Showing papers in "Proceedings of The Royal Society B: Biological Sciences in 1967"

Crystallographic Studies of the Activity of Hen Egg-White Lysozyme

Abstract: The chemical evidence for the enzymic activity of lysozyme will be discussed in detail by other speakers at this meeting, but in order to describe our crystallographic studies of the interactions between the enzyme and its substrates it is necessary to summarize briefly what was known about them at the beginning of our work. Simultaneously with his discovery of lysozyme Fleming (1922) discovered a Gram-positive species of bacteria, Micrococcus lysodeikticus , which is particularly susceptible to the action of the enzyme. It was not until much later, however, that Salton (1952) demonstrated that the substrate is located entirely within the bacterial cell wall and it is only very recently that its chemical constitution has been established. Valuable early experiments (for example, by Meyer, Palmer, Thomson & Khorazo 1936; Meyer, Hahnel & Steinberg 1946; and by Epstein & Chain 1940) showed that lysozyme releases N -acetyl-amino sugars from M. lysodeikticus , but the first indication of the type of linkage attacked by lysozyme came when Berger & Weiser (1957) showed that lysozyme also degrades chitin, the linear polymer of N -acetylghicosamine.

On the Conformation of the Hen Egg-White Lysozyme Molecule

Abstract: Crystals of hen egg-white lysozyme, grown at pH 4.7 (Alderton & Fevold 1946), are tetragonal with a = b = 79.1 A, c = 37.9 A, space group P 43212 (Palmer, Ballantyre & Galvin 1948; Blake, Fenn, North, Phillips & Poljak 1962). Each of the eight asymmetric units in the cell comprises a single lysozyme molecule, molecular weight about 14 600, together with 1 M sodium chloride solution which constitutes some 33.5% of the weight of the crystal (Steinrauf 1959). The structure of these crystals has been determined by X-ray analysis by the method of multiple isomorphous replacement developed in the studies of haemoglobin (Green, Ingram & Perutz 1954; Blow 1958; Perutz, Rossmann, Cullis, Muirhead, Will & North 1960) and myoglobin (Kendrew, Dickerson, Strandberg, Hart, Davies, Phillips & Shore 1960). Anomalous scattering data were used in conjunction with the isomorphous replacement intensity differences (North 1965) to form a joint probability distribution for the phase of each reflexion. The position of the centroid of each probability distribution gave a phase angle and weighting factor for each reflexion from which the electron density map with minimum r.m.s. error was calculated (Blow & Crick 1959). A large number of different heavy atom derivatives were studied (Poljak 1963; Blake, Koenig, Mair, North, Phillips & Sarma 1965) and three proved satisfactory for calculating an electron density map at 2 A resolution. They contained respectively ortho -mercuri hydroxytoluene para -sulphonic acid, UO2F53- and an ion derived from UO2(NO3)2, probably UO2(OH)n(n-2)-

The Release of Acetylcholine from Nerve Endings by Graded Electric Pulses

Abstract: 1. The effect of brief depolarizations focally applied to a motor nerve ending was studied. Particular attention was paid to the relation between (i) strength and duration of the pulse and (ii) the size and latency of the resulting end-plate potential. 2. The release of acetylcholine lags behind the depolarization which causes it. If pulses of less than 4 ms duration are used (at 5 degrees C), the release starts after the end of the pulse. 3. Within a certain range, lengthening the pulse increases the rate of the ensuing transmitter release. 4. Unexpectedly, lengthening the depolarizing pulse also increases the latency of the transmitter release. This finding is discussed in detail. It is regarded as evidence suggesting that entry into the axon membrane of a positively charged substance (external Ca$^{2+}$ ions or a calcium compound CaR$^{+}$) is the first step leading to the release of acetylcholine packets from the terminal.

The Ferrier Lecture: Neuroglial Cells: Physiological Properties and a Potassium Mediated Effect of Neuronal Activity on the Glial Membrane Potential

