Presynaptic Nicotinic Receptors Stimulate Increases in Intraterminal Calcium of Chick Sympathetic Neurons in Culture

Abstract: Stimulation of chick sympathetic neurons in culture by the cholinergic agonists acetylcholine, nicotine, and 1,1-dimethyl-4-phenylpiperazinium (all at 10–1,000 µmol/L) induced concentration-dependent increases of free calcium levels measured by fura 2 fluorescence in neuronal processes. The response evoked by acetylcholine had both nicotinic and muscarinic components, whereas that induced by 1,1-dimethyl-4-phenylpiperazinium was purely nicotinic. Tetrodotoxin (0.3 µmol/L) blocked completely the increase of intraterminal free calcium level evoked by electrical stimulation. On the other hand, stimulation with 1,1-dimethyl-4-phenylpiperazinium still evoked 20–25% of the control response in the presence of tetrodotoxin. The concentration-response relationship of 1,1-dimethyl-4-phenylpiperazinium stimulation did not differ in the absence and in the presence of tetrodotoxin. The nicotinic antagonists d -tubocurarine (10 µmol/L) and mecamylamine (10 µmol/L), but not α-bungarotoxin (125 nmol/L), prevented the increase of intraterminal free calcium level evoked by 1,1-dimethyl-4-phenylpiperazinium (100 µmol/L) in the presence of tetrodotoxin. These observations indicate the presence of nicotinic receptors on neuronal processes that increase the intraterminal concentration of free calcium and probably modulate transmitter release. Their pharmacological properties are similar to those of nicotinic receptors located on neuronal cell bodies.     

Neural and smooth muscle development in the chicken gizzard

The development of the autonomic ganglia of Auerbach's plexus and gizzard smooth muscle was studied in chicken embryos. Nervous system and smooth-muscle-specific antibodies were employed in immunofluorescence stainings on tissue sections to investigate the temporal and spatial frame of neural and muscular differentiation in relation to each other. Subserosal clusters of neural cells were clearly demonstrable at embryonic day 5 (ED5), the earliest stage analysed, with the monoclonal antibody El (SGIII-1). Fine nerve fibres (ED6) and, later, large axon bundles projecting from subserosal neuron clusters towards the lumen were followed and found to reach the luminal border by ED11. Already in early development the area of the future laminar tendons on the ventral and dorsal surface of the gizzard was devoid of neuroblasts, and nerve fibres were not extending to the muscle-tendon borderline until ED16. Double stainings with antibodies to smooth muscle myosin (SMM) and El revealed that SMM expression, taken as an indicator for muscle differentiation, followed neural growth. It was first detectable in close apposition to the differentiating neuroblasts in the caudal and cranial portion of the gizzard at ED6. With further development, myosin expression proceeded inward towards the lumen in a wave which followed the ingrowth of E1-positive nerve fibres from the prospective Auerbach plexus. Neuromuscular differentiation deviated from this pattern in the lateral tendon area where nerve growth was delayed and myosin expression preceeded the arrival of E1-positive nerve fibres. The findings suggest that the gizzard could serve as a model system for the analysis of potential early nervous system imprints on smooth premuscle mesenchyme differentiation.     

Mortality associated with moderate intakes of wine,beer, or spirits.

OBJECTIVE--To examine the association between intake of different types of alcoholic drinks and mortality. DESIGN--Prospective population study with baseline assessment of alcohol intake, smoking habit, income, education, and body mass index, and 10-12 years'' follow up of mortality. SETTING--Copenhagen city heart study, Denmark. SUBJECTS--6051 men and 7234 women aged 30-70 years. MAIN OUTCOME MEASURE--Number and time of cause-specific deaths from 1976 to 1988. RESULTS--The risk of dying steadily decreased with an increasing intake of wine--from a relative risk of 1.00 for the subjects who never drank wine to 0.51 (95% confidence interval 0.32 to 0.81) for those who drank three to five glasses a day. Intake of neither beer nor spirits, however, was associated with reduced risk. For spirits intake the relative risk of dying increased from 1.00 for those who never drank to 1.34 (1.05 to 1.71) for those with an intake of three to five drinks a day. The effects of the three types of alcoholic drinks seemed to be independent of each other, and no significant interactions existed with sex, age, education, income, smoking, or body mass index. Wine drinking showed the same relation to risk of death from cardiovascular and cerebrovascular disease as to risk of death from all causes. CONCLUSION--Low to moderate intake of wine is associated with lower mortality from cardiovascular and cerebrovascular disease and other causes. Similar intake of spirits implied an increased risk, while beer drinking did not affect mortality.     

