Why does rem sleep occur? A wake-up hypothesis.

Brain activity differs in the various sleep stages and in conscious wakefulness. Awakening from sleep requires restoration of the complex nerve impulse patterns in neuronal network assemblies necessary to re-create and sustain conscious wakefulness. Herein I propose that the brain uses rapid eye movement (REM) to help wake itself up after it has had a sufficient amount of sleep. Evidence suggesting this hypothesis includes the facts that, (1) when first going to sleep, the brain plunges into Stage N3 (formerly called Stage IV), a deep abyss of sleep, and, as the night progresses, the sleep is punctuated by episodes of REM that become longer and more frequent toward morning, (2) conscious-like dreams are a reliable component of the REM state in which the dreamer is an active mental observer or agent in the dream, (3) the last awakening during a night's sleep usually occurs in a REM episode during or at the end of a dream, (4) both REM and awake consciousness seem to arise out of a similar brainstem ascending arousal system (5) N3 is a functionally perturbed state that eventually must be corrected so that embodied brain can direct adaptive behavior, and (6) cortico-fugal projections to brainstem arousal areas provide a way to trigger increased cortical activity in REM to progressively raise the sleeping brain to the threshold required for wakefulness. This paper shows how the hypothesis conforms to common experience and has substantial predictive and explanatory power regarding the phenomenology of sleep in terms of ontogeny, aging, phylogeny, abnormal/disease states, cognition, and behavioral physiology. That broad range of consistency is not matched by competing theories, which are summarized herein. Specific ways to test this wake-up hypothesis are suggested. Such research could lead to a better understanding of awake consciousness.

Free will debates: Simple experiments are not so simple.

The notion that free will is an illusion has achieved such wide acceptance among philosophers and neuroscientists that it seems to be acquiring the status of dogma. Nonetheless, research in this area continues, and this review offers a new analysis of the design limitations and data interpretations of free-will experiments. This review presents 12 categories of questionable conclusions that some scholars use to promote the idea that free will is an illusion. The next generation of less ambiguous experiments is proposed.

Radioanalytical data interpretation when the ratio reading/median is lognormally distributed.

The purpose of this paper is to assist those who might be confronted by non-normal and non-homoscedastic error distributions representable by continuous probability density functions. Methods are presented to demonstrate how mathematical algorithms can be developed to obtain a "best fit" calibration line and how uncertainty ranges in interpretations of unknowns can be obtained from the calibration. The data used to demonstrate these methods were obtained from Brookhaven National Laboratory fission track analysis data for plutonium in urine. Examination of the variability in the fission track analysis data, during the period of time that the demonstration data were collected, revealed that the deviations from the mean were neither normal nor lognormal, but the ratios of tracks divided by the median at each plutonium level were lognormally distributed. Consequently, the differences between the logarithms of observed tracks and the median were normally distributed. The new "best fit" line was obtained by minimizing a reduced chi-square statistic made up of the squared differences in logarithms, divided by the variance in logarithms and degrees of freedom. Thus, to detect a worker urine sample to be above the 58-person "control" population 95 percentile [about 3.2 microBq (85 aCi)] at the 95% probability level (0.05 Type H error) would now require an average of about 11 microBq (300 aCi) per sample, compared to 5 microBq per sample (132 aCi per sample) in a previous paper. This paper presents the algorithms used to obtain the new calibration line and the uncertainty distributions of interpretations at various analyte levels. The importance of maintaining process control over the statistical interpretation of bioassay data as well as for the radiochemical procedures for achieving the lowest feasible level of detection is demonstrated.

Behavioral arrest: in search of the neural control system.

