Biomedical subjects

Andrew T Smith

Publications and source records attributed to Andrew T Smith.

17 recordsLinked to original sources

PubMed2006

A comment on the severity of the effects of non-white noise in fMRI time-series.

Analysis of fMRI time-series data is usually performed within the general linear model (GLM), and conclusions are based on the probability (p) of a false-positive voxel. In its simplest form, this process assumes that temporal noise in the time-series is random (white); if it is not, then an error in the estimation of p will occur. Various methods for correcting the problem have been developed, but they are not always used. Analysis packages vary in their ability to deal with this problem, and there is a lack of consistent advice to non-experts concerning whether this issue is serious or is small enough to be ignored. The variability of expert opinion partly reflects the fact that the magnitude of the error that occurs when estimating p in fMRI data is largely undocumented. In this discussion, aimed more at "neuroscience users" than methods experts, we try to clarify the position by documenting the scale of the error in a simple block-design experiment. Based on a fixed-effects group analysis of null datasets (8 participants scanned while at rest), we show that the magnitude of the problem can be very substantial. Without correction, it was necessary to set a nominal probability threshold of about p=0.000001 in order to achieve an actual probability of a false positive of 0.001. This means that if task-related activations were present in such a dataset, only if they reached the former, very low nominal p value could they be relied upon. With the use of standard correction methods, the error was largely removed. Our results suggest that some form of pre-whitening or correction for effects of non-white noise is essential in single-subject GLM analyses and fixed-effects group analyses of fMRI data. It is less important but probably prudent to apply a correction at the first level of random-effects group analyses. We also evaluate the effect of high-pass filtering. We find that the number of false positives in a fixed-effects analysis increases markedly as the filter cut-off frequency is increased. The sensitivity benefits derived from high-pass filtering may thus be partially offset by a significant increase in the number of false positives.

Adult↗

PubMed2006

FMRI adaptation reveals separate mechanisms for first-order and second-order motion.

A key unresolved debate in human vision concerns whether we have two different low-level mechanisms for encoding image motion. Separate neural mechanisms have been suggested for first-order (luminance modulation) and second-order (e.g., contrast modulation) motion in the retinal image but a single mechanism could handle both. Human functional magnetic resonance imaging (fMRI) has not so far convincingly revealed separate anatomical substrates. To examine whether two separate but co-localized mechanisms might exist, we used the technique of fast fMRI adaptation. We found direction-selective adaptation independently for each type of motion in the motion area V5/MT+ of the human brain. However, there was a total absence of cross-adaptation between first-order and second-order motion stimuli. This was true in both of the two subcomponents of MT+ (MT and MST) and similar results were found in V3A. This pattern of adaptation was consistent with psychophysical measurements of detection thresholds in similar stimulus sequences. The results provide strong evidence for separate neural populations that are responsible for detecting first- and second-order motion.

Adaptation, Physiological↗

PubMed2006

Assessment of stimulus-induced changes in human V1 visual field maps.

Visual cortex contains a set of field maps in which nearby scene points are represented in the responses of nearby neurons. We tested a recent hypothesis that the visual field map in primary visual cortex (V1) is dynamic, changing in response to stimulus motion direction. The original experimental report replicates, but further experimental and analytical investigations do not support, the interpretation of the results. The V1 map remains invariant when measured using stimuli moving in different directions. The measurements can be explained by small and systematic response amplitude differences that arise when probing with stimuli moving in different directions.

Algorithms↗

PubMed2006

Time perception: manipulation of task difficulty dissociates clock functions from other cognitive demands.

Previous studies suggest the involvement in timing functions of a surprisingly extensive network of human brain regions. But it is likely that while some of these regions play a fundamental role in timing, others are activated by associated task demands such as memory and decision-making. In two experiments, time perception (duration discrimination) was studied under two conditions of task difficulty and neural activation was compared using fMRI. Brain activation during duration discrimination was contrasted with activation evoked in a control condition (colour discrimination) that used identical stimuli. In the first experiment, the control task was slightly easier than the time task. Multiple brain areas were activated, in line with previous studies. These included the prefrontal cortex, cerebellum, inferior parietal lobule and striatum. In the second experiment, the control task was made more difficult than the time task. Much of the differential time-related activity seen in the first experiment disappeared and in some regions (inferior parietal cortex, pre-SMA and parts of prefrontal cortex) it reversed in polarity. This suggests that such activity is not specifically concerned with timing functions, but reflects the relative cognitive demands of the two tasks. However, three areas of time-related activation survived the task-difficulty manipulation: (i) a small region at the confluence of the inferior frontal gyrus and the anterior insula, bilaterally, (ii) a small portion of the left supramarginal gyrus and (iii) the putamen. We argue that the extent of the timing "network" has been significantly over-estimated in the past and that only these three relatively small regions can safely be regarded as being directly concerned with duration judgements.

