Magnetic resonance imaging of masticatory muscles in basset hounds.

OBJECTIVES: To describe low- and high-field MRI characteristics of occult masticatory muscle lesions in basset hounds. MATERIALS AND METHODS: Retrospective review of the MR images of the heads of 44 basset hounds that had been imaged for reasons unrelated to suspected muscle disease. RESULTS: In most basset hounds, there was reduced bulk of the superficial part of the temporalis and masseter muscles together with bilaterally symmetrical T2W and T1W hyperintense signal with reduced contrast enhancement compared to normal muscle. Interpretation of various pulse sequences suggested that the affected muscle areas contained adipose tissue with no evidence of inflammation, and this was confirmed on biopsy in one dog. This pattern of changes is different from MRI changes in inflammatory myopathies. CLINICAL SIGNIFICANCE: Poor muscle bulk and MRI signal changes in the superficial parts of the temporalis and masseter muscles in basset hounds appear to be normal findings in this breed and should not be misinterpreted as evidence of an inflammatory myopathy.

Use of a rotary battery-powered device for the collection of bone marrow in dogs and cats.

Bone marrow biopsies form an important part of investigations into a large number of conditions in veterinary patients. In the human field, powered drivers specifically designed for the collection of bone marrow have become available. These systems have been shown to obtain better quality bone marrow, more quickly and with less pain. This study showed that a powered rotary bone marrow collection system could be used in dogs and cats to collect bone marrow, easily and without complications. Good quality bone marrow aspirates were obtained in the majority of patients; where aspirates could not be collected it was not possible to obtain aspirates from alternative sites using a standard manual technique. Bone marrow core samples were of increased size (mean useable area 16.5 mm(2)), compared to samples obtained using a standard manual technique (mean useable area 4.1 mm(2), P<0.001). Samples obtained using the powered system were also judged to be of significantly better diagnostic quality. The use of a powered system for bone marrow collection may aid the collection of better quality bone marrow samples in small animal patients.

Early brain proton magnetic resonance spectroscopy and neonatal neurology related to neurodevelopmental outcome at 1 year in term infants after presumed hypoxic-ischaemic brain injury.

This study investigated the accuracy of prediction of neurodevelopmental outcome at 1 year using cerebral proton magnetic resonance spectroscopy (MRS) and structured neonatal neurological assessment in term infants after presumed hypoxic-ischaemic brain injury. Eighteen control infants and 28 infants with presumed hypoxic-ischaemic brain injury underwent proton MRS investigation. Studies were carried out as soon as possible after the cerebral insult, most within 48 hours. Infants had an early structured neurological assessment at a median of 19 hours (range 0 hours to 9 days) from the presumed hypoxic-ischaemic insult and a late assessment at a median of 7 days (range 3 to 25 days) during recovery. The maximum cerebral peak-area ratio lactate:N-acetylaspartate measured by proton MRS accurately predicted adverse outcome at 1 year with a specificity of 93% and positive predictive value of 92%. Neurological assessment had a tendency for false-positive predictions. However, both early and late neurological examination can be used as a reliable indicator for a favourable outcome at 1 year having negative predictive values of 100% and 91% respectively.

Proton magnetic resonance spectroscopy of the brain during acute hypoxia-ischemia and delayed cerebral energy failure in the newborn piglet.

