A vascular subtraction method for improving the variability of evoked tympanic membrane displacement measurements.

OBJECTIVE: Evoked tympanic membrane displacement (TMD) measurements show a correlation with intracranial pressure (ICP). Attempts to use these measurements for non-invasive monitoring of ICP in patients have been limited by high measurement variability. Pulsing of the tympanic membrane at the cardiac frequency has been shown to be a significant source of the variability. In this study we describe a post processing method to remove the cardiac pulse waveform and assess the impact of this on the measurement and its repeatability. APPROACH: Three-hundred and sixteen healthy volunteers were recruited for evoked TMD measurements. The measurements were quantified by V m, defined as the mean displacement between the point of maximum inward displacement and the end of the stimulus. A sample of spontaneously pulsing TMDs was measured immediately before the evoked measurements. Simultaneous recording of the ECG allowed a heartbeat template to be extracted from the spontaneous data and subtracted from the evoked data. Intra-subject repeatability of V m was assessed from 20 repeats of the evoked measurement. Results with and without subtraction of the heartbeat template were compared. The difference was tested for significance using the Wilcoxon sign rank test. MAIN RESULTS: In left and right ears, both sitting and supine, application of the pulse correction significantly reduced the intra-subject variability of V m (p value range 4.0 × 10-27 to 2.0 × 10-31). The average improvement was from 98 ± 6 nl to 56 ± 4 nl. SIGNIFICANCE: The pulse subtraction technique substantially improves the repeatability of evoked TMD measurements. This justifies further investigations to assess the use of TMD measurements in clinical applications where non-invasive tracking of changes in ICP would be useful.

A model of in-vivo hydrocephalus shunt dynamics for blockage and performance diagnostics.

The accumulation of excess cerebrospinal fluid in the ventricles of the brain results in hydrocephalus, a condition that is fatal if left untreated. The usual remedy is to insert a shunt into the ventricles of the brain, which drains excess fluid away, moderated by a pressure dependent valve. It is important that the system functions properly so that a reasonable intracranial pressure is maintained. Unfortunately, pressure measurements in the ventricles are highly invasive, while pressure measurements in the shunt outside the skull may not detect any blockage in the catheter inside. Here we develop a model primarily aimed at detecting in vivo a blockage and other shunt malfunction using non-invasive measurements, so that shunt valves can be adjusted accordingly. The system offers a clear insight into how currently available clinical measurements may be utilized. We then extend this to investigate the phenomenon of 'chatter' (rapid opening and closing) and other mechanisms including intracranial pressure pulsatility. Although simple, the model offers a clear indication of what is required for successful regulation of both intracranial pressure and shunt flow.

A mitochondrial DNA sequence variant associated with schizophrenia and oxidative stress.

We have previously reported a changed mitochondrial (mt) gene expression in brain from patients with schizophrenia [Schizophr. Res. 14 (1995) 203]; now, we describe the distribution in the mtDNA from lymphocytes of a heteroplasmic sequence variation that was originally found in the mtDNA from the postmortem brain of a patient with schizophrenia. The variant is m.12027T>C and results in the change from isoleucine to threonine at position 423 of the ND4 subunit of NADH-ubiquinone reductase. Using a PCR-RFLP method, we have determined the heteroplasmy as the ratio of variant to total (variant ratio) at m.12027 in 184 controls and 181 patients with schizophrenia as well as 24 postmortem brain samples. The distribution of variants is bimodal having peaks at variant ratios of 0.262 and 0.732. The variant-rich fraction is very significantly associated with schizophrenia in males (47%), while there is only 18% in control males. There are significantly more variant-rich control females (36%) than control males (18%), suggesting that the female population is less sensitive to the presence of a variant in terms of liability to schizophrenia. In variant-rich samples from postmortem brain originating from both sexes, there is an increased superoxide production, suggesting that the variation contributes to oxidative stress. Antioxidant glycosides, such as quercetin rutoside, quench the superoxide production without (in contrast to neuroleptic drugs) interfering with the electron transfer activity of the reductase.

Sonotubometry findings in children at high risk from middle ear effusion.

