Differentiation of fiber types in wing muscles during embryonic development: effect of neural tube removal

The embryonic precursors of the avian slow (type I and III) and fast (type II) fibers can be distinguished from each other early in muscle formation (stage 28, V. Hamburger and H. L. Hamilton, J. Morphol, 88, 49-92, 1951) on the basis of the differential sensitivity of their myosin ATPases. To test the neural dependence of fiber type differentiation, the source of motor innervation was eliminated by excision of the brachial neural tube at stages 16-18 before muscles are innervated. Removal of the brachial neural tube did not affect the number of primary myotubes in a sample muscle of the forelimb (ulnimetacarpalis dorsalis, UMD) up until stage 36. Myosin ATPase staining at a variety of pHs revealed the typical patterns of fiber types in muscles of neural-tube free embryos in stages 35-37. These muscles included the anterior latissimus dorsi, brachialis, and UMD which showed presumptive type III staining (type IIIEMB), the pronator superficialis and flexor carpi ulnaris which showed embryonic type II staining (type IIEMB), and the triceps brachii muscles which showed characteristic arrangements of both type IEMB and type IIEMB fibers. The normal patterns of type IEMB and type IIEMB myotubes were also seen in muscles containing a heterogeneous mixture of fiber types such as the biceps brachii, extensor metacarpi radialis, and adductor indicis muscles, although the intensity of acid-stable ATPase staining of the type IEMB myotubes in these muscles was lower than in innervated muscles. It is concluded that the earliest differentiation of muscle fiber types is independent of the nervous system.     

Successful cardiac allografts in syngeneic radiation chimeras

Summary Successful cardiac allografts were accomplished across the major histocompatibility complex of rats. LEW and F344 (Ag-B ²) rats were lethally irradiated and grafted with WF (Ag-B ¹) hearts on day 0. Either on day 0 or day 2, the hosts were repopulated with syngeneic hemopoietic cells. The best results were obtained (86%) when a mixture of 3.0 × 10⁷ non-adherent syngeneic bone marrow and thymus cells were used to repopulate the recipients. In contrast, all of the WF to LEW heart grafts were rejected within 30 days if syngeneic thoracic duct and bone marrow cells were used to repopulate the host.Supported by Grant HL18186 from the National Institutes of HealthRecipient of a Research Career Development Award CA70879 from the National Institutes of Health     

Variation in nuclear DNA in the genus Secale

Estimates of the 4C DNA amount per nucleus in 16 taxa of the genus Secale made by Feulgen microdensitometry ranged from 28.85 picograms (pg) in S. silvestre PBI R52 to 34.58 pg in S. vavilovii UM 2D49, compared with 33.14 pg in S. cereale cv. Petkus Spring which was used as a standard. Giemsa C-banding patterns showed considerable interspecific and intraspecific variation and several instances of polymorphism for large telomeric C-bands. The proportion of telomeric heterochromatin in the genome ranged from about 6% in S. silvestre and S. africanum to about 12% in cultivated rye. A detailed comparison of nine taxa showed no overall relationship between 4C DNA amount and the proportion of telomeric heterochromatin in the genome. However, evidence is presented which strongly supports the notion that the major evolutionary change in chromosome structure in Secale has involved the addition of heterochromatin at, or close to, the telomeres. It is suggested that saltatory amplification events at telomeres were initially responsible for each large increase in DNA amount. Subsequently unequal crossing over between homologues may have played an important secondary role by extending the range of variation in the amount of heterochromatin at a given telomere, while crossing over between non-homologues may have provided a useful mechanism allowing an increase in the DNA amount at one telomere to be distributed between chromosomes.     

Nature of the antigenic determinants of T locus antigens

The nature of the antigenic specificities of several antigens associated with the T/t complex in the mouse were analyzed by means of glycosidase and haptene inhibition studies. Results indicate that on testicular cells sugar residues are involved in at least six different T/t antigenic determinants. The immunodominant sugar appears to be different for each of the specificities. The specificity for the following T/t antigens resides predominantly in the sugars indicated: T:sialic acid; t12:beta-D-galactose; tw32:beta-D-galactose; t0:L-fucose; tw1:N-acetyl-D-galactosamine; tw18:L-fucose. It seems probable that these sugars are found at the terminal reducing ends of the carbohydrate portion of T/t-bearing moleculse. These studies imply that at least some of the genes in the T locus code for glycosyltransferases or regulators of glycosyltransferases which modigy oligosaccharide structures and impart specificity to the T/t antigens by alteration of their terminal sugar residues.     

