Biomedical subjects
Publications and source records attributed to A M Thomson.
Deoxyribonuclease activity was surveyed in 22 strains belonging to 12 species of rumen bacteria, with lambda bacteriophage DNA as substrate. Activity was readily detected in broken cell preparations from 15 of these strains. Particularly high levels of activity were present in cells and culture supernatant of all 5 strains of Bacteroides succinogenes, and 2 out of 6 strains of Bacteroides ruminicola, examined.
In isolated slices of hypothalamus, suprachiasmatic nucleus (SCN) neurons were recorded intracellularly. Blockade of Ca++ channels increased spike duration, eliminating an early component of the afterhyperpolarization (AHP) that followed evoked spikes. The duration and reversal potential of AHPs were, however, unaffected, suggesting that only an early, fast component of the AHP was Ca(++)-dependent. Unlike other central neurons that exhibit pacemaker activity, therefore, SCN neurons do not display a pronounced, long-lasting Ca(++)-dependent AHP. Extracellular Ba++ and intracellular Cs+ both revealed slow depolarizing potentials evoked either by depolarizing current injection, or by repolarization following large hyperpolarizations. They had different effects on the shape of spikes and the AHPs that followed them, however. Cs+, which blocks almost all K+ channels, dramatically reduced resting potential, greatly increased spike duration (to tens of milliseconds), and blocked AHPs completely. In contrast, Ba++ had little effect on resting potential and produced only a small increase in spike duration, depressing an early Ca(++)-dependent component and a later Ca(++)-independent component of the AHP. The relatively weak pacemaker activity of SCN neurons appears to involve voltage-dependent activation of at least one slowly inactivating inward current, which brings the cells to firing threshold and maintains tonic firing; both Ca(++)-dependent and Ca(++)-independent K+ channels, which repolarize cells after spikes and maintain interspike intervals; and Ca++ channels, which contribute to activation of Ca(++)-activated K+ currents and may also contribute to slow depolarizing potentials. In the absence of powerful synaptic inputs, SCN neurons express a pacemaker activity that is sufficient to maintain an impressively regular firing pattern. Slow, repetitive activation of optic input, however, increases local circuit activity to such an extent that the normal pacemaker potentials are overridden and firing patterns are altered. Since SCN neurons are very small and have large input resistances, they are particularly susceptible to synaptic input.
Frequencies of greater than 10(5) transformants per microgram DNA were achieved in Bacteroides ruminicola F101 by electroporation of cells under anaerobic conditions, using the 19.5 kbp tetracycline resistance plasmid pRRI4. Similar procedures gave frequences of 10(6) erythromycin resistant transformants per microgram DNA with the shuttle plasmid pDP1 (19 kbp) in Bacteroides uniformis. Transformation of B. uniformis occurred at a far lower frequency (10(3) micrograms) when pDP1 DNA was derived from E. coli rather than B. uniformis.
One class of excitatory amino-acid receptors, the N-methyl-D-aspartate (NMDA) receptors, mediates transmission at a small, but important, group of synapses in the neocortex. These receptors are implicated in neuronal plasticity during development in young mammals and in memory acquisition in adults. Recently, responses of isolated membrane patches to NMDA were shown to be greatly enhanced by glycine. This, together with the demonstration that the strychnine-insensitive glycine-binding site is distinct from, but linked to, the NMDA receptor has excited intense interest in glycine as a synaptic modulator. Before proposing a physiological function, however, it is important to determine whether glycine could enhance synaptic responses to NMDA receptor activation in intact, adult tissue. An earlier study failed to demonstrate enhancement of NMDA responses when glycine was applied and it was proposed that in intact tissue the high-affinity glycine site was already saturated by endogenous glycine. It remained possible that glycine concentrations can be maintained at low levels close to synaptic receptors. We have examined responses of neurons in slices of adult neocortex to focal applications of excitatory amino acids and glycine and report enhancement by glycine of NMDA receptor-mediated excitatory postsynaptic potentials.
