Characterization of a developmentally transient adrenergic depolarizing response in cultured locus coeruleus neurons

The effects of iontophoretically applied noradrenaline have been tested on intracellularly recorded locus coeruleus neurons grown in explant cultures from neonatal mice. In addition to hyperpolarizing responses mediated by alpha 2-adrenergic receptors, as observed in locus coeruleus neurons in vivo and in brain slices from adult animals, alpha 1-mediated depolarizations were observed to succeed the initial hyperpolarizations in some cultures. It was shown that the depolarizing responses were only present in younger cultures, i.e., less than 26 days in vitro. In cultures less than 20 days old, all cells displayed the biphasic hyperpolarizing-depolarizing responses. Both components of the response appear to be direct, since they were present when synaptic transmission was blocked by including tetrodotoxin or by altering divalent cations in the perfusate. The depolarizing responses were frequently reduced in solutions with altered divalent cation content, and this might reflect a calcium dependency of this response. The hyperpolarizing and depolarizing components of the responses to noradrenaline were progressively blocked by increasing concentrations of the selective antagonists yohimbine and prazosin, respectively, in the dose ranges of 100 mM - 1 microM (yohimbine) and 20-200 nM (prazosin). Recent results from electrophysiological studies of locus coeruleus neurons in brain slices suggest that similar changes occur in the animal as well as in culture. It is possible that the transient depolarizing responses reflect a developmentally important enhanced responsiveness of locus coeruleus neurons during the early postnatal period.     

Distribution and hydraulic significance of large woody debris in a lowland Australian river

The line-intersect technique was used to measure the loading of large woody debris in a 1.8 km reach of the Thomson River, Victoria (catchment area of 3540 km²). A debris census (measuring every item present) was done over 0.775 km of this reach. The transect technique over-estimated the actual loading revealed by the census. The loading of debris 0.01 m in diameter for the total 1.8 km reach was 0.0172 m³ m^–2, which is higher than that measured in many headwater streams in other parts of the world. The volume loading of debris measured from low level aerial photographs was only 4.8% of the value estimated by the line-intersect technique. The line-intersect estimates were biased due to non-random orientation of debris in the stream (causing estimated errors of +8% for volume loading and +16% for surface area loading). It is recommended that to avoid this problem, when using the line-intersect transect technique in lowland rivers, each line should comprise at least two obliquely-angled transects across the channel. The mean item of debris (0.1 m in diameter) had a trunk basal diameter of 0.45 m, a length of 7.4 m, and volume of 0.7 m³. The riparian trees and the in-channel debris were of similar dimensions. The debris tended to be close to the bed and banks and was oriented downstream by the flow at a median angle of 27°. Because of this orientation, most debris had a small projected cross-sectional area, with the median value being only 1 m². Thus, the blockage ratio (proportion of projected area of debris to channel cross-sectional area) was also low, ranging from 0.0002 to 0.1, with a median value of 0.004. The average item of debris, which occupied only 0.4% of the cross-section, would have minimal influence on banktop flow hydraulics, but the largest items, which occupied around 10%, could be significant. Judicious re-introduction of debris into previously cleared rivers is unlikely to result in a large loss of conveyance, or a detectable increase in flooding frequency.     

Role of the tubulin-microtubule system in lymphocyte activation

The role of the tubulin-microtubule system was examined in human peripheral blood leukocytes after activation with phytohemagglutinin (PHA). Soluble tubulin and microtubules were measured with a [(3)H]colchicine-binding assay. It was found that the tubulin content of PHA-activated lymphocytes was consistently increased relative to total protein content after 36 h of culture. There was no increase in the proportion of total tubulin synthesis which was present as microtubules at 36 h. Nevertheless, as a result of increased tubulin synthesis, there was a two-to three-fold increase in total microtubular mass. Colchicine, which disrupts microtubles, was used to assess the role of microtubule assembly in the sequence of events which follow lymphocyte activation, namely lymphokine release, protein synthesis, RNA synthesis, and DNA synthesis. Colchicine consistently inhibited DNA synthesis but did not inhibit release of the lymphokine, osteoclast activating factor (OAF). Protein and RNA syntheses were inhibited much less than DNA synthesis. The fact that some effects of PHA on lymphocytes appear to require intact microtubules and at least one does not suggest that the microtubule dependent step in PHA-stimulated lymphocyte activation occurs at a stage after propagation of the signal from the membrane to the cell interior.     

