Abolished circadian rhythm of melatonin secretion in sedated and artificially ventilated intensive care patients.

BACKGROUND: Sleep disturbance is common in intensive care patients. Aside from its unpleasantness, there is a correlation with intensive care unit (ICU) syndrome/delirium. Reasons for sleep deprivation appear to be multifactorial, including the underlying illness, an acute superimposed disturbance, medications, and the ICU environment itself. There are reasons to believe that alterations of the 'biological clock' might contribute. Melatonin secretion is one reflection of this internal sleep/wake mechanism. Melatonin levels are normally high during the night and low during daytime, being suppressed by bright light. METHODS: Melatonin levels in blood and urine were studied over 3 consecutive days in eight critically ill patients during deep sedation and mechanical ventilation. Sedation was assessed with the sedation-agitation (SAS) scale and bispectral index (BIS) monitor. RESULTS: The circadian rhythm of melatonin release was abolished in all but one patient, who recovered much more quickly than the others. There was no correlation between melatonin levels and levels of sedation. CONCLUSIONS: This study indicates that dyssynchronization of the melatonin secretion rhythm is common in critically ill and mechanically ventilated patients. It could be hypothesized that an impairment of the melatonin rhythm may play a role in the development of sleep disturbances and delirium in intensive care patients, and that melatonin supply could reduce the incidence of these phenomena.

Chronic ethanol exposure enhances activating protein-1 transcriptional activity in human neuroblastoma cells.

This study demonstrates a method for studying the effects of ethanol on transcription mediated by activating protein-1 (AP-1). The effects of ethanol on AP-1 activity and on the signaling cascades in this process were investigated by using a reporter gene technique with secreted alkaline phosphatase as the reporter gene coupled to nine DNA AP-1-binding elements. Long-term ethanol exposure (48-72 h) dose dependently enhanced AP-1 transcriptional activity in SH-SY5Y cells. Shorter exposure periods with ethanol did not influence AP-1 transcriptional activity compared with findings for control cells. Inhibition of protein kinase C (PKC) dramatically decreased AP-1 activity in both control and ethanol-exposed cells and abolished the ethanol enhancement. This finding suggests a pivotal role for PKC-coupled signaling in AP-1 transcriptional activity. Phorbol ester stimulation of AP-1 transcriptional activity was not influenced by long-term ethanol exposure. This finding indicates that signaling events upstream of PKC are the targets for ethanol. Mitogen-activated protein kinases ERK and p38 may play a role in ethanol-enhanced AP-1 activity because inhibitors of both enzymes partly reduced the enhancement. The inhibitors also partly blocked phorbol ester-induced AP-1 activation, which demonstrates a function of these mitogen-activated protein kinases downstream of PKC.

Role of nitric oxide in ethanol-induced up-regulation of muscarinic acetylcholine receptors in SH-SY5Y cells.

BACKGROUND: Ethanol abuse has been shown to up-regulate muscarinic acetylcholine receptors (mAChRs) in the central nervous system, both in vivo and in vitro, but the detailed mechanisms for this action are not known. Recent studies suggest that the actions of ethanol in several biological systems involve nitric oxide (NO) pathways. The aim of this work was therefore to determine whether the effects of long-term ethanol treatment on mAChRs involved actions upon NO pathways in a well defined cell culture system. METHODS: Human neuroblastoma SH-SY5Y cells were used as an in vitro model system. The time- and dose-dependent effects of ethanol on endogenous NO production, as well as the effects of the NO donor 3-morpholino-sydnonimine-chloride and of the neuronal NO synthase (nNOS) inhibitor N(omega)-propyl-L-arginine on mAChR number and on ethanol effects upon these receptors, were studied. RESULTS: Ethanol time- and dose-dependently decreased the production of NO in the cells. Exogenous NO decreased the number of mAChRs and totally blocked the effects of ethanol upon these receptors. Inhibition of nNOS up-regulated the number of mAChRs, but this effect was not additive to the effects of ethanol. CONCLUSIONS: The results of this study suggest that the number of cell-surface mAChRs in SH-SY5Y cells may be correlated with changes in NO levels. The number of cell surface mAChRs decreased with NO-elevating treatment and increased with NO-lowering treatment. Because ethanol, which is known to up-regulate mAChRs in SH-SY5Y cells, also decreased NO levels and because nNOS inhibition and ethanol effects on mAChRs were not additive, it is conceivable that ethanol-induced up-regulation of mAChRs involves inhibition of nNOS.

