Cell surface changes in preimplantation mouse embryos during compaction investigated using FITC conjugated lectins after proteolytic enzyme treatment

Summary Fluorescein-conjugated lectins were used to examine the reappearance of glycoproteins on the surface of 8-cell mouse embryos after treatment with proteolytic enzymes. Embryos were decompacted in calcium free medium, treated with various proteases and the process of recompaction monitored. The most effective enzymes in delaying recompaction were subtilopeptidase A and proteinase K at 1 mg/ml; the initiation of recompaction was delayed by about 5 h and 90% recompaction by 14–18 h. Papain and -chymotrypsin were only effective in the absence of calcium. The reappearance of receptors for fluorescein-conjugated Con-A, MPA, RCA-I, FBP, BSL-II and DBA was examined photometrically at 0,8–10 and 17–18 h after proteinase K treatment. There was an increase in binding of MPA, RCA-I, FBP and BSL-II in control embryos during the period of the experiment, between approx. 61 and 80 h post coitum in which embryos passed from the 8-cell stage to the 16–32 cell stage. Con-A binding remained the same and that of DBA decreased. By the time that 50% of enzyme treated embryos had recompacted (8–10 h) binding of Con-A was similar to control embryos. Binding of FBP had almost reached control levels while that of BSL-II, DBA, RCA-I and MPA had reached 60–85% of control levels. When embryos were fully compact (17–18 h) Con-A, FBP and DBA were bound in equal or slightly greater amounts to enzyme treated as to control embryos, and receptors for BSL-II, MPA and RCA-I had recovered almost to control levels. The results clearly show that the recovery of glycoproteins on the surface of 8–16 cell embryos parallels recompaction, providing further evidence for the role of these molecules in compaction.     

Computer-linked HPLC system for feedback control of fermentation substrates

Summary A data acquisition/control microcomputer system was interfaced to a commercial HPLC data transmission module. Control of substrate (ethanol) levels for four 7.5 L fermenters containing 100 g/L wet weight of the yeastCandida norvegensis was accomplished by employing intermittent, automated HPLC monitoring and a BASIC-encoded proportional integral policy for controlling substrate feed rates. Ethanol levels were maintained at 0.25, 0.50, 0.75 and 1.00% w/v.     

The utility of photo-affinity labels as `mapping' reagents. A study of sub-populations of a specific rabbit antibody by using structurally related photo-affinity reagents

A specific rabbit antibody against the 4-azido-2-nitrophenyl determinant was photo-labelled by the homologous hapten ε-(4-azido-2-nitrophenyl)-l-lysine, and by the close structural isomer ε-(5-azido-2-nitrophenyl)-l-lysine. The extents of covalent labelling of the antibody-binding site were assessed by using radioactive haptens and exhaustive displacement dialysis, which leaves the unlabelled sites empty but largely intact. A single photolysis of hapten–antibody complex suffices to label those sites that are capable of being labelled. Although there is considerable overlap among sub-populations of antibody that will bind the two haptens non-covalently, sites that can be covalently labelled by one reagent cannot be labelled by the other.     

Comparison of the effects of ultraviolet light and ethylmethanesulphonate upon the frequency of mitotic recombination in yeast

Summary Host-cell reactivation of gamma-irradiated phage T1 in strains of E. coli K-12 has been compared with HCR of UV-irradiated phage in these same strains and with the radiation sensitivities of these strains (Fig. 1–4). The pattern of the HCR of gammairradiated phage in these strains is like that of the HCR of UV-irradiated phage. HCR in strains whose genotype is uvr ⁺rec^- is like that of the wild type; whereas, HCR is minimal in strains which are uvr ^-. It is suggested that some type of gamma-ray-induced base damage in phage DNA is repaired in uvr ⁺ strains.This work was supported by the United States Atomic Energy Commission Contract No. AT(11-1)-1686. — This is report No. COO-1686-6.Supported in part by the United States Public Health Service Training Grant No. 5T1 RH-80-02(67).     

The integument of the Queensland fruit fly,Dacus tryoni (Frogg.)

Summary Testes of the Japanese freshwater turtle Clemmys japonica Temmnick et Schlegel were fixed in 3% potassium permanganate buffered to pH 7.2 with Veronal-acetate buffer, and thin sections of the tissue, embedded in epoxy Epon resin, were studied under the electron or light microscope.At the early stage of differentiation of the spermatid, the cytoplasm contains a few mitochondria provided with cristae which are oriented transversely or longitudinally. As the differentiation of spermatids proceeds, the mitochondrion has been modified into a cupshaped body with a wall consisting of several concentric layers. Such body has been referred to the mitochondrial lamellar body. The formation of such a body is mainly attributed to the mitochondrial cristae, and subsequently to the membrane system of the endoplasmic reticulum. In a more advanced stage of differentiation, the mitochondrial lamellar bodies appear wrapped around a bundle of tail filaments, and seem to present a very wide surface available for the localization of organized enzyme systems to facilitate the motion of spermatozoa.Prior to the formation of the mitochondrial lamellar bodies, the Golgi apparatus has been reorganized into a peculiar body with a floral appearance, consisting of numerous tubular elements, and revealing to be positive in PAS-reaction. The body has been designated as the tubular body which has never been demonstrated in any spermatogenic cells through animal kingdom.One to three tubules oval in cross section, approximately 430 × 700 Å in diameter, have been found in the nucleoplasm along the longitudinal axis of a greatly elongated, cone-shaped nucleus of the spermatid. The tubules open on the apex surface of the nucleus, but they are not encountered in the acrosome. A possible physiological significance of the tubules has been discussed in view of the function of the acrosomal tubules in the decapod and other species spermatozoa as well as on the basis of the metabolism of nucleus.This study was supported by Grant GM-8327 from the United States Public Health Service.We wish to express our gratitude to Dr. B. A. Afzelius, Wenner-Gren Institute, University of Stockholm, for his valuable suggestion to the present work.     