Scientists have spent hundreds of years trying to understand how the brain controls movement. Why has there been so little interest in knowing how the brain STOPS movement? This review calls attention to behavioral phenomena in which an animal or human undergoes temporary total-body arrest of movement, that is, behavioral arrest (BA). These states can be actively induced by visual stimuli, by body and limb manipulations, and by drugs. Historically, these states have been considered as unrelated, and their literature does not cross-connect. What is known about the causal mechanisms is scant, limited mostly to implication of the brainstem in manipulation-induced BA and dopaminergic blockade in the striatum in the case of drug-induced BA. The possibility has not been experimentally tested that all of these states share with each other not only an active global immobility in which awkward postures are maintained, but also underlying neural mechanisms. This review identifies key brainstem, diencephalic, and basal forebrain areas that seem to be involved in causing BA. We review the evidence that suggest a possible role in BA for the following brain structures: entopeduncular nucleus, medullary and pontine reticular zones, parabrachial region, pedunculopontine nucleus and nearby areas, substantia nigra, subthalamic nucleus, ventromedial thalamic nucleus, and zona incerta. Such areas may operate as a BA control system. Confirmation of which brain areas operate collectively in BA would require testing of several kinds of BA in the same animals with the same kinds of experimental tests. Areas and mechanisms might be elucidated through a strategic combination of the following research approaches: imaging (fMRI, c-fos), lesions (of areas, of afferent and efferent pathways), chemical microstimulation, and electrical recording (of multiple units and field potentials, with an emphasis on testing coherence among areas). We suggest the working hypothesis that BA is created and sustained by coherent, perhaps oscillatory, activity among a group of basal forebrain and brainstem areas that collectively disrupt the normal spinal and supraspinal sequencing controls of reciprocal actions on the extensors and flexors that otherwise produce movement.

A simple method of target preparation for the bulk analysis of powder samples by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS).

A simple and rapid procedure using a glue technique has been developed for the preparation of stable targets from powder samples for bulk analysis by LA-ICP-MS. The procedure was evaluated for the analysis of trace elements in SiC, of rare-earth elements in different types of silicate (rocks, sediments, and soils), and of Au and platinum-group elements in geological silicates. The test analysis was conducted using an IR laser in combination with a quadrupole mass spectrometer. The recommended preparation procedure offers the possibility of different types of calibration, for example application of certified reference samples in combination with prepared spiked samples on a base of a natural or synthetic matrix, or addition calibration. The resulting calibration functions are linear over a range of several decades. The trueness of the results was evaluated by use of certified reference samples. Analytical concentration ranges, detection limits, and the relative standard deviations are reported.

Coherent EEG indicators of cognitive binding during ambiguous figure tasks.

We tested the hypothesis that perception of an alternative image in ambiguous figures would be manifest as high-frequency (gamma) components that become synchronized over multiple scalp sites as a "cognitive binding" process occurs. For 171 combinations of data from 19 electrodes, obtained from 17 subjects and 10 replicate stimuli, we calculated the difference in correlation between the response to first seeing an ambiguous figure and when the alternative percept for that figure became consciously realized (cognitively bound). Numerous statistically significant correlation differences occurred in all frequency bands tested with ambiguous-figure stimulation, but not in two kinds of control data (a reaction-time test to sound stimuli and a no-task, mind-wandering test). Statistically significant correlation changes were widespread, involving frontal, parietal, central, and occipital regions of both hemispheres. Correlation changes were evident at each of five frequency bands, ranging up to 62.5 Hz. Most of the statistically significant correlation changes were not between adjacent sites but between sites relatively distant, both ipsilateral and contralateral. Typically, these correlation changes occurred in more than one frequency band. These results suggest that cognitive binding is a distinct mental state that is reliably induced by ambiguous-figure perception tasks. Coherent oscillations at multiple frequencies may reflect the mechanism by which such binding occurs. Moreover, different coherent frequencies may mediate different components of the total cognitive-binding process.

Biological water and its role in the effects of alcohol.

Alcohol and water compete with each other on target membrane molecules, specifically, lipids and proteins near the membrane surface. The basis for this competition is the hydrogen bonding capability of both compounds. But alcohol's amphiphilic properties give it the capability to be attracted simultaneously to both hydrophobic and hydrophilic targets. Thus, alcohol could bind certain targets preferentially and displace water, leading to conformational consequences. This article reviews the clustering and organized character of biological water, which modulates the conformation of membrane surface molecules, particularly receptor protein. Any alcohol-induced displacement of biological water on or inside of membrane proteins creates the opportunity for allosteric change in membrane receptors. This interaction may also prevail in organelles, such as the Golgi apparatus, which have relatively low concentrations of bulk water. Target molecules of particular interest in neuronal membrane are zwitteronic phospholipids, gangliosides, and membrane proteins, including glycoproteins. FTIR and NMR spectroscopic evidence from model membrane systems shows that alcohol has a nonstereospecific binding capability for membrane surface molecules and that such binding occurs at sites that are otherwise occupied by hydrogen-bonded water. The significance of these effects seems to lie in the need to learn more about biological water as an active participant in biochemical actions. Proposed herein is a new working hypothesis that the molecular targets of ethanol action most deserving of study are those where water is trapped and there is little bulk water. Proteins (enzymes and receptors) certainly differ in this regard, as do organelles.

Determination of acetaldehyde in blood by solid phase extraction and high performance liquid chromatography.