Studies of the brains of severely birth-asphyxiated infants using proton (1H) magnetic resonance spectroscopy (MRS) have shown changes indicating a rise in cerebral lactate (Lac) and a fall in N-acetylaspartate (Naa). The aim of this study was to test two hypotheses: 1) that these changes can be reproduced in the newborn piglet after transient reversed cerebral hypoxiaischemia, and their time course determined; and 2) that changes in Lac peak-area ratios are related to changes in phosphorylation potential as determined by phosphorus (31P) MRS. Eighteen piglets aged < 24 h were anesthetized and ventilated. Twelve underwent temporary occlusion of the carotid arteries and hypoxemia, and six served as sham-operated controls. 1H and 31P spectra were acquired alternately, both during the insult and for the next 48 h, using a 7-tesla spectrometer. During hypoxiaischemia, the median Lac/total creatine (Cr) peak-area ratio rose from a baseline of 0.14 (interquartile range 0.07-0.27), to a maximum of 4.34 (3.33-7.45). After resuscitation, Lac/Cr fell to 0.75 (0.45-1.64) by 2 h, and then increased again to 2.43 (1.13-3.08) by 48 h. At all stages after resuscitation Lac/Cr remained significantly above baseline and control values. Naa/Cr was significantly reduced below baseline and control values by 48 h after resuscitation. The increases in the Lac peak-area ratios were concomitant with the falls in the [phosphocreatine (PCr)*]/ [inorganic phosphate (Pi)] ratio, during both acute hypoxiaischemia and delayed energy failure. The maximum Lac/Naa during delayed energy failure correlated strongly with the minimum [nucleotide triphosphate (NTP)]/[exchangeable phosphate pool (EPP)] (r = -0.94, p < 0.0001). We conclude that both hypotheses have been confirmed.

Mild hypothermia after severe transient hypoxia-ischemia reduces the delayed rise in cerebral lactate in the newborn piglet.

This study tested the hypothesis that mild hypothermia after severe transient hypoxia-ischemia reduces the subsequent delayed rise in cerebral lactate peak-area ratios as determined by proton (1H) magnetic resonance spectroscopy (MRS) in the newborn piglet. Nine piglets aged < 24 h underwent temporary occlusion of the common carotid arteries and hypoxemia. Resuscitation was started when cerebral [phosphocreatine]/[inorganic phosphate] had fallen close to zero and [nucleotide triphosphate (NTP)]/[exchangeable phosphate pool (EPP)] was below about a third of baseline. On resuscitation rectal and tympanic temperatures were lowered to 35 degrees C for 12 h after which normothermia (38.5 degrees C) was resumed. 1H MRS data collected over 48 or 64 h after resuscitation were compared with concurrently established data from 12 piglets similarly subjected to transient cerebral hypoxia-ischemia, but maintained normothermic, and six sham-operated controls. The severity of the primary insult (judged from the time integral of depletion of [NTP]/[EPP]) was similar in the hypothermic and normothermic groups. The maximum lactate/N-acetylaspartate ratio observed between 24 and 48 h after resuscitation in the hypothermic group was 0.10 (0.05-0.97), median (interquartile range), which was significantly lower than that observed in the normothermic group, 1.28 (0.97-2.14), and not significantly different from that observed in the control group, 0.08 (0.06-0.11). Similar results were obtained for lactate/choline and lactate/total creatine. We conclude that mild hypothermia after a severe acute cerebral hypoxic-ischemic insult reduces the delayed elevation in lactate peak-area ratios, thus reflecting reduced lactate accumulation.

Neonatal Screening for Haemoglobinopathies in South East London: findings and comparisons with Jamaica

INTRODUCTION: Neonatal screening for sickle cell disorders has been shown to reduce mortality and morbidity. Methods of screening vary but in 1994, the local Health Authority funded universal neonatal screening across the whole of Lambeth, Southwark and Lewisham. This paper will report the findings of the first three years of operation of the programme and compare findings with the screening programme currently operating in Jamaica. METHOD: since May 1994 dried bloo[d] spots of all infants have been screened at King's College Hospital and screen positive cases followed up by counsellors. Infants are followed up at four sites (Guy's, King's, Lewisham and St.Thomas') according to parental preference. Minimum standards for follow-ups have been agreed by clinicians across all four sites and information of the success in achieving these standards is now being collected. RESULTS: Overall there have been 122 affected infants detected in the first two years 10 months of the programmes operation. This is made up of 83 HbSS, 35 HbSC, 4HbSBThal. This gives a birth preference of 23.4 per 1000 total population (2.3HbSS, 1.0 HbSC). Allowing for a termination rate of 20 percent this indicates that the expected birth prevalence in the district would be 4.2 per 1000 births. This compares with birth prevalence of of 0.3 per 1000 for congenital hypothyroidism, 06. per 1000 for cystic fibrosis and 0.1 for phenylketonuria. The distribution of the births is unevem with 57 in Southwark, 40 in Lambeth and 25 in Lewisham. The paper will report on the follow-up and outcome of care provided for this population to date. Discussion: South East London has the highest prevalence of sickle cell disorders of any district in the UK. Sickle cell disorder is now as common in South East London as it is in Jamaica. The follow-up arrangements established in South East London provide an opportunity for colloboration with the West Indies which may help to determine some of the reasons for differences in the natural course of the disease in these populations. The population based approach established should allow monitoring of the impact of community education and antenatal screening programmes on the birth prevalence over time. (AU)