The functioning of the eustachian tube has an important role to play in the development of middle ear disease. It would be useful if a clinical test could assist in the identification of eustachian tube dysfunction, particularly if this is shown to be an indicator of persistent middle ear effusion. The aim of this study was to compare the results of sonotubometry using the MMS-10 instrument in children at high risk from middle ear effusion with a group of normal subjects. Forty-one subjects (age range 5-6 years) were allocated to one of two groups (experimental group, 21 subjects; control group 20 subjects) based on a questionnaire designed to identify subjects at high risk from middle ear effusion. The test protocol allowed each subject to swallow three times for each of two pure-tones (7 and 8 kHz) delivered by the nasal probe. Sonotubometry indicated opening of the eustachian tube on swallowing in around 80% of subjects. The incidence of positive findings varied greatly amongst subjects across both groups. In the control group, the mean increase in sound pressure level on swallowing was 11.5 dB (+/- 4.3) and 9.8 dB (+/- 2.5) for 7 and 8 kHz, respectively. The corresponding means for duration were 118 ms (+/- 47.9) and 137 ms (+/- 61.8). Sonotubometry failed to demonstrate a difference between the two groups of subjects. Hence, the clinical application of sonotubometry to identify subjects at high risk from middle ear effusion is not supported.

Quantitative assessment of intracranial pressure by the tympanic membrane displacement audiometric technique in children with shunted hydrocephalus.

The objective of this prospective study was to compare the clinical features at presentation, tympanic membrane displacement test results and direct intracranial pressure measurements in children with shunted hydrocephalus to procure a quantitative measure of the intracranial pressure by tympanic membrane displacement test. A prospective comparative evaluation of 61 clinical episodes of shunt malfunction was assessed by volume displacement of the tympanic membrane and direct intracranial pressure measurements in 40 patients with shunted hydrocephalus between January 1995 and June 1996. The volume displacement of the tympanic membrane (Vm) on stapedial contraction was inward for raised intracranial pressure in 27 episodes and ranged from -120 nl to -506 nl (mean = -250 nl). This was confirmed by direct intracranial pressure monitoring, which ranged from 23 to 40 mm Hg (mean = 29 mm Hg). The tympanic membrane displacement test measurement in 30 episodes of low intracranial pressure ranged from +263 nl to +810 nl (mean = +530 nl), and this was corroborated by direct intracranial pressure measurement ranging from 1 to 6 mm Hg (mean = 3.8 mm Hg). The normal baseline Vm values obtained when the subjects were asymptomatic ranged from +58 nl to +175 nl (mean = +115 nl). The tympanic membrane displacement test as a non-invasive diagnostic tool in predicting changes in intracranial pressure had a sensitivity of 93% and specificity of 100%. The predictive value of the test was 100%, and the negative predictive value was 73%. The kappa statistical analysis was used to measure the agreements between the groups. The strength of the agreement was very good, kappa = 0.88 and the P value was < 0.001. The objective measure of intracranial pressure by tympanic membrane displacement test with the Vm value of -200 nl and more negative was indicative of raised intracranial pressure and a Vm value of +200 nl and greater, for low intracranial pressure. The intracranial pressure measurements made on an individual subject basis were reliable and accurate. The test can therefore be used for regular assessment of shunted hydrocephalics to enable correlation of intracranial pressure with symptoms in individual patients.

Superoxide, neuroleptics and the ubiquinone and cytochrome b5 reductases in brain and lymphocytes from normals and schizophrenic patients.

The effects of the neuroleptic flupenthixol on the expression of the genes coding for the mitochondrial ubiquinone and cytochrome b5 reductases have been studied because of the importance of these enzymes in energy metabolism, oxidative stress and also because similar but oppositely directed changes have been previously observed in the cerebral cortex from schizophrenics. The neuroleptic flupenthixol reduces the expression in rats of the gene coding for NADH-cytochrome b5 reductase as measured by in situ hybridisation and its enzymic manifestation. Flupenthixol also reduces the enzymic activity of the mitochondrial NADH-ubiquinone reductase, and it has been previously shown that mRNA from the mitochondrially coded parts of the enzyme are reduced by the drug. Both the cis- and therapeutically less active trans-flupenthixol were found to produce these changes in rats. Post-mortem brain tissue from schizophrenics who have received neuroleptic medication have reduced levels of both reductases as measured enzymically, Lymphocyte samples from schizophrenics also have reduced levels of both reductases compared with normals. The superoxide anion O2- is the principle agent of oxidative stress and both the cytochrome b5 and the ubiquinone reductase enzymes were semi-purified from sheep liver and shown to produce appreciable amounts of superoxide. Superoxide production is reduced in brain homogenates from rats treated with flupenthixol. Its production is also reduced in brain tissue and lymphocytes from schizophrenics receiving neuroleptic medication. We conclude that neuroleptic medication reduces the expression of both the ubiquinone and cytochrome b5 reductase and among the effects of this reduction is a decrease in the production of neurotoxic superoxide.

