Total Cellular Volume Research Articles

Stereological study of the sexual dimorphism in mouse submandibular glands.

The sexual dimorphism of the mouse submandibular glands was studied by stereological methods. Albino mice of both sexes aged 140 days were used. Their average weight was 34.7 g por males and 26.0 g for females. The following morphometric dimensions were evaluated for the acini, intercalated ducts, convoluted granular tubules, striated ducts, excretory ducts and stroma: volume density, total volume, surface density, total external surface, surface-to-volume ratio, nuclear and cellular volume, and absolute number of cells. We also determined the mean diameters and estimated the total length of striated ducts and granular tubules in the gland. Analysis of the results showed that sexual dimorphism is present in all the morphological compartments of the mouse submandibular glands.

From Cell Growth to Leaf Growth: I. Coupling Cell Division and Cell Expansion

AbstractAn area of plant growth modeling that appears to have been neglected is incorporation of current knowledge regarding cell expansion and cell division into the growth models. The objective of this research was to incorporate this knowledge into a quantitative description of plant growth. A general cell growth module is described that combines known aspects of cell expansion and cell division. Cell expansion is governed by water flux equations. The cells start at a fixed minimum volume and expand to a final (fixed) volume. The wall extensibility of a cell is considered to be a function of its volume. Cell division is described through use of a daughter ratio vs. time function that categorizes the cells resulting from a division as either proliferative or nonproliferative. Temperature is coupled to the water flow equations through its influence on cell cycle time. The daughter ratio vs. time function is of paramount importance. The influence of a number of daughter ratio functions on the rate of cell production and the rate of total cell volume accumulation are considered. Exponential, linear, and sigmoidal increases in both cell numbers and total cellular volume are shown to result from the various daughter ratios examined. A comparison was made of simulations using a constant wall extensibility to those made using the wall extensibility vs. volume function. The difference between these two cases was much less than the differences obtained by varying the daughter ratio function.

Effect of benzoic acid on metabolic fluxes in yeasts: a continuous-culture study on the regulation of respiration and alcoholic fermentation.

Addition of benzoate to the medium reservoir of glucose-limited chemostat cultures of Saccharomyces cerevisiae CBS 8066 growing at a dilution rate (D) of 0.10 h-1 resulted in a decrease in the biomass yield, and an increase in the specific oxygen uptake rate (qO2) from 2.5 to as high as 19.5 mmol g-1 h-1. Above a critical concentration, the presence of benzoate led to alcoholic fermentation and a reduction in qO2 to 13 mmol g-1 h-1. The stimulatory effect of benzoate on respiration was dependent on the dilution rate: at high dilution rates respiration was not enhanced by benzoate. Cells could only gradually adapt to growth in the presence of benzoate: a pulse of benzoate given directly to the culture resulted in wash-out. As the presence of benzoate in cultures growing at low dilution rates resulted in large changes in the catabolic glucose flux, it was of interest to study the effect of benzoate on the residual glucose concentration in the fermenter as well as on the level of some selected enzymes. At D = 0.10 h-1, the residual glucose concentration increased proportionally with increasing benzoate concentration. This suggests that modulation of the glucose flux mainly occurs via a change in the extracellular glucose concentration rather than by synthesis of an additional amount of carriers. Also various intracellular enzyme levels were not positively correlated with the rate of respiration. A notable exception was citrate synthase: its level increased with increasing respiration rate. Growth of S. cerevisiae in ethanol-limited cultures in the presence of benzoate also led to very high qO2 levels of 19-21 mmol g-1 h-1. During growth on glucose as well as on ethanol, the presence of benzoate coincided with an increase in the mitochondrial volume up to one quarter of the total cellular volume. Also with the Crabtree-negative yeasts Candida utilis, Kluyveromyces marxianus and Hansenula polymorpha, growth in the presence of benzoate resulted in an increase in qO2 and, at high concentrations of benzoate, in aerobic fermentation. In contrast to S. cerevisiae, the highest qO2 of these yeasts when growing at D = 0.10 h-1 in the presence of benzoate was equal to, or lower than the qO2 attainable at mu(max) without benzoate. Enzyme activities that were repressed by glucose in S. cerevisiae also declined in K. marxianus when the glucose flux was increased by the presence of benzoate.(ABSTRACT TRUNCATED AT 400 WORDS)