A simple method has been developed for the measurement of acetaldehyde in blood. Samples were treated with perchloric acid to precipitate proteins. After centrifugation, the supernatant, together with the internal standard (crotonaldehyde), were reacted with 2,4-dinitrophenylhydrazine reagent. The derivatized acetaldehyde was then isolated by solid phase extraction and followed by high performance liquid chromatography quantitation. This method had a minimal detectable concentration of 0.28 microM, and displayed an intra-assay precision of 2.23% and an inter-assay precision of 5.18%. The recoveries of acetaldehyde at added concentrations of 1.42 and 7.14 microM were 106.7% and 101.9%, respectively.

Variations of equine urinary volatile compounds during the oestrous cycle.

Equine urine was analysed by capillary gas chromatography. The volatile profiles from oestrous and dioestrous samples were compared to establish any qualitative or quantitative difference that may have potential value in olfactory communication. Forty-five different volatile compounds were detected. Of these, 17 major compounds were common to all chromatograms. The chemical profile of oestrous urine was distinguished by the presence of a unique peak that was not present in dioestrous samples. Numerous constituents exhibited endocrine dependence: while the concentrations of seix peaks increased at oestrus, the concentrations of another five peaks decreased at the same time. Since oestrous urine, but not dioestrous urine, has been shown to elicit sexual behaviour in the stallion, the unique peak, together with the peaks that were present in increased concentration at oestrus, may represent important chemical signals that stallions use to detect urinary 'oestrous odours'. Statistical analyses also indicated that the relative ratios (normalized peak areas) of many peaks changed significantly across the oestrous cycle: the rations increased in nine peaks, decreased in six peaks, and remained constant in two peaks at oestrus.

Volatile compounds of bovine milk as related to the stage of the estrous cycle.

Previous reports on behavioral assays with trained dogs suggested that milk samples from cows at diestrus, proestrus, and estrus had different odors. To identify the odor differences, volatile compounds in milk were isolated and analyzed by gas chromatography and mass spectrometry. About 80 peaks were detected in each chromatogram, of which 59 were present in all samples, and 23 were tentatively identified. The major identified compounds included the following six structurally distinct classes: ester, aldehyde, ketone, alcohol, fatty acid, and lactone. Although no unique peaks were found to be specific to samples taken at diestrus, proestrus, or estrus, 36 compounds exhibited significant differences in concentration among the three reproductive stages. These quantitative differences may account for the variation of milk odors during the estrous cycle. In order to investigate the quantitative differences systematically, multivariate discriminant techniques were used to relate the gas chromatographic profiles with the three stages of the estrous cycle. Stepwise discriminant analysis indicated that 15 of the 59 peaks in each chromatogram could best be used to reveal the differences among milk samples taken at diestrus, proestrus, and estrus stages. The discriminant function based on the 15 key peaks could classify all of the samples into their original categories at a total accuracy of 97.9%. Canonical analysis indicated that milk samples from different stages were clearly separated from each other in a two-dimensional space.

Amphiphilic binding site of ethanol in reversed lipid micelles.

Proton and phosphorous NMR spectroscopy were used to study a model membrane system consisting of reversed lipid micelles to test the hypothesis that alcohol and anesthetics compete with water for the same hydrogen bonding sites on lipid surfaces. When low concentrations of water and ethanol were added in equal parts in the absence of lipid and nonpolar solvent, the NMR spectrum consisted of a combination of all water and ethanol peaks, except for the ethanol OH peak. Compared with pure water, the bulk water peak became broader and shifted downfield to 5.1. In reversed micelles made of water, DPPC, and nonpolar solvent, the addition of ethanol caused a conspicuous upfield shift of the bulk water peak and also broadened and decreased its height. This effect was magnified as ethanol concentration increased, indicating that alcohol alters the organization of water and moves water protons into a new domain where nearby atoms are more able to shield water protons. Water shifted the P-31 resonant frequency of DPPC downfield, and the effect magnitude varied with water concentration. Ethanol did not cause such a shift, suggesting that only water was interacting in the phosphorous region. Two-dimensional nuclear Overhauser effect (NOESY) spectroscopy indicated that the ethanol methylene is adjacent to the methylene next to the carbonyl of the DPPC fatty acid moiety, at least in some configurations. Interaction at this point is also indicated by the transformation from an apparent pentet to a doublet of triplets at certain ethanol/water ratios.

Cyclic changes in volatile constituents of bovine vaginal secretions.

