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Biomedical subjects

J F Morrison

Publications and source records attributed to J F Morrison.

At least 19 recordsLinked to original sources
PubMed1992

Human dihydrofolate reductase: reduction of alternative substrates, pH effects, and inhibition by deazafolates.

The kinetics of the NADPH-dependent reduction of 7,8-dihydrofolate, folate, and 7,8-dihydrobiopterin by human dihydrofolate reductase have been examined over the pH range from 4.0 to 9.5. The V and V/K profiles obtained with the three substrates indicate that a single ionizing residue at the active site of the enzyme must be protonated for catalysis. Both the maximum velocity of the reactions and the rate of interaction of the substrates with the enzyme-NADPH complex decrease in the order dihydrofolate greater than dihydrobiopterin much greater than folate. From the pK values of the V/K profiles, it can be concluded that, while dihydrofolate behaves as a sticky substrate and dihydrobiopterin exhibits slight stickiness, folate is not a sticky substrate. Further support for this conclusion comes from the results of deuterium isotope effects. The pK values obtained from both the V and V/Kfolate profiles are similar to the intrinsic pK value of 5.6 for both the free enzyme and the enzyme-NADPH complex. The folate analogue, 5-deazafolate, is not a substrate, but it undergoes strong interaction with the enzyme. This interaction, which is enhanced by the presence of NADPH, is due to protonation of the bound ligand that does not involve the single ionizing group at the active center of the enzyme. Difference spectra yield evidence for the protonation of bound 5-deazafolate and show that, on binding to the enzyme-NADPH complex, the pK of the N-8 atom is raised to about 10 from a value of about 4 in solution. The results are in accord with those of a recent paper on the three-dimensional structure of the enzyme-5-deazafolate complex [Davies, J.F., Delcamp, T.J., Prendergast, N.J., Ashfors, V.A., Freisheim, J.H., & Kraut, J. (1990) Biochemistry 29, 9467-9479] which indicate that there is hydrogen bond formation between N-8 of the ligand and the carbonyl group of Ile-7. However, the present findings do not support the idea that bound 5-deazafolate resembles the transition-state complex for folate reduction. Quinazolines also interact strongly with the enzyme but in a pH-independent manner. The dissociation constants for the binary complexes are an order of magnitude lower than that for the binding to the enzyme of unprotonated 5-deazafolate. This difference reflects the hydrophobic nature of the amino acid residues at the active site that are near the N-5 and N-8 nitrogens of bound pterins.

Folic Acid↗
PubMed1992

Response to nebulized ipratropium bromide and terbutaline in acute severe asthma.

Outpatient studies on asthmatics have shown that inhaled anti-cholinergic agents decrease in efficacy as FEV1 falls. To determine whether there are changes in response to inhaled anti-cholinergics during acute bronchoconstriction we have examined the effects of nebulized ipratropium and terbutaline in nine hospitalized patients recovering from acute severe asthma. At 6 a.m. each day throughout the admission, baseline PEFR was recorded. Ipratropium bromide, 1 mg, was nebulized and PEFR measured again 1 h later. Following this, terbutaline, 5 mg, was nebulized with further measurement of PEFR 15 min after nebulization. Results were analysed by paired t-tests. Mean baseline PEFR rose from 157 l m-1 on patients worst day to 300 l m-1 on their best day (P less than 0.01). Ipratropium improved mean PEFR by 55 l m-1 and 42 l m-1 on patients worst and best days respectively (P less than 0.01). Subsequent terbutaline improved mean PEFR on patients worst day by 23 l m-1 (P less than 0.01) but only by a non-significant 4 l m-1 on their best day (P = 0.09). Hence, ipratropium produced 96% of total bronchodilatation when baseline was highest, but achieved only 71% of total response when baseline was lowest, a highly significant change in response (P less than 0.01). We conclude that in acute severe asthma as baseline PEFR rises response to inhaled ipratropium improves, compared with total response to combined ipratropium followed by terbutaline.

Acute Disease↗
PubMed1992

Oesophageal motility, luminal pH, and electrocardiographic-ST segment analysis during spontaneous episodes of angina like chest pain.