One class of excitatory amino acid (EAA) receptor, the N-methyl-D-aspartate (NMDA) receptor, has excited enormous interest because of its unique pharmacological and physiological properties. Yet another property of this receptor/channel complex has recently emerged: its activation is greatly enhanced by and may even be dependent on, sub-micromolar concentrations of glycine.
1. In slices of rat neocortex, spike triggered averaging was employed to record in one neurone the excitatory postsynaptic potential (e.p.s.p.) generated by a spike in another, neighbouring neurone. When recorded at different membrane potentials, some of these e.p.s.ps exhibited a voltage relation typical of neuronal responses to N-methyl-D-aspartate (NMDA). 2. Selective NMDA antagonists reduced the amplitude of these e.p.s.ps, but had little effect on their early rising phase. In contrast, a less selective excitatory amino acid antagonist reduced all phases of the e.p.s.p. 3. By analyzing single axon e.p.s.ps we have been able to establish that the synaptic input to one cortical cell, delivered by a single presynaptic cortical cell, operates simultaneously via NMDA and non-NMDA amino acid receptors.
1. In rat isolated thalamic slices, intracellular recordings were made of responses to electrophoretically applied gamma-aminobutyric acid (GABA) in the absence and presence of sodium pentobarbitone (NaPb). 2. Responses to electrophoretically applied GABA were biphasic in the majority of neurones studied. They consisted of an early, negative-going and later, positive-going phase, when recorded close to reversal potential. 3. An increase in the GABA ejection current caused an enhancement of the late positive-going phase, along with a shift in the reversal potential of the whole response to a more positive value. These changes were accompanied by an increase in GABA-induced conductance (decrease in resistance) and an increase in the duration of the response. 4. Application of NaPb to the GABA response produced similar effects, namely an enhancement of the late positive-going phase, a shift in the reversal potential to a more positive value, an increase in conductance and an increase in duration. 5. GABA-mediated inhibitory postsynaptic potentials (i.p.s.ps) were evoked by electrical stimulation of the nucleus reticularis thalami. Application of NaPb caused an increase in the duration of the i.p.s.p. and a shift in its reversal potential to a more positive value.
In isolated slices of rat thalamus, inhibitory postsynaptic potentials evoked by electrical stimulation of the nucleus reticularis, were recorded intracellularly in relay neurons in the anterior part of the thalamus. These inhibitory postsynaptic potentials were found to have reversal potentials close to the resting potential of the recorded cell, to reduce neuronal excitability and to be sensitive to electrophoretic application of the GABA antagonists bicuculline and picrotoxin, indicating that they were GABA-activated, chloride mediated events. Voltage sensitive responses of relay neurons evoked by current injection and by inhibitory postsynaptic potentials were then compared. Hyperpolarizing current pulses and hyperpolarizing inhibitory postsynaptic potential trains elicited from membrane potentials positive to -70 mV resulted in rebound slow spike activation on repolarization. Depolarizing current pulses and depolarizing inhibitory postsynaptic potential trains evoked slow spikes when elicited from membrane potentials negative to -60 mV. There was, however, one major difference, the slow spikes evoked by inhibitory postsynaptic potentials were always delayed to the end of the train. Reversal potentials of evoked inhibitory postsynaptic potentials were found to depend on the potential at which the membrane was held immediately before the inhibitory postsynaptic potential was evoked, indicating that passive distribution of chloride ions contributes to their equilibrium potential. Evoked inhibitory postsynaptic potentials consisted of at least two components with different reversal potentials although current voltage relations indicated that similar decreases in membrane resistance were associated with both components and that they shifted approximately in parallel when inhibitory postsynaptic potentials were evoked from different holding potentials. Trains of GABA-mediated inhibitory postsynaptic potentials, similar to those recorded during spindling, will evoke slow spikes in almost all thalamic relay neurons irrespective of other synaptic inputs. This response will effectively synchronize burst firing in all cells receiving the same inhibitory input.