Abstract: Neuroglial cells constitute a separate class of cells in the nervous system; they have been studied intensively since their original description by Virchow in 1846. As a rule anatomists find no difficulty in recognizing them by their staining properties, their shape and configuration as well as by their characteristic location between and around neurons. Electron microscopy has in recent years added much important subcellular detail and has shown how intermingled neurons and glial cells are, being separated from each other by narrow clefts 100 to 200 A wide (figures 1 A, B and 5, plates 1, 2 and 4). These studies have not changed the well-established grouping of mammalian glial cells into two main classes, the oligodendrocytes and the astrocytes . It is customary to state that glial cells outnumber neurons by 10 to 1 in the vertebrate nervous sytem. They are, however, smaller and according to some rough estimates they make up as much as 50% of the volume of mammalian brains. That glial cells differ significantly from neurons was clear from the beginning because they do not possess axons and, unlike mammalian neurons, they retain their ability to divide throughout life. The possible role of the large mass of glial cells in our nervous system has been of continued interest. During the past decade this interest in the physiology of neuroglia has been reinforced, largely under the stimulus of electron-microscopic and chemical studies of the nervous system. Among the numerous recent reviews and symposia only a few will be mentioned (Windle 1958; Nakai 1963; Mugnaini & Walberg 1964). The recent studies of the physiology of neuroglial cells have been reviewed by Kufller & Nicholls (1966) and a biblio­graphy on neuroglia has been compiled by Little & Morris (1965).

Tetrodotoxin and neuromuscular transmission

Abstract: 1. The puffer fish poison, tetrodotoxin ( T . T .) was applied to eliminate impulse propagation in nerve and muscle fibre, and the physiological properties of the neuromuscular junction were studied under this condition. 2. Spontaneous miniature end-plate potentials of normal frequency and amplitude were recorded in the T . T .-paralysed muscle. 3. Depolarization of motor nerve endings by locally applied current still produces the usual increase in the frequency of miniature end-plate potentials (e. p. ps). 4. When brief current pulses are applied to the nerve endings e. p. ps can be evoked, whose size varies with the intensity of the current. The responses are composed, like normal e. p. ps, of a statistically varying number of miniature potentials. The response fails when calcium is removed from the bath. 5. When two identical pulses are applied at varying intervals, facilitation of the second e. p. p. occurs, similar to that observed normally with pairs of nerve impulses. 6. It is concluded that tetrodotoxin while blocking electric excitation in nerve and muscle does not interfere with the release of acetylcholine from nerve endings nor with its local action on the muscle fibre.

Prenatal and Postnatal Growth and Development of the Central Nervous System of the Pig

Abstract: The growth and chemical development of the brain and spinal cord of the pig have been studied between the 52nd day of gestation and 3 years of age. The brain increased in weight faster than the spinal cord. Its most rapid period of growth lasted from about 50 days before birth to about 40 days afterwards and was made up of an early period characterized by DNA-P deposition and a later one of lipid deposition. The percentage of water in each part of the central nervous system fell during development and the concentrations of total N, phospholipid-P and cholesterol rose. The concentration of DNA-P fell gradually over the period studied in the brain as a whole, but in the cerebellum and brain stem the fall was preceded by a rise to a peak concentration at about 95 days of gestation. The cerebellum was the only part of the central nervous system examined in which the absolute amount of DNA-P attained a mature value before three years of age. The spinal cord had no early growth spurt and for this reason there was no time when the absolute amounts of cholesterol and phospholipid-P were rising conspicuously rapidly. There was, however, a period when the concentration of cholesterol increased most rapidly and this coincided with the one in other parts of the central nervous system. The bearing of these results on the effects of stress on the development of the central nervous system is discussed.

The Projection of the Lateral Geniculate Nucleus Upon the Cortex in the Cat

Abstract: The retrograde cell degeneration in the lateral geniculate nucleus of the cat has been studied after lesions of the visual and adjoining areas of the cortex. Following lesions which are limited to area 17, the medium and small cells of the main laminae of the nucleus degenerate; damage restricted to area 18 does not result in any localized, severe degeneration, but combined destruction of areas 17 and 18 causes all the cells-large, medium and small-of the main laminae and the central interlaminar nucleus to degenerate. Cellular change in the medial interlaminar nucleus is only found after involvement of area 19. When the cortex of the middle suprasylvian gyrus is removed in addition to these areas the degeneration in the lateral geniculate nucleus is much more severe, and there is loss of the laminar pattern due to severe gliosis in the central interlaminar nucleus. There is a well-defined topical organization in the geniculo-cortical projection, and in the antero-posterior dimension it is the same to all areas of the visual cortex, anterior parts of the nucleus projecting anteriorly and posterior parts posteriorly. Medial parts of the main laminae are related to the lateral part of area 17 and to the medial part of area 18, and lateral parts of the laminae project to the medial part of area 17 and to the lateral part of area 18. After partial lesions which involve both areas 17 and 18 the cellular degeneration affects the laminae differentially along their antero-posterior extent, that in lamina A being the most anterior and that in lamina B the most posterior; in sagittal sections such a band, or column, of degeneration passes from antero-superior to postero-inferior at right angles to the plane of the laminae.