Lactate, N-acetylaspartate, choline and creatine concentrations, and spin-spin relaxation in thalamic and occipito-parietal regions of developing human brain.

Previous studies of the brains of normal infants demonstrated lower lactate (Lac)/choline (Cho), Lac/creatine (Cr), and Lac/ N-acetylaspartate (Naa) peak-area ratios in the thalamic region (predominantly gray matter) compared with occipitoparietal (mainly unmyelinated white matter) values. In the present study, thalamic Cho, Cr, and Naa concentrations between 32-42 weeks' gestational plus postnatal age were greater than occipito-parietal: 4.6 +/- 0.8 (mean +/- SE), 10.5 +/- 2.0, and 9.0 +/- 0.7 versus 1.8 +/- 0.6, 5.8 +/- 1.5, and 3.4 +/- 1.1 mmol/kg wet weight, respectively: Lac concentrations were similar, 2.7 +/- 0.6 and 3.3 +/- 1.3 mmol/kg wet weight, respectively. In the thalamic region, Cho and Naa T2s increased, and Cho and Lac concentrations decreased, during development. Lower thalamic Lac peak-area ratios are principally due to higher thalamic concentrations of Cho, Cr, and Naa rather than less Lac. The high thalamic Cho concentration may relate to active myelination; the high thalamic Naa concentration may be due to advanced gray-matter development including active myelination. Lac concentration is higher in neonatal than in adult brain.

Proton magnetic resonance spectroscopy of the brain in normal preterm and term infants, and early changes after perinatal hypoxia-ischemia.

The aims of this study were 1) to define normal perinatal maturational changes in proton metabolite peak-area ratios in two regions of the neonatal brain, the thalamic and occipitoparietal regions, and 2) to investigate abnormalities of these ratios after perinatal hypoxia-ischemia. Fifty-four infants were studied: 35 normal control infants at 31-42 wk of gestational plus postnatal age, and 19 "asphyxiated" infants suspected of cerebral hypoxic-ischemic injury. Proton spectra were collected at 2.4 tesla from (2 cm)3 voxels using the point-resolved spectroscopy technique with a 270-ms echo time. Lactate was detected in all infants studied. In the normal infants, lactate relative to N-acetylaspartate (NAA), choline and creatine was significantly greater in the occipitoparietal region than in the thalamus, and fell with increasing maturity in both regions, whereas NAA/ choline increased. The 19 asphyxiated infants were studied on a total of 34 occasions during the 1st wk of life (median age 1.8 d), at gestational plus postnatal ages of 27-41 wk. Maximum lactate/NAA was above 95% confidence limits for the control data in one or both regions in 11 of the 19 infants. Minimum NAA/choline was below 95% confidence limits in only one asphyxiated infants, who was later found to have congenital hypothyroidism. SD scores for lactate, relative to NAA, choline, and creatine, were higher in both regions in the asphyxiated infants compared with the normal infants, particularly in the thalamus. Early results of 1-y follow-up examinations indicate that raised lactate/NAA carries a poor long-term prognosis.

Cerebral energy metabolism in isovaleric acidaemia.