In isolated thalamic slices, responses of relay neurons to electrophoretically applied GABA were recorded intracellularly and compared with inhibitory postsynaptic potentials evoked by electrical stimulation of the reticularis nucleus of the thalamus. Both reduced the excitability of thalamic neurons and were biphasic in the majority of neurons studied, consisting of an early, negative-going and a later, positive-going component, when recorded close to reversal potential (mean reversal potentials -66.6 and -57.7 mV). Bicuculline and picrotoxin applied electrophoretically reduced conductance increases evoked by GABA in all neurons. The later, positive-going component was more sensitive to these antagonists (applied with submaximal doses) than the early component. Current-voltage relations for responses to GABA, like those for inhibitory postsynaptic potentials, were non-linear in the majority of neurons. In particular, there was a region of reduced slope resistance close to the reversal potential. Holding the membrane at a conditioning potential was found to change the subsequent response and its reversal potential. Positive holding potentials shifted reversal potentials in the positive direction only when GABA was applied during the conditioning period. Negative holding potentials were effective whether GABA was applied during the conditioning period or not. Recovery from these effects followed a similar time course at all membrane potentials tested. Injection of Cl- produced a positive shift in the reversal potential for both components of the response to GABA and of the evoked inhibitory postsynaptic potential. Inhibitory postsynaptic potentials evoked in thalamic relay neurons by stimulation of the nucleus reticularis resembled responses to GABA in their biphasic nature, reversal potentials and sensitivity to antagonists and to changes in intracellular chloride.
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Tetracycline resistance was transferred at frequencies between 10(-7) and 10(-6) per recipient cell in anaerobic matings between two strains of the strictly anaerobic rumen bacterium Bacteroides ruminicola. The donor strain, 223/M2/7, was a multiple-plasmid-bearing tetracycline-resistant strain from the ovine rumen, and the recipient, F101, was a rifampin-resistant mutant of B14, a bovine strain belonging to B. ruminicola subsp. brevis. Resistance transfer could occur in the presence of DNase, but not in dummy mating mixtures in which filtrate from a donor culture replaced donor cells. Acquisition of tetracycline resistance by the recipient was accompanied by the appearance of a 19.5-kilobase pair plasmid (designated pRRI4) which was homologous with a plasmid of similar size and restriction pattern present in the donor strain. A transconjugant (F115) carrying pRRI4 was also able to act as a donor of tetracycline resistance and plasmid DNA in matings with another recipient. Derivatives of F115 that had spontaneously lost tetracycline resistance lacked detectable plasmid DNA. It is concluded that pRRI4 mediated the transfer of tetracycline resistance. Transfer of resistance was not detectably enhanced by pregrowth of the donor in medium containing tetracycline. Transfer of tetracycline resistance was not detected from 223/M2/7 to a strain, 23 belonging to B. ruminicola subsp. ruminicola.
1. Using isolated slices of rat cingulate and sensorimotor cortex, intracellular recordings were obtained from pyramidal neurons in layer III. Simultaneous extracellular recordings were obtained from neurons in ventral layer III and layer IV. Spike-triggered averaging was employed to investigate synaptic connections from neurons in layers III/IV to pyramidal cells in layer III. 2. Of 701 simultaneously recorded pairs of neurons, comprising 699 extracellularly and 128 intracellularly recorded neurons, synaptic connections were demonstrated in 30 pairs. Of these, 29 were excitatory postsynaptic potentials (EPSPs) and 1, an inhibitory postsynaptic potential (IPSP). Single-axon EPSPs with a wide variety of amplitudes were recorded: the range recorded at membrane potentials between -68 and -72 mV was 0.079-2.3 mV. Comparing recordings obtained from different cells, EPSP amplitude was found to be independent of both the membrane resistance of the postsynaptic neuron and the EPSP time course; i.e., the largest EPSPs were not necessarily those recorded from neurons with the highest input resistance, nor those with the briefest time course. 3. Shape indices: width at half amplitude and rise-time, indicative of both proximal and distal synaptic locations were obtained. Normalized rise-times were between 0.1 and 2 times the membrane time constant and half-widths between 0.8 and 20 times. 4. The majority of postsynaptic neurons displayed nonlinear voltage relations typical of pyramidal neurons, and the contribution to EPSP shape of voltage-dependent currents was investigated. EPSP amplitude and duration were found to be dependent on membrane potential. The majority of single-axon EPSPs (26 of 29), increased in amplitude and duration with membrane depolarization over the range -95 - -50 mV, despite the significant decrease in driving force for the EPSP that would be expected to accompany such large depolarizations. This increase coincided with an increase in the amplitude of voltage responses to small injected current pulses. 5. It is concluded that the amplitude and time course of single-axon EPSPs recorded in cortical pyramidal somata are affected not only by the amplitude of the postsynaptic current and the location(s) of the synapse(s) relative to the soma, but also by voltage-dependent currents. The possibility that the increase in amplitude and duration of these EPSPs with membrane depolarization is due to N-methyl-D-aspartate receptor involvement is discussed.