The frequency of nerve action potentials generated by applied currents

Abstract: The steady frequency of nerve action potentials generated by maintained current stimuli (frequency-current curve) for the Hodgkin-Huxley equations was calculated both for a uniformly polarized membrane and a point polarized cable. The possible range of steady frequencies that can be maintained with a point polarized cable at 6$\cdot $3 degrees C is very limited and the range is even smaller at higher temperatures. Several alterations to the Hodgkin-Huxley equations were considered, but it was concluded that further mechanisms were probably required to explain the wide range of frequencies displayed by some nerve cells. The frequency range was correlated with the time course of the afterpotentials produced by a propagated action potential. The excitability of the cable to a second current stimulus also followed the afterpotentials closely. The frequency-current curve is markedly non-linear, though, because of the non-linear current-voltage curve, the frequency is quite linearly related to the steady voltage produced if regenerative activity is blocked. The results are compared with simplified models and the experimental literature. It is likely that further mechanisms are required to explain the adaptation, variability and wide range of steady frequencies displayed by some nerve cells.

The Accumulation of Noradrenaline in Constricted Sympathetic Nerves as Studied by Fluorescence and Electron Microscopy

Abstract: The effects of constricting post-ganglionic sympathetic nerves have been studied in the cat splenic nerve and guinea-pig hypogastric nerve. The results obtained using a fluorescence method for the histochemical localization of noradrenaline have been compared with electron microscopic findings. A close correlation was found between the accumulation of fluorescent material, attributable to noradrenaline, and of vesicles with an electron dense core (granular vesicles) believed to contain noradrenaline, proximal to the constriction in these nerves. This accumulation of noradrenaline was visible by 1 h after operation and increased rapidly in amount during the succeeding hours. It apparently reached a maximum after approximately 2 days and was found in what appeared to be newly formed axons 3 to 4 days after operation. Reserpine reduces the fluorescence and the number of vesicles with electron dense cores which accumulate proximal to the constriction. It is suggested, (1) that the fluorescent material is due, at least in part, to the presence of the granular vesicles, and (2) that the constriction has blocked the normal proximo-distal movement of noradrenaline which is believed to occur in post-ganglionic sympathetic axons.

The Dynamics of Burrowing in Ensis (Bivalvia)

Abstract: The digging activity of Ensis arcuatus shows six stages, together termed the 'digging cycle', which are repeated cyclically and are similar to those of other burrowing bivalves. A digging cycle involves the integration of pedal protraction and retraction with the opening and closing of the valves, much of the musculature of the body playing a part in each cycle. Extension and probing of the foot involves only the intrinsic pedal musculature which generates low pressures (10 cm) in the pedal haemocoele. The hinged shell acts as the basis of a fluid/muscle system which allows the strength of adduction to be used in digging and consists of two separate fluid filled chambers, the haemocoele and the mantle cavity. Adduction of the valves generates high pressures (100 cm) in each equally and simultaneously. In the haemocoele this pressure gives rise to the characteristic bulbous form of the foot which ensures a secure anchorage so that at pedal retraction the shell is drawn down. In the mantle cavity the pressure produces powerful jets of water which assist movement of the shell by loosening the adjacent sand. Estimates of the muscle tensions indicate that the pressures recorded may be obtained at tensions of not more than 2 Kg/cm$^{2}$.

The Direct Synthesis of Phosphoenolpyruvate from Pyruvate by Escherichia coli

Abstract: Extracts of Escherichia coli are shown to contain an enzyme system which in the presence of Mg 2+ catalyses the direct formation of phospho enol pyruvate from pyruvate and ATP with concomitant formation of AMP and inorganic phosphate. This enzyme, which has been designated 9phospho enol pyruvate synthase9 ( PEP -synthase) has been purified 80-fold and is free of pyruvate kinase activity; PEP synthesis proceeded most rapidly at pH 8 to 8.5. At pH values between 6.2 and 7.5 the enzyme can catalyse the formation of ATP and pyruvate from PEP , AMP and inorganic phosphate; if arsenate is used instead of phosphate, pyruvate and ADP are produced instead. Studies of the enzymic formation of PEP with ATP specifically labelled with 32 P, and of the reverse reaction with [U -14 C] AMP , suggest that the PEP -synthase reaction involves the transfer of a pyrophosphoryl-group. The physiological role of PEP -synthase has been demonstrated with mutants of E. coli devoid of the enzyme: in contrast to wild-type organisms, such mutants neither grow on pyruvate, lactate or alanine, nor form glycogen from lactate. It is thus concluded that PEP -synthase plays an important role in the anaplerotic and the biosynthetic reactions which enable the organisms to grow on pyruvate as sole carbon source.