Tympanic membrane displacement testing in regular assessment of intracranial pressure in eight children with shunted hydrocephalus.

OBJECT: The authors assessed the accuracy and repeatability of the tympanic membrane displacement (TMD) test, an audiometric technique that is used to evaluate changes in intracranial pressure (ICP) in children with shunted hydrocephalus. METHODS: A prospective comparative evaluation of 31 clinical episodes of shunt malfunction was made by using the serial TMD test and direct ICP measurement in eight children with shunted hydrocephalus between January 1995 and February 1996. The volume displacement of the tympanic membrane (Vm) on stapedial contraction was inward for raised ICP in 11 instances and ranged from -120 to -539 nl (mean -263.5 nl). This was confirmed by direct ICP monitoring, which showed values ranging from 20 to 30 mm Hg (mean 26 mm Hg). The TMD test measurement (Vm) in 18 instances of low ICP ranged from 263 to 717 nl (mean 431.3 nl); this was corroborated by direct ICP measurement, which ranged from 3 to 7 mm Hg (mean 4.2 mm Hg). The normal baseline Vm values obtained when patients were asymptomatic ranged from 98 to 197 nl (mean 110 nl). As a noninvasive diagnostic tool used in predicting changes in ICP, the TMD test had a sensitivity of 83% and specificity of 100%. The positive predictive value of the test was 100% and the negative predictive value was 29%. CONCLUSIONS: The TMD test can be used on a regular basis as a reproducible investigative tool in the assessment of ICP in children with shunted hydrocephalus, thereby reducing the need for invasive ICP monitoring. The equipment necessary to perform this testing is mobile. It will provide a useful serial guide to ICP abnormalities in children with shunted hydrocephalus.

Why monitor perilymphatic pressure in Menière's disease?

A cohort of 39 patients with confirmed raised intracranial pressure was investigated. Of these patients, 24 (62%) complained of tinnitus and 11 (28%) suffered from paroxysmal rotary vertigo. Intracranial hypertension can occur without the usual headache and visual symptoms. In such cases, the patient may be referred to the otolaryngological clinic and the condition may be mistaken for Menière's disease or a labyrinthine disorder. The Tympanic Membrane Displacement (TMD) technique now provides a non-invasive method of monitoring the intracranial and perilymphatic pressures. This study provides recommendations for the use of TMD techniques in the otolaryngological clinic for screening, diagnosing and monitoring treatment of patients presenting with raised perilymphatic pressure.

Mitochondrial involvement in schizophrenia and other functional psychoses.

Gene expression has been studied in post-mortem frontal cortex samples from patients who had suffered from schizophrenia and depressive illness. mRNA was extracted and characterised by translation and separation of the products by 2D gel electrophoresis. Post-mortem artefacts and the agonal experience did not affect the size distribution or amount of specific translation products. Four expression products were specifically reduced in samples from schizophrenics compared with normals. The expression of six products was altered in affective disorder, one in common with schizophrenia, two the same as in schizophrenia but increased. cDNA libraries were produced from the mRNA samples and 5 clones present at abnormal levels in schizophrenia identified by differential screening, isolated and sequenced. All the sequences encode mitochondrial transcripts; four encode mitochondrial rRNA and one the amino acid sequence of cytochrome oxidase sub-unit II. Increased cytochrome oxidase transcripts were found in a further set of mRNA extracts from schizophrenic patients including two who had not received neuroleptic medication. The effects of neuroleptic administration as exemplified by alpha-flupenthixol compared with the ineffective beta-flupenthixol were studied in experimental animals. It was found that 13 out of 28 clones whose levels were altered were mitochondrial in origin including rRNA, COX I & II and the NADH-Q reductase. Those encoding respiratory enzymes were at abnormally low levels as a result of alpha-flupenthixol administration. Measurements of the enzymic activity of cytochrome c oxidase in post-mortem frontal cortex of schizophrenics did not indicate any differences in overall activity but there was a decreased sensitivity to azide that was abolished by neuroleptics. Studies on NADH-cytochrome c reductase showed that schizophrenics whether medicated or not had a reduced rotenone sensitive activity that was compensated for by increased rotenone insensitive activity. We conclude that changes in mitochondrial gene expression are involved in schizophrenia and probably other functional psychoses.

Intracranial pressure at high altitude and acute mountain sickness.