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Colchicine induced multivalent formation is used to examine the effects of multivalent formation on chiasma frequency in Senecio squalidus. The increases observed are interpreted as the results of increases in numbers of pairing segments or decreases of chiasma interference following pairing partner exchange.
Using intracellular recordings from pyramidal neurons in isolated slices of rat cerebral cortex epileptiform discharges evoked (1) in the presence of gamma-aminobutyric acid antagonists, and (2) in the absence of Mg2+ were compared. Depolarization shift responses recorded in the presence of bath applied picrotoxin, or electrophoretically applied picrotoxin or bicuculline, were similar in many respects to depolarization shifts reported previously, except that they could be evoked by stimuli subthreshold for evoking discernible postsynaptic potentials in these experiments. Large depolarizations evoked by repetitive activation of an N-methylaspartate receptor mediated synapse in the absence of Mg2+, displayed several properties similar to those of depolarization shifts evoked in the presence of gamma-aminobutyric acid antagonists, i.e. similar shape, latency, inability to follow high repetition rates and a similar voltage relation, suggesting activation of the same cellular mechanism. "Slow spikes" evoked as part of the response to electrophoretically applied N-methylaspartate were augmented, i.e. they were replaced by larger, longer, more complex events, when gamma-aminobutyric acid antagonists were applied. The potentiated response, evoked in the absence of Mg2+, was dependent on the activation of an N-methylaspartate receptor mediated synapse and was blocked by N-methylaspartate antagonists. In contrast, depolarization shifts could be evoked in the presence of large doses of N-methylaspartate antagonists, when gamma-aminobutyric acid antagonists were applied. Spontaneous depolarizations similar to depolarization shifts were recorded when cells were exposed to low, tonic, electrophoretic applications of excitatory amino acids under control conditions. In addition, some potentiation of the N-methylaspartate receptor mediated excitatory postsynaptic potential was achieved in the presence of Mg2+ when cells were depolarized by 10-20 mV. Depolarization shifts evoked when bicuculline was applied electrophoretically to different parts of the dendritic field, some hundreds of microns from the soma, differed in shape, latency and time course and the depolarization shift evoked when bicuculline was applied at one site summed with the depolarization shift evoked when it was applied elsewhere. We conclude that different inputs are required to activate the responses evoked in the presence of gamma-aminobutyric acid antagonists and in the absence of Mg2+. The possibility that both involve activation of dendritic Ca2+ currents and that the magnitude of the response depends on the proportion of the dendritic field activated, is discussed.