The relation between oesophageal motility, luminal pH, and spontaneous pain events in 47 patients with recurrent angina like pain and normal coronary arteries was investigated. Preliminary investigation by conventional station pull through manometry (SPTM), was followed by a 24 hour period of ambulatory oesophageal motility and luminal pH monitoring. Computerised analysis of motility and pH data recorded during chest pain was then compared with pre-elected control samples taken before and after symptoms. Concurrent real time electrocardiographic (ECG)-ST segment analysis was performed to catalogue any ECG-STT wave changes indicative of myocardial ischaemia. SPTM showed a high group percentage incidence of simultaneous contractions (mean (SD) 11.1 (2.3)%) and a raised lower oesophageal sphincter tone (57.4 (15.2) mm Hg). During ambulatory monitoring, 35 patients experienced one or more episode of angina, providing a total of 59 pain events, although no significant change in group motility and reflux parameters peculiar to episodes of chest pain were found. Ischaemic ECG changes were detected in 10 (21%) patients, but were accompanied by pain in only two. Independent analysis of the ECG traces corresponding to these purported ischaemic ECG events determined them unequivocal in three patients and probable in a further two. No apparent correlation was noted, however, between these ECG events and corresponding patterns of motility or reflux.

Adult↗
PubMed1992

Diurnal variation in FEV1 after heart-lung transplantation.

We have examined the diurnal variation in forced expiratory volume in one second (FEV1) in 25 heart-lung transplantation patients over a four week period in order to study the pathophysiological mechanisms underlying the increased mortality and morbidity which occurs at night in asthma. These patients do not have pulmonary autonomic nervous reflexes, but often have muscarinic receptor hypersensitivity. They also develop mixed cell infiltration of the lung tissue in the course of infection or rejection. Thus, they show many features in common with asthma. Seventeen patients (68%) showed a significant diurnal variation in airway calibre (mean amplitude of FEV1 was 4.6% (SD 3.7%)), which is similar to that reported in normal adults. One patient had a diurnal variation of 34.5% during an episode of rejection. This variation fell to 6.9% after steroid therapy, a change often seen in asthma. There was a correlation between increased amplitude of the variation and the presence in transbronchial biopsies of airway submucosal eosinophils and lymphocytes, associated with a histological diagnosis of acute rejection and with epithelial damage. No association was seen with muscarinic receptor sensitivity. The variation in FEV1 showed no alteration from the normal day/night synchronization, and the peak values were around 1300 h. We conclude that the diurnal variation in FEV1 after heart-lung transplantation is not dependent on autonomic nerve reflexes or muscarinic receptor sensitivity, but is related to the consequences of inflammation described above.

Adult↗
PubMed1992

Assessing physiological benefit from domiciliary nebulized bronchodilators in severe airflow limitation.

In steroid resistant chronic obstructive pulmonary disease (COPD) we assessed the effect of q.i.d. domiciliary nebulized fenoterol (F) 1.25 mg and ipratropium (I) 0.5 mg for three weeks in a placebo-controlled, randomized, double-blind, crossover study. The twenty patients studied (mean forced expiratory volume in one second (FEV1) 0.8 l) all showed less than 20% increase in FEV1 to 200 micrograms inhaled salbutamol (S) and less than 20% increase in peak expiratory flow rate (PEFR) after 2 weeks prednisolone therapy. Respiratory function tests, 5 min walking distance (5 MWD), visual analogue scales (VAS) for breathlessness, oxygen cost diagrams and reversibilities were performed weekly for three weeks with patients on their usual therapy, after three weeks domiciliary F+I, after three weeks saline and, finally, after a further three weeks on usual therapy again. Primary end-points, selected prior to unblinding, were mean home twice daily PEFR, trapped gas volume, FEV1 and 5 MWD. Home PEFR rose from 164 l.min-1 on saline to 196 l.min-1 on F+I (p = 0.0001). Secondary end-point analysis revealed a fall in home inhaler usage and a rise in VAS. Using the criterion of +15% and greater than 20 l.min-1 increase in home PEFR, 11 out of 20 patients had a "positive" trial. We suggest that such patients, but not others, benefit from long-term, nebulized beta 2-agonist and ipratropium. Trials using home PEFR recordings should be used to identify them.

Aged↗
PubMed1991

Effects of visceral distension on the activities of neurones receiving cutaneous inputs in the rat lumbar dorsal horn; comparison with effects of remote noxious somatic stimuli.