The Vascular Circulation in the Brain, Optic Lobes and Thoracic Ganglia of the Crab Carcinus

Abstract: Ink injected into the blood system of the crab reveals a rich capillary system within all the central ganglia. The capillaries link afferent blood vessels with efferent vessels in such a way that the afferent vessels carry the blood deep into the neuropile and the capillaries take the blood through the synaptic areas and out to the surface efferents. The efferent vessels carry blood through the ganglion sheath and into the haemocoele. The branches of the capillary system in the structured neuropile areas of the optic lobe are regularly spaced and proliferation occurs at known synaptic areas. Capillaries in the diffuse brain neuropile are not geometrically arranged. Blood flows initially to the synaptic regions and thence past the nerve somata. With the exception of the retinula cell axons, main nerve bundles and axon chiasmata are poorly supplied with capillary systems. The demonstrated association of an extensive blood supply with synaptic areas in the optic lobe is correlated with previous experimental findings that integrated visual reflexes fail very shortly after interruption of the blood supply but primary receptor cell activity is relatively unaffected.

The 24-hour periodicity of microfilariae: biological mechanisms responsible for its production and control.

Abstract: An explanation is presented which covers most of the experimental data about the mechanisms by which the periodicity of microfilariae is maintained: During the night-time (with Wuchereria bancrofti and similar filariae) the microfilariae are evenly distributed throughout the blood and they are thus available for ingestion and transmission by mosquitoes. During the day-time they accumulate in the small vessels of the lungs, and hence they are few in the peripheral blood; this phase is probably adapted to allow the microfilariae to enjoy favourable physiological conditions in the lungs. The accumulation is due to an active reflex by the microfilariae themselves; and it probably depends on a sideways migration through the precapillary network of arterioles. The factor in the lungs which holds up the passage of the microfilariae so that they accumulate there (in preference to the capillaries of other organs), is the great increase in oxygen tension, which may be termed the 'oxygen barrier'. The 24 h cycle of the microfilariae is orientated to the 24 h cycle of the host; and some rhythmic change in the host acts as a cue to the microfilariae. Each microfilaria has a weak endogenous circadian rhythm of its own, but the rhythms of the individual microfilariae are dominated by that of the host, so that all the different individuals do approximately the same thing at the same time, and they do it at the right time (i.e. right for transmission). Different species of microfilariae respond differently to the same stimuli, and they depend on different arrangements for the maintenance of their rhythms. Three main groups of periodic microfilariae may be recognized. (a) W. bancrofti, Brugia malayi, etc. These depend upon the absolute size of the venous-arterial (V A) difference in oxygen tension ('oxygen barrier') which is lower by night (e.g. 40 mmHg) than it is by day (e.g. 55 mmHg) and so the microfilariae pass through the lungs by night but accumulate there by day. If at night the patient is caused to breathe oxygen, the arterial oxygen rises; or if he is caused to take vigorous muscular exercise, the venous oxygen tension falls; in both cases the V A difference becomes greater and the microfilariae accumulate in the lungs. (b) Loa loa of man, Edesonfilaria malayensis of monkeys in Thailand and Monnigofilaria setariosa of East African mongooses. In this group the sensitivity of the microfilariae to the oxygen barrier is greatly increased or decreased by the 24 h changes in the body temperature of the host. Accordingly, the cycle of microfilariae of this group indirectly depends upon the temperature cycle of the host. (c) Dirofilaria immitis, D. repens of dogs, D. aethiops (corynodes) of monkeys, etc. These microfilariae are probably sensitive only to the lower range of oxygen tensions, e.g. 30 to 60 mmHg. On the whole their cycle depends on day-night changes in the oxygen barrier as with W. bancrofti, but under special circumstances (as explained in the text) administration of oxygen may cause liberation of microfilariae from the lung instead of accumulation. The mechanism controlling the Pacific type of W. bancrofti cannot yet be identified, since the experimental evidence is insufficient. The behaviour of microfilariae is adapted to promote transmission by arranging the maximum number of microfilariae in the peripheral blood at times when the arthropod vector is likely to bite. The most sophisticated arrangement to achieve this is by a 24 h rhythm-the classical 'periodicity'. A less sophisticated arrangement is illustrated by various filariae of rodents, e.g. Litomosoides carinii and Dipetalonema witei, in which the parasites are transmitted by mites or ticks which suck blood in the nest or burrow, and the microfilariae are stimulated to swarm in the peripheral blood by a fall in body temperature when the animal sits quietly in its nest. Filariae which are still less sophisticated, e.g. Acanthocheilonema perstans and Dipetalonema gracile, do not possess any arrangements for adjusting the supply of microfilariae in the peripheral blood to the feeding habits of the vectors. Furthermore, some microfilariae, e.g. those of D. immitis and D. repens, are adjusted to their vectors on an annual variation as well as on a 24 h one and they are most numerous in the blood during July and August (when mosquitoes are most numerous in temperate zones). Some hosts (e.g. dogs) have a less marked 24 h rhythm than other hosts (e.g. man and monkey) and the cycles of their microfilariae are similarly less marked.

