Human Y-chromatin : II. DNA replication

The DNA synthesis of the human Y-chromatin in its various morphological configurations was studied by labeling with tritiated thymidine (³H-TdR) and autoradiography. Continuous terminal and pulse labeling studies revealed that while condensed, the Y-body lagged in DNA synthesis behind the rest of the nucleus. The highest rate of incorporation of ³H-TdR by the Y-body occurred when it was dispersed. These observations are consistent with the replication characteristics of the Y chromosome as determined by conventional late labeling studies and strongly suggest that the Y-body uncoils at the time it replicates its DNA.     

Thermal phase transitions in deuterated lecithin bilayers.

Differential Thermal Analyses and Optical Density measurements show that perdeuteration of the fatty acid residues in phosphatidylcholines causes a 4–5°C decrease in the phase transition temperatures of bilayer dispersions prepared from these deuterated phospholipids. The implications of these findings on the use of deuterated phospholipids in membrane research will be briefly discussed.     

Melanogenesis in cultures of peripheral nervous tissue. II. Environmental factors determining the fate of pigment-forming cells

Spinal ganglia from 4- to 7-day [Stage 23–30; Hamburger and Hamilton (1951) J. Morphol.88, 49–92] chicken embryos were cultured in vitro to investigate the effect of various environmental conditions on cell differentiation. Culture morphology (i.e., degree of dispersion of the explanted ganglia, survival of neurons, and outgrowth of axons) was observed to depend upon several factors including: (1) the age of the explanted ganglia, (2) the presence or absence of nerve growth factor (NGF), and (3) the nature of the substratum on which the cultured tissue resides. These observations enabled us to disturb the association of neurons with the other cells in ganglion cultures and thereby modulate the differentiation of adventitious melanocytes. Thus, in medium permissive for melanogenesis, melanocytes appear when the association between neurons and small stellate nonneuronal cells in the ganglion is disrupted. This disruption is most extensive (1) when young (Stage 26–27, 5-day) ganglia are explanted on plastic substrata, in the initial absence of NGF, and (2) when cells from enzyme-dissociated ganglia are cultured on plastic substrata. In comparable media, pigment cell differentiation is not observed when the association between neurons and small stellate cells is preserved. Such associations tend to endure (1) in developmentally older (Stage 30+, 7- to 8-day) ganglia or (2) when ganglia are cultured on agar or fibroblast substrata. We conclude that loss of association between neurons and the nonneuronal cells in young ganglia is necessary for the latter to undergo melanogenesis in vitro.     

Analogs of natural lipids. II. Polymorphic behavior of the tris-Homo acyl derivatives of cyclopentane-1,2,3-triols

The polymorphic behavior of the three series of tris-homoacyl (C_14:0, C_16:0 and C_18:0) cyclopentane-1,2,3-triol analogs of the natural saturated triglycerides has been studied using differential thermal analysis, Fourier transform infrared spectroscopy, and X-ray diffraction. It was found that the triglyceride analogs derived from the 1,2,3/0 and 1,2/3 cyclopentanetriols exhibit different polymorphic behavior than that of the natural triglycerides. The analogs derived from 1,3/2 cyclopentanetriol, however, were found to parallel the polymorphic behavior of the natural triglycerides quite closely. This polymorphic behavior is discussed in terms of the different configurations which the chains assume in each of the triglyceride analogs.     

Corpus allatum regulation during the metamorphosis of Periplaneta americana: Axon pathways

The anatomy of the retrocerebral complex was studied after supravital staining with methylene blue, and axonal tracts within the corpora allata (CA) were traced after applying the CoCl₂ technique together with Timm's sulfide-silver enhancement. Cobalt chloride fills of the nerves to and from the CA revealed two major sources of innervation: the brain and the subesophageal ganglion. Three cell clusters in the brain contribute axons that reach each nervus corporis allati I (NCA I) and, apparently, pass to or beyond the CA. These are: a cluster of 8 to 12 cells in the contralateral pars lateralis, a cluster of 16 to 20 cells in the ipsilateral pars lateralis, and a cluster of 50 to 60 cells in the contralateral pars intercerebralis. PAF-stained sections of other brains revealed a corresponding number of PAF-positive cells in these same regions. The medial and lateral neurons arborize in the neuropile adjacent to the pars intercerebralis, and may associate there. The lateral group also arborizes extensively in the neuropile surrounding the pedunculus of the mushroom body. At least four cell bodies located antero-ventrad in the subesophageal ganglion send axons to the CA via each nervus corporis allati II (NCA II).To determine possible inhibitory pathways to the CA, the NCA I, NCA II, and postallatal nerves of last instar larvae were severed; either singly, or in combination. Additional experiments were performed on last instar larvae to substantiate that superlarvae were a direct result of an enhanced or sustained juvenile hormone titre. These experiments included: implanting two or more CA, extirpating one CA, or applying 100 μg of Altosid topically onto allatectomized larvae. The experiments indicated that only NCA I is an inhibitory pathway and that superlarvae were a direct consequence of CA activation. NCA II does not seem to provide the CA with an essential excitatory innervation; when it and NCA I are severed a supernumerary apolysis will still result. Some of the cells in the brain stainable by the CoCl₂ method are most probably identical to those that are PAF-positive. These cells may inhibit the CA in last instar larvae via neurosecretomotor junctions.     