1. Raised intracranial pressure has been noted in severe forms of acute mountain sickness and high-altitude cerebral oedema, but the role of intracranial pressure in the pathogenesis of mild to moderate acute mountain sickness is unknown. 2. Serial measurements of intracranial pressure were made indirectly by assessing changes in tympanic membrane displacement in 24 healthy subjects on rapid ascent to 5200 m. 3. Acute hypoxia at 3440 m was associated with a rise in intracranial pressure, but no difference was found in pressure changes at 4120 or 5200 m in subjects with or without symptoms of acute mountain sickness. 4. Raised intracranial pressure, though temporarily associated with acute hypoxia, is not a feature of acute mountain sickness with mild or moderate symptoms.

Astrocyte transplants alleviate lesion induced memory deficits independently of cholinergic recovery.

Basal forebrain tissue fragments taken from embryonic day 15 were separated into primary astrocytes and primary neurons in culture and grafted to rats with alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid lesions to the nucleus basalis and medial septal regions. The two cell types were compared in two experimental paradigms for their behavioural, biochemical and histochemical effects; standard transplants of whole basal forebrain and sham transplants served as positive and negative controls, respectively. Each transplant cell type was characterised by in vitro immunocytochemistry to assess content and purity. Memory deficits produced by the lesions in a spatial win-stay T-maze task (Experiment 1) and a spatial plus associative radial maze task (Experiment 2) were significantly improved by the astrocyte, but not by the neuronal, primary cell transplants. The astrocyte graft groups performed as well as standard cholinergic rich basal forebrain groups, reaching control levels on both tasks, while the neuronal transplant groups were not significantly different to lesioned (sham transplanted) rats. There was no recovery in choline acetyltransferase activity in brain regions containing astrocyte grafts whereas activity in the neuronal graft regions was increased (often to control levels), similar to recovery produced by basal forebrain grafts. Grafts in all groups survived, transplanted neurons displaying similar morphology and placement in the host brain to unseparated basal forebrain grafts, while astrocytes showed evidence of migration. The cultured astrocytes were estimated to be > 95% pure, showing positive staining for all astrocyte markers and an absence of staining for neuronal markers. The results indicate that the restoration of cognitive function following fetal grafting is not dependent upon a restoration of cholinergic neuronal activity but is more likely mediated via diffuse graft-host communication, with trophic secretion a probable factor. This study emphasizes the usefulness of astrocytes in the repair of central nervous system injury and has implications for therapeutic potential.

Genetic linkage analysis of schizophrenia using chromosome 11q13-24 markers in Israeli pedigrees.

It is generally agreed that there is a genetic component in the etiology of schizophrenia which may be tested by the application of linkage analysis to multiply-affected families. One genetic region of interest is the long arm of chromosome 11 because of previously reported associations of genetic variation in this region with schizophrenia, and because of the fact that it contains the locus for the dopamine D2 receptor gene. In this study we have examined the segregation of schizophrenia with microsatellite dinucleotide repeat DNA markers along chromosome 11q in 5 Israeli families multiply-affected for schizophrenia. The hypothesis of linkage under genetic homogeneity of causation was tested under a number of genetic models. Linkage analysis provided no evidence for significant causal mutations within the region bounded by INT and D11S420 on chromosome 11q. It is still possible, however, that a gene of major effect exists in this region, either with low penetrance or with heterogeneity.

A study of altered gene expression in frontal cortex from schizophrenic patients using differential screening.

In order to examine the molecular basis of schizophrenia we have employed a sequential differential hybridisation protocol to isolate mRNAs whose abundances are altered in schizophrenic compared to normal frontal cortex. Five cDNAs present at abnormal levels in the schizophrenic brain have been isolated by this method. The sequences were identified on the basis of homologies in the EMBL and Genbank databases. All the sequences encode mitochondrial transcripts; one encodes part of the 12s rRNA, three encode parts of the 16s rRNA region of the mitochondrial genome whilst the fourth encodes part of the amino acid sequence of cytochrome oxidase sub-unit II. It was established that mitochondrial sequences were not over-represented in the library and that this could therefore not account for the isolation of five mitochondrial transcripts by this procedure. Increased levels of cytochrome oxidase mRNA were detected in a further set of extracts from the frontal cortex of eight schizophrenic patients and five controls. The amount of mt-DNA was measured in these samples but there was no difference between schizophrenic and control. These results indicate a possible abnormality of mitochondrial function in the schizophrenic frontal cortex.

Fimbria-fornix lesions impair spatial performance and induce epileptic-like activity but do not affect long-term potentiation in the CA1 region of rat hippocampal slices.