Identification of a common molecular basis for combined 17α-hydroxylase/17,20-lyase deficiency in two Mennonite families

Summary During the course of studies to characterize mutations of the CYP17 gene that cause the 17-hydroxylase-deficient form of congenital adrenal hyperplasia we have discovered two ostensibly unrelated Mennonite families in which affected individuals are homozygous for the same mutation. The defect is a four-base duplication in exon 8 of the CYP17 gene, which alters the reading frame encoding the C-terminal 26 animo acids of cytochrome P450₁₇.     

ATP-Evoked Ca2+ Mobilisation and Prostanoid Release from Astrocytes: P2-Purinergic Receptors Linked to Phosphoinositide Hydrolysis

Astrocyte cultures prelabelled with either [3H]inositol or 45Ca2+ were exposed to ATP and its hydrolysis products. ATP and ADP, but not AMP and adenosine, produced increases in the accumulation of intracellular 3H-labelled inositol phosphates (IP), efflux of 45Ca2+, and release of thromboxane A2 (TXA2). Whereas ATP-stimulated 3H-IP accumulation was unaffected, its ability to promote TXA2 release was markedly reduced by mepacrine, an inhibitor of phospholipase A2 (PLA2). ATP-evoked 3H-IP production was also spared following treatment with the cyclooxygenase inhibitor, indomethacin. We conclude that ATP-induced phosphoinositide (PPI) breakdown and 45 Ca2+ mobilisation occurred in parallel with, if not preceded, the release of TXA2. Following depletion of intracellular Ca2+ with a brief preexposure to ATP in the absence of extracellular Ca2+, the release of TXA2 in response to a subsequent ATP challenge was greatly reduced when compared with control. These results suggest that mobilisation of cytosolic Ca2+ may be the stimulus for PLA2 activation and, thus, TXA2 release. Stimulation of alpha 1-adrenoceptors also caused PPI breakdown and 45 Ca2+ efflux but not TXA2 release. The effects of ATP and noradrenaline (NA) on 3H-IP accumulation were additive, but their combined ability to increase 45Ca2+ efflux was not. Interestingly, in the presence of NA, ATP-stimulated TXA2 release was reduced. Our data provide evidence that functional P2-purinergic receptors are present on astrocytes and that ATP is the first physiologically relevant stimulus found to initiate prostanoid release from these cells.     

Effect of starvation and sampling time on plasma alkaline phosphatase activity and calcium homeostasis in the rat

The effect of starvation and sampling time on plasma alkaline phosphatase activity, total plasma calcium concentration and whole blood ionized calcium concentration was determined in the rat. Starvation caused a significant fall in total and ionized calcium concentrations as well as in alkaline phosphatase activity. These changes were accompanied by a fall in whole blood pH and an increase in the anion gap and a decrease in urinary excretion of calcium. These indices were restored to normal following refeeding. There was no change in serum 25-OH vitamin D concentrations following starvation for 3 days. Alkaline phosphatase activity showed a pattern compatible with the presence of a circadian rhythm when sampling took place between 0800 and 1800 h. Total and ionized calcium concentrations did not show such a rhythm when animals were fed the present diet.     

A long-term perspective of dissolved organic carbon transport in Sycamore Creek, Arizona, USA

Dissolved organic carbon (DOC) dynamics were examined over five years (1989–1993) in Sycamore Creek, a Sonoran Desert stream, specifically focusing on DOC concentration in surface and hyporheic waters, and rates of export. In 1989 and 1990, the years of lowest stream discharge (0.08 and 0.04 m³ s^–1 annual mean of daily discharge, respectively), DOC was high, averaging 7.37 and 6.22 mgC l^–1 (weighted annual means). In contrast, from 1991 through 1993, a period of increased flow (1.1, 1.2 and 4.3 m³ s^–1), concentration was significantly lower (P<0.001) with annual mean concentrations of 3.54, 3.49 and 3.39 mgC l^–1. Concentration exhibited little spatial variation between two sampling stations located 6 km apart along the mainstem or between surface and hyporheic waters. Annual export of DOC from Sycamore Creek varied 100-fold over the five-year period from a mean rate of only 24 kgC d^–1 in 1990 to 2100 kgC d^–1 in 1993. Ninety percent of DOC was exported by flows greater than 2.8 m³ s^–1, and 50% during flows greater than 27 m³ s^–1; flows of 2.8 and 24 m³ s^–1 occurred only 9 and 1% of the time. The export of organic matter in Sycamore Creek appears to be coupled to El Niño-Southern Oscillation phenomena. The years of highest export, 1991–1993, had El Niño conditions while 1989 and 1990 had medial conditions.     

Anemophilous plants select pollen from their own species from the air

In wind-tunnel experiments, Niklas (1985) has demonstrated the ability of anemophilous plants to select pollen from their own species from the airstream. However, there have been no field experiments to establish whether this operates in nature. We surveyed the pollents on the stigmas of four different, coextensive, dioecious anemophilous species surrounded by a Pinus radiata plantation. The alien pine pollen was overrepresented relative to background levels on only one of the four species. For all three indigenous species with both male and female plants in the area, the highest proportion — in all cases more than 40% — of the pollen found on their stigmas came from their own species. One indigenous species lacked male plants in the area; consequently, the results from this species are difficult to interpret. However, for all four species, there was at least 15% pine pollen, and some pollen of other indigenous species. These results suggest that there is some pollen selection, but that the mechanisms are may be not as effective as Niklas has suggested.