(1) Unitary extracellular recordings were made from 92 lumbar dorsal horn neurones in urethane-anaesthetised rats. These neurones were classed as 'noxious-only' (4), 'non-noxious-only' (33) or 'convergent' (55) by their responses to stimulation of their cutaneous receptive fields on the ipsilateral hindpaw. (2) Distension of abdominal viscera (colon, urinary bladder) depressed the activities of the vast majority (93%) of the convergent neurones but of only one other cell (a non-noxious-only neurone). Similarly, noxious stimulation of widespread somatic structures depressed activity in all but one of the convergent neurones but in only 3 other cells (one non-noxious- and two noxious-only neurones). One or other of these procedures also excited 3 cells--one convergent neurone responding to distension of the colon, another to stimulation of widespread somatic structures and one non-noxious-only neurone being excited by stimulation on the contralateral hindpaw. (3) The inhibitory effects of the noxious somatic stimuli were very like those described previously and termed 'diffuse noxious inhibitory controls' (DNIC) and it seems likely that the effects of the visceral stimuli were also manifestations of DNIC, particularly in view of their similar, nearly total, specificity to convergent neurones. There were however, some small differences in the extent and temporal evolution of the inhibitory effects of the visceral and of the somatic stimuli--the visceral stimuli generally producing weaker inhibitions with slower rates of onset and recovery. It is proposed that these differences may have reflected different amounts and patterns of activity in the relevant primary afferent fibres rather than being due to different central neural mechanisms. (4) These results and the likely explanation that the effects of the visceral stimuli were mediated by a diffuse mechanism should be taken into account when interpreting the results of other studies in which inhibitory effects are produced by visceral stimulation.

Afferent Pathways↗
PubMed1991

pH dependency of the reactions catalyzed by chorismate mutase-prephenate dehydrogenase from Escherichia coli.

The variation with pH of the kinetic parameters associated with the mutase and dehydrogenase reactions catalyzed by chorismate mutase-prephenate dehydrogenase has been determined with the aim of elucidating the role that ionizing amino acid residues play in binding and catalysis. The pH dependency of log V for the dehydrogenase reaction shows that the enzyme possesses a single ionizing group with a pK value of 6.5 that must be unprotonated for catalysis. This same group is observed in the V/Kprephenate, as well as in the V/KNAD, profile. The V/Kprephenate profile exhibits a second ionizing residue with a pK value of 8.4 that must be protonated for the binding of prephenate to the enzyme. For the mutase reaction, the V/Kchorismate profile indicates the presence of three ionizing residues at the active site. Two of these residues, with similar pK values of about 7, must be protonated, while the third, with a pK value of 6.3, must be unprotonated. It can be concluded that all three groups are concerned with the binding of chorismate to the enzyme since the maximum velocity of the mutase reaction is essentially independent of pH. This conclusion is confirmed by the finding that the Ki profile for the competitive inhibitor, (3-endo,8-exo)-8-hydroxy-2-oxabicyclo[3.3]non-6-ene-3,5-dicarboxylic acid, shows the same three ionizing groups as observed in the V/Kchorismate profile. By contrast, the Ki profile for carboxyethyldihydrobenzoate shows only one residue, with a pK value of 7.3, that must be protonated for binding of the inhibitor.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗
PubMed1991

Kinetic studies on chorismate mutase-prephenate dehydrogenase from Escherichia coli: models for the feedback inhibition of prephenate dehydrogenase by L-tyrosine.

Kinetic studies have been undertaken to elucidate the mechanism of the allosteric inhibition by tyrosine of the prephenate dehydrogenase activity of the bifunctional dimeric enzyme chorismate mutase-prephenate dehydrogenase. The effect of tyrosine on the initial velocity of the reactions in the presence of both prephenate and the alternative substrate, 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate, have been determined. In addition, investigations have been made of the effect of tyrosine on the inhibition of the reaction by the inhibitory analogues of prephenate, (4-hydroxyphenyl)pyruvate, and (carboxyethyl)-1,4-dihydrobenzoate. The results of the double inhibition experiments indicate clearly that the enzyme possesses a distinct allosteric site for the binding of tyrosine. The initial velocity data obtained with both substrates have been fitted to the rate equations that describe a wide range of models. From a comparison of the results obtained from studies with the two substrates, and with a knowledge of the value for the dissociation constant of the tyrosine-enzyme complex, definitive conclusions have been reached about the mechanism of the allosteric inhibition. It is concluded that tyrosine combines twice at allosteric sites and in an antisynergistic fashion, while prephenate reacts at both active sites of the dimeric enzyme as well as weakly at one of the allosteric sites. It appears that the latter is simple competition reaction that affects neither the binding of prephenate at the active site nor the rate of product formation. The model also predicts the formation of an active tyrosine-enzyme-prephenate complex that yields product at a much slower rate than does the enzyme-prephenate complex.(ABSTRACT TRUNCATED AT 250 WORDS)

Allosteric Regulation↗
PubMed1991

Further evidence for the absence of a descending cholinergic projection from the brainstem to the spinal cord in the rat.