S100B as a predictor of size and outcome of stroke after cardiac surgery.

BACKGROUND: Stroke after cardiac surgery is a clinical problem with often fatal or disabling outcome. To assess severity and probable outcome in affected patients only from clinical and radiological examinations is difficult. The glial-derived protein S100B has been suggested to be a marker of cerebral ischemia, and increased blood concentrations of S100B have been shown to correlate with size of lesion and prognosis after stroke. We studied the validity of S100B as a predictor of size of brain lesion and median term outcome in a consecutive group of patients suffering from stroke after cardiac surgery. METHODS: During a period of 17 months, 20 patients with clinical signs of postoperative stroke were investigated with S100B measurement, sampled at 5, 15 and 48 hours after surgery. All patients were examined with computed tomography or magnetic resonance imaging to confirm the diagnosis, and the size of cerebral infarction was estimated from the radiological examinations. The patients were followed up for survival 24 to 39 months after surgery. RESULTS: S100B concentration in blood 48 hours after surgery correlated with the size of infarcted brain tissue (r = 0.68, p < 0.001). Nine patients had S100B levels exceeding 0.5 microg/L and a 2-year mortality of 78%, whereas the 11 patients with S100B below 0.5 microg/L had a mortality of 18%. CONCLUSIONS: Increased S100B in patients with a stroke following cardiac surgery correlate with the size of infarcted brain tissue. High S100B levels 48 hours after surgery have a negative predictive value for median term survival.

Brain-specific NSE and S-100 proteins in umbilical blood after normal delivery.

BACKGROUND: To determine normal blood levels of brain-specific proteins S-100 and neuron specific enolase (NSE) in healthy newborns and their mothers following uncomplicated birth. METHODS: Umbilical artery and vein blood and maternal venous blood was collected at 112 consecutive uncomplicated deliveries. Venous blood samples were taken from 18 of the neonates 3 days after birth. S-100 and NSE were analyzed quantitatively by double antibody immunoluminometric assay (Sangtec Medical AB, Sweden). RESULTS: Compared with adults, healthy neonates had higher levels of both S-100 and NSE. For S-100, median levels (range) were 1.10 microg/l (0.38-5.50 microg/l and 0.98 microg/l (0.43-2.70 microg/l) in umbilical artery and vein, respectively. For NSE, median levels (range) in umbilical artery blood and vein were 27 microg/l (10-140 microg/l) and 10.75 microg/l (8.80->/=200 microg/l) respectively. The maternal venous blood levels of both S-100 and NSE were significantly lower than in their infants. At 3 days of life, neonatal venous levels of the proteins were still high: S-100, 0.48-9.70 microg/l; NSE, 17->/=200 microg/l. In contrast to adults, haemolysis affected the S-100 levels in umbilical blood significantly. CONCLUSION: Concentrations of both S-100 and NSE in blood are greater in newborns after normal birth than in healthy adults. The higher levels in umbilical artery blood than in umbilical vein blood are consistent with a fetal origin of these proteins. High levels in venous blood at 3 days of life suggest that the high levels at birth are not related to the birth process but reflect a high activity of these proteins during fetal development.

Phosphatidylethanol in post-mortem blood as a marker of previous heavy drinking.

Phosphatidylethanol (PEth) is an ethanol-phospholipid adduct, formed via non-oxidative metabolism of ethanol. PEth was measured in femoral blood from 85 consecutive forensic autopsies and was detected in 35 of the cases at concentrations ranging from 0.8 to 22.0 micromol/l. Of the PEth positive cases, 12 did not have significant levels of ethanol in the blood. Two cases (both suicides involving hanging) had detectable ethanol, but no PEth present in the blood. We conclude that measurements of PEth provide indications of previous alcohol abuse in cases where this may not otherwise be evident.

Effects of ethanol on phosphoinositide hydrolysis and muscarinic acetylcholine receptor number in SH-SY5Y cells.