The Binding and Cleavage by Lysozyme of N-acetylglucosamine Oligosaccharides

Abstract: In the first part of this Discussion the beautiful crystallographic results obtained at the Royal Institution were described (Blake, Mair et al ., p. 365, Blake, Johnson et al ., p. 378). Particular attention was paid to the nature of certain lysozymesaccharide complexes, some involving oligosaccharides of N -acetylglucosamine. This paper describes reactions of these same sugars, namely, their binding and cleavage by the enzyme.

Prey Detection by Chaetognatha via a Vibration Sense

Abstract: Chaetognaths (arrow worms) have two types of tufts along the body, not previously distinguished. The bristles seen in life are non-sensory processes of epithelial cells. The ciliary tufts visible in sections are groups of sensory neurons each of which bears a non-motile cilium at the tip of its dendrite. The high sensitivity to vibrations set up by a nearby oscillating source in the dark is attributed to the non-motile cilia. The example analysed, Spadella cephaloptera, makes an accurate feeding movement towards any source vibrating at 9 to 20 c/s with an amplitude of 100 to 500 $\mu $m at a distance of 1 to 3 mm. Other vibrations are ignored or cause an escape response.

The uptake of amino acids by mouse cells (strain LS) during growth in batch culture and chemostat culture: the influence of cell growth rate

Abstract: The uptake of thirteen essential amino acids by mouse LS cells in suspension culture was determined by bacteriological assay methods. Chemostat continuous-flow cultures were used to determine the effect of different cell growth rates on the quantitative amino acid requirements for growth. The growth yields of the cells (Y = g cell dry weight produced/g amino acid utilized) were calculated for each of the essential amino acids. A mixture of the non-essential amino acids, serine, alanine and glycine increased the cell yield from the essential amino acids. The growth yields from nearly all the essential amino acids in batch culture were increased when glutamic acid was substituted for the glutamine in the medium. The growth yields from the amino acids in batch culture were much less at the beginning than at the end of the culture. The highest efficiencies of conversion of amino acids to cell material were obtained by chemostat culture. When glutamic acid largely replaced the glutamine in the medium the conversion of amino acid nitrogen to cell nitrogen was 100% efficient (that is, the theoretical yield was obtained) at the optimum growth rate (cell doubling time, 43 h). The maximum population density a given amino acid mixture will support can be calculated from the data. It is concluded that in several routinely used tissue culture media the cell growth is limited by the amino acid supply. In batch culture glutamine was wasted by (1) its spontaneous decomposition to pyrrolidone carboxylic acid and ammonia, and (2) its enzymic breakdown to glutamic acid and ammonia, but also glutamine was used less efficiently than glutamic acid. Study of the influence of cell growth rate on amino acid uptake rates per unit mass of cells indicated that a marked change in amino acid metabolism occurred at a specific growth rate of 0$\cdot $4 day$^{-1}$ (cell doubling time, 43 h). With decrease in specific growth rate below 0$\cdot $4 day$^{-1}$ there was a marked stimulation of amino acid uptake rate per cell and essential amino acids were consumed increasingly for functions other than synthesis of cell material.

The peduncle lobe: a visuo-motor centre in octopus.

Abstract: The peduncle lobe, the olfactory lobe and the optic gland constitute the peduncle complex. The two former structures are nervous, with a central neuropil surrounded by a cell layer, but the latter is an endocrine organ. The peduncle lobe contains two regions, the spine and the basal zone. In the spine the cells are 'small' (less than 5 $\mu $m), and the neuropil comprises a parallel array of axons, oriented with their long axes horizontal and parallel to the antero-posterior axis of the body. The axons form at least three efferent tracts, which leave the lobe via the basal zone, and run to higher motor centres in the central brain. In the basal zone the majority of cells are 'large' (greater than 5 $\mu $m) but about one third are 'small'. The large cells send axons to higher motor centres in the central brain. The small cells in the basal zone project into the peduncle spine. The basal zone receives afferent fibres from the ipsilateral optic lobe, the contralateral peduncle lobe and the ipsilateral basal lobes and is presumed to provide an area for interaction between information from these areas. The organization of the neuropil is less obvious here than it is in the spine, but it is possible to discern optico-pedunculo afferents, redistributing in a regular way in the horizontal plane. Both regions of the peduncle lobe project to motor areas in the central brain. The olfactory lobe, however, does not project to motor areas. It is composed of three distinct lobules: the posterior two lobules receive the olfactory nerve from the olfactory organ. The lobe is connected with the contralateral olfactory lobe and the ipsilateral dorsal basal lobe, a 'silent' area. Its neuropil is confluent with that of the peduncle lobe and they are assumed to influence one another closely. Stimulation of the nervous parts of the peduncle complex produces-in acute preparations-motor responses including colour changes, arm movements, and movements of the whole animal. After surgical removal of the nervous parts of both complexes there is locomotor dysfunction. Removal of the olfactory organs alone has no apparent effect on the behaviour of blind Octopus or Sepia and these organs are no more sensitive to the chemical stimuli used than other parts of the body. Anatomical, physiological and behavioural evidence suggest that the nervous part of the peduncle complex, probably the peduncle lobe alone, is part of a motor-control system. Evidence is presented that it regulates motor activity on the basis of visual information derived from the optic lobe.