The electrical activity of mechanoreceptive and neurosecretory neurons in the stick insectCarausim morosus

Summary Action potentials have been recorded from the neurosecretory cells which lie on the link nerve inCarausius morosus. The neurosecretory cells are spontaneously active in completely isolated preparations, firing with a regular but low frequency (<1 imp/s) or in small bursts (12 imp/s). The action potentials recorded extracellularly from the neurosecretory fibres are characteristically of long duration (2 to 10 ms), whereas those of motor or sensory fibres are of shorter duration (0.6 to 0.8 ms). The neurosecretory action potentials are also characterised by their slow conduction velocity (0.15 to 0.25 m/s) compared to those from motor and sensory fibres (0.54 to 0.7 m/s). The action potentials are propagated from the region of the cell body towards the terminals and have been recorded passing along all the major nerves in the periphery.Recordings from three of the non-neurosecretory cells which lie on peripheral nerves show that they respond to stretching of the nerves upon which they lie or of nerves which branch in the immediate vicinity. The action potentials are propagated away from the cell body towards the central nervous system. The neurons are termed peripheral nerve stretch receptors.We are grateful to the Science Research Council for financial support.     

Activation of human prothrombin by a procoagulant fraction from the venom of Echis carinatus. Identification of a high molecular weight intermediate with thrombin activity.

In the presence of a procoagulant fraction (Echis carinatus procoagulant) isolated from the venom of the saw-scaled viper Echis carinatus sochureki, purified human prothrombin (P1) is completely converted to thrombin. The first step is the removal of an NH2-terminal peptide (F1) representing approximately one-third of the prothrombin molecule. The remaining peptide (P2) is then cleaved by the action of E.c. procoagulant to yield a two-chain, disulfide-bridged protein (P'2) which has the same molecular weight as P2. P'2 has enzymic (thrombin) activity, as evidence by incorporation of radiolabeled diisopropylphosphate into its heavy chain (TB), hydrolysis of p-toluenesulfonylarginine methyl ester, and clotting of fibrinogen. Relative to thrombin, its esterolytic activity greatly exceeds its clot-promoting activity. Examination of the polypeptide chains obtained by reducing P'2 has shown that its larger chain (TB) is indistinguishable from the heavy chain of thrombin. Its other chain (F2TA) consists of the light chain (TA) of thrombin bound by peptide linkage to the protion of the prothrombin molecule which had been adjacent to F1. Removal of this portion (F2) is catalyzed by thrombin (and, evidently, by P'2), but not by the E.c. procoagulant. When F2 is removed from P'2, the remaining two-chian protein is indistinguishable from thrombin by any of the criteria applied--molecular weight, subunit chain composition, or enzymic activity. Polyacrylamide gel electrophoresis was carried out in sodium dodecyl sulfate before and after disulfide reduction of samples generated in the presence and in the absence of diisopropylphosphorofluoridate, which inhibits thrombin but not the E.c. procoagulant. Such experiments showed that thrombin (and probably P'2), as well as E.c. procoagulant, catalyzes the release of F1. Furthermore, thrombin brings about the cleavage of F1 to yield a two-chain, disulfidebridged protein (F'1). These observations, particularly those made in the course of characterizine P'2, have led to the conclusion that cleavage of the peptide bond connecting the TA and TB portions of the prothrombin molecule (or its derivatives) produces a serine active center and, hence, a molecule possessing thrombin activity. This cleavage is catalyzed by the E.c. procoagulant but not by thrombon itself.