Previous studies suggest that the effects of ethanol on carbachol-stimulated I(1,4,5)P3 formation and on the number of mAChRs may be independent of each other. The aim of this work was to further study this hypothesis. Human neuroblastoma SH-SY5Y cells were used as a model system. Acute exposure of the cells to 100 mM ethanol induced a decrease in [3H]N-methylscopolamine ([3H]NMS) binding at 30 seconds which was of lower magnitude and of shorter duration than the previously described ethanol-induced inhibition of the peak of carbachol-stimulated I(1,4,5)P3 formation. Long-term ethanol treatment of the cells induced a time- and concentration-dependent increase in [3H]NMS binding. Three hours of 100 mM ethanol treatment were sufficient to increase the number of mAChRs at the cell surface but these receptors were not immediately functionally active, suggesting that they may be newly synthesized. Furthermore, the ethanol-induced potentiation of carbachol-stimulated I(1,4,5)P3 formation, after two days, was, for all ethanol concentrations tested, of higher magnitude than the ethanol-induced increase in mAChR number. Together, these data indicate that both acute and chronic ethanol-induced changes in carbachol-stimulated I(1,4,5)P3 formation may not only be explained by changes in mAChR density at the cell surface but may rather be the consequence of actions of ethanol down-stream of the receptor.

Changes of cerebrospinal fluid monoamine metabolites during long-term antidepressant treatment.

This study describes the changes in cerebrospinal fluid (CSF) monoamine metabolites during antidepressant treatment for more than 6 months. Eight patients, who received antidepressant treatment after attempted suicide and then underwent lumbar punctures every 3 or 4 months, were included. Plasma drug concentrations and the clinical outcome were also measured. Consistent with previous reports about antidepressant treatment for between 3 and 6 weeks, both 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG) and 5-hydroxyindoleacetic acid (5-HIAA) were significantly decreased after treatment for a mean of 15 weeks compared to pretreatment. However, after continued treatment for a mean of 30 weeks the MHPG concentration remained significantly lower than at pretreatment while 5-HIAA had returned to the pretreatment level. The clinical outcome was significantly correlated to the pretreatment 5-HIAA/MHPG ratio. These results suggest that the frequently reported reduction in CSF 5-HIAA after antidepressant treatment does not remain during long-term treatment.

Normalization rate and cellular localization of phosphatidylethanol in whole blood from chronic alcoholics.

Phosphatidylethanol (PEth) is an abnormal phospholipid which is formed in the presence of ethanol, via the action of phospholipase D (PLD). PEth in blood is a potential marker of alcohol abuse. The present study was made to determine the compartmentalization and the elimination rate of PEth in human whole blood. PEth was assayed by an improved HPLC technique, with evaporative light-scattering detection. Blood from six alcoholic males was separated into different blood cell fractions. The PEth concentration in whole blood was 2.5+/-0.9 and 1.9+/-1.1 micromol/l in erythrocytes. Only one subject had detectable PEth in the mononuclear cells. Fifteen patients (13 men, two women) with chronic alcoholism, were followed as inpatients, after admission to an alcohol detoxification clinic. PEth, carbohydrate-deficient transferrin (CDT) and gamma-glutamyltransferase (GGT) were measured on days 1, 3, 5 and 7 after admission. Linear regression analysis of logarithmic PEth values in individuals, with measurable PEth at day 1, gave a good fit (P<0.001) with the one-compartment elimination model. The half-life was calculated as 4.0+/-0.7 days. A weak significance (P<0.05) was observed in the correlation of PEth at day 1 and half-life values of the same subjects.

Neuron-specific enolase increases in plasma during and immediately after extracorporeal circulation.

BACKGROUND: Minor cerebral complications are common after cardiac surgery. Several biochemical markers for brain injury are under research; one of these is neuron-specific enolase (NSE). The purpose of this study was to investigate the release of this enzyme into the blood during and immediately after extracorporeal circulation and to evaluate the effect of hemolysis on this release. METHODS: Sixteen patients scheduled for elective heart surgery were included in the study. Blood samples for analysis of NSE and free hemoglobin in plasma were drawn before, during, and up to 48 hours after the end of extracorporeal circulation. The release of NSE from erythrocytes and its correlation to the release of free hemoglobin was studied by serial dilution and hemolysis in vitro. RESULTS: The peri- and postoperative course was uneventful in all patients. Extracorporeal circulation initiated a release of NSE that reached a maximum 6 hours after the end of perfusion. Thereafter, the levels declined with an estimated t1/2 of 30 hours. The concentration of free hemoglobin increased during the perfusion, with maximum levels at the end of perfusion, after which they fell rapidly to normal values. The in vitro study showed a strong linearity between the release of NSE and free hemoglobin after induced hemolysis. CONCLUSIONS: The increased levels of enolase at the end of cardiopulmonary bypass can, to a major part, be explained by the release from hemolysed erythrocytes. The value of NSE as a marker for brain injury in these situations is therefore doubtful.