A newborn infant with an acute metabolic encephalopathy caused by isovaleric acidaemia had severe impairment of cerebral energy metabolism. This was detected by phosphorus and proton magnetic resonance spectroscopy. After treatment she made excellent clinical recovery, her spectroscopic abnormalities resolved, and she was neurologically normal at the age of 1 year.

Quantitation of phosphorus metabolites in newborn human brain using internal water as reference standard.

A new method for noninvasive, in vivo quantitation of cerebral phosphorus (31P) metabolites is described. The technique employs point-resolved spectroscopy (PRESS) to obtain both 31P-metabolite and proton (1H) water spectra: brain water is used as an internal concentration reference. Spin-spin relaxation times (T2s) of cerebral 31P metabolites are much longer than the minimum echo time (TE) usable on a spectrometer equipped with actively shielded gradient coils. With short-TE (approximately 10 ms) 31P PRESS, T2 relaxation is minimal and phase modulation of the nucleotide triphosphate (NTP) multiplets can be accounted for 1H water spectra were acquired using several TEs so that extra- and intracellular water signals could be separated from that due to cerebrospinal fluid. Prior calibration of the 31P and 1H spectrometer channels and an assumed brain-water concentration enabled estimations of metabolite concentrations. Using this method, mean 31P metabolite concentrations in the brains of eight normal infants of gestational plus postnatal age 34 to 39 wk were: phosphomonoester (PME) 5.6 (SD 0.9); inorganic phosphate 1.4 (0.4); mobile phosphodiester 2.3 (0.6); phosphocreatine 2.9 (0.3); nucleotide triphosphate 2.8 (0.6); and total mobile phosphate 21.4 (2.8) mmol/kg wet.

The estimation of local brain temperature by in vivo 1H magnetic resonance spectroscopy.

Brain temperature may be important for investigating pathology and cerebroprotective effects of pharmaceuticals and hypothermia. Two methods for estimating temperature using 1H magnetic resonance spectroscopy are described: a partially water-suppressed binomial sequence and non-water-suppressed point-resolved spectroscopy. Relative to N-acetylaspartate (Naa), water chemical shift (delta H2O-Naa) in piglet brain depended linearly on temperature from 30 degrees to 40 degrees C: temperature was 286.9-94.0 delta H2O-Naa degrees C. Thalamic temperature in six normal infants was 38.1 degrees +/- 0.4 degree C indicating that local brain temperature could be estimated with adequate sensitivity for studying pathologic and therapeutic changes.

Mild hypothermia after severe transient hypoxia-ischemia ameliorates delayed cerebral energy failure in the newborn piglet.

Severely birth-asphyxiated human infants develop delayed ("secondary") cerebral energy failure, which carries a poor prognosis, during the first few days of life. This study tested the hypothesis that mild hypothermia after severe transient cerebral hypoxia-ischemia decreases the severity of delayed energy failure in the newborn piglet. Six piglets underwent temporary occlusion of the common carotid arteries and hypoxemia. Resuscitation was started when cerebral [phosphocreatine (PCr)]/[inorganic phosphate (Pi)] as determined by phosphorus magnetic resonance spectroscopy had fallen almost to zero and [nucleotide triphosphate (NTP)]/[exchangeable phosphate pool (EPP)] had fallen below about 30% of baseline. Rectal and tympanic temperatures were then reduced to 35 degrees C for 12 h after which normothermia (38.5 degrees C) was resumed. Spectroscopy results over the next 64 h were compared with previously established data from 12 piglets similarly subjected to transient cerebral hypoxia-ischemia, but maintained normothermic, and six sham-operated controls. The mean severity of the primary insult (judged by the time integral of depletion of [NTP]/[EPP]) was similar in the hypothermic and normothermic groups. In the normothermic group, [PCr]/[Pi] and [NTP]/[EPP] recovered after the acute insult and then fell again. Minimum values for these variables observed between 24 and 48 h were significantly higher in the hypothermic group and not significantly different from the control values (p < 0.05, analysis of variance). A large reduction in secondary energy failure relative to the extent of the primary insult was shown and no further fall in either [PCr]/[Pi] or [NTP]/[EPP] took place up to 64 h in the hypothermic piglets.(ABSTRACT TRUNCATED AT 250 WORDS)

Brain-metabolite transverse relaxation times in magnetic resonance spectroscopy increase as adenosine triphosphate depletes during secondary energy failure following acute hypoxia-ischaemia in the newborn piglet.