The mechanisms of hydrolysis of glycosides and their revelance to enzyme-catalysed reactions.

Abstract: Glycoside hydrolysis can be formally represented by (I). where the group O R ( R is alkyl, aryl or a carbohydrate) is displaced from the C1 or C2 atom of an aldose or ketose sugar (or sugar derivative) respectively; the ring system being either five- or six-membered. Like the analogous hydrolysis of acetals, glycoside hydrolysis shows specific catalysis by hydrogen ions and, except where R is an aryl residue, base catalysis is absent. A number of enzymes* catalysing the hydrolysis of particular glycosides have been characterized: all show a high degree of specificity towards the carbohydrate structures present in the substrates. Both the acid-catalysed and the enzyme-catalysed reactions have been much studied, but whereas the former are now reasonably well understood, the mechanisms involved in the latter are still largely unknown.

The Chemical Structure of Lysozyme Substrates and Their Cleavage by the Enzyme

Abstract: The isolation of a disaccharide, N-acetyl-$\beta $-D-glucosaminyl-(1 $\rightarrow $ 4) N-acetylmuramic acid (NAG-NAM) and a corresponding tetrasaccharide, from lysozyme digests of Micrococcus lysodeikticus cell walls, is described. These compounds have been used for the study of the enzymic activity of lysozyme. Digestion of the tetrasaccharide into disaccharide has been followed by (a) paper chromatography, (b) colorimetry, using the Morgan-Elson reaction, and (c) polarimetry. Lysozyme was found to catalyse transglycosylation in addition to hydrolysis, and it is proposed that hydrolysis of the tetrasaccharide does not proceed by direct cleavage, but by a transfer mechanism, via long chain oligosaccharides. N-acetyl-glucosamine and closely related oligosaccharides strongly inhibit the enzymic activity of lysozyme. Fluorescence studies show that these inhibitors interact with the tryptophan residues in the active site of the enzyme.

The effect of undernutrition on the postnatal development of the brain and cord in pigs.

Abstract: Sucking pigs about 2 weeks old were held back by undernutrition so that they weighed only 5 to 6 kg when they were a year of age. The brain and cord developed during this time to the size to be expected in a normal pig about 10 weeks old but, although they remained immature for their chronological age, the effect on the various constituents was not uniform. The accumulation of cholesterol was less retarded than that of DNA-P or the increase in brain weight. During rehabilitation on a highly satisfactory diet the final body weight reached at 3$\frac{1}{2}$ years was 80% of that to be expected in an adult pig and was equivalent only to that of a normal pig two years old. The central nervous system grew to the appropriate size for the body. The percentage of cholesterol in the central nervous system rose during rehabilitation, but, particularly in the forebrain, brain stem and spinal cord, remained subnormal for the chronological age. The deficiency of DNA-P in the rehabilitated brain was even greater, and the absolute amount finally corresponded to that found in the brain of a normal animal only one year of age.

An investigation of some properties of endothelium related to capillary permeability