The clinical value of serum S-100 protein measurements in minor head injury: a Scandinavian multicentre study.

PURPOSE: This study of patients with minor head injury was designed to investigate the relation of S-100 protein measurements to computed tomograpy (CT) findings and patients outcomes. Increased serum levels of this protein were hypothetized to predict intracranial pathology and increased frequency of post-concussion symptoms. METHODS: One hundred and eighty-two patients were studied with Glasgow Coma Scale scores of 13-15. The study recruited patients from three Scandinavian neurotrauma centres. Serum levels of S-100 protein were measured at admittance and CT scans of the brain were obtained within 24 hours postinjury in all patients. Outcome was evaluated with the Rivermead Postconcussion Symptoms Questionnaire (RPQ) 3 months after the injury. RESULTS: Increased serum level of S-100 protein was detected in 69 (38%) patients, and CT scan demonstrated intracranial pathology in 10 (5%) (brain contusion in seven, epidural haematoma in two, traumatic subarachnoid haemorrhage in one). The proportion of patients with detectable serum level was significantly (p < 0.01) higher among those with intracranial pathology (90%) compared to those without (35%). The negative predictive value of an undetectable S-100 level was 0.99. Sixty-two per cent reported one or more post-concussion symptoms at follow-up. A trend was observed towards an increased frequency of post-concussion symptoms among patients with detectable serum levels. CONCLUSIONS: Undetectable serum level of S-100 protein predicts normal intracranial findings on CT scan. Determination of S-100 protein in serum may be used to select patients for CT scanning. Increased S-100 serum levels may be more related to post-concussion symptoms caused by mild traumatic brain injury than to symptoms of psychological origin.

Elimination of S100B and renal function after cardiac surgery.

OBJECTIVES: To determine the biologic half-life of the S100B protein and to investigate if the elimination of S100B depends on glomerular filtration rate (GFR). DESIGN: Prospective human study. SETTING: University hospital. PARTICIPANTS: Sixteen patients who underwent cardiac surgery with cardiopulmonary bypass. INTERVENTIONS: Shed mediastinal blood (autotransfusion) was returned to the patients postoperatively and used to study the kinetics of S100B. Iohexol was infused simultaneously to estimate GFR. S100B was measured at 0, 20, 40, 60, and 180 minutes after infusion. Iohexol was measured at 180 and 240 minutes after infusion. MEASUREMENTS AND MAIN RESULTS: S100B followed first-order kinetics, and the biologic half-life for S100B was determined to be 25.3 +/- 5.1 minutes. GFR was determined to be 63.8 +/- 34.4 mL/min. No correlation was found between GFR and S100B half-life. CONCLUSIONS: The elimination of S100B after cardiac surgery is faster than reported earlier and not affected by a moderate decrease in GFR. This finding is important when evaluating S100B levels after cardiac surgery.

Reduced cerebrospinal HVA concentrations and HVA/5-HIAA ratios in suicide attempters. Monoamine metabolites in 120 suicide attempters and 47 controls.

Dysfunctions of central monoaminergic systems are important elements of the leading biological hypotheses of suicide and depression. The purpose of the present paper was to study the levels and the relationships between the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA), the dopamine metabolite homovanillic acid (HVA) and the norepinephrine metabolite 3-methoxy-4-hydroxyphenylglycol (MHPG) in the cerebrospinal fluid (CSF) in 120 hospitalised suicide attempters and 47 controls (healthy volunteers or patients admitted for minor surgery). The suicide attempters showed significantly lower HVA levels (174+/-82 vs. 216+/-96 nmol/L, P=0.004), HVA/5HIAA ratios (1.6+/-0.5 vs. 2.1+/-0.6, P=0.0001) and HVA/MHPG ratios (4.2+/-2.1 vs. 4.8+/-1.7, P=0.02) than the controls. The correlations between the monoamine metabolites were markedly lower in patients than in controls. CSF 5-HIAA showed no significant differences between patients and controls (107+/-40 vs. 108+/-51 nmol/L) or between violent and non-violent attempters (112+/-58 vs. 105+/-33 nmol/L). The monoamine metabolites showed no significant differences between survivors and patients who subsequently completed suicide, or between suicide attempters subgrouped by psychiatric diagnoses. The results suggest that low HVA levels and altered relationships between the monoamine metabolites are associated with suicidal behaviour.