Adolescent↗

PubMed2006

Transfer of interfered motor patterns to self from others.

Previous behavioural and neuroimaging data on humans demonstrated that kinematics and the level of brain activity vary according to whether participants reach towards and grasp a target object presented in isolation or flanked by a distractor object. Here we seek to explore whether a differential activation can be revealed by the mere observation of another person grasping an object in isolation or alongside a distractor. To this end we used event-related functional magnetic resonance imaging to localize neural activity related to action observation that was influenced by the presence of a distractor object. We found that observing a human model reaching-to-grasp a three-dimensional target alongside a distractor elicits a differential level of activation in a network of areas typically involved during action observation: the dorsal sectors of the premotor cortex and the inferior frontal gyrus. Whereas our previous understanding of the human action observation system has been restricted to actions directed to single objects, we provide compelling evidence that areas within this network modulate with respect to the context in which the observed action takes place. This may prove to be a fundamental process for our understanding of how others' actions can be represented at a neural level.

Adult↗

PubMed2006

When gaze turns into grasp.

Previous research has provided evidence for a neural system underlying the observation of another person's hand actions. Is the neural system involved in this capacity also important in inferring another person's motor intentions toward an object from their eye gaze? In real-life situations, humans use eye movements to catch and direct the attention of others, often without any accompanying hand movements or speech. In an event-related functional magnetic resonance imaging study, subjects observed videos showing a human model either grasping a target object (grasping condition) or simply gazing (gaze condition) at the same object. These two conditions were contrasted with each other and against a control condition in which the human model was standing behind the object without performing any gazing or grasping action. The results revealed activations within the dorsal premotor cortex, the inferior frontal gyrus, the inferior parietal lobule, and the superior temporal sulcus in both "grasping" and "gaze" conditions. These findings suggest that signaling the presence of an object through gaze elicits in an observer a similar neural response to that elicited by the observation of a reach-to-grasp action performed on the same object.

Adult↗

PubMed2005

Attentional modulation in the human visual cortex: the time-course of the BOLD response and its implications.

Throughout the visual areas of the brain, the sensory response to a stimulus is enhanced by attending to the stimulus. Neurophysiological studies in primates show that such enhancement is marked in posterior parietal cortex and some anterior occipital areas, but much more modest in the earliest processing stages, such as the primary visual cortex (V1). In contrast, human fMRI studies show large and robust attentional modulation in all visual areas, including V1. We investigate the possibility that, in the case of fMRI, the BOLD (blood oxygen level dependent) response may be increased not only by local attention-related increases in neural activity, but also by local blood-flow increases caused by remote control systems that anticipate an impending need for oxygen at the attended location. Such changes could be much more rapid than the rather slow response to oxygenation change that typifies the BOLD response. We have employed a paradigm that isolates the component of the BOLD response due to attentional modulation and the component due to the mere presence of a visual stimulus. The results show that the temporal profiles of the BOLD responses in human V1 to the onset of a stimulus and to the onset of attention are extremely similar. The time-course of the attention-related BOLD response is not consistent with the action of remote, anticipatory control mechanisms and suggests that the modulatory effect of attention seen in human V1 with fMRI probably reflects genuine changes in local neural activity that are considerably larger than in non-human primates.

Adult↗

PubMed2005

Spectroscopic and kinetic properties of the horseradish peroxidase mutant T171S. Evidence for selective effects on the reduced state of the enzyme.