The adenosine triphosphate (ATP)-dependent sodium/potassium pump extrudes intracellular sodium in exchange for extracellular potassium. Low ATP causes pump dysfunction increasing both intracellular sodium and water thereby enhancing metabolite mobility. This should be detectable by proton magnetic resonance spectroscopy (MRS) as increased metabolite transverse relaxation times (T2s). During secondary cerebral energy failure in the newborn piglet, proton and phosphorus MRS showed large increases in the T2s of choline, creatine, N-acetylaspartate, and lactate that correlated with ATP depletion. These results provide insight into factors affecting metabolite T2s and show that T2s may be useful for studying cellular oedema.

Delayed ("secondary") cerebral energy failure after acute hypoxia-ischemia in the newborn piglet: continuous 48-hour studies by phosphorus magnetic resonance spectroscopy.

Phosphorous (31P) spectra from the brains of severely birth-asphyxiated human infants are commonly normal on the first day of life. Later, cerebral energy failure develops, which carries a serious prognosis. The main purpose of this study was to test the hypothesis that this delayed ("secondary") energy failure could be reproduced in the newborn piglet after a severe acute reversed cerebral hypoxic-ischemic insult. Twelve piglets were subjected to temporary occlusion of the common carotid arteries and hypoxemia [mean arterial PO2 3.1 (SD 0.6) kPa]. Mean cerebral phosphocreatine concentration [PCr]/inorganic orthophosphate concentration [Pi] decreased from 1.40 (SD 0.29) to 0.01 (SD 0.02), and nucleotide triphosphate concentration [NTP]/exchangeable phosphate pool concentration [EPP] decreased from 0.19 (SD 0.02) to 0.06 (SD 0.04) (p < 0.001 for each decrease). On reperfusion and reoxygenation of the brain, mean [PCr]/[Pi] and [NTP]/[EPP] returned to baseline. Observations continuing for the next 48 h showed that [PCr]/[Pi] again decreased, in spite of normal arterial PO2, mean arterial blood pressure, and blood glucose, to 0.62 (SD 0.61) at 24 h (p < 0.01) and 0.49 (SD 0.37) at 48 h (p < 0.001). [NTP]/[EPP] also decreased, but to a lesser degree. Intracellular pH remained unchanged. These findings appeared identical with those seen in birth-asphyxiated human infants. No changes in cerebral metabolite concentrations took place in six control piglets. The severity of secondary energy failure, as judged by the lowest [PCr]/[Pi] recorded at 24-48 h, was directly related to the extent of acute energy depletion, obtained as the time integral of reduction in [NTP]/[EPP] (p < 0.0001). This animal model of secondary energy failure may prove useful for testing cerebroprotective strategies.

Detection of propan-1,2-diol in neonatal brain by in vivo proton magnetic resonance spectroscopy.

Cerebral in vivo proton magnetic resonance spectroscopy of 13 newborn infants displaying seizures and receiving phenobarbitone, in one case supplemented by phenytoin, showed signals from propan-1,2-diol (the injection vehicle for both these anticonvulsants). Subsequent in vitro spectroscopy of cerebro spinal fluid (CSF) from one of these infants also showed signals from this substance. The estimated in vivo propan-1,2-diol concentration (approximately 3 mM) was less than that measured in the CSF sample (14.4 mM). These observations suggest that propan-1,2-diol may accumulate in cerebral tissue and misidentification of its signals in both in vivo and in vitro proton spectra may confuse diagnoses of metabolic or other disorders.

Increased apoptosis in the cingulate sulcus of newborn piglets following transient hypoxia-ischaemia is related to the degree of high energy phosphate depletion during the insult.