[The biological mechanisms underlying alcohol dependence]. / De biologiska mekanismerna bakom beroende av alkohol.

Recent advances in neuroscience have enabled alcohol dependence to be investigated at cellular and molecular levels. Alcohol exerts biological effects by interacting with cell membranes and receptors, and modifies the function of proteins which regulate signal transduction, intracellular pathways, and gene expression. Adaptation to the acute effects of alcohol constitutes a major determinant of the development of increased tolerance, withdrawal syndrome and dependence. Important targets for alcohol include transmitter-regulated ion channels, receptors coupled to guanosine triphosphate-binding proteins, second messengers, and gene transcription factors. Studies of rodents bred for alcohol sensitivity or resistance have uncovered molecules of importance for the development of alcohol dependence, and certain ethanol-sensitive genes have been identified.

[The biological mechanisms underlying alcohol dependence]. / De biologiska mekanismerna bakom beroende av alkohol.

Recent advances in neuroscience have enabled alcohol dependence to be investigated at cellular and molecular levels. Alcohol exerts biological effects by interacting with cell membranes and receptors, and modifies the function of proteins which regulate signal transduction, intracellular pathways, and gene expression. Adaptation to the acute effects of alcohol constitutes a major determinant of the development of increased tolerance, withdrawal syndrome and dependence. Important targets for alcohol include transmitter-regulated ion channels, receptors coupled to guanosine triphosphate-binding proteins, second messengers, and gene transcription factors. Studies of rodents bred for alcohol sensitivity or resistance have uncovered molecules of importance for the development of alcohol dependence, and certain ethanol-sensitive genes have been identified.

S100beta after coronary artery surgery: release pattern, source of contamination, and relation to neuropsychological outcome.

BACKGROUND: S100beta has been suggested as a marker of brain damage after cardiac operation. The aim of this study was to characterize the early S100beta release in detail and relate it to neuropsychological outcome. METHODS: Three groups of patients were investigated. All patients underwent coronary artery bypass surgery (CABG) with extracorporeal circulation. In group A, 110 patients had sampling of S100beta for the first 10 postoperative hours and also underwent neuropsychological testing. In group B, 14 patients were examined for the effect of autotransfusion on S100beta levels. Eight patients in group C had their intraoperative bleeding processed with a cell-saving device. RESULTS: Group A had a heterogeneous release pattern with several rapid elevations in S100beta concentration. In group B, high concentrations of S100beta were found in the autotransfusion blood (range 0.2 to 210 microg/L) with a concurrent elevation of serum S100beta levels after transfusion of shed blood. In group C, high levels of S100beta were found in the blood from the surgical field (12.0+/-6.0 microg/L) and decreased (1.1+/-0.64 microg/L) after wash. Group C had significantly lower S100beta values at the end of cardiopulmonary bypass compared to group A (0.53+/-0.35 microg/L versus 2.40+/-1.5 microg/L). S100beta values were corrected for extracerebral contamination with a kinetic model. With this correction, an association was found between adverse neuropsychological outcome and S100beta release in group A (r = 0.39, p < 0.02). CONCLUSIONS: A significant amount of S100beta is found both in the blood from the surgical field and in the shed mediastinal blood postoperatively. Infusion of this blood will result in infusion of S100beta into the blood and interfere in the interpretation of early systemic S100beta values.

Chronic effects of ethanol on muscarinic acetylcholine receptors are modulated by protein kinase C.