Groups of rats were given bilateral fimbria-fornix lesions and one month later grafted into the hippocampus with fetal cholinergic and non-cholinergic (hippocampal) neural tissue. Three weeks and 3 months after transplantation the animals were trained to find and then to retain the location of a hidden platform in the Morris water maze. After the final behavioral testing phase, electrophysiological studies of the short- and long-term potentiation (STP and LTP) and epileptiform activity of evoked responses were performed in vitro in the CA1 region of the hippocampus. The lesions produced a marked deficit in spatial function in the early testing phase which showed some recovery at the three month time point. Neither the cholinergic nor the non-cholinergic grafts improved spatial performance; indeed, on some measures these groups showed a significantly greater deficit than the lesion-alone group. Epileptiform activity, which was defined as the ratio of the sum of amplitudes of second and third population spikes to the amplitude of the first, before tetanization was not significantly different for all groups. After tetanization of the radiatum input, however, the epileptiform activity in the FFL group was significantly higher in comparison to that of the control groups. Grafting of cholinergic tissue decreased this parameter to the control level, but non-cholinergic grafts did not modify the lesion-induced epileptiform activity. Epileptiform activity after tetanization of the oriens input was approximately equal for all groups. There were no significant differences between surgical groups in STP and LTP for both the oriens and radiatum inputs.(ABSTRACT TRUNCATED AT 250 WORDS)

The lymphoblast beta-adrenergic receptor in bipolar depressed patients: effect of chronic incubation with lithium chloride.

We have recently reported a study of beta-adrenergic receptor binding characteristics in lymphoblast cell lines derived from patients with bipolar disorder (BD) and healthy, matched control subjects. In the present study we have investigated the effects of incubating cells from the same subjects with lithium chloride (1 mM) for 7 days prior to assay. There was no difference in beta-adrenergic receptor number between control and BD cell lines and incubation with lithium had no effect on receptor number in either group. Exposure of the cells to isoprenaline (1 nM) for 24 h immediately prior to assay caused significantly less down-regulation in BD cells (15 +/- 5%) than control cells (39 +/- 4%), as described previously. Incubation with lithium significantly increased the down-regulation response to isoprenaline in BD cells (39 +/- 6%) but not in control cells (30 +/- 7%). After lithium, the agonist-induced decrease in beta-AR number in BD cells was no longer significantly different from that in control cells. We conclude that lithium selectively enhanced the agonist down-regulation of beta-adrenergic receptors in cells derived from patients with bipolar disorder. The functional significance of this result and the potential biochemical mechanisms responsible for this effect are discussed.

Immunoidentification of cellular brain proteins associated with cognitive recovery in brain transplants.

In adult, lesion-impaired rat brain receiving embryonic day 15 (E15) fetal transplants, the level of expression of glial fibrillary acidic protein (GFAP) correlates positively with choline acetyltransferase (ChAT) levels and also with measurements of successful behavioural recovery. These results suggest that glial cells may play a pivotal role in the cognitive success of so-called cholinergic-rich transplants. The objective of this study was to investigate the association between GFAP- and ChAT-staining antigens in or around cholinergic-rich fetal grafts transplanted in adult cortex. An immunohistochemical fluorescent double-labelling technique was used to simultaneously identify GFAP- and ChAT-staining cells to assess whether there was a different type or distribution of cells present in these successful transplants. On brain sections of transplant area, GFAP-staining glial cells did not co-label with ChAT-staining cells. The transplant area, therefore, did not reveal a different type of cell from those seen in comparable normal cortical brain but rather a greater concentration of both GFAP- and ChAT-positive staining cells.

Heterosynaptic short-term depression of population spike amplitude in the pyramidal layer of the CA1 hippocampal region evoked by a theta-like tetanization.

Heterosynaptic short-term depression (STD) of the stratum radiatum and stratum oriens inputs to the CA1 region was studied in rat hippocampal slices. STD was evoked by trains of 1050 impulses with interstimulus interval (ISI) variable from 10 to 700 ms. The STD was found to be very pronounced for tetanizations with ISI around 200 ms, and almost absent for ISI less than 50 ms or more than 500 ms. These data show that theta-like tetanization is an effective pattern not only for induction of the long-term potentiation (LTP), as has been shown previously, but for production of the heterosynaptic STD as well. This implies that heterosynaptic STD can effectively modulate induction of LTP by theta-like tetanization, and plays an important role in differentiation of potentiated pathways. It is discussed that the theta-like tetanization-induced release of ACh is a possible mechanism of the STD.