Studies on horseradish peroxidase C and other haem peroxidases have been carried out on selected mutants in the distal haem cavity providing insight into the functional importance of the distal residues. Recent work has demonstrated that proximal structural features can also exert an important influence in determining the electronic structure of the haem pocket. To extend our understanding of the significance of proximal characteristics in regulating haem properties the proximal Thr171Ser mutant has been constructed. Thr171 is an important linking residue between the structural proximal Ca2+ ion and the proximal haem ligand, in particular the methyl group of Thr171 interdigitates with other proximal residues in the core of the enzyme. Although the mutation induces no significant changes to the functional properties of the enzyme, electronic absorption and resonance Raman spectroscopy reveal that it has a highly selective affect on the reduced state of the enzyme, effectively stabilizing it, whilst the electronic properties of the Fe(III) state unchanged and essentially identical to those of the native protein. This results in a significant change in the Fe2+/Fe3+ redox potential of the mutant. It is concluded that the unusual properties of the Thr171Ser mutant reflect the loss of a structural restraint in the proximal haem pocket that allows 'slippage' of the proximal haem ligand, but only in the reduced state. This is a remarkably subtle and specific effect that appears to increase the flexibility of the reduced state of the mutant compared to that of the wild-type protein.

Horseradish Peroxidase↗

PubMed2004

Negative BOLD in the visual cortex: evidence against blood stealing.

The positive BOLD (blood oxygen level-dependent) response elicited in human visual cortex by a localized visual stimulus is accompanied by a reduction in the BOLD response in regions of the visual cortex that represent unstimulated locations in the visual field. We have suggested previously that this negative BOLD reflects attention-related suppression of neural activity, but it might also be explained in terms of "blood stealing," i.e., hemodynamic changes that have no neural correlate. We distinguish two possible hemodynamic effects of this type: (1). blood flow reduction caused by locally reduced pressure in vessels that share their blood supply with nearby dilated vessels; and (2). blood flow reduction caused by active constriction of vessels under neural control. The first is ruled out as an explanation of negative BOLD by showing that a visual stimulus that stimulates primary visual cortex in one hemisphere can cause extensive suppression in the other hemisphere i.e., it is not a local phenomenon. Negative BOLD most likely reflects suppression of neural activity, but could also reflect an active blood flow control system.

Attention↗

PubMed2003

Effects of phthalic anhydride modification on horseradish peroxidase stability and activity.

Phthalic anhydride (PA) modification stabilizes horseradish peroxidase (HRP) by reversal of the positive charge on two of HRP's six lysine residues. Native and PA-HRP had half-inactivation temperatures of 51 and 65 degrees C and half-lives at 65 degrees C of 4 and 17 min, respectively. PA-HRP was more resistant to dimethylformamide at room temperature and tetrahydrofuran at 60 degrees C and to unfolding by heat, guanidine chloride, EDTA, and the reducing agent tris(2-carboxyethyl)phosphine hydrochloride. Binding of the hydrophobic probe Nile Red to the native enzyme and to PA-HRP was similar. The kinetics of both HRPs with the substrates ABTS, ferrocyanide, ferulic acid, and indole-3-propionic acid were measured, as was binding of the inhibitor benzhydroxamic acid. Small improvements in the catalytic properties were detected.

Binding Sites↗

PubMed2003

Evolutionary relationships among strains of Mycobacterium tuberculosis with few copies of IS6110.

Molecular typing of Mycobacterium tuberculosis by using IS6110 shows low discrimination when there are fewer than five copies of the insertion sequence. Using a collection of such isolates from a study of the epidemiology of tuberculosis in London, we have shown a substantial degree of congruence between IS6110 patterns and both spoligotype and PGRS type. This indicates that the IS6110 types mainly represent distinct families of strains rather than arising through the convergent insertion of IS6110 into favored positions. This is supported by identification of the genomic sites of the insertion of IS6110 in these strains. The combined data enable identification of the putative evolutionary relationships of these strains, comprising three lineages broadly associated with patients born in South Asia (India and Pakistan), Africa, and Europe, respectively. These lineages appear to be quite distinct from M. tuberculosis isolates with multiple copies of IS6110.

Africa↗

PubMed2003

Surround modulation measured with functional MRI in the human visual cortex.