We have previously demonstrated that long-term ethanol treatment increased the number and function of muscarinic acetylcholine receptors (mAChRs) in human neuroblastoma cells, but the molecular mechanisms involved in these changes are unknown. In the present study, the effect of protein kinase C (PKC) on these events was investigated in human neuroblastoma SH-SY5Y cells. Following exposure to 100 mM ethanol for 2 days, both [³H]N-methylscopolamine binding and carbachol-stimulated I(1,4,5)P3formation were increased. When cells were cultured in the presence of 12-O-tetradecanoylphorbol 13-acetate (TPA), a potent activator of PKC, the effects of ethanol on mAChR number were totally inhibited but ethanol still potentiated carbacholstimulated I(1,4,5)P3 formation in TPA treated cells. TPA dose-dependently inhibited carbochol-stimulated I(1,4,5)P3 formation and this effect appeared to be independent of PKC phosphorylating activity. On the other hand, PKC inhibitors mimicked ethanol effects on mAChR number and function. Selective inhibition of classical PKC isozymes with 1 µΜ Gö 6976 for 2 days caused an increase in mAChR number and function, suggesting a role for these isozymes in ethanol-induced upregulation of mAChRs. These data indicate that longterm ethanol treatment may upregulate the number of mAChRs by counteracting PKC-mediated phosphorylation. The effects of ethanol on receptor-coupled phosphoinositide hydrolysis appear to be independent of PKC activity.

Phosphatidylethanol in blood as a marker of ethanol consumption in healthy volunteers: comparison with other markers.

Phosphatidylethanol is a "pathological" phospholipid, formed via the action of phospholipase D only in the presence of ethanol. The present study was made to elucidate how different levels and patterns of alcohol intake affect blood levels of phosphatidylethanol in comparison with other markers of abuse. We used a new HPLC-evaporative light-scattering detection technique for phosphatidylethanol quantitation. This method had a total coefficient of variation of <20% at the detection limit of 0.2 nmol, equaling 0.8 micromol/liter of whole blood. Two groups were studied. (a) Five healthy volunteers were given 32 to 47 g of ethanol in a single dose, to give blood ethanol levels of approximately 25 mmol/liter after 30 to 60 min. Phosphatidylethanol, carbohydrate-deficient transferrin (CDT), and blood ethanol were measured before and after the intake. (b) Twelve student volunteers were studied during a 3 week period of prolonged alcohol consumption (total estimated intake: 1334 +/- 488 g, mean +/- SD) and phosphatidylethanol, serum-CDT, gamma-glutamyltransferase, and blood ethanol were measured at the start of the period (day 1) and twice at the end of the period (days 18 and 21). In group (a), no phosphatidylethanol was detected at any time after ethanol dosage/intake. In group (b), no blood phosphatidylethanol or blood ethanol could be demonstrated at the start, and serum-CDT was below the discrimination limit (1.3%) in all persons. No phosphatidylethanol was detected in those four persons with the lowest intake (742 +/- 150 g). However, the remaining eight persons had detectable levels of phosphatidylethanol (1.0 to 2.1 micromol/liter), and these had a higher total intake (1630 +/- 389 g). There was a statistically significant (p = 0.02) increase in serum CDT for 3 weeks. However, only 3 of 12 persons increased above the discrimination limit. The present results indicate that a substantial alcohol intake is needed to elevate blood phosphatidylethanol. In comparison with serum-CDT, blood phosphatidylethanol appears more sensitive.

Significance of serum S100 release after coronary artery bypass grafting.

BACKGROUND: S100 protein has been suggested to be a serum marker for cerebral complications after cardiac operation and extracorporeal circulation. The aim of this study was to characterize the S100 release pattern after extracorporeal circulation in 515 consecutive patients undergoing coronary artery bypass grafting. METHODS: Clinical variables and outcome were prospectively registered. The cerebral outcome was determined by clinical examination. S100 was measured at the end of extracorporeal circulation, and after 5, 15, and 48 hours. RESULTS: After operation, 13 patients had stroke, 12 had delayed awakening, and 17 had encephalopathy. Early S100 release, immediately after extracorporeal circulation, was associated with age and perfusion time, but not with cerebral outcome. However, S100 release after 5 to 48 hours was associated with cerebral complications and risk factors for such outcome. Patients with stroke had higher S100 levels after 15 to 48 hours. A subset of patients with renal failure had overall higher S100 levels at 5 hours. CONCLUSIONS: Early and late S100 release indicate different mechanisms for release and emphasizes the potential power of this new biochemical marker for cerebral damage.