Headspace gas chromatography was used to examine the volatile components of bovine vaginal secretions in three consecutive estrous cycles from each of four cows. Results indicated that there was an interesting peak with a retention of 2.7 min. It showed a successive rise and fall zero to three days before estrus. Gas chromatographic evidence and mass spectral data confirmed that this peak was acetaldehyde. Another unidentified compound with a retention time of 27 min was found to be unique to proestrus. It occurred two to three days prior to estrus, but was absent at any other time of the cycle.

Acute ethanol decreases NMR relaxation times of water hydrogen protons in fish brain.

The traditional belief about ethanol's mechanism of action is based on ethanol's lipophilicity and capability to penetrate and disorder lipid bilayers. This traditional belief is now being supplanted by growing evidence that ethanol has relatively selective actions on certain synaptic receptors, such as those for NMDA, serotonin, and GABA. It was recently argued that these receptor specificities are secondary to a preferential ability of ethanol to displace membrane bound water in the domains of certain receptors. The data obtained in this study are consistent with the original hypothesis: any disorganization of cellular water by ethanol will be detectable by proton nuclear magnetic resonance (NMR) spectroscopy. In particular, the relaxation times of water hydrogen protons reflect how constrained water molecules are by the macromolecules within cells. The relaxation time of "bulk" water is lengthened relative to water molecules that are under the influence of electromagnetic fields of macromolecular surfaces within cells. Here, we tested this hypothesis in living fish, which dosed themselves by swimming in water that had added ethanol. Estimates of brain alcohol at 5 min after initial exposure revealed that the brain concentration was only about 1/3 that of the water in which they were swimming. The average value of the NMR relaxation time T1, but not T2, was decreased at 5 min (when brain concentrations were on the order 100 mM) and reached statistical significance at 10 and 30 min after initial exposure.(ABSTRACT TRUNCATED AT 250 WORDS)

A possible feline model for human blepharospasm.

'Benign essential blepharospasm' is a human eyelid disorder of unknown aetiology characterized by involuntary, bilateral, and disabling spasmodic contracture of the orbicularis oculi muscle. Treatments are frequently disappointing. Here we report what might be a first step toward developing an animal model for exploring mechanisms of the disorder and potential treatments. We surgically implanted stimulating electrodes into brain areas known to supply input to the lateral division of the facial nerve nucleus to induce blinking by electrical stimulation. Single-pulse stimuli at or near the facial, parabrachial, red, and interstitial nuclei produced consistent stimulus-induced eyelid contractions. Responses were ipsilateral to stimulation, except for the interstitial nucleus where contralateral responses occurred. Little or no other movements of the face, head, or body occurred at eye-blink threshold voltages. When these sites were stimulated with pulse trains, eyelid closures followed stimulus frequency and tended to fuse into constant closure. Thresholds at each stimulus site remained constant during three days of testing. Drug treatments produced no consistent effect on eye-blink threshold from any stimulation site, even when general behaviour was affected. We conclude that these input pathways to the facial nucleus may contribute to blepharospasm and that future neurochemical and electrophysiological study of these pathways may produce a suitable animal model for understanding this disorder.

Innervation of the feline eyelids.

The innervation of the eyelids is incompletely understood. This is a particular problem for those who wish to develop animal models of eyelid dysfunction in humans. Blepharospasm, for example, is a disease of uncontrolled eyelid spasm that is difficult to manage clinically because the aetiology is not understood. The anatomical literature on eyelid innervation is sparse and even conflicting. We attempted to study eyelid innervation, both sensory and motor, with injection of horseradish peroxidase (HRP) into the superior eyelid, inferior eyelid, and bulbar conjunctiva. We used 13 anesthetized weanling cats. Shape and structure of the facial nucleus varied along its rostrocaudal extent, but there was a clear demarcation of lateral and medial division. HRP-filled facial nucleus cells were ipsilateral to the injection site, and label appeared throughout the rostrocaudal length. All injection sites, including bulbar conjunctiva, labelled facial nucleus neurons located with overlapping distribution, predominantly in the dorsal part of the lateral division. Likewise, heavy labelling occurred throughout the entire ipsilateral cranial cervical ganglion and the trigeminal ganglion in all kittens. Injection of upper or lower eyelids caused some labelling in the second through the fourth cervical spinal ganglia.

FTIR evidence for alcohol binding and dehydration in phospholipid and ganglioside micelles.