Abstract: The muscle capillaries of hearts taken directly from rats and mice, or perfused in vitro through the coronary vessels, were investigated by electron microscopy. The electron-dense marker particles ferritin and saccharated iron oxide were introduced into the blood or perfusion fluid. Versene, enzymes, surface active agents, and inhibitors of metabolism were added to the perfusion fluid, and perfusion was also carried out at 0 degrees C. Caveolae and vesicles were present in the endothelium of perfused hearts after all these measures; in damaged cells they disappeared in parallel with the cell and nuclear membranes. Particles entered caveolae and vesicles from the lumen under all conditions of perfusion. With saccharated iron oxide it was not certain what proportion of particles entering caveolae from the lumen passed right through the cell, since some of the particles in external caveolae could have come from accumulations in the basement membrane. Small numbers of ferritin molecules seemed to traverse the cell by the caveolae and vesicles in perfused hearts and in the whole animal. Groups or chains of communicating caveolae and vesicles were often seen. The cells remained closely apposed and the junctions intact in all preparations. Saccharated iron oxide passed into the junctions. In perfused hearts a variable amount left the vessels by this route, which was probably the main source of accumulations in the basement membrane and extravascular structures. Very little ferritin entered the junctions either in the intact animal or in vitro. The basement membrane was largely removed by papain and by impure preparations of collagenase but was not clearly affected by the other enzymes used. The removal of calcium from the perfusion fluid caused the basement membrane to separate from the endothelium.

Monocular fixation in human eye movement

Abstract: The monocular eye movements associated with the maintenance of fixation have been recorded using the contact lens/optical lever system. The records, in analogue form on magnetic tape, were subsequently converted to a digital form and analysed on a computer. It is found that there is reasonable agreement between the responses of the same subject on different days, Analysis of the magnitudes and preferred directions of saccades and drifts and their interrelationships show that there is an elliptical overall fixation area which is sub-divided into a series of overlapping short-period fixation areas. The results indicate that saccades occurring during fixation have one of two functions, either (a) to recentre the retinal image on the short-period mean fixation position, or (b) to move the short-period fixation area, possibly to avoid retinal receptor fatigue. An organizational model of the fixation control system has been developed from the analysis of results.

The Release of Vasopressin without Oxytocin in Response to Haemorrhage

Abstract: Blood samples were collected from anaesthetized cats during haemorrhage or stimulation of the peripheral end of the vagus. Vasopressin and oxytocin were estimated in the samples by assaying alcohol extracts for antidiuretic activity in the water loaded rat and for milk-ejecting activity in the lactating guinea-pig. Haemorrhage caused vasopressin to be released into the blood with out detectable amounts of oxytocin. A similar result was obtained with vagal stimulation provided that the fall of blood pressure which it produced exceeded a critical value of about 80 mmHg. Failure to detect oxytocin in blood samples containing vasopressin was not due to the presence of adrenaline or any other inhibitory substance in the extracts blocking the response of the mammary gland to oxytocin. The stimulus for the independent release of vasopressin by haemorrhage appears to be the associated fall in arterial blood pressure.

Hypothalamic pathways for independent release of vasopressin and oxytocin.

Abstract: By means of a simple method for extraction of blood samples, reliable recoveries of vasopressin and oxytocin were obtained. This method was used in anaesthetized cats for estimation of these hormones, released into external jugular venous blood, after localized electrical stimulation in the hypothalamus. Stimulation of the supraoptic nucleus or supraoptico-hypophysial tract released large amounts of vasopressin without oxytocin. Stimulation of the paraventricular nucleus or tractus paraventricular is cinereus (Greving) released moderate amounts of vasopressin alone. Stimulation at points in the tuberal region of the hypothalamus released oxytocin with or without vasopressin. These could lie on the caudal tract from the paraventricular nucleus (Laqueur).

Psychological refractory period and the length of time required to make a decision.

Abstract: Three groups of men were tested in a situation involving reaction to two successive stimuli at a short known time interval. Each reaction required the choice of one of two reaction keys, depending upon the light stimulus delivered. For one group of subjects, the first reaction was made easy by making the left key correct for the left light; for the second group, the first reaction was made slower by making the left key correct for the right light. In the third group, one of the two possible stimuli for the first reaction occurred more often than the other, so that the first reaction was sometimes fast and sometimes slow. The second reaction was delayed if the interval between stimuli was shorter than the first reaction time; in conditions where the first reaction time was longer, the second reaction time was delayed by an even greater amount. This result supports the view that a common decision-making mechanism of limited capacity is occupied by the first reaction and so is unable to deal with the second one; two other theories of the effect seem inconsistent with the present data.

Relationship between chemical structure and biological activity of hen egg-white lysozyme and lysozymes of different species.

Abstract: Fleming discovered an enzyme which he called lysozyme; it had ‘the power of rapidly dissolving certain bacteria, which is possessed by many animal and vegetable tissues and secretions and to a very marked degree by egg-white' (Fleming 1922). Hen egg-white lysozyme was crystallized by Abraham & Robinson in 1937; its homogeneity was verified by different chromatographic and physical methods, which have recently been reviewed (Jolles, P. 1960, 1964). Since 1958, Jolles and others have been engaged in elucidating the primary structure; they published a preliminary formula in 1961 (Jolles, J. & Jolles, P. 1961), followed in 1963 by a detailed study (Jolles, J., Jauregui-Adell, Bernier & Jolles, P. 1963); their structure differed slightly from that of Canfield (1963). The positions of the four disulphide bonds were also established (see table 1), and the complete formula is indicated in figure 1.