An increase in the number of cells undergoing apoptosis was observed in the cingulate sulcus of newborn piglets 48 h after a global hypoxic-ischaemic insult. Apoptotic death was identified morphologically (by light and electron microscopy) and by DNA fragmentation, detected by in situ end labelling. The number of apoptotic cells was directly related to the degree of high-energy phosphate depletion during hypoxia-ischaemia, measured using continuous 31P magnetic resonance spectroscopy. These results may have implications for the understanding and treatment of perinatal hypoxic-ischaemic brain injury.

Effects of N-omega-nitro-L-arginine methyl ester on the cerebral circulation of newborn piglets quantified in vivo by near-infrared spectroscopy.

The effects of N-omega-nitro-L-arginine methyl ester (L-NAME) on basal cerebral vascular tone, the vasodilatory effects of acetylcholine (ACh), and the cerebrovascular response to alterations in arterial carbon dioxide tension (CBVR) were investigated using near-infrared spectroscopy. Seven newborn piglets were anesthetized and mechanically ventilated; mean arterial blood pressure (MAP) was monitored and near-infrared spectroscopy used to measure changes in total cerebral Hb concentration. At the beginning of the experiment, CBVR was measured and then 10, 20, 30, and 100 mg.kg-1 L-NAME were administered sequentially; ACh (1, 2, 3, and 5 micrograms) was given before and after each injection of L-NAME. At the end of this sequence, CBVR was measured again and finally sodium nitroprusside (1.5 mg.kg-1) was administered. Ten and 20 mg.kg-1 L-NAME caused a significant decrease in total cerebral Hb concentration of -0.59 (-3.21 to -0.02) and -1.46 (-6.50 to -0.15) mumol.L-1 (median and range), respectively (Wilcoxon p < 0.05), but subsequent injections did not. Ten, 20, and 100 mg.kg-1 L-NAME caused an increase in MAP (Wilcoxon p < 0.05). ACh caused an increase in total cerebral Hb concentration and a decrease in MAP that was impaired but not abolished by L-NAME (ANOVA p < 0.05). CBVR was not affected by L-NAME. Sodium nitroprusside caused a reduction in mean (SD) MAP of 4.7 (1.6) kPa, and a slower rise in [tHb] of 13.44 (2.03) mumol.L-1. Postmortem examination of three animals revealed NADPH-diaphorase staining in neurons, cerebral blood vessels, carotid artery, and jugular vein.(ABSTRACT TRUNCATED AT 250 WORDS)

Allosteric interactions of glycogen phosphorylase b. A crystallographic study of glucose 6-phosphate and inorganic phosphate binding to di-imidate-cross-linked phosphorylase b.

The binding to glycogen phosphorylase b of glucose 6-phosphate and inorganic phosphate (respectively allosteric inhibitor and substrate/activator of the enzyme) were studied in the crystal at 0.3 nm (3A) resolution. Glucose 6-phosphate binds in the alpha-configuration at a site that is close to the AMP allosteric effector site at the subunit-subunit interface and promotes several conformational changes. The phosphate-binding site of the enzyme for glucose 6-phosphate involves contacts to two cationic residues, Arg-309 and Lys-247. This site is also occupied in the inorganic-phosphate-binding studies and is therefore identified as a high-affinity phosphate-binding site. It is distinct from the weaker phosphate-binding site of the enzyme for AMP, which is 0.27 nm (2.7A) away. The glucose moiety of glucose 6-phosphate and the adenosine moiety of AMP do not overlap. The results provide a structural explanation for the kinetic observations that glucose 6-phosphate inhibition of AMP activation of phosphorylase b is partially competitive and highly co-operative. The results suggest that the transmission of allosteric conformational changes involves an increase in affinity at phosphate-binding sites and relative movements of alpha-helices. In order to study glucose 6-phosphate and phosphate binding it was necessary to cross-link the crystals. The use of dimethyl malondi-imidate as a new cross-linking reagent in protein crystallography is discussed.