Melanogenesis in cultures of peripheral nervous tissue: I. The origin and prospective fate of cells giving rise to melanocytes

Chick embryo spinal ganglia, peripheral nerves, and connective tissue usually associated with ganglia were cultured separately using several combinations of media and substrata. Melanocytes appear in cultures of both ganglia and peripheral nerves. The only cell type common to both the ganglion and peripheral nerve that could account for the observed pigment cells was the population of small cells with intensely staining nuclei that normally associates closely with nerve cell bodies and fibers. These cells could be distinguished morphologically from fibroblastic cells, which originated in the connective tissue capsule and did not undergo melanogenesis. We conclude that these small cells are supportive (Schwann, satellite, and perineurial) cell precursors and are one source of melanocytes in cultured peripheral nervous tissue.     

Alteration of liver plasma membrane protein conformation by bovine growth hormone in vitro

The response of liver plasma membranes prepared from hypophysectomized, bovine growth hormone-treated hypophysectomized, and normal rats to addition of bovine growth hormone in vitro were studied by circular dichroism and spectrofluorometry. Membranes of hypophysectomized but not normal or treated rats showed increased negative ellipticity in the presence of bovine growth hormone (10^?9) without any change in trough position; at higher concentrations (10^?7?10^?6, m) there was less negative ellipticity. Membranes of hypophysectomized, normal, and growth hormone-treated rats all showed decreased emission of fluorescence and a small shift of the emission peak from 333 to 338 nm in the presence of bovine growth hormone (10^?17?10^?7, m). The excitation of fluorescence was quenched by bovine growth hormone. Polarization of excitation of fluorescence was unchanged.     

Correlation between DNA synthesis and intracellular NAD in cultured human leukemic lymphocytes.

The NAD⁺ level in lymphocytes obtained from an individual with acute monocytic leukemia increased five-fold and then remained constant when the cells were adapted to growth in suspension culture. When the NAD⁺ level of established cells was lowered by means of a nicotinamide-poor medium or by the action of 1-methyl-1-nitrosourea, there was a concomitant decrease in the rate of DNA synthesis. These results indicate that there is a direct correlation between intracellular NAD⁺ and the synthesis of DNA in cultured leukemic lymphocytes. However, the exact nature of the relationship remains speculative.     

Ca2+-stimulated ATPase during the early development of parthenogenetically activated eggs of the sea urchin Paracentrotus lividus

A Ca²⁺-stimulated ATPase shows fluctuations in the activity in parthenogenetically activated sea urchin eggs very similar to those described earlier for fertilized eggs. Besides activity peaks in the first part of a cell cycle the enzyme activity increases when the mitotic apparatus (MA) or MA-like structures like monasters or cytasters are formed. A possible function of the enzyme in the assembly of the MA is discussed.     

Programmed autophagocytosis accompanying conjugation in the ciliate Stylonychia mytilus

Conjugation and postconjugant development in Stylonychia is accompanied by a period of approximately 80 hr during which the cells are unable to ingest food. This period is one of considerable synthetic activity, encompassing important portions of the development of the new macronucleus. Light- and electron-microscopic observations of conjugating pairs and exconjugant cells reveal a process of autophagocytosis that may provide the supplies of energy (and precursors) necessary for postconjugant developmental events. Small autophagosomes (AP) form in conjugants; degenerating mitochondria are prominent among the inclusions observed in these bodies. Soon after separation of pairs, large dense AP appear, apparently by coalescence and condensation of the smaller AP. These “mature” AP give only a slight acid phosphatase reaction. The number of AP slowly declines during postconjugant development; about 60 hr after separation all the AP have been digested. Several observations suggest that this autophagocytosis is part of the developmental program initiated soon after mating begins: (1) The first indications of AP formation occur while conjugating pairs are still able to feed, and thus cannot be attributed to the stress of starvation; (2) formation of large numbers of AP is rather abrupt, whereas their dissolution is very gradual, covering most of the nonfeeding period; and (3) the pattern of AP formation and dissolution is similar in cells whose new macronuclear development has been prevented by brief hydroxyurea treatment.     

