Integument and sensory nerve differentiation ofDrosophila leg and wing imaginal discs in vitro

Summary An ultrastructural analysis is presented of the cuticular and neural structures formed by the prothoracic leg and wing imaginal discs of maleDrosophila melanogaster larvae during culture in vitro with 0.2 g/ml of -ecdysone. A pupal cuticle, and subsequently an imaginal cuticle with a well-defined epicuticle and a laminated endocuticle is formed. The ultrastructure of the epidermis and of cuticular structures such as bristles, trichomes, apodemes, and tracheoles is very similar to that found in situ. Dendrites and nerve cell bodies are formed in vitro, and sensory axons form nerve bundles similar to those of normal appendages in situ, despite their isolation from the central nervous system. It is concluded that at the ultrastructural level, differentiation in vitro closely parallels the normal course of development.     

Reconstitution of model membranes from phospholipid and outer membrane proteins of Proteus mirabilis. Role of proteins in the formation of hydrophilic pores and protection of membranes against detergents.

Outer membrane proteins extracted from isolated cell walls of Proteus mirabilis were able to combine the cell wall phospholipids in a model membrane system. The presence of outer membrane proteins in vesicular model membranes mediated the release of previously entrapped [14C]sucrose while [3H]inulin was retained. Incorporation of lipopolysaccharide from the same cell walls was not required for the formation of such selectively permeable membranes. Three major outer membrane proteins of apparent molecular weights 39000, 36000 and 17000 were isolated using acetic acid and sodium deoxycholate solution as solvents and avoiding the strongly denaturing sodium dodecyl sulfate. The isolated proteins were assayed for their ability to form hydrophilic pores in reconstituted membranes. The trypsin-sensitive 39000-Mr protein and the peptidoglycan-associated 36000-Mr protein were equally effective in this function whereas the 17000-Mr protein mediated little penetration of low molecular weight solute. The 39000-Mr and 36000-Mr proteins also protected reconstituted membrane vesicles from disruption by detergent while 17000-Mr protein was ineffective in this regard.     

The genetical analysis of covariance structure.

The analysis of covariance structures (J?reskog, 1973) is adapted to the simultaneous maximum likelihood estimation of genetical and environmental factor loadings and specific variances. The goodness of fit is tested by chi square and standard errors of parameter estimates can be obtained. Any linear model used in univariate genetical analyses can be extended to the multivariate case. Most biological hypotheses about the relationships between variables can be specified by a variety of factor models. Individual parameters can be given fixed values or set to zero and hypotheses concerning the congruence of genetical and environmental correlations can be tested. The method is illustrated with published twin data on cognitive abilities.     

Cytochemical and electrophoretic studies of haemoglobin synthesis in the fat body of a midge, Chironomus thummi

The fat body of developing mid- and late fourth instar larvae of a midge, Chironomus thummi, has been investigated by means of the benzidine reaction for the localization of haemoglobin within cells. In the subepidermal fat body the reaction deposits of the haemoglobin pseudo-peroxidase activity appear predominantly in the intracisternal cavities of ER and the Golgi, and later, in the pharate pupal stage, in small dense granules (0.5–1 μm in. diameter).All the major protein bands of fat body extracts, which are resolved in electrophoresis, give the benzidine reaction and show incorporation of ¹⁴C-amino levulinic acids, in this case a specific marker for haemoglobin synthesis. In addition, labelled proteins show identical electrophoretic mobility as the haemoglobins of the haemolymph, suggesting that haemoglobins are synthesized in the fat body. Two types of fat body cells seem to differ with respect to their rôle in haemoglobin metabolism.     

Molecular dynamics of the local anesthetic tetracaine in phospholipid vesicles

Upon introduction into phosphatidylcholine vesicles, the 13C magnetic resonance peaks of the aromatic resonances of tetracaine are broadened while the T1 relaxation times show little change. Addition of tetracaine to vesicles containing 30% cholesterol produces a similar broadening in the 13C NMR spectrum of tetracaine. Nuclear magnetic resonance parameters of phosphatidylcholine in vesicles which are unchanged by the addition of equimolar tetracaine include 13C T1 relaxation time and 31P linewidth, T1 relaxation time, and nuclear Overhauser effect enhancement. These results are interpreted as indicating a hydrophobic interaction between hydrocarbon portions of the anesthetic and phospholipid bilayer. The rotational correlation time of tetracaine about its long axis in the vesicles has been calculated from the 13C NMR spin lattice relaxation times to be about 10(-10.3) s and is unchanged by incorporation into the phospholipid bilayer. The positively charged ammonium group of tetracaine interacts with the negatively charged phosphate group of the vesicle lipids. Using shift reagents and 31P NMR, tetracaine has been shown to displace cations from the bilayer surface, and does not undergo fast flip-flop across the vesicle bilayer.     