Intracellular recordings from pyramidal neurones in isolated slices of rat cerebral cortex allowed a comparison of postsynaptic potentials and responses to electrophoretic application of excitatory amino acids. The responses of all neurones to N-methylaspartate displayed an unusual voltage relation and were associated with an apparent increase in membrane resistance, properties that were dependent on the presence of extracellular Mg2+. Responses to N-methylaspartate could be elicited only when the electrophoretic pipette was positioned close to the cell soma and were associated with generation of slow spikes which triggered bursts of fast spikes. In contrast, in the majority of neurones, responses to glutamate, aspartate, cysteate and cysteine sulphinate demonstrated a conventional voltage relation, were associated with a decrease in membrane resistance, evoked no slow spikes, were insensitive to extracellular Mg2+ concentrations between 1 and near 0 mM and could be evoked with small currents of amino acids when the electrophoretic pipette was greater than 100 micron from the cell soma. Responses to the five amino acids were tested with the N-methylaspartate antagonist 2-amino-5-phosphonovaleric acid. In the majority of cells, this antagonist blocked responses to N-methylaspartate at doses that had only a small effect on responses to the other amino acids. In these experiments it was also possible to confirm previous reports that ketamine and cyclazocine act as selective N-methylaspartate antagonists. In 4/63 neurones, responses to glutamate and aspartate and in 1/5 neurones one component of the response to cysteate, displayed properties similar to those of responses to N-methylaspartate. In one other neurone, large applications of cysteine sulphinate or glutamate could evoke slow spikes and fast spike bursts, a firing pattern that was sensitive to 2-amino-5-phosphonovalerate. The present experiments therefore demonstrate that all four naturally occurring amino acids tested can activate N-methylaspartate receptors and indicate that any one of these could be the transmitter at the N-methylaspartate receptor-mediated synapse on cortical pyramidal neurones. However, in the majority of neurones, these putative transmitters activated other receptor types preferentially. The possibility that this may result from the greater accessibility of non-N-methylaspartate receptors to electrophoretically applied agonists, is discussed.
Consultation between the clinicians and epidemiologists responsible for the Perinatal Mortality Surveys in Scotland and in the Northern Regional Health Authority in England showed that the classification of perinatal death introduced more than 30 years ago by Sir Dugald Baird still retained its utility, but that unintentional differences in the way cases were being classified had threatened the validity of temporal or geographical comparisons. To overcome this problem an effort has now been made to define the main terms used in this classification more precisely. To preserve continuity, the main structure of the original groupings has been retained; but the opportunity has been taken to adjust certain minor groups in conformity with recent ideas, and also to modify definitions to take into account the greatly improved prognosis for babies of very low birthweight. Otherwise, it is thought that subclassification of the main groups offers a better method of exploring new hypotheses than any radical alteration of the main groups themselves.
In isolated slices of rat cerebral cortex, intracellular recordings were obtained from pyramidal cells that were predominantly in layers II/III. These cells could be antidromically activated from the underlying white matter and had resting potentials of greater than -75 mV, action potentials with amplitudes of greater than 70 mV (measured from threshold), overshoots of 20-30 mV, and thresholds 20-30 mV positive to the resting potential. The responses of these cells to short (1-2 s) pulses of electrophoretically applied N-methylaspartate (NMA) decreased in amplitude with membrane hyperpolarization between -40 and -120 mV, and were associated with an apparent increase in membrane resistance when recorded in the presence of 1 mM-Mg2+. However, in the absence of Mg2+, responses to NMA increased progressively in amplitude with hyperpolarization and were associated with a decrease in membrane resistance. In addition to conventional excitatory post-synaptic potentials (e.p.s.p.s) and inhibitory post-synaptic potentials (i.p.s.p.s), electrical stimulation of the underlying white matter evoked a novel e.p.s.p. This e.p.s.p. displayed a similar voltage relation to the response evoked by NMA and was associated with an apparent increase in membrane resistance. When Mg2+ was removed from the bathing medium, the properties of the novel e.p.s.p. changed, and it displayed a conventional voltage relation and was associated with a decrease in membrane resistance. In the absence of Mg2+, novel e.p.s.p.s showed potentiation on low frequency repetitive stimulation (0.5-2 Hz). A fully potentiated response could evoke bursts of slow potentials each of which could evoke a burst of fast spikes. In contrast, the more conventional e.p.s.p.s and i.p.s.p.s evoked in pyramidal neurones were unaffected by reducing the Mg2+ concentration from 1.0 to near 0 mM and conventional e.p.s.p.s showed no potentiation on repetitive, low frequency repetition, even after several hours exposure to Mg2+-free medium. The NMA antagonists: 2-amino-5-phosphonovaleric acid, ketamine and cyclazocine, applied electrophoretically at doses that blocked responses to NMA, but which had little effect on responses to glutamate, blocked the novel e.p.s.p. and its potentiation.