Role of equilibration before rapid freezing of mouse embryos

The time requirements for permeation by glycerol and dehydration by sucrose before rapid freezing of Day-3 mouse embryos by direct transfer to -180 degrees C were studied. When the embryos were equilibrated in 2.0, 3.0, or 4.0 M-glycerol + 0.25 M-sucrose for 2.5 to 40 min, the post-thaw viability increased (P less than 0.001) with the length of equilibration period at 4 degrees C. At 20 degrees C the volume of embryos increased with the duration of equilibration up to 20 min (P less than 0.001), but the post-thaw viability was not affected. The effect of equilibration in glycerol-sucrose was determined at 20 degrees C for embryos which were previously permeated by glycerol, dehydrated by sucrose or left in PBS + 5% FCS. The survival of previously permeated embryos was not affected by equilibration for 1-16 min in glycerol-sucrose. The maximum survival rate was attained after shorter equilibration in glycerol-sucrose for embryos without pretreatment (4 min) than for those previously dehydrated (8 min). It is concluded that increases in the intracellular glycerol level are beneficial for the viability of rapidly frozen mouse embryos and previous or concomitant exposure to sucrose unfavourably affects glycerol permeation.     

Taurine Levels in Discrete Brain Nuclei of Rats

Concentrations of taurine have been measured in 44 microdissected rat brain nuclei or areas. Taurine is ubiquitously present and distributed unevenly in the rat brain: the ratio of the highest (pyriform cortex) to lowest (midbrain reticular formation) concentrations is 4.7:1. High taurine levels were found in cerebral cortical areas, caudate-putamen, cerebellum, median eminence, and supraoptic nucleus. Acute pain stress reduced taurine levels in the hypothalamus and the lower brainstem nuclei but not in cortical areas. Increased locomotor and behavioral activities following a high dose of amphetamine elevated taurine concentrations significantly in the substantia nigra and locus ceruleus.     

Enhancive and suppressive effects of cytomegalovirus on human lymphocyte responses in vitro.

Virus-specific lymphocyte proliferation in the presence of cytomegalovirus (CMV) without and with monocytes was studied in healthy persons. Three categories of lymphocyte response could be distinguished: seropositive low responders, naturally high responders, and lymphocyte populations responding well to CMV antigen in the presence of added CMV-incubated autologous monocytes. This latter category could be identified by preincubating autologous monocytes with CMV. CMV-seronegative persons were nonresponders. Early CMV antigens were produced in monocytes but not in lymphocytes by all CMV isolates. Infection of monocytes as detected by antibody to early viral protein did not appear to abort the antigen-presenting ability. The virus-specific responding lymphocytes were mainly of the T4+ phenotype. In contrast, addition of CMV to polyclonal mitogens significantly suppressed total lymphocyte DNA synthesis. CMV thus may have an enhanced virus-specific stimulatory effect on lymphocytes together with monocytes but a suppressive effect on the total lymphocyte population.     

Amiloride potentiates atrial natriuretic factor inhibitory action by increasing receptor binding in bovine adrenal zona glomerulosa

The interaction of atrial natriuretic factor (ANF) with the diuretic amiloride was studied in bovine adrenal zona glomerulosa. Amiloride enhances 2 to 3-fold high affinity binding of [125I] ANF to zona glomerulosa membrane receptor with an ED50 of 10 microM. This effect is due to a recruitement of high affinity receptor sites and to an increase of their affinity from a Kd of 23 to 8 pM. This enhancing effect is almost equipotently elicited by guanabenz, while clonidine is 20-fold less potent and arginine is inactive. ATP reduces by 30 to 50% [125I] ANF binding with an IC50 of 50 microM. Amiloride and ATP opposite effects on [125I] ANF binding are mutually competitive. Low concentrations of amiloride (less than 100 microM) potentiate the inhibitory effect of ANF in hormone-stimulated steroid secretion with a 3-fold decrease in ANF IC50 at 10 microM amiloride. Higher concentrations of amiloride (greater than 100 microM) directly inhibit aldosterone secretion with an IC50 of 500 microM and a maximum of 80 to 100% reversal of stimulation by various secretagogues. These results indicate that amiloride synergistically potentiates ANF inhibitory action by altering ANF receptor binding properties. They also suggest a role for sodium transport and for phosphorylation-dephosphorylation mechanisms in the mode of action of ANF.     

How does morphine work on colonic motility? An electromyographic study in the human left and sigmoid colon

The effect of morphine on colonic motility was investigated by recording the colonic myoelectric spiking activity by means of a 50 cm long silastic tube equipped with 4 bipolar AgAgCl ring electrodes fixed at 10 cm intervals that was introduced into the left colon in 8 healthy subjects by flexible sigmoidoscopy. Tracings were obtained for 1 hour in the fasting state and for another 1 hour after i.m. injection of morphine sulphate 0.15 mg/kg. The different types of spike bursts were compared before and after morphine injection. The control tracings showed that the spiking activity of the colon was made of 2 types: 1)- Rhythmic Stationary Spike Bursts (RSB), that were seen at only one electrode site; 2)- Sporadic Bursts, that were either propagating over all 4 electrodes (SPB) or non propagating (SNPB). Injection of morphine was followed by 1)- a considerable increase in the number of RSB from 107 +/- 43 bursts/hour (mean +/- SEM) to 491 +/- 23 bursts/hour; 2)- the complete disappearance of the SPB dropping from 7.3 +/- 2.0 bursts/hour to 0.3 +/- 0.2 bursts/hour; 3)- no significant change in SNPB (from 52 +/- 4 bursts/hour to 57 +/- 5 bursts/hour). These results indicate that 1)- stimulation of colonic smooth muscle activity by morphine seems to result from an increase in the number of rhythmic stationary bursts; 2)- however inhibition of colonic transit may be related to the decrease in the number of sporadic propagating bursts.     