A fine-structural study of embryonic and larval development in the gymnoblastic hydroid Pennaria tiarella.

1. The pregastrulation blastomers contain electron-dense granules which become localized after gastrulation in the apices of the developing epithelio-muscle cells and persist throughout larval development. The cytoplasm of the blastomeres is organized into anucleate, membrane-delimited lobules. The lobules, which persist until six hours of development, come to contain a single, peripherally located cisterna of granular endoplasmic reticulum. Microvilli are present at the earliest stages examined and persist throughout development. Cilia are first detected at four hours. 2. Gastrulation, marked by the appearance of the mesoglea, occurs between six and eight hours of development. Basal foot processes of epithelio-muscle cells are detected by eight hours, but myonemes cannot be detected until later in development. 3. Immediately following gastrulation, mucous cells begin their differentiation from dividing cells located near the apex of the ectoderm. During their differentiation, the cells elongate toward the mesoglea. 4. By 16 hours post-fertilization, a third cell type can be detected in the ectoderm. The cell, which contains no granules, has an unusual cytoplasmic organization in which fused membranes divide the cytoplasm into parallel compartments containing a single cisterna of granular endoplasmic reticulum. 5. The findings of the present study are correlated with those of previous studies of development in Pennaria and other hydroids. The possible functional roles of the Type I granules, the cytoplasmic lobules, and the nongranular cell are discussed.     

Biological and biochemical studies of cells transformed by simian virus 40 temperature-sensitive gene A mutants and A mutant revertants.

The growth properties of hamster cells transformed by wild-type Simian virus 40 (SV40), by early SV40 temperature-sensitive mutants of the A complementation group, and by spontaneous revertants of these mutants were studied. All of the tsA mutant-transformed cells were temperature sensitive in their ability to form clones in soft agar and on monolayers of normal cells except for CHLA-30L1, which was not temperature sensitive in the latter property. All cells transformed by stable revertants of well-characterized tsA mutants possessed certain growth properties in common with wild-type-transformed cells at both temperatures. Virus rescued from tsA transformants including CHLA30L1 was temperature sensitive for viral DNA replication, whereas that rescued from revertant and wild-type transformants was not thermolabile in this regard. T antigen present in crude extracts of tsA-transformed cells including CHLA30L1, grown at 33 degreeC, was temperature sensitive by in vitro immunoassay, whereas that from wild-type-transformed cells was relatively stable. T antigen from revertant transformants was more stable than the tsA protein. Partially purified T antigen from revertant-transformed cells was nearly as stable as wild-type antigen in its ability to bind DNA after heating at 44 degrees C, whereas T antigen from tsA30 mutant-transformed cells was relatively thermolabile. These results further indicate that T antigen is a product of the SV40 A gene. Significantly more T antigen was found in extracts of CHLA30L1 grown to high density at the nonpermissive temperature than in any other tsA-transformed cell similarly grown. This is consistent with the suggestion that the amount of T antigen synthesized in CHLA30L1 is large enoughto allow partial expression of the transformed phenotype at the restrictive temperature. Alternatively, the increase in T antigen concentration may be secondary to one or more genetic alterations that independently affect the transformed phenotype of these cells.     

Interactions of thyrotropin releasing hormone and morphine sulfate in rodents.

Thyrotopin releasing hormone (TRH) produces “wet dog shakes” in rats similar to those observed during morphine withdrawal. The shaking behavior precipitated by morphine abstinence can be exacerbated by TRH administration while the other components of the morphine withdrawal syndrome remain unchanged. Morphine, chlorpromazine, apomorphine, and Δ⁹-tetrahydrocannabinol effectively block shakes induced by either TRH administration or morphine withdrawal. These results suggest the possibility that endogenous TRH may be associated with the “wet dog shakes” observed as a portion of morphine's abstinence syndrome in rats. However, TRH is unable to alter the stereospecific binding of morphine $\text{in}$$\text{vivo}$ or $\text{in}$$\text{vitro}$, and naloxone fails to potentiate the number of TRH-induced shakes. TRH has no antinociceptive properties, and it cannot alter those of morphine. These data suggest that more than one neuromechanism may be responsible for shaking behavior in rats.