Visual context profoundly influences 1) the responses of mammalian visual neurons and 2) the perceptual sensitivity of human observers to localized visual stimuli. We present data from functional MRI studies demonstrating contextual modulation in the human visual cortex. Subjects viewed a circular grating patch that was continuously present. A surround grating was added in an ON-OFF block design to reveal its effect on the central region. Stimulus-correlated activation was quantified and visualized on a flattened map of the occipital gray matter. Modulation was measured in a region of interest activated by the central grating alone. The observed effects were predominantly suppressive, consistent with the effects typically found in single neurons and perception. Suppression was greatest when the surround and center had the same orientation and was reduced or absent when it was orthogonal. When spatial phase was manipulated, suppression was greatest for in-phase center/surround gratings and much reduced or reversed (facilitation) for opposite-phase stimuli. With eccentric stimulus presentation, suppression was reduced and facilitation became more common. The findings provide a direct demonstration of the existence of powerful and stimulus-specific surround effects in human visual cortex.

Adult↗

PubMed2002

Structural analysis of the two horseradish peroxidase catalytic residue variants H42E and R38S/H42E: implications for the catalytic cycle.

The crystal structures of horseradish peroxidase C (HRPC) active-site mutants H42E and R38S/H42E co-crystallized with benzhydroxamic acid (BHA) and ferulic acid (FA), respectively, have been solved. The 2.5 A crystal structure of the H42E-BHA complex reveals that the side-chain O atoms of Glu42 occupy positions that are very similar to the positions of the two side-chain N atoms of the distal histidine in the wild-type HRPC-BHA structure. The mutation disturbs the hydrogen-bonding network extending from residue 42 to the distal calcium ion and results in the absence of the water molecule that is usually ligated to this ion in plant peroxidases. Consequently, the distal calcium ion is six- rather than seven-coordinated. In the 2.0 A R38S/H42E structure the position of Glu42 is different and no FA is observed in the distal haem pocket. This is a consequence of the absence of the Arg38 side chain, which limits the flexibility of the Glu42 side chain and modulates its acidity, making it unsuitable as a general acid-base catalyst in the reaction cycle. The water ligated to the distal calcium ion is present, showing that the wild-type distal hydrogen-bonding network is preserved. These results show why a glutamic acid residue can substitute for the conserved distal histidine in HRPC and that Arg38 plays a significant role in controlling the positioning and ionization state of the residue at position 42. Furthermore, these structures indicate that changes in the distal cavity are conveyed through the distal hydrogen-bonding network to the distal calcium site.

Amino Acid Sequence↗

PubMed2002

The catalytic pathway of horseradish peroxidase at high resolution.

A molecular description of oxygen and peroxide activation in biological systems is difficult, because electrons liberated during X-ray data collection reduce the active centres of redox enzymes catalysing these reactions. Here we describe an effective strategy to obtain crystal structures for high-valency redox intermediates and present a three-dimensional movie of the X-ray-driven catalytic reduction of a bound dioxygen species in horseradish peroxidase (HRP). We also describe separate experiments in which high-resolution structures could be obtained for all five oxidation states of HRP, showing such structures with preserved redox states for the first time.

Catalysis↗

PubMed2002

Spectroscopic characterization of mutations at the Phe41 position in the distal haem pocket of horseradish peroxidase C: structural and functional consequences.

Three mutants of horseradish peroxidase isoenzyme C (HRPC) have been constructed in which the conserved distal aromatic residue Phe(41) has been substituted by Trp, Val or Ala and the properties of the mutant proteins have been compared with that of the wild-type. The ferric and ferrous states have been studied by resonance Raman, electronic absorption and Fourier-transform infrared spectroscopies, together with their respective fluoride and CO complexes as probes for the integrity of the distal haem-pocket hydrogen-bonding network. The catalytic properties of the mutants, most notably the HRPC-mutant Phe(41)-->Trp (F41W) variant, were also affected. Structural modelling suggests that the bulky indole group of the F41W mutant blocks the distal cavity, inhibiting the binding of fluoride and CO to the haem iron, severely impairing the reaction of the enzyme with H(2)O(2) to form Compound I. Substitution with the smaller side-chain residues Val or Ala resulted in a 2-fold increase in the affinity of the mutants for the aromatic donor benzhydroxamic acid (BHA) compared with the wild-type, whereas the sterically hindered F41W mutant was not able to bind BHA at all. All the mutations studied increased the amount of a ferric six-coordinate aquo-high-spin species. On the other hand, the similarity in the Fe-Im stretching frequencies of the mutants and wild-type protein suggests that the distal haem-pocket mutations do not cause any substantive changes on the proximal side of the haem. Spectra of the HRPC mutant Phe(41)-->Ala-CO and the HRPC mutant Phe(41)-->Val-CO complexes strongly suggested a weakening of the interaction between CO and Arg(38) due to a secondary rearrangement of the haem relative to helix B. The effects observed for these HRP mutants were somewhat different from those noted recently for the analogous Coprinus cinereus peroxidase (CIP) mutants, particularly the Trp mutant. These differences can be reconciled in part as being due to the smaller size of the distal cavity of HRP compared with that of CIP.