Serotonergic and catecholaminergic neurons are known to project from the brainstem to the spinal cord. However, evidence for a bulbo-spinal projection that is cholinergic is sparse despite immunocytochemical and physiological evidence for a cholinergic influence on the cord. In this study we examined the possibility of a direct cholinergic bulbo-spinal projection in the rat using a combination of retrograde axonal tracing techniques and choline acetyltransferase immunocytochemistry. Although many cells were found to project to the cord from the brainstem, none were identified as being cholinergic, confirming previous evidence that the cholinergic innervation of the cord is intrinsic.

Animals↗
PubMed1991

Differential neuropeptide expression after visceral and somatic nerve injury in the cat and rat.

The expression of neuropeptides galanin, vasoactive intestinal polypeptide (VIP) and substance P was compared after injury to somatic (sciatic, pudendal) and visceral (pelvic) nerves. Studies in normal rats and the mutant rat 'mutilated foot' suggested that galanin increases in sensory but not sympathetic fibres after sciatic nerve injury, while VIP appears to increase in both sensory and sympathetic fibres, and substance P to decrease in sensory fibres. A direct comparison of neuropeptide changes after somatic and visceral nerve injury was made in the cat dorsal sacral spinal cord, where both pudenal (somatic) and pelvic (visceral) afferents terminate. Four weeks after pudendal nerve transection in the cat there was an increase of VIP and galanin but decrease of substance P in the dorsal sacral cord, similar to the changes in lumbar dorsal cord after sciatic nerve section in the rat. In contrast, 4 weeks after pelvic nerve transection in the cat, galanin was unchanged in the ipsilateral dorsal sacral spinal cord, whereas VIP is known to decrease markedly and substance P to remain unchanged. There is thus differential peptide expression before and after injury in somatic and visceral systems, which may be regulated in part by the target organ. We have proposed that the neuropeptide changes occur in neurons that regulate development, maintenance and repair after injury, processes that may differ in somatic and visceral systems.

Animals↗
PubMed1991

Characterization of tightly bound substrates in pure preparations of dihydrofolate reductase: implications for studies on enzymes.

Investigations have been made to determine the identity and binding characteristics of the pterins that are bound tightly to dihydrofolate reductases which are isolated from vertebrate sources by a well established procedure. This procedure involves the binding of enzyme to a Sepharose-methotrexate column, elution with dihydrofolate and removal of free dihydrofolate by dialysis or by size-exclusion chromatography. Addition to such preparations of NADPH results in oxidation of the nucleotide and from the progress curves so obtained, it is possible to identify the bound pterin and to calculate the stoichiometry of binding. The data indicated that stoichiometric amounts of dihydrofolate or folate plus dihydrofolate were bound to the dihydrofolate reductases. It can also be concluded that binding occurs at the active sites of the enzymes. Enzyme preparations from which bound pterin had been removed by isoelectric focussing reacted with folate or dihydrofolate to form 1:1 enzyme-pterin complexes from which the pterin could not be removed by dialysis or by size-exclusion chromatography. From the magnitude of the dissociation constants for the enzyme-pterin complexes and the concentration of enzyme present in fractions after the step involving affinity chromatography on Sepharose-methotrexate, it could be concluded that the presence of bound folate and/or dihydrofolate in pure preparations of dihydrofolate reductase is simply a consequence of an association-dissociation reaction. Preparations of the enzyme from bacterial sources were also found to contain bound pterins. The findings have implications with respect to kinetic and thermodynamic studies on dihydrofolate reductase and other enzymes which are isolated by affinity chromatography techniques.

Chromatography↗
PubMed1991

Reversibility tests in chronic obstructive airways disease: their predictive value with reference to benefit from domiciliary nebuliser therapy.