A tetrapeptide isolated from hamster embryo with central opiate properties on gastrointestinal motility but not on pain perception

The effects of centrally administered kentsin (H-Thr-Pro-Arg-Lys-OH) on intestinal motility and on pain perception were investigated in rats chronically equipped with lateral ventricle catheters. Intestinal motility was recorded electromyographically from electrodes placed on the duodeno-jejunum; analgesia was evaluated by the hot-plate and tail-flick tests. Kentsin (4.0 ug/kg), injected intracerebroventricularly (ICV) 2 hours after the beginning of a meal, restores the "fasted" i.e. the migrating myoelectric complex of intestinal motility, while a 5 times higher dose administered subcutaneously was inactive. The ICV effect of kentsin was blocked by previous ICV administration of naloxone (400 ug/kg). In contrast, kentsin administered ICV (40 ug/kg) or SC (200 ug/kg) did not affect significantly (P greater than 0.05) the time latency in the two analgesic tests during 90 minutes after its administration and did not significantly modify the analgesic effects of (D5-Ala2, Met5) enkephalinamide. We conclude that kentsin when centrally administered acts on opiate receptors to alter gastrointestinal motility but without effects on pain perception.     

Effect of pertussis toxin on the heart muscarinic-cholinergic receptors and their function

Administration of pertussis toxin to rats induced a significant increase in heart rate that was evident as soon as 24 hours after the administration of the toxin and that persisted for at least 15 days. Electrical stimulation of the vagus decreased dramatically the heart rate of control animals but was unable to do it so in rats treated with pertussis toxin. In cardiac membranes muscarinic agonists decreased adenylate cyclase activity (approximately equal to 20-25%); no effect was observed in membranes obtained from toxin-treated animals. Agonist displacement of antagonist binding [( 3H] Quinuclidinyl benzilate) indicated that treatment with pertussis toxin decreased the proportion of receptors in the high affinity state for agonists. All these data suggest that blockade of the parasympathetic tone plays a key role in the induction of tachycardia by pertussis toxin.     

Molecular cloning and characterization of the trpC gene from Penicillium chrysogenum

Summary We cloned the Penicillium chrysogenum trpC gene from a genomic library by complementation of an Escherichia coli trpC mutant lacking phosphoribosylanthranilate isomerase activity. The gene ecodes a 2.7 kb poly(A)⁺ RNA. We localized the gene by sequence analysis in a 2.9 kb DNA insert found in the smallest plasmid selected from the library. Sequence data strongly suggest that the organization of the gene is similar to that described in other Ascomycetes. We found that a DNA fragment which codes only for the carboxy-terminal protion of the polypeptide is sufficient for complementation of the E. coli trpC9830 mutation.     

Membrane lipid changes in root cells of rape (Brassica napus) as a function of water-deficit stress

Rape seedlings ( Brassica napus L. cv. Brink) were exposed to repeated water-deficit stress. The water-stress program started after 19 days of growth and consisted of three 24 h stress periods interspersed with 24 h rewatering periods. After the third stress period the seedlings were harvested and the membrane lipids of the roots were extracted, isolated and quantified. The stress caused an increased ratio of dry weight roots/shoot. Furthermore, the total amount of acyl lipids as well as phospholipids decreased drastically. However, the relative distribution of individual phospholipids was constant and independent of stress. Free and esterified sterols showed only a small decrease in response to water stress. As a consequence the ratio free sterols/phospholipids increased from 0.07 in the control root cells to 0.15 in the stressed cells. The lipid changes are discussed in relation to membrane activity.     

Cell Surfaces in Plant-Microorganism Interactions : VI. Elicitors of Ethylene from Colletotrichum lagenarium Trigger Chitinase Activity in Melon Plants

Treatment of melon leaves or seedlings with elicitors of Colletotrichum lagenarium, a fungal pathogen of melon, increases chitinase activity. In treated leaves, chitinase is enhanced within the first 6 hours and becomes 2 to 10 times higher than in control leaves after 24 hours. Ethylene is increased simultaneously and is correlated with chitinase elicitation. In the presence of aminoethoxyvinylglycine, an inhibitor of ethylene synthesis, both elicitor-induced ethylene and elicitor-induced chitinase are inhibited. This inhibition is overcome by added exogenous ethylene. On the other hand, 1-aminocyclopropane-1-carboxylic acid the direct precursor of ethylene, triggers chitinase activity. Chitinase elicitation is thought to be a protein synthesis dependent process, as it does not occur in the presence of cycloheximide.