The Role of Lymphocytes in Antibody Formation: II. The Influence of Lymphocyte Migration on the Initiation of Antibody Formation in the Isolated, Perfused Spleen

Abstract: The aim of these studies was to examine the possible immunological significance of the process of lymphocyte recirculation. For this purpose a technique for perfusing the isolated spleen of the rat was developed. Antigen (sheep erythrocytes) was added to the perfusate and, after 3 to 6 h periods of perfusion, the spleens were cut into fragments and implanted into the peritoneal cavities of X-irradiated syngeneic recipients to determine the magnitude of the haemolysin response. The lymphocyte concentration in the blood perfusing the spleen was varied within wide limits to determine whether a migration of lymphocytes through the spleen influenced its ability to respond to antigenic stimulation. Normal spleens perfused for 3 h with blood containing a normal concentration of lymphocytes gave substantial haemolysin titres in the recipients when sheep erythrocytes had been added initially to the perfusate. Spleens depleted of lymphocytes in vivo by either X-irradiation or chronic drainage from a thoracic duct fistula produced very low titres after perfusion with lymphocyte-free blood and antigen, but their responses were restored by adding lymphocytes to the perfusate. The antibody response of normal spleens was significantly depressed following perfusion for 3 h with lymphocyte-free blood to which antigen had been added initially. However, normal responses were obtained in this experiment if the addition of antigen to the perfusate was delayed for 5 h and perfusion then continued for one further hour. The delay allowed lymphocytes to migrate from the normal spleens into the perfusate and to build up the concentration in the initially lymphocyte-free blood to normal levels. The conclusion drawn from this paradoxical finding was that the magnitude of the haemolysin response depended upon the concentration of lymphocytes in the perfusate at the time of antigen addition and not upon the total number of lymphocytes in the spleen. The rate of migration of small lymphocytes from the blood into the spleen was found to be directly proportional to the concentration of lymphocytes in the perfusate. The most acceptable explanation of all the data is that the magnitude of the haemolysin response is proportional to the concentration of lymphocytes in a limited compartment of the splenic lymphoid tissue into which lymphocytes have recently migrated from the blood. This compartment is probably located in the central area of the periarteriolar lymphocyte sheath.

Modification of transmitter release by electrical interference with motor nerve endings

Abstract: 1. A hyperpolarizing pulse applied through an external microelectrode to a nerve terminal during the falling phase of its action potential can suppress transmitter release; a depolarizing pulse applied during the same period can potentiate the release. 2. The facilitating action of calcium on transmitter release was studied by synchronizing the arrival of the nerve impulse with a rapid increment of the external calcium concentration. Using ionophoretic pulses of calcium, an effect could be produced with very little delay; sometimes the calcium pulse was effective when it preceded the arrival of the nerve impulse by only 1 to 2 ms.

The Classification of Modifiable Synapses and Their Use In Models For Conditioning

Abstract: Modifiable synapses are analysed theoretically. It is proved that they fall into three classes (called A, B and C), such that any two members of the same class can, with the aid of non-remembering elements that perform simple logical operations, replace one another in any net; but a member of class A cannot replace one of class B or C. A member of class B can replace one of class A or C only if non-logical elements, for example noise generators, are included in the net. Models are designed that use modifiable synapses and show the principal features of classical and operant conditioning. If the models are to be given the property of extinction, classical conditioning requires modifiable synapses of classes A and B or of class C, and operant conditioning of class B or C. It is argued that if the conditioning and memory mechanisms of the nervous system store information by means of modifiable synapses, some of these must be of class B or C.

The Repair of X-Ray Damage to the Deoxyribonucleic Acid in Micrococcus radiodurans: A Study of the Excision Process

Abstract: Following non-lethal anoxic X-irradiation the bacterium Micrococcus radiodurans suffers a delay in deoxyribonucleic acid ( DNA ) synthesis and during this delay components are released from the cellular DNA into the cytoplasm and into the surrounding culture medium. These events are subsequent to the radiochemical damage suffered by the macromolecular structure of the DNA which can be detected by physico-chemical means (Dean, Feldschreiber & Lett 1966). The rate at which material is released (or excised) from the DNA is inde­pendent of the X-ray dose and the kinetics of the process suggest that an enzyme is effecting the majority of the release and that it is present in limiting amounts. Studies of the excision products indicate that the enzyme is an exonuclease.