Anthramycin inhibition of restriction endonuclease cleavage and its use as a reversible blocking agent in DNA constructions.

Anthramycin can form a stable complex with DNA which does not dissociate upon repeated ethanol precipitations. The complex forms in less than one hour at pH 5.5. Bound anthramycin seems to be located in the minor groove of the DNA helix in the anthramycin DNA complex, since methylation of adenosine residues at N-3 by dimethylsulfate is reduced. The anthramycin-DNA complex is resistant to digestion by an excess of a number of restriction enzymes. Anthramycin can be removed from DNA by incubation at acid pH. The released DNA can then be cleaved by restriction enzymes. Anthramycin-DNA complexes can be acted upon by T4 polynucleotide ligase to form longer DNA molecules. The ability of anthramycin to form a stable but reversible complex which is not cleaved by restriction enzymes but can engage in joining reactions may allow a wider variety of DNA fragments to be more readily constructed in vitro.     

The effects of short-term isolation on systolic blood pressure and heart rate in rats

The effect of short-term isolation on the systolic blood pressure and heart rate of rats has been studied. Five days continuous isolation in glass metabolic cages caused systolic arterial hypertension in all animals. Isolation in standard cages for this time period caused hypertension, but only in 55% of the animals. Both forms of isolation caused an initial tachycardia. Handling and contact with other animals for 1 hr daily prevented the development of hypertension in some animals but did not alter the blood pressure once the hypertension had developed. Group-housing of animals after a 3 week period of isolation restored blood pressure to control levels within 24 hr. It is possible that stress imposed by isolation activated the sympatho-adrenal system and thereby caused these changes.     

Evidence against the release of prostaglandin-like material from isolated intestinal tissue by pure cholera toxin.

Prostaglandin-like material was released from finely cut guinea-pig ileum or human intestinal mucosa during incubation with Krebs solution. The tissue inactivated some significant change in release of prostaglandin-like material when pure cholera toxin was incubated with guinea-pig ileum or human intestinal mucosa. The work is discussed in relation to the action of cholera toxin in vivo.     

Association of spectrin with its membrane attachment site restricts lateral mobility of human erythrocyte integral membrane proteins

Interactions between spectrin and the inner surface of the human erythrocyte membrane have been implicated in the control of lateral mobility of the integral membrane proteins. We report here that incubation of “leaky” erythrocytes with a water-soluble proteolytic fragment containing the membrane attachment site for spectrin achieves a selective and controlled dissociation of spectrin from the membrane, and increases the rate of lateral mobility of fluorescein isothiocyanate-labeled integral membrane proteins (> 70% of label in band 3 and PAS-1). Mobility of membrane proteins is measured as an increase in the percentage of uniformly fluorescent cells with time after fusion of fluorescent with nonfluorescent erythrocytes by Sendai virus. The cells are permeable to macromolecules since virus-fused erythrocytes lose most of their hemoglobin. The membrane attachment site for spectrin has been solubilized by limited proteolysis of inside-out erythrocyte vesicles and has been purified (V). Bennett, J Biol Chem 253:2292 (1978). This 72,000-dalton fragment binds to spectrin in solution, competitively inhibits association of ³²P-spectrin with inside-out vesicles with a K_i of 10^?7M, and causes rapid dissociation of ³²P-spectrin from vesicles. Both acid-treated 72,000-dalton fragment and the 45,000 dalton-cytoplasmic portion of band 3, which also was isolated from the proteolytic digest, have no effect on spectrin binding, release, or membrane protein mobility. The enhancement of membrane protein lateral mobility by the same polypeptide that inhibits binding of spectrin to inverted vesicles and displaces spectrin from these vesicles provides direct evidence that the interaction of spectrin with protein components in the membrane restricts the lateral mobility of integral membrane proteins in the erythrocyte.