The Sertoli cell of the water buffalo (Bubalus bubalis) during the spermatogenic cycle

Ultrastructural features and morphometric evaluations of buffalo Sertoli cells are reported for the six phases of the spermatogenic cycle. The phases of the tubular seminiferous epithelium are identified according to characteristic cellular associations with completed spermiation as demarcation between two cycles. Average tubular diameter (245 μm) and epithelial height (61 μm) do not vary significantly during the cycle. The relative Sertoli cell volume in the seminiferous epithelium varies between 30% (phase 4) and 39% (phase 8). The calculated volume of a single Sertoli cell increases from a nadir of 7118 μm3 in phase 3 abruptly to a maximum of 8968 μm3 in phase 4 and is then gradually reduced during the following phases. The Sertoli cell surface area shows a similar trend: it amounts to 11105 μm2 in phase 3 and to 14260 μm2 in phase 4. The contact area of the Sertoli cell with adjacent cells and structures is subject to characteristic changes; from the expansion of basal Sertoli-Sertoli contacts it is concluded that the blood-testis barrier in the buffalo is particularly tight during phases 8, 1 and 2. The irregularly contoured nucleus contains a vesicular nucleolus, has a calculated volume from 465 μm3 to 543 μm3 and occupies ∼5 to 7% of the cell. Volume percentages of mitochondria (4%), Golgi apparatus and lysosomal bodies are rather constant during the cycle. Whorls and orderly arranged aggregates of the smooth endoplasmic reticulum occur in basal location as well as in close association with elongating spermatids. Smooth ER is the organelle that exhibits the most prominent changes during the Sertoli cell cycle: it occupies 5.79% in phase 3 and 20.9% in phase 4 of the total cellular volume. Phagocytosis of residual bodies is insignificant in this species and a lipid cycle is absent in buffalo Sertoli cells.

Novel origins of lineage founder cells in the direct-developing sea urchin Heliocidaris erythrogramma

The lineage and fate of each blastomere in the 32-cell embryo of the direct-developing sea urchin Heliocidaris erythrogramma have been traced by microinjection of tetramethylrhodamine-dextran. The results reveal substantive evolutionary modifications of the ancestral cell lineage pattern of indirect sea urchin development. Significant among these modifications are changes in the time and order of cell lineage segregation: vegetal ectodermal founder cells consistently arise earlier than during indirect development, while internal founder cells generally segregate later and in a different sequence. Modifications have also arisen in proportions of the embryo fated to become various cell types and larval structures. Ectodermal fates, particularly vestibular ectoderm, comprise a greater proportion of the total cellular volume in H. erythrogramma. Among internal cell types, coelom consumes more and endoderm less of the remaining cellular volume than during indirect sea urchin development. Evolutionary modifications are also apparent in the positional origin of larval cell types and structures in H. erythrogramma. These include an apparent tilt in the axis of prospective cell fate relative to the animal-vegetal axis as defined by cleavage planes. Together these evolutionary changes in the cell lineage of H. erythrogramma produce an accelerated loss of dorsoventral symmetry in cell fate relative to indirect development. The extent and diversity of rearrangements in its cell lineage indicate that the nonfeeding larva of H. erythrogramma is a highly modified, novel form rather than a degenerate pluteus larva. These same modifications underscore the evolutionarily flexible relationship between cell lineage, gene expression, and larval morphology in sea urchin development.

High resolution 23Na-nuclear magnetic resonance study of stroke-prone spontaneously hypertensive rat erythrocytes.

The intracellular Na+ content of washed erythrocytes from stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar-Kyoto normotensive rats (WKY) was measured by a high resolution 23Na-nuclear magnetic resonance (NMR) technique using a non-permeant aqueous shift reagent, dysprosium triethylenetetramine hexaacetic acid, Dy(TTHA)3-. The initial intracellular Na+ of freshly isolated and washed erythrocytes was very low (approximately 5 mmol/l) and increased progressively with prolonged incubation in isotonic salt solution at 37 degrees C. There was no significant difference in the erythrocyte Na+ concentration between SHRSP and WKY over the entire period of measurement, nor was any difference detected in their osmotic fragility or total cellular volume, although the osmotic fragility decreased with incubation time. The high energy phosphate metabolites were also studied in the same erythrocytes by 31P-NMR. The level of intracellular ATP decreased with incubation at 37 degrees C but showed no difference between the SHRSP and WKY samples. Inclusion of 1 mmol/l ouabain in the incubation medium substantially retarded the breakdown of intracellular ATP and resulted in a concomitant increase in intracellular Na+. However, neither the ouabain-sensitive nor the ouabain-insensitive component of Na+ influx altered in SHRSP erythrocytes compared with WKY erythrocytes in paired experiments. Our results do not support the hypothesis that altered Na+ transport, resulting in an increase in erythrocyte Na+ concentration, is associated with spontaneous hypertension.