Fine structure of dense-cored vesicles in glomus cells of rat carotid body after fixation with permanganate or glutaraldehyde

Summary Glomus (Type I) cells of the carotid body of adult rats were studied electron microscopically after fixation with potassium permanganate or with glutaraldehyde and osmium tetroxide. Two permanganate fixation methods (using Krebs-Ringer-glucose, pH 7.0, or acetate buffer, pH 5.0) were compared. Numerous dense-cored vesicles were observed only in about one tenth of the glomus cells when neutral permanganate was used for fixation, although all glomus cells showed such vesicles after fixation with glutaraldehyde and osmium tetroxide. Numerous vesicles with a dense core were observed in about one third of the cells after fixation with acid potassium permanganate. With this fixation, small dense-cored vesicles similar to those in adrenergic nerve terminals were occasionally seen in the cytoplasm of glomus cells. It is tentatively concluded that the amine-storing vesicles of the carotid body are different from those in the small intensely fluorescent (SIF) cells and those in adrenergic nerve terminals.     

Sequence homology and recombination between the genomes of morphologically dissimilar bacteriophages LP 52 and theta

Summary A restriction fragment map of Bacillus licheniformis temperate phage LP 52 DNA (molecular weight 38.5×10⁶) was established, using restriction endonucleases BamHI (8 target sites), BglI (10 sites), BglII (13 sites) and EcoRI (22 sites). The map is linear, with well-defined ends, without any signs of circular permutation. The DNA of a related phage, LP 51, produced identical restriction fragments. At least 62% DNA of LP 52 has been found homologous to the DNA of the recently discovered, morphologically quite dissimilar, phage , as demonstrated by hybridization of electrophoretically separated restriction fragments of DNA. Under the same conditions, the DNAs of LP 52 and of the morphologically similar Bacillus subtilis phage 105 did not cross-hybridize. The homologous regions in the genomes of phages LP 52 and have been shown to be colinear. Comparison of the cleavage maps of phages LP 52 and has shown that, within the regions of homology, not a single restriction fragment and few restriction sites have been conserved during divergent evolution. Three major regions of heterology were defined; the longest one, covering the right-hand end of the map (73±2.75% up to 100% LP 52 genome length) appeared to contain genes coding for structural proteins of the virions; a shorter region at the left-hand end of the map (coordinates zero to 10.3±3.3% LP 52 genome length) and a very short central region (coordinates 41.8–43.9%) could be identified, the latter apparently containing a regulatory locus responsible for the heteroimmune behavior of the two phages. Recombinants between phages LP 52 and were isolated. Mapping of recombinant genomes has indicated mutual substitution of allelic pieces of LP 52 and DNAs upon strict conservation of overall genome length.     

Determination and postnatal development of fructokinase activity in swine liver]

Starting from the spectrophotometric method, described in ADELMAN et al., optimal reaction conditions for the measurement of fructokinase (ketohexokinase) in pig liver were systematically studied. It was necessary to increase the concentration of the substrate and further to lower the concentration of ATP for an optimal Mg: ATP-radio of 2:1. Using the optimized method fructokinase activity was determined in pig liver in relation to age, beginning from the last days of pregnancy to puberty. In liver of fetuses and newborn piglets during the first two days of life no or only a minute activity of the fructokinase was recorded. Therefore, the high level of fructose in fetal blood results from the inability of the fetus to metabolize fructose synthezised in the placenta or the fetal organs. At the end of the first week of life the activity of fructokinase was 10 times, after the second week 15-20 times higher than at birth. This high level remains constant during the suckling period and after weaning. For this reason, piglets after the first week of life are able to metabolize fructose and after the third week to form glucose and to release it into circulation. In adult pigs the activity of fructokinase in the liver decreases slightly. It corresponds-as in rat and human-to the elimination rate of experimentally applied fructose from the circulation. Therefore, this enzyme even in pigs is of significant importance for the utilization of fructose.     

An efficient procedure for serial nucleic acid hybridizations by titration analysis.

A simplified and efficient procedure has been developed for performing hybridization reactions and analyses of the percentage of hybridization on glass plates. This method is as accurate as conventional methods, while considerably reducing the time and costs to perform the hybridization in titration hybridization experiments.