The role of short-term tests of reversibility in selecting patients with COAD for long-term nebuliser therapy is uncertain. In a double-blind placebo-controlled crossover study we have examined the correlation between short-term reversibility and response to a home nebuliser. We studied 20 patients with severe COAD (mean age 66, mean FEV1 0.81 l) and little reversibility (less than 20% increase in FEV1 post-inhaled salbutamol 200 micrograms and less than 25% increase in peak expiratory flow rate, PEFR, on oral steroids). PEFR, spirometry, lung volumes and airways conductance were recorded before and 1 h after a mixture of nebulised ipratropium 0.5 mg and fenoterol 1.25 mg. Patients then recorded twice-daily PEFR at home while they received nebulised ipratropium plus fenoterol, or saline placebo, four times a day for three week blocks using a double-blind cross over protocol. Mean PEFR on home nebuliser rose from 164 l m-1 (placebo) to 196 l m-1 (ipratropium plus fenoterol), paired t-test P = 0.0001. Correlation coefficients between short-term response for PEFR, spirometry and lung volumes, and improvement in home PEFR on nebulised ipratropium plus fenoterol, were all poor (R = -0.37-0.35, P = 0.83-0.11). We conclude that in severe COAD, reversibility tests of PEFR, spirometry and lung volumes do not correlate with response to a home nebuliser. Home measurements of PEFR are probably the best objective method of assessing response to a home nebuliser in such patients.

Aged↗
PubMed1991

The parasympathetic nervous system and the diurnal variation of lung mechanics in asthma.

In a placebo-controlled double-blind experiment the effects of cholinergic blockade with 30 micrograms kg-1 atropine administered by intravenous injection have been studied in nocturnal asthma. Cholinergic blockade at night reversed the changes in indices of airflow in small airways and of gas trapping to the values seen after administration of atropine during the daytime. However, the improvement in indices of airflow in large airways was not complete, reaching only the daytime placebo levels. No changes were seen in any aspect of breathing pattern. The results are discussed in relation to the literature.

Adult↗
PubMed1991

Effects of H1-receptor blockade with terfenadine in nocturnal asthma.

1. In a single-blind placebo controlled study we have measured peak flow (PEFR) at 04.00 h and 16.00 h in eight asthmatics 6 h after placebo or terfenadine 120 mg, to determine if diurnal variation in histamine mediated effects contribute to nocturnal bronchoconstriction in asthma. 2. On placebo there was a significant diurnal variation in mean PEFR of 41 l min-1 (P less than 0.05). Terfenadine improved 04.00 h baseline mean PEFR from 242 to 278 l min-1 (P less than 0.05) but a 38 l min-1 diurnal variation in mean PEFR persisted (P less than 0.05). 3. We conclude that H1-receptor blockade with terfenadine may produce modest nocturnal bronchodilatation but does not influence the diurnal variation in PEFR in asthma suggesting that H1-receptor mediated effects are not important in the pathogenesis of nocturnal asthma.

Adult↗
PubMed1991

Dose response to inhaled salbutamol in chronic obstructive airways disease.

High dose inhaled salbutamol is increasingly used in the management of chronic obstructive airways disease. To determine the range of doses to achieve optimal bronchodilatation and the proportion of patients requiring high dose therapy we have studied 23 patients with chronic obstructive airways disease. Cumulative dose responses were measured to six incremental doses of salbutamol (0.2 to 1.2 mg) delivered by metered dose inhaler. Results were analysed by polynomial regression to calculate the smallest dose required to produce 90% maximal bronchodilatation in each patient. While 5/23 (22%) required greater than 1 mg the majority, 14/23 (61%), achieved 90% maximal bronchodilation with salbutamol 0.6 mg or less. The 8 patients with severe airflow limitation (FEV1 less than or equal to 1 litre) showed a similar pattern of response. We conclude that in chronic obstructive airways disease there are wide individual variations in the dose of inhaled salbutamol producing 90% maximal bronchodilatation with only a minority requiring high dose therapy.

Administration, Inhalation↗
PubMed1991

Platelet activation in nocturnal asthma.

Platelet activation may be a factor in the bronchial hyperresponsiveness that characterises asthma. As hyperresponsiveness is increased at night, changes in platelet activation over 24 hours were related to the diurnal changes in peak expiratory flow and plasma catecholamine concentrations in five subjects with asthma and five normal subjects. The effect of muscarinic receptor blockade with intravenous atropine at 0400 hours on these measurements was also studied. Platelet activation, assessed as the ration of beta thromboglobulin to platelet factor 4, was highest when the peak expiratory flow rate was at its lowest in the asthmatic subjects. There was no correlation between platelet activation and plasma catecholamine concentrations. Intravenous atropine did not alter the ratio of beta thromboglobulin to platelet factor 4, suggesting that parasympathetic activity is not the cause of the increased platelet activation at night.

Adult↗