Functional changes in luteinizing hormone-secreting cells from pre- and postpartum ewes.

Luteinizing hormone (LH)-containing cells from ovine pituitaries obtained during gestation and at various times after parturition were examined to determine whether the ability to store and secrete LH in vitro was correlated with morphological changes. Pituitaries collected on days 50 and 140 of gestation and on days 2, 13, 22, and 35 after parturition were enzymatically dissociated and the resulting cells cultured in media containing estradiol (12 pg/ml), cortisol (12 ng/ml), or no steroid. After 4, 7, or 10 days of culture, cells were washed and basal LH, gonadotropin-releasing hormone (GnRH)-stimulated release of LH, and cellular content of LH were determined. The content of LH (ng/10(6) cells) was lowest on day 140 of gestation (2.7 +/- 0.3) and day 2 postpartum (2.2 +/- 0.6) and then increased (P less than 0.05) on days 13 (36.6 +/- 8.3), 22 (59.9 +/- 14.4), and 35 (54.6 +/- 19.3) postpartum. The percentage of pituitary cells containing immunoreactive LH nearly doubled (P less than 0.05) between days 2 (5.6 +/- 0.2%) and 35 (10.6 +/- 1.1%) postpartum. Moreover, LH-containing cells were smaller, and the percent total cellular volume occupied by secretory granules was less on day 2 than on days 22 and 35 after parturition. Secretion of LH after 4, 7, or 10 days of culture reflected the cellular content of LH and was not influenced by the presence of steroids in the media. These data indicate that decreased synthesis of LH during gestation is associated with hypoplasia of the LH-secreting cells. These cells are reactivated during the postpartum period and their capacity to synthesize LH gradually returns to normal.

The passive electrical properties of guinea-pig ventricular muscle as examined with a voltage-clamp technique.

1. A voltage-clamp technique was developed for stable recording of small currents in guinea-pig ventricular muscle. Small cylindrical preparations were impaled with three micro-electrodes, one for measuring the feed-back potential and two for injecting current. 2. The longitudinal potential profile resulting from current injection at one point was measured. It agreed well with the theoretical predictions for a linear cable which is sealed at both ends ('healing over'), with a length constant (lambda) of 580 +/- 145 micron. 3. When the clamp current was injected symmetrically into each half of the preparation via two electronic current pumps a spatially homogeneous clamp could be achieved in preparations with a diameter of less than or equal to 250 micron and a length of less than or equal to 2 lambda. 4. The membrane capacity and the membrane resistance of the preparations at the resting potential were measured with small voltage-clamp pulses. Assuming a specific membrane capacity (Cm) of 1 microF/cm2 a specific membrane resistance (Rm) of 6.7 +/- 1.8 k omega cm2 was obtained in Tyrode solution containing 3 mM-K. 5. The total surface area was calculated from the measured capacity of the preparation assuming a Cm of 1 microF/cm2. The total cellular volume was estimated from optical measurement of the external dimensions of the preparation assuming an extracellular space of 25%. From these data the average surface/volume ratio of individual cells was calculated to be 7200 cm2/cm3. 6. From the measured electrical constants the specific resistance of the intracellular space (Ri) was calculated to be 200-250 omega cm. With small constant current pulses a membrane time constant of 6.6 +/- 1.3 ms was measured. 7. The influence of the extracellular potassium concentration ([K]o) on Rm was studied in the range 1.5-6 mM-[K]o. Rm was found to depend on [K]o less than predicted by the constant field theory.

Pineal cells enhance choline acetyltransferase activity in sympathetic neurons.