We theorize that intoxicants and modern anesthetics bind at the membrane-water interface and displace (dehydrate) bound water molecules by breaking the hydrogen bonds. We tested this hypothesis by examining the effect of butanol on the binding of water to the polar regions of lipids in reversed micelles. Understanding the mechanisms of intoxication requires studies in physiologically relevant systems such as systems containing sialoglycoconjugates, especially gangliosides, which concentrate in the synapses of neural tissue. Therefore, we compared butanol effects on phospholipid with effects on ganglioside. Hydrogen-bond breaking activity of 1-butanol was studied in reversed micelles made of dipalmitoylphosphotidylcholine (DPPC), ganglioside (GM1 and GT1b) or the lipid mixture in a D2O-CCl4 medium. Fourier transform infrared spectroscopy (FTIR) data indicated that 1-butanol binds to DPPC and to gangliosides. Adding GM1 to the DPPC micelles introduces a new binding site for the alcohol. GT1b binds more butanol than GM1, because of more binding sites provided by extra sialic acid moieties. Spectral red shifts indicate that both water and butanol bind to the C = O group of sialic acid. Butanol partially releases the surface-bound water by disrupting hydrogen bonds, as indicated by an appearance of a sharp new free OD stretching band of the released D2O molecules. However, control studies with lipid-free systems in CCl4 revealed that a free OD peak could occur from a deuterium exchange reaction between D2O and 1-butanol(ol-h).(ABSTRACT TRUNCATED AT 250 WORDS)

Are there EEG correlates of mental states in animals?

The thesis of this paper emerges from the fact that mental states are generated by neural processes that also produce an associated electroencephalogram (EEG). Thus, it is logical to expect correlations between mental state and EEG. The corollary is that the EEG can serve as an index of mental state, which can be particularly useful for studies in animals, where mental states are much less accessible for objective study than in humans. Herein, I briefly review the traditional approaches that have informed our attitudes about animal mental states. Virtually all of our conclusions about mental states in animals are drawn by inference from behavioral observation, a process that is highly and unavoidably subject to anthropomorphism. Traditionally, the electroencephalogram (EEG) has been used in a crude way as an objective indication of physical and behavioral state in animals. This, however, has led to substantial controversy, because there are several situations in which EEG patterns and behavior seem to be dissociated. We not only fail to understand these dissociated states, but there are also important humane animal-welfare issues that remain unresolved because we do not fully understand the extent to which the EEG can reflect mental state. At issue is whether EEG-behavioral dissociations, to the extent that they exist, are proof that the EEG is dissociated from mental states. Powerful new EEG methods, such as topographical EEG mapping, wavelet analysis, and testing for nonlinear ('chaotic') dynamical properties and short-term serial dependencies, are now available for studying the extent to which the EEG can index thinking and feeling in humans and, by extrapolation, in animals. Critics who have become disenchanted with the utility of the EEG should at least concede that fresh approaches to old problems are now available and should therefore be thoughtfully considered. If such research does nothing more than improve the rigor of the debate over animal welfare and rights issues, it will be worth the effort.

Ethanol effects on total sialic acid of various brain regions and visceral organs.

These experiments were designed to extend our earlier observations on acute ethanol effects on brain sialic acid (SA), an acidic sugar component of membrane gangliosides and many glycoproteins. Here, we tested for differential effects of ethanol on total sialic acid in various brain regions and for effects on other organs, such as liver, kidney, heart, and skeletal muscle. Subjects were young adult, male rats. The first experiment compared two commonly accepted analytical methods for brain SA on peripheral tissues. Consistently higher levels were evident with the resorcinol method in all tissues, especially in liver, compared with the thiobarbiturate method. Resorcinol-measured levels in the liver and kidney were on the order of 350 micrograms/g, wet weight. In the brain of controls, the resorcinol method revealed total SA levels to be on the order of 550-650 micrograms/g in the basal ganglia, cerebral cortex, cerebellum, and hippocampus. Brainstem levels were significantly lower. Ethanol, given IP at 2 g/kg, seemed to decrease total SA in all brain regions at four hours after injection, with statistically significant decreases in the hippocampus and brainstem. With 3 g/kg, only the cerebellum showed a significant decrease at four hours, compared to saline-injected controls, but the decrease was large (25%). Analysis of the other organs showed that, compared to saline-injected controls, ethanol decreased SA in the liver. There was a small, but significant, decrease in heart SA at four hours after 3 g/kg. In skeletal muscle, ethanol significantly increased total SA at 2 g/kg, but not at 3 g/kg.(ABSTRACT TRUNCATED AT 250 WORDS)