component woody species, and the cover of canopy layer of the sample stands. Two different sequences of sample stands were found. One was from deciduous scrubs, through Larix kaempferi forests and Abies forests, to Tsuga diversifolia forests, and the other from Abies-Tsuga thickets to Abies forests. Through analyses of the forest structure and composition, soil survey and identification of fallen logs, the former sequence was recognized as the primary sere and the latter as a regeneration sere following gap formation. During forest succession, basal area reached a maximum in the seral phase with a multi-layered structure. The Tsuga forests, whose understory is restricted to a moss layer, were regarded as the climax. The death or fall of Tsuga stems resulted in gaps, which were subsequently occupied by Abies-Tsuga thickets. The second Abies forests were distinguished from the ones in the primary sere by the occurrence of Dryopteris and Cacalia and the lack of Rhododendron in the understorey. Both Abies forest types included Tsuga saplings. Thus, a cyclic relation is supposed between Abies and Tsuga.Nomenclature follows Ohwi (1975) and Nakaike (1982) for vascular plants, Iwatsuki & Noguchi (1973) for mosses, Inoue (1981) for hepaticae, Kashiwadani (1981) for lichens, respectively. Abies veitchii, A. mariesii were lumped as Abies spp.I wish to express my sincere gratitude to Prof. Toshio Hamaya, Tokyo, for the cordial guidance and encouragement. I also thank Prof. M. Numata and Dr. M. Ohsawa, Chiba, Prof. K. Okutomi, Tokyo, Dr. K. Suzuki, Tokyo, Dr. M. Suzuki, Kanazawa, and Mr. H. Taoda, Kumamoto, for their valuable advice and discussions.     

Gap junction reductions precede tissue hyperplasia following temperature-upshift in theDrosophila ts-mutantl(3)c43 hs1

Summary The wing discs of the temperature-sensitiveDrosophila mutantl(3)c43 ^hs1 become hyperplastic when larvae are reared at the restrictive temperature of 25° C or above (Martin et al. 1977). We have previously shown that reductions in gap junctions are correlated with the hyperplasia (Ryerse and Nagel 1984a). We report here that reductions in gap junction surface density, number and percent of the lateral plasma membrane area precede the onset of tissue hyperplasia as defined by the gross appearance of tissue overgrowth in the wing pouch and an increase in cell number. Gap junction reductions begin soon after temperature upshift and become significantly different from non-shifted controls by 16 h. Direct cell counts indicate that there is no difference in the total number of cells in experimental vs control discs until after 16 h when the 28° C discs begin to grow rapidly with a cell doubling time of about 6 h as compared with about 13 h for the 20°C controls. The finding that gap junction reductions precede the onset of tissue hyperplasia is consistent with the idea that gap junctions play a regulatory role in growth control and pattern formation and strengthens our hypothesis (Ryerse and Nagel 1984b) that a minimum number and a specific distribution of gap junctions are required for normal development.     

Climate and plant distribution at global and local scales

This paper investigates, with predictive models, the utility of ecophysiological responses to climate as predictors of plant distribution. At the global scale responses to extreme minimum temperatures and to the hydrological budget effectively predict the distribution limits of the major vegetation types of the World. A minimum temperature of -15°C, for example, appears critical in controlling the poleward spread of vegetation that is dominated by evergreen broadleaved species; however, the presence or absence of more frost resistant species, such as those that are deciduous broadleaved, is not obviously explained in terms of extremes of climate. In such cases, predicting the competitive relationships between species is necessary and dependent on the climatic sensitivity of population dynamics.We are grateful to Hans de Kroon for discussions on the application of matrices to ecology.     

Dye coupling in the organ of Corti

Summary Dye-coupling in an in vitro preparation of the supporting cells of the guinea-pig organ of Corti was evaluated by use of the fluorescent dyes, Lucifer Yellow, fluorescein and 6 carboxyfluorescein. Despite the presence of good electrical coupling in Hensen cells (coupling ratios >0.6) the spread of Lucifer yellow was inconsistent. Hensen cells are very susceptible to photoinactivation, i.e., cell injury upon illumination of intracellular dye; and this in conjunction with Lucifer Yellow's charge and K+-induced precipitability may account for its variability of spread. Fluorescein and 6 carboxyfluorescein, on the other hand, spread more readily and to a greater extent than Lucifer Yellow, often spreading to cell types other than those of Hensen. Dye spread is rapid, occurring within a few minutes. These results suggest that molecules of metabolic importance also may be shared by the supporting cells of the organ of Corti.