Cells derived from the neonatal rat pineal gland were cocultured with cells derived from neonatal rat superior cervical ganglia (SCG) in an attempt to determine whether a sympathetic target organ with only adrenergic properties could enhance the development of adrenergic transmitter properties in sympathetic neurons in tissue culture. Choline acetyltransferase was measured as an index of cholinergic differentiation, and tyrosine hydroxylase was measured as an index of adrenergic differentiation. As indices of total cell number and cellular volume, DNA and protein, respectively, were also measured. We found that the pineal-SCG cocultures contained ten times greater choline acetyltransferase activity than sister neuronal cultures cultured without pineal cells, thus indicating that the pineal cells enhanced cholinergic properties in the sympathetic neurons. This cholinergic enhancement was dependent upon the presence of nerve growth factor and could not be obtained with pineal-conditioned medium. Tyrosine hydroxylase activity, measured on cultures sister to those mentioned above, was low in all cultures and decreased somewhat in SCGs cultured alone. TH activity in the pineal-SCG cocultures, however, increased slightly. Some tyrosine hydroxylating activity developed in pineals cultured alone, however, and may have been responsible for the small increase in tyrosine hydroxylase activity noted in the pineal-SCG cocultures. The implications of these results for a determination of the role that target organ plays in the development of the transmitter properties of sympathetic neurons are discussed.

Nuclear gene dosage effects on mitochondrial mass and DNA.

In order to assess the effect of nuclear gene dosage on the regulation of mitochondria we have studied serial sections of a set of isogenic haploid and diploid cells of Saccharomyces cerevisiae, growing exponentially in the absence of catabolite repression, and determined the amount of mitochondrial DNA per cell. Mitochondria accounted for 14% of the cytoplasmic and 12% of the total cellular volume in all cells examined regardless of their ploidy or their apparent stage in the cell cycle. The mean number of mitochondria per cell was 22 in the diploid and 10 in the haploids. The volume distribution appeared unimodal and identical in haploids and diploids. The mitochondrial DNA accounted for 12.6 +/- 1.2% and 13.5 +/- 1.3% of the total cellular DNA in the diploid and haploid populations, respectively. These values correspond to 3.6 x 10(-15) g, 2.2 x 10(9) daltons, or 44 genomes (50 x 10(6) daltons each) per haploid and twice that per diploid cell. On this basis, the average mitochondrion in these cells contains four mitochondrial genomes in both the haploid and the diploid.

Na + pool and Na + concentration in epidermis of frog skin

I. Estimations of intracellular [Na +] c in epithelial cells of non-short-circuited skins of Rana pipiens in sulfate Ringer's were made at varying [Na +] 0 by two indirect methods. (a) Intracellular Na + content (μequiv/cm 2) was determined by the “Na +-pool method”. From this and estimations of total cellular and extracellular volume an average value for [Na +] e was calculated. (b) [Na +] e was determined by using a modified Nernst equation, assuming Na + permselectivity for the “outer border” of skin. Both methods yielded identical values for [Na +] e which varied in the same manner with [Na +] o . 2. From variations of [Na +] 8 [Na +] in the subcorneal space) and [Na +] e with changing [Na +] o , E 00 (potential difference across outer border) was calculated and the results compared to the experimentally found change of 35 mV for a io-fold change in [Na +] 0 . The calculation s based on [Na +] s and [Na +] e gave the correct slope, but the absolute potential differences (PD) were too low when compared to known measurements of the intracellular PD values. 3. From this discrepancy it is suggested that [Na +] e in the cells of the “first reacting cell layer” is lower, and [Na +] e in the remaining cells is higher than [Na +] e . This is illustrated by a sample calculation. Values for [Na +] e , first reacting cell layer, are 3–13 mM for [Na −] 0 = 7−110mM . This supports the hypothesis of passive entry of Na + into the epidermis. By computer simulation it is shown that a model can be constructed that maintains in a steady state such [Na +] gradients and yields flux characteristics in agreement with typical laboratory observations.