Imipramine associations with plasma components and its uptake by cultured human cells

It has been proposed that in vivo variability in response to certain hydrophobic chemicals or drugs, such as imipramine, may be due in part to the varying plasma lipid levels in patients. The distribution of [3H]imipramine into the lipoproteins of human plasma was therefore studied. Differential density centrifugation of plasma containing [3H]imipramine resulted in flotation of very low density, low density and high density lipoproteins (VLDL, LDL, HDL) and approximately one-third of the total 3H radioactivity. Twelve percent of the radioactivity was present in the sedimented fraction which included most of the plasma proteins. There appeared to be little specific binding of [3H]imipramine to VLDL or LDL, as shown by ultracentrifugation, dialysis and column chromatography. [3H]Imipramine was readily incorporated into cultured human fibroblasts;o no differences were observed in cellular uptake whether it was added to the medium in plasma, LDL or HDL. Also, no differences in uptake of [3H]imipramine by LDL-receptor positive and receptor negative cells were noted. These experiments indicate that LDL is not a major vehicle for the transport of this drug and that both the bound and free fractions are available for cellular uptake.     

Differential regulation of the N-methyl transferases responsible for phosphatidylcholine synthesis in Saccharomyces cerevisiae

The enzymes catalyzing the conversion of phosphatidylethanolamine to phosphatidylcholine were assayed by measuring the incorporation of label from [¹⁴C-CH₃]-S-adenosyl-methionine into the endogenous phospholipids of particulate, cell-free preparations from S. cerevisiae grown in the presence of N-methylethanolamine, N,N-dimethylethanolamine, or choline. The results indicate that each base in the growth medium results in reduced levels of all the N-methyltransferase activity involved in the formation of the phosphatidyl ester of the given base. By following the conversion of exogenous [³²P]-phosphatidyldimethylethanolamine to [³²P]-phosphatidylcholine it has been shown that the activity of the third methyl transfer is 90% lower in particles prepared from choline grown cells than in particles prepared from cells grown without choline. The results suggest that there are at least two enzymes involved in the conversion of phosphatidylethanolamine to phosphatidylcholine and that their levels can be regulated individually.Supplementing the growth medium with any of the three methylated aminoethanols results in markedly increased cellular levels of their corresponding phosphatidyl esters and decreased levels of the precursor phosphatidyl esters. The fatty acid composition of phosphatidylcholine also changes when the medium is supplemented with choline suggesting that the proportions of the molecular species of this phosphatide depends on whether synthesis is via methylation of phosphatidylethanolamino or from the supplemented aminoethanol.     

Isolation and analysis of neutral glucans from Ecklonia radiata and Cystophora scalaris

Neutral glucans were isolated from the stipes and fronds of Eklonia radiata and Cystophora scalaris. Partial acid hydrolysis revealed the presence of gentiobiose and laminara-oligosaccharides. Methylation analysis, periodate oxidation, and enzyme studies indicated that the glucans contain β-(1→3) and β-(1→6) linkages. Methylation studies showed that branching in these glucans occurs via a 1,3,6-tri-O- substituted residue with a frequency of one branch point per seven glycosyl residues. In contrast to laminaran from Laminaria digitata, the intrachain (1→3)- and (1→6)- glucopyranoside occur in a molar ratio of 1:1. Enzymic hydrolysis confirmed the absence of long segments of (1→3)-linked residues in the glucans.     

Comparative evaluation of quantum theory of nerve excitation

Deliberate evaluation of the quantum theory of nerve excitation is made by comparing it with Hill's theory in fitting the experimental data on threshold-frequency relation, optimum frequency (v₀) for nerve excitation and strength-duration relation. Decrease of v₀ and increase of all the time constants (Hill's λ andk, Wei'sT ₂ and spike durationw) with decreasing temperature are interpreted on the basis of the dipole relaxation timeT ₂ but inexplicable from Hill's theory or any other existing theory. The closeness ofk,T ₂ andw values is explained. A variety of experimental results obtained by others is discussed. Finally, a comparison is made between the Hodgkin-Huxley equations and the quantum theory. Most of the facts (electrical and non-electrical) tend to support the thesis that nerve excitation is a macroscopic expression of quantum transitions of dipoles between energy states.     

T-cell neoplasm induced by subcutaneous transplantation of a plasmacytoma: characterization of tumor cells.

Thymocytes, isolated 6 days following subcutaneous (sc) transplantation of BALB/c MOPC-315 plasmacytoma into F₁ (BALB/c × C57BL) hybrid mice, when injected sc into normal syngeneic mice, caused the development of a solid sc tumor. The cells of the newly developed tumor were of a mixed population of θ⁺F₁ (BALB/c × C57BL) and θ^? BALB/c cells (approximately 1:1), which represents a new type of mixed T cell-plasma cell neoplasm. Efforts were made to isolate the transformed thymocytes (θ⁺) from the plasmacytoma (θ^?) cells in the new tumor, exploiting differences in their surface properties. Treatment of the mixed tumor cell population with peanut agglutinin (PNA) revealed that only the T tumor cells were agglutinated. The agglutinated cells were recovered after dispersing the clumps with d-galactose (0.15 M) and consisted of 95% θ⁺ cells. The PNA-agglutinated cells were found to induce a similar tumor (85% θ⁺ cells) when injected sc into F₁ (BALB/c × C57BL) mice.