Amino Acid Substitution↗

PubMed2002

Reactions of the class II peroxidases, lignin peroxidase and Arthromyces ramosus peroxidase, with hydrogen peroxide. Catalase-like activity, compound III formation, and enzyme inactivation.

The reactions of the fungal enzymes Arthromyces ramosus peroxidase (ARP) and Phanerochaete chrysosporium lignin peroxidase (LiP) with hydrogen peroxide (H(2)O(2)) have been studied. Both enzymes exhibited catalase activity with hyperbolic H(2)O(2) concentration dependence (K(m) approximately 8-10 mm, k(cat) approximately 1-3 s(-1)). The catalase and peroxidase activities of LiP were inhibited within 10 min and those of ARP in 1 h. The inactivation constants were calculated using two independent methods; LiP, k(i) approximately 19 x 10(-3) s(-1); ARP, k(i) approximately 1.6 x 10(-3) s(-1). Compound III (oxyperoxidase) was detected as the majority species after the addition of H(2)O(2) to LiP or ARP, and its formation was accompanied by loss of enzyme activity. A reaction scheme is presented which rationalizes the turnover and inactivation of LiP and ARP with H(2)O(2). A similar model is applicable to horseradish peroxidase. The scheme links catalase and compound III forming catalytic pathways and inactivation at the level of the [compound I.H(2)O(2)] complex. Inactivation does not occur from compound III. All peroxidases studied to date are sensitive to inactivation by H(2)O(2), and it is suggested that the model will be generally applicable to peroxidases of the plant, fungal, and prokaryotic superfamily.

Catalase↗

PubMed2002

Expression of Drosophila melanogaster xanthine dehydrogenase in Aspergillus nidulans and some properties of the recombinant enzyme.

Recent crystal structures of xanthine dehydrogenase, xanthine oxidase and related enzymes have paved the way for a detailed structural and functional analysis of these enzymes. One problem encountered when working with these proteins, especially with recombinant protein, is that the preparations tend to be heterogeneous, with only a fraction of the enzyme molecules being active. This is due to the incompleteness of post-translational modification, which for this protein is a complex, and incompletely understood, process involving incorporation of the Mo and Fe/S centres. The enzyme has been expressed previously in both Drosophila and insect cells using baculovirus. The insect cell system has been exploited by Iwasaki et al. [Iwasaki, Okamoto, Nishino, Mizushima and Hori (2000) J. Biochem (Tokyo) 127, 771-778], but, for the rat enzyme, yields a complex mixture of enzyme forms, containing around 10% of functional enzyme. The expression of Drosophila melanogaster xanthine dehydrogenase in Aspergillus nidulans is described. The purified protein has been analysed both functionally and spectroscopically. Its specific activity is indistinguishable from that of the enzyme purified from fruit flies [Doyle, Burke, Chovnick, Dutton, Whittle and Bray (1996) Eur. J. Biochem. 239, 782-795], and it appears to be more active than recombinant xanthine dehydrogenase produced with the baculovirus system. EPR spectra of the recombinant Drosophila enzyme are reported, including parameters for the Fe/S centres. Only a very weak "Fe/SIII" signal (g(1,2,3), 2.057, 1.930, 1.858) was observed, in contrast to the strong analogous signal reported for the enzyme from baculovirus. Since this signal appears to be associated with incomplete post-translational modification, this is consistent with relatively more complete cofactor incorporation in the Aspergillus-produced enzyme. Thus we have developed a recombinant expression system for D. melanogaster xanthine dehydrogenase, which can be used for the production of site-specific mutations of this enzyme.

Animals↗