The Axon Reaction: A Review of the Principal Features of Perikaryal Responses to Axon Injury

This chapter provides an overview of the principal features of perikaryal responses to axon injury. The neuron is an unusual cell. Its axon terminals may be situated at what in cellular terms is an enormous distance from the cell body (perikaryon); the volume of the latter may be but a small fraction of the total cellular volume. Yet the neuronal processes are maintained and their substance is constantly renewed from the perikaryon. The separation of an axon from its cell body results (in vertebrates) in the degeneration of the separated portion and is followed by a series of morphological changes in the perikarya. The most conspicuous of these is the disintegration, redistribution, and apparent disappearance from the cell body of cytoplasmic basophil material. Changes in axotomized neurons are generally assessed by comparison with the corresponding contralateral neurons of the experimental animal.

Sodium, Potassium, and Chloride Concentrations in the Schwann Cell and Axon of the Squid Nerve Fiber

Sodium, potassium, and chloride concentrations were determined in the sheath cells and axoplasm of the nerve fiber of the squid Sepioteuthis sepioidea. The sheaths were obtained by slitting the nerve fiber, the extracellular electrolytes were washed out in isotonic sucrose solution, and the concentrations in the cells were determined after different soaking times in the sucrose solution. Values for the Schwann cell were calculated by extrapolation to zero time from the plots of the logarithms of the concentrations in the cells as a function of soaking time in sucrose solution. The Schwann cells made up 84 per cent of the sheath's total cellular volume. The Schwann cell concentrations in millimols per liter, are: 312 (404-241) for sodium, 220 (308-157) for potassium, and 167 (208-138) for chloride. The concentrations in the axoplasm (mean +/- SE), in millimols per liter are: 52 +/- 10 for sodium, 335 +/- 25 for potassium, and 135 +/- 14 for chloride. The possibility that some fraction of the Schwann cell electrolytes, especially of sodium, is bound, cannot be discarded.

The relationship between sodium, potassium, and chloride in amphibian muscle.

The Na,(+) Cl(-), and K(+) content of toad plasma and the sartorius muscle has been determined. Although the Na(+) and Cl(-) level of the muscles in the living animal varied greatly (0 to 38.0 m.eq. per kg., and 0 to 31.8 m.eq. per kg. respectively) the K(+) level was subject to a smaller variation (76.5 to 136 m.eq. per kg.). There was a direct relationship between Na(+) and Cl(-), which was independent of the K(+) level. There is a closely related gain of Na(+) and Cl(-) when muscle is soaked in normal Ringer. These gains are not related to the K(+) loss, frequently found on soaking. The relationship between the three ions was studied in a large series of 124 muscles in normal Ringer. As found in vivo, there was a correlation between Na(+) and Cl.(-) This correlation was independent of K(+) content, except when this was abnormally low. Alteration of the external NaCl level produced concomitant changes in the internal levels of these ions. Alteration of the external KCl level produced an increase in internal Cl(-) similar to that found with high NaCl solutions, but the amount of K(+) entering the cell was approximately one-third of the external increase. Removal of K(+) from the external solution did not result in a loss of K(+) from the cell, although there was an adequate amount of Cl(-) present to accompany it. The results cannot be reconciled with either a Donnan concept for the accumulation of K(+), or a linked carrier system. A theory is proposed to account for the ionic differentiation within the cell. The K(+) is assumed to be adsorbed onto an ordered intracellular phase. The normal metabolic functioning of the cell is necessary to maintain the specificity of the adsorption sites. There is another intracellular phase, which lacks the structural specificity for K(+), and which contains Na(+), Cl(-), and K(+) in equilibrium with the external solution. The dimensions of the free intracellular phase will vary from cell to cell, but it will be smaller in the intact animal, and will increase on soaking in normal Ringer, until it is approximately one-third of the total cellular volume. The increase in this phase may be ascribed to a decrease in the energy available to maintain the ordered phase.

Further Observations on the Effect of Cortisone and Thyroxin on the Blood Picture of Hypophysectomized Rats

Abstract Adult female rats of the Wistar strain were hypophysectomized and divided into 3 groups, the first of which received no treatment for 100 days, the other 2 groups receiving no treatment for 50 days followed by daily subcutaneous injections of 0.6 mg. of cortisone alone or in combination with 0.005 mg. of thyroxin for another 50 days. Another group of rats served as normal unoperated controls. Hypophysectomy induced a decrease in the number of nucleated cells in the bone marrow, the erythroid elements being the cells responsible for this decrease; the myeloid elements remained normal in number. These marrow changes were reflected in a decrease in reticulocyte percentage and subsequent anemia, while the total white cell count remained normal. A combined thyroxin-cortisone therapy induced a decrease in myeloid elements in the bone marrow, which resulted in a decrease in the peripheral white cell count. It increased the number of erythroid elements in the bone marrow to a level above that of the hypophysectomized animals with no treatment but not to normal levels. This is undoubtedly due to a very rapid production and release of these cells, for the reticulocyte count was 72 per cent above normal and the anemia in these animals completely eliminated. Blood volume studies showed that these changes in the peripheral blood were not due to a hemoconcentration. Cortisone therapy alone induced changes in the myeloid elements in the bone marrow similar to that following the dual therapy, and produced a similar decrease in peripheral white cell count. This therapy did not, however, eliminate the post-hypophysectomy anemia as did the combined therapy. Although erythropoiesis in the marrows appeared stimulated and the reticulocyte percentage was elevated, the total cellular volume in these animals was not elevated. A combined thyroxin and cortisone therapy will not only prevent post-hypophysectomy anemia, as was found previously, but will eliminate the anemia once it has become established.

The cadmium electrode for storage battery testing

Previously, all biological measurements of intracellular electric fields (E fields), using voltage dyes or patch/voltage clamps, were confined to cellular membranes, which account for <0.1% of the total cellular volume. These membrane-dependent techniques also frequently require lengthy calibration steps for each cell or cell type measured. A new 30-nm “photonic voltmeter”, 1000-fold smaller than existing voltmeters, enables, to our knowledge, the first complete three-dimensional E field profiling throughout the entire volume of living cells. These nanodevices are calibrated externally and then applied for E field determinations inside any live cell or cellular compartment, with no further calibration steps. The results indicate that the E fields from the mitochondrial membranes penetrate much deeper into the cytosol than previously estimated, indicating that, electrically, the cytoplasm cannot be described as a simple homogeneous solution, as often approximated, but should rather be thought of as a complex, heterogeneous hydrogel, with distinct microdomains.

An Address ON MIND AND MOTIVE: SOME NOTES ON CRIMINAL LUNACY: Delivered before the Section of Forensic Medicine of the Seventeenth International Congress of Medicine

<b>1469</b> <b>Objectives: </b>We have previously demonstrated that low concentrations (0.1 mg/ml) of 2-Deoxyglucose (2-DG) have significant therapeutic effects on a variety of human cancer cell lines. In order to understand the mechanisms underlying the effects of 2-DG we evaluated in-vitro exogenous glucose utilization and cellular proliferation during 2-DG treatment. <b>Methods: </b>Two NSCLC cell lines (H3255 and HCC4006) with different degrees of sensitivity to 2-DG treatment were studied. To measure the effects of 2-DG treatment on glucose utilization and cellular proliferation, 1 x 105 cells were plated in media with 2 mg/ml glucose and uptakes of F18-fluorodeoxyglucose (FDG) and F18-fluorothymidine (FLT) were measured before and after treatment with 0.1 mg/ml 2-DG for three days. 2-DG was not present during the FDG/FLT uptake experiments. The intracellular radiotracer concentration was calculated by dividing the total sample FDG or FLT uptake by the total cellular volume, which was determined by measurements of cell diameters and total cell number with a Beckman Coulter Counter. The degree of growth inhibition was measured by counting the number viable, trypan blue negative treated cells, relative to controls. <b>Results: </b>As expected from previous experiments, 2-DG treatment was more effective in HCC4006 than in H3255 cells (reduction in the number of viable cells 50% vs. 35%, respectively). This corresponded to an approximately 2-times higher baseline FDG-uptake in HCC4006 as compared to H3255 cells. However, 2-DG treatment had only a minor impact on FDG-uptake. Treatment with 0.1 mg/ml 2-DG for 3 days reduced FDG uptake by 14% in HCC4006 (p=NS) and by 24% in H3225 (p=NS). In contrast, 2-DG treatment markedly reduced FLT uptake by 49% in HCC4006 (p&lt;0.001) and by 22% in H3255 (p=0.05). <b>Conclusions: </b>These results imply that in cells with a high baseline FDG uptake, 2-DG at a low concentration (1/10th of the normal plasma concentration) that only minimally inhibits glucose metabolism significantly inhibits cellular proliferation. These findings warrant further testing of 2-DG treatment in